The Peter Attia Drive - #33 - Rudy Leibel, M.D.： Finding the obesity gene and discovering leptin

00:00:00.000 Hey everyone, welcome to the Peter Atiyah Drive. I'm your host, Peter Atiyah.

00:00:10.140 The Drive is a result of my hunger for optimizing performance, health, longevity, critical thinking,

00:00:15.600 along with a few other obsessions along the way. I've spent the last several years working with

00:00:19.840 some of the most successful, top-performing individuals in the world, and this podcast

00:00:23.620 is my attempt to synthesize what I've learned along the way to help you live a higher quality,

00:00:28.360 more fulfilling life. If you enjoy this podcast, you can find more information on today's episode

00:00:33.000 and other topics at peteratiyahmd.com.

00:00:41.260 Hey, welcome to this episode of The Drive. On this episode, I interview an amazing scientist and a

00:00:47.480 very dear friend, Dr. Rudy Leibel. Rudy's a professor at Columbia University, where his work has focused

00:00:56.300 primarily on type 2 diabetes and obesity. I've known Rudy for quite a while, probably about six

00:01:03.020 years now, maybe a little longer. We worked together very closely back at the Nutrition Science Initiative,

00:01:09.240 where he was one of the very important collaborators in one of the more theoretical experiments that we

00:01:14.980 did. Rudy and I have always just had a ton of fun just hanging out over great meals and great wine and

00:01:20.880 great beer talking about science. And so I thought it might be fun to try to reproduce some of those

00:01:26.900 discussions here. A couple of things. First, we recorded this on a Friday afternoon on the Upper East

00:01:33.080 Side, and you'll probably notice a little more noise than usual just on account of the street noise. So

00:01:39.560 apologies for that. Second point is, we spend quite a bit of time, probably about the first hour and

00:01:46.000 10 hour and 15 minutes, just talking about the discovery of leptin, for which Rudy played arguably

00:01:54.000 the single most important role. Obviously, many people played a role in the discovery of leptin. But

00:01:58.880 you know, in many ways, Rudy was sort of the chief architect of that. And certainly inside of the

00:02:03.500 scientific community, he is largely regarded as that, though, unfortunately, history may write that

00:02:10.160 story a little bit differently. So for those of you listening to this who don't really,

00:02:14.320 really get off on geeky science and knowing the nuts and bolts of all of the experiments and how

00:02:20.320 one found this gene and the difference between a southern blot and a northern blot and a DNA sequence,

00:02:26.480 you might just want to skip to about an hour 15 when we get to the other stuff. But that said,

00:02:32.340 if you really are interested in science, I think there's a lot to be gained from listening to these

00:02:36.420 discussions because, you know, that's certainly something I plan to do a lot of as interview scientists.

00:02:41.260 And part of that is kind of understanding their stories. Again, a lot of what we talk about in

00:02:45.180 this post is technical. I'm hopeful that in the show notes, we'll be able to provide a lot of sort

00:02:49.940 of the necessary glossary to make it a little easier to ingest. I don't do a great job. I apologize

00:02:55.020 of trying to clarify terms. Occasionally, I remember that not everybody knows what hyperphagia means.

00:03:01.340 And I explained that that means eating too much. But again, I apologize for that. And just look to the

00:03:06.280 show notes if you want clarification on anything. We get into a lot of discussion around obesity,

00:03:11.440 the genetics of obesity and the regulation of obesity. How much of this is regulated centrally,

00:03:16.160 meaning in the brain, versus peripherally, everywhere outside of the brain. We talk a lot

00:03:20.500 about energy expenditure. And for those of you who are really interested in this science,

00:03:24.460 Rudy gives a great explanation of the two techniques that are used to measure energy expenditure,

00:03:28.420 known as indirect calorimetry and doubly labeled water. We talk about a number of genes that are

00:03:33.840 known to predispose to obesity, some very acutely and some very crudely, like the FTO gene. And we,

00:03:40.540 of course, talk a lot about insulin resistance, which in many ways is sort of a bit of a paradox,

00:03:45.040 because on the one hand, if you're insulin resistant, you technically shouldn't be able

00:03:48.460 to get fat. But of course, many insulin resistant people are able to get fat. They do accumulate

00:03:53.440 adipose tissue. What else can I say about this? I guess those are kind of the most important things

00:03:57.820 I'd say to guide you. So in summary, if you really don't want to hear about how leptin was

00:04:01.800 discovered, skip to an hour 15. If you do sit back and relax. The other highlight of this was,

00:04:07.900 it was the first time I ever gave Rudy a Topo Chico. He loved it. And I think I've got another

00:04:13.320 convert. So anyway, hope you enjoy this. And without further ado, here is my lovely discussion

00:04:20.140 with the amazing Dr. Rudy Lively.

00:04:25.620 All right, Rudy, how are you?

00:04:27.500 I'm very well, thank you.

00:04:28.680 Thanks so much for trekking over to the east side.

00:04:31.820 Well, thanks for inviting me.

00:04:35.460 This will be one of the first times in a while that we hang out without a drink,

00:04:39.880 including my favorite beer, which you're one of the few people I've shared it with.

00:04:44.720 Exactly.

00:04:46.140 I don't tell anybody what that beer is because I, and I just found 19 bottles of it in Florida

00:04:52.360 last week. So I'm pretty psyched about that. I'll continue to share with you, of course.

00:04:58.000 It's still in Florida.

00:04:59.380 No, it's actually in San Diego right now. Yeah, it just arrived. There's so much I want to talk

00:05:04.100 about. I almost don't even know where to begin. I'm guessing that a number of the people listening

00:05:08.140 to this won't actually know who you are necessarily, but I think by the end of this,

00:05:12.320 they'll be super interested in learning more about you. During the introduction, I will have

00:05:17.900 explained sort of a lot of stuff about you, but tell me a little bit about what you do and,

00:05:22.660 more importantly, why you do it.

00:05:24.580 I've been very interested since the past 30 years or so in biology of the regulation of body weight

00:05:34.180 in both animals and humans. But the studies of animals are designed primarily to shed light on

00:05:43.760 what's the basis for the control of body weight in humans. I'm a physician and have been interested

00:05:49.840 in the clinical problem of obesity for even longer than I've been doing research on it.

00:05:57.420 And I have taken, over the years, various approaches, beginning with studies of human

00:06:05.700 adipose tissue aspirated using various needles from various subcutaneous depots in humans, meaning

00:06:13.960 around the rear end and in the front of the abdomen. And then became very interested in the

00:06:21.780 genetics of obesity and have done some studies trying to identify various genes that are related

00:06:28.680 to that and continue to do so. And have also done a large number of studies in mice designed to

00:06:35.560 look at this problem. And very recently, or at least within the past five or 10 years, have begun to focus

00:06:44.360 on the use of stem cells to try to understand, again, the biology of brain cells that regulate body weight

00:06:52.160 and the cells in the pancreas that produce insulin. Because obesity and diabetes go hand-in-hand

00:07:00.480 clinically. And this is not an accident. They're very tightly related in a number of really very

00:07:07.560 interesting ways. So that my studies of obesity have led more or less inexorably towards the study

00:07:16.560 of diabetes as well. So now my laboratory really does some of both. And you're at Columbia now. You

00:07:23.760 were at Rockefeller before that. But originally, you were in Boston, correct? That's correct. My clinical

00:07:29.300 training. I'm trained as a pediatrician and an endocrinologist. And that training took place

00:07:34.420 in Boston at both the Massachusetts General Hospital and the Children's Hospital. And then for

00:07:41.260 five or six years, I worked at both the Mass General and a community hospital in the Boston area,

00:07:48.600 the Cambridge City Hospital. Again, doing both general pediatrics and some endocrinology.

00:07:56.040 So what got you interested in obesity?

00:07:57.600 I was interested in obesity, or at least in the part of the brain that regulates body weight,

00:08:04.160 even as a medical student. We didn't study it specifically. It wasn't emphasized the role of

00:08:10.300 the hypothalamus, which turns out to be very important in that regard. But I had the opportunity

00:08:15.280 as a medical student, actually a first-year medical student, to work at the Walter Reed Army

00:08:21.880 Institute of Research with a neuroscientist named Harvey Carton, who was a physician who was interested

00:08:30.380 actually in the study of bird brains. And he took me on for a summer, actually it ended up being two

00:08:40.480 summers, to work in his laboratory, primarily helping with histological studies of the bird brain.

00:08:47.540 But the laboratory that he was in was run by a man named Wally J. H. Nauda, who was recognized at the

00:08:57.220 time, and subsequently perhaps even more so, as one of the great neuroanatomists with regard to the

00:09:03.880 hypothalamus. So actually I was indoctrinated a bit just by hanging around the other people who were

00:09:10.780 working there, including Dr. Carton, in terms of the importance of the hypothalamus in a number of

00:09:16.260 areas of physiology.

00:09:18.780 Where does the hypothalamus sit in relation to the pituitary, or even more grossly, like other main

00:09:24.380 structures in the brain?

00:09:25.260 Yeah, I think you can sort of picture it as lying between, or being triangulated in a sense, or at the

00:09:32.280 crosshairs of a line drawn between the eyes and the side of the forehead, and it's situated just

00:09:39.900 above the pituitary gland, if you know where that is.

00:09:43.500 Yeah, which is sort of behind the optic chiasm.

00:09:46.140 Yes, that's correct. And it's a very small organ, or at least it's a very small part of the brain,

00:09:53.640 about the tip of the small finger, but does play a role in the regulation of many important

00:10:00.640 physiological functions, including things like blood pressure, body temperature, and from my point of

00:10:09.200 view, very importantly, body weight, and also increasingly apparent that it plays an important

00:10:14.860 role in blood sugar control as well. So it's been known for many, many years as being a critical

00:10:21.700 part of the brain, a so-called vegetative brain, not under conscious control.

00:10:27.720 Work that's gone on over the past 30, 40, 50 years has increasingly raised the level of sophistication

00:10:37.260 with which we understand the function of that part of the brain.

00:10:42.240 About 21 years ago, or maybe 20 years ago, whenever it was, but early in medical school,

00:10:47.060 I remember when we did neuroanatomy, the professor said, if you have to sacrifice any part of your brain,

00:10:53.500 and you're sort of prioritizing, the last thing you want to sacrifice is the hypothalamus. If you're only

00:11:00.160 allowed to keep a couple square centimeters of brain, keep the hypothalamus.

00:11:05.220 You know, I mean, we could get into the weeds on exactly...

00:11:09.060 That's a poor man's problem when you're...

00:11:11.160 That's right. I think that would be important. The other thing you'd want to hang on to is the part of

00:11:15.900 the brain that regulates respiration. Because even with the hypothalamus, if you're not breathing,

00:11:22.060 you're not going to get very far. But I agree.

00:11:24.840 I think he was excluding the brain stem for this.

00:11:27.100 I understand. I understand. I would... The hypothalamus is right up there.

00:11:30.280 Yeah. Yeah. At some point, I want to come back to it. I want to have you tell us all about Zucker

00:11:35.020 rats, because that's sort of what got me interested in this idea of how can this part of the brain,

00:11:40.980 when subjected to so many different types of insults and lesions, produce so many,

00:11:46.380 I mean, seemingly wild and disparate phenotypes that seem so out of whack. But before we go there,

00:11:52.800 let's go back to... You're a junior pediatrician schlepping along, taking care of overweight kids,

00:11:59.040 and we're in the, what, the late 70s? What time of year is this? What time of your life is this?

00:12:04.400 Yeah, we're in the mid-70s.

00:12:05.880 Mid-70s. So there's not that many obese kids, are there?

00:12:09.840 There are plenty.

00:12:10.980 Still in back then, yeah?

00:12:12.240 Oh, yeah.

00:12:12.700 Okay. So what are you doing for them? How are you helping them?

00:12:15.840 So back in that day, the conventional view was that obesity was largely due to imbalances of

00:12:24.100 hormones, at least potentially, in addition to whatever other sort of behavioral issues might

00:12:29.360 be implicated. But almost all children with obesity were referred, if it was significant and severe

00:12:38.780 enough to either an endocrinologist or a psychiatrist. That was sort of the bifurcation point for referral

00:12:45.920 of these children. I don't think either the endocrinologist or the psychiatrist really could

00:12:51.320 do very much for these children. I certainly couldn't. And it was an evening in the fall in the city of

00:12:59.160 Cambridge in a small office that I used for the very small number of referrals that I actually saw.

00:13:06.220 Most of my time was spent with the medical students trying to teach them various aspects of pediatrics.

00:13:12.320 But one evening, I saw a young boy about seven or eight years old with his mother and examined him,

00:13:19.960 weighed him, and so forth, and determined. It's quite clear that he was very obese. He had no

00:13:24.760 other stigmata of the sorts of things that we looked for, meaning very severe problems with thyroid

00:13:31.880 or adrenal gland, which can sometimes produce severe obesity. And he didn't have any of the stigmata of the

00:13:38.080 single genetic type of disorders that were known about at the time, like Prader-Willi syndrome or

00:13:44.020 Part A. Beatle. And I said to the mother, your son has severe obesity. I can't tell you what the

00:13:52.560 etiology of it is. It's clear that there's something going on. There's problems that we don't really

00:13:59.340 fully understand or understand very well at all. And my only advice is to try to restrict the number of

00:14:07.140 calories that he eats and increase his physical activity, which is, I think, unobjectionable

00:14:13.660 advice in any circumstance like that. And the mother turned to the boy and said,

00:14:22.200 Randall, let's get out of here. This doctor doesn't know shit.

00:14:29.080 I love that you remember his name.

00:14:31.080 Oh, yes, I do remember his name. And she took him by the hand and they left. And I remember

00:14:36.880 sitting behind the desk and thinking to myself, Randall's mother, you've got a point.

00:14:44.240 And it was that experience, actually, I'd been interested in some of the physiological aspects

00:14:49.980 of obesity and adipose tissue that led me to decide that if I was going to do anything about

00:14:58.720 this, I was going to have to train myself to do the kind of research that might be helpful in terms

00:15:04.720 of understanding the disease. And it was shortly after that, that I actually moved to New York,

00:15:10.660 to the Rockefeller, to begin to try to do those kinds of experiments.

00:15:14.580 Yeah, that story, which obviously we've discussed before, and I get such a kick out of it. But at

00:15:20.660 the same time, it's actually a remarkable example of something that medicine is a privilege. It gives

00:15:25.840 you a privilege and it's up to you to do anything with it, right? Which is patients generally provide

00:15:31.020 the appropriate physician with the right level of humility. And I think, look, it's a testament to you

00:15:37.000 that you sort of could reflect on what Randall's mom said, because I think there are other people

00:15:41.560 who couldn't, right? There are other people who could say, well, screw you. That's the advice I've

00:15:45.160 got. And that's like the best I can do. And, you know, wouldn't skip a beat kind of moving on. But

00:15:50.320 when I talk to a lot of great physician scientists, that seems to be the one thing that they all have

00:15:54.980 in common. They may have many things in common, but this one is important, which is it's the ability

00:15:59.540 to sort of pause in your tracks and say, wait a minute, there is a clinical situation and I cannot

00:16:04.940 for the life of me explain it. And I have to know the answer. So it's so amazing that one

00:16:11.160 chance encounter like that basically altered the entire course of your career and put you on this

00:16:17.140 path to do some really amazing stuff as we'll, I think, discuss in the next hour or so.

00:16:23.580 I think you're probably right that one of the great, I think, privileges of a medical education

00:16:30.500 is that it does give you a very broad perspective on the human condition, if you want to call it that,

00:16:36.340 all the way from psychological issues to very fundamental biological ones. And physicians

00:16:43.460 as a group do have this, as you pointed out, rather unique opportunity to look at a problem

00:16:51.700 that afflicts human beings, sometimes at the level of basic metabolism and endocrinology,

00:16:59.320 other times on a more systemic or even social level. And I'm sure we could name some of the

00:17:06.720 names of people who have operated all these levels. But I think you're right. I've always felt that it

00:17:13.580 was really somehow an enormous opportunity and a gift to be able to pursue this down to the level

00:17:22.340 that I've been able to do in the past 30 years or so. And I must say, in addition to whatever my

00:17:29.780 previous training and experience did to make that possible, I also have to acknowledge that my family

00:17:38.020 was very supportive in terms of allowing me to basically back out of a more classical sort of

00:17:46.140 academic medical track and retrain myself to the level that I thought was required in order to do

00:17:53.840 this research or basically move from a very nice home in Brookline, Massachusetts to an 800 square foot

00:18:01.340 apartment in Manhattan with a wife and two small children and lived there for 15 or 16 years while I

00:18:11.340 sort of got my act together. There's plenty of great research that can happen at Harvard. Why did you

00:18:16.620 come to New York? I came to New York actually after looking around in the Boston area for someone who

00:18:22.940 could mentor me in this kind of activity. And it turned out there were, I mean, there's some interesting

00:18:28.780 stories about that. But the advice that I got from the people who were not in a position to help me in the

00:18:34.840 way that I thought I needed help was that I might consider going to the Rockefeller and working with a

00:18:40.420 physician scientist named Jules Hirsch, who at the time was very interested in the biology of adipose

00:18:47.160 tissue from a number of perspectives. And I had actually communicated with Dr. Hirsch in the literature

00:18:55.080 by commenting on some of the work that he had done with regard to adipocytes and signals that might come

00:19:02.840 from adipocytes that affect potentially food intake and body weight. This was an area that he was quite

00:19:08.840 interested in. And I was too, based on some of the thinking I had done about it while I was in the

00:19:15.080 training in Boston. And he and I actually had had a communication in the literature on some of the

00:19:21.360 work that he had done. And I went actually to visit his laboratory while I was in New York for an entirely

00:19:27.520 different reason. And it was very shortly after that that I moved. Which is another great example of

00:19:33.460 the types of common threads we see in great scientists, whether they be scientists or physician

00:19:38.880 scientists, which is mentorship, right? Again, every person I talk to who's done something remarkable in

00:19:44.140 science can point to mentors, a mentor or mentors. And obviously for you, Jules was probably the most

00:19:51.900 important mentor, I'm guessing. That's right. I had very important other mentors in the area of

00:19:58.060 clinical medicine. And the one who trained me in endocrinology, an individual, again, a physician

00:20:04.720 scientist, more clinically oriented named Jack Crawford, himself was very interested in the role

00:20:11.520 of body fat and its interaction with other physiological systems in the body, most notably the

00:20:19.400 onset of puberty. And it was he that actually put me in contact with another scientist at Harvard named

00:20:26.140 Rose Frisch, who also was very interested in why, for example, young women who lost weight as a result of

00:20:34.300 anorexia, or more commonly in her experience, due to very vigorous physical training for things like

00:20:42.040 distance running or bicycling, swimming, became amenorrheic. In other words, their period stopped. And Rose was

00:20:49.520 very curious about why this was and whether there was a communication between fat and parts of the

00:20:56.120 brain that regulate the gonadal axis, which is the hypothalamus again. And I remember having many

00:21:03.640 conversations with Rose about where the signal might be coming from. She thought it actually might be

00:21:09.360 coming from adipose tissue or adipocytes in the bone marrow. And it was always encouraging me to

00:21:14.940 study bone marrow as a source of whatever this signal might be.

00:21:20.480 So what year did you arrive at Rockefeller?

00:21:22.900 1978.

00:21:23.460 At that point in time, was the adipose tissue, was adipose cell, was this regarded as an inert

00:21:33.240 sort of storage depot for fatty acid, or was it considered an endocrine organ? Like what was the

00:21:39.480 state of thinking about fat?

00:21:42.420 At that time, the view of the role of adipose tissue was very strongly in the direction of the

00:21:51.960 sort of former characterization that you made, which is that it was a passive depot for fat

00:21:58.200 in the form of storing free fatty acids as triglycerides so that they were hydrophobic,

00:22:05.980 so that you could pack a lot of calories into an organ or into a cell without having a lot of water

00:22:11.940 there. So it's a very efficient way of storing energy. Actually, as you know, up to probably around

00:22:18.740 nine calories per gram. And its role as a signaling device or an endocrine organ was really people

00:22:29.340 were thinking about it, but there was no firm evidence in this regard, other than I think the

00:22:36.740 increasing sense that the size of the adipose mass was doing something to the levels of circulating

00:22:44.620 insulin, that somehow insulin rises when the fat mass rises and the mechanism or the relationship

00:22:52.280 of these two...

00:22:52.560 Not the other way around? Was it clear which direction?

00:22:55.700 So at the time, I think the view was pretty much that as fat mass increases, the concentration of

00:23:02.000 insulin rises to meet the consequences of so-called insulin resistance, which occurred not only in

00:23:10.280 adipose tissue, but maybe more importantly in liver and muscle, and so that there was some relationship

00:23:18.500 between fat mass and this endocrine. But fat mass is a secretory organ itself of molecules that might

00:23:27.620 have some of these effects. It was really a pretty new idea that nobody really had any direct,

00:23:34.980 at least at the time, evidence for.

00:23:37.140 Obviously, we're going to get to the story of leptin, which I think is just another great example

00:23:41.780 of incredible persistence, an incredible focus and drive. But obviously, before getting to the

00:23:49.200 punchline, which occurred several decades later, what was your first insight into a hormone that

00:23:57.140 would go on to be leptin? Where did you first figure this, that there was something that was being

00:24:01.600 secreted by fat cells? I think the first evidence in this regard, or at least some of the most important

00:24:09.700 evidence, came from experiments that were done by an investigator at the Jackson Laboratories in Maine

00:24:17.460 named Douglas Coleman. And Coleman was a biochemist, physiologist, who was interested in some of the rare

00:24:29.960 mouse mutations that led to very severe obesity. And he was one of the first to study

00:24:38.760 the OB-OB or obese mouse, which was a spontaneous mutation that arose in the Jackson colony.

00:24:48.360 And he also was one of the close students of another mutation that arose there a number of

00:24:54.840 years later in the so-called diabetes or DB gene.

00:24:59.260 So what was the phenotype of this OB-OB mouse?

00:25:02.560 The OB-OB mouse was...

00:25:05.640 That's not to be confused with the OB-1 Kenobi mouse. That's a very different phenotype, right?

00:25:09.660 That's correct.

00:25:10.120 Yeah.

00:25:10.440 The OB-OB mouse was noted to be very severely obese very early in life. So these animals were

00:25:21.900 clearly obese by the time shortly after weaning, which is three or four weeks of age. You could

00:25:30.560 tell that they were obese. And if you left them alone and let them eat as much as they wanted,

00:25:35.600 they would eat up to the point of becoming pretty heroically obese, meaning these animals could reach

00:25:43.320 weights of 70 or 80 grams. Whereas a normal so-called wild type mouse at maximum weight would

00:25:51.780 be somewhere around 40 or 50 grams much later in life. An OB mouse could get up to 60 or 70 grams

00:25:59.920 very early in life. The animals were infertile.

00:26:03.400 So how did you, how did they keep breeding them?

00:26:05.380 So the way they would breed these animals is breed the parents, the heterozygous animals that were able

00:26:12.820 to generate.

00:26:13.920 So get two heteros to make homos.

00:26:15.720 Correct.

00:26:16.480 So the phenotype centered around hyperphagia?

00:26:18.940 It's centered around two phenotypes actually that Dr. Coleman was able to show. Primarily the

00:26:25.600 obesity was due to hyperphagia, but he also was able to demonstrate by a mechanism.

00:26:30.600 Sorry, I should just clarify for listeners. That means excessive appetite.

00:26:34.380 Yes, by excess food intake. That was the primary cause of their obesity. But he also,

00:26:41.140 by using a technique called pair feeding, where he would only feed the animal the amount that a

00:26:46.000 normal weight animal would eat, that those animals tended to store more of their excess calories as

00:26:52.260 fat. We sometimes refer to this as partitioning. And by another set of experimental maneuvers was

00:26:58.860 able to show that their metabolic rate was slower. So these animals had what you might call a trifecta

00:27:05.860 for obesity. They ate more, spent less, and whatever they stored was preferentially stored as fat.

00:27:14.180 And these were fascinating animals. I mean, people tried for many years to figure out what was wrong

00:27:20.140 with them. And they had many metabolic consequences of their obesity, but nothing about their physiology

00:27:27.020 necessarily pointed to what the primary mechanism was, where the genetics clearly indicated that this

00:27:35.840 was a single gene, or very likely to be a single.

00:27:39.300 And this, of course, is long before you've got your PCR technology that can very easily help you

00:27:45.380 sequence this. So two questions, though. Lifespan was what?

00:27:49.560 Lifespan, not severely affected, actually. They could live to a fairly normal lifespan,

00:27:55.300 somewhat short. Two and a half, three years?

00:27:58.060 Not quite. They don't last quite that long, but certainly up to 18 months or two years.

00:28:03.620 Important to note, because of the question of whether fat makes something or not, Coleman

00:28:08.440 not only observed these OB mice, but also this second mutation that arose later at the Jackson Lab and was

00:28:17.180 named the diabetes mouse. That's because that animal looked a lot like an OB mouse, but got diabetes

00:28:24.240 very early in life, unlike the OB mouse, which seemed to be obese, but not necessarily particularly

00:28:32.880 diabetes prone. And it was clear that the mutation in the gene, again, another single gene that caused

00:28:39.800 the diabetes mouse, was not the OB gene. It was on a different chromosome. Coleman was able to show

00:28:46.680 that. So here you had animals that looked a great deal like each other, at least in terms of the

00:28:53.600 overeating and the low energy expenditure. One was prone to get diabetes and the other not. And what

00:29:00.180 Coleman ultimately did was to join the circulation of the OB mouse to a wild-type animal and the DB or

00:29:11.280 diabetes mouse to a wild-type animal. So like a live parabiosis. It's a lot. That's the technical

00:29:17.220 term for it is a parabiosis. And he showed that if you hooked an OB mouse up to a wild-type mouse in

00:29:24.080 this way, that the OB mouse would correct its hyperphagia, its excess food intake, and actually

00:29:32.240 begin to lose weight. So something presumably in the wild-type mouse, some humoral factor, some hormone,

00:29:38.920 some something, must have been missing in the OB-OB mouse that, when given to it, corrected its

00:29:45.660 phenotype. Correct. And then what happened when you did the parabiosis with the DB mouse? With the DB

00:29:51.000 mouse, you got the opposite situation, where the wild-type animal began to lose weight and actually

00:29:59.600 stopped eating and would die of starvation, basically. Whereas the DB mouse just went merrily on its way,

00:30:06.480 eating and remaining obese. So Coleman thought, based on these two experiments, that maybe the OB mouse

00:30:17.300 was missing something that the wild-type mouse produced, and the DB mouse, that looked an awful

00:30:22.780 lot like an OB mouse, was missing the ability to respond to whatever that molecule or molecules

00:30:30.320 were that were required to regulate body weight. And this was around the time that it was becoming

00:30:36.900 clear that hormones had receptors that were specific to those hormones, like thyroid, insulin,

00:30:43.220 growth hormone. So Coleman hypothesized that the OB mouse was missing what we refer to as the ligand,

00:30:50.780 the circulating hormone, and the DB mouse was missing the receptor for it. And it turns out

00:30:57.680 that Dr. Coleman was right. Everything you've just said was known by what point in time?

00:31:03.640 I would say by the late 1970s. Okay, so just as you're coming on the scene, the whippersnapper who

00:31:10.020 doesn't know shit. Yes. The guy up in Jackson, up in Maine, knows this, and you can't wait to get all over

00:31:17.680 this. Correct. So I began by actually studying adipose tissue to see whether there was something

00:31:26.240 that might be regulated by adipose tissue that would fit in this sort of general category of

00:31:33.960 something that might produce a signal or might be related to the anatomy of the adipose tissue.

00:31:41.880 At the time, Jules Hirsch was particularly interested in the fact that it appeared that what happens when

00:31:48.720 people get obese is that their fat cells expand up to a sort of maximum size, and then new fat cells

00:31:55.900 begin to appear. Whether they were made de novo or had been resting in the area of the other adipocytes

00:32:02.180 wasn't clear at the time. But what happens as people get more and more obese is that more and more fat

00:32:08.060 cells are recruited so that if you look at a very obese individual, they have what is referred to in

00:32:15.300 the literature as hyperplastic adipose tissue. They have more fat cells. And Jules had actually figured

00:32:20.620 out a way to count the fat cells of a human being. And you can look under a microscope and see how big

00:32:27.140 the fat cells are. So he was very interested in what it is that permits a big fat cell to call up,

00:32:35.060 if you will, the other fat cells that are going to be needed once it reaches a critical size,

00:32:40.660 and whether these fat cells were generating something that might act like a signal.

00:32:46.180 Are there any organelles inside a fat cell? Oh, yeah. Fat cell has the complete repertoire

00:32:50.820 of organelles that any other cell does. So it's got a quote-unquote regular nucleus. It's got

00:32:57.260 mitochondria. Oh, yeah. Yeah. It's got the whole... It's just its cytoplasm is dominated by these lipid

00:33:03.460 droplets. Correct. It's designed to be able to hold these huge lipid droplets, but otherwise is a

00:33:08.640 perfectly respectable cell. It has all the other components. And what Jules had been doing, along

00:33:17.200 with another investigator in the lab, a guy named Irv Faust, was extirpating adipose tissue from rats.

00:33:24.640 They were working primarily with rats, although there were some OB mice around. I'd actually seen

00:33:30.500 those when I was at Harvard in Boston. An endocrinologist who trained me used to talk to me

00:33:35.880 about them. But Jules and Irv were extirpating adipose tissue depots from rats and looking to

00:33:44.640 see what happened. And one of the things that they were struck by is that when they took a fat pad out,

00:33:50.560 if they waited long enough and just let the animal eat whatever it wanted to eat, it would eat its way

00:33:56.780 back up to restoring that fat pad. Through de novo creation of more fat cells?

00:34:02.280 So you leave the capsule of the depot there and it is able to regenerate new fat cells and they fill

00:34:11.100 up and they don't fill up to three times the size that they were before. They go right back to the

00:34:16.360 appropriate size for that depot. And one of the things that Jules and Irv were very interested in

00:34:24.360 is how does the animal know that it should eat a little bit more to be able to generate the fat

00:34:32.040 that's been extirpated? And did this experiment only work when you started with obese rats or did

00:34:38.580 it work if you took a lean rat as well? It worked with lean rats just as well. As a matter of fact,

00:34:44.040 most of the work they did was with lean rats or not genetically obese rats.

00:34:49.120 Now speaking of rats, you explained to us what the OB-OB and the DB-DB-DB-OB.

00:34:56.860 What about the Zucker rat, which preceded a lot of this stuff but had some interesting and similar

00:35:01.040 features, right? Right. So the Zucker rat was discovered or identified, if you will, by a woman

00:35:08.520 named Lois Zucker, the rats named after her and her husband. And it had characteristics very similar to

00:35:15.280 these mice that we were talking about, the OB and the DB mice. They were hyperphagic. They

00:35:21.520 appeared to have a little bit lower energy expenditure and would become hugely obese. I

00:35:28.260 mean, these animals got up to heroic size. Yeah, we're going to include some pictures of the

00:35:33.520 Zucker mice in this, in the notes. The Zucker rats.

00:35:36.340 That's right, the Zucker rats in this show.

00:35:37.760 Got very, again, got very obese. And many, many studies were done of the Zucker rat trying to,

00:35:45.320 again, understand what the mechanism behind its obesity was. Again, it appeared to be due to a

00:35:53.400 single gene. In other words, like the OB or DB mouse, there was a single mutant gene. When the animal

00:35:59.320 had two copies of the abnormal or low activity gene, the animals would become very obese.

00:36:05.940 And one of the reigning hypotheses about the Zucker rat, again, at around the late 70s, early 80s,

00:36:15.540 was that the fat itself was making too much of an enzyme called lipoprotein lipase. And this is an

00:36:24.680 enzyme which is produced by adipose tissue, by adipocytes, which breaks down circulating triglycerides

00:36:32.500 and allows the free fatty acids that are released to be taken up by the adipocytes. And the glycerol

00:36:40.280 backbone, so to speak, remains in the circulation. So the idea was that the Zucker rat, for some reason,

00:36:46.980 had hyperactive lipoprotein lipase. And the adipose tissue was acting like a vacuum cleaner,

00:36:53.820 in a sense. It was sucking up the circulating fat, the triglyceride, and storing it. And that's how

00:37:01.900 the animals got obese. And in association with that observation was the idea that maybe the process of

00:37:09.100 sucking up substrate from the circulation does something to drive or increase food intake. So

00:37:18.120 this model was often referred to as a pull model for the development of obesity. That is that the

00:37:26.260 adipose tissue was actually acting as a pulling mechanism for substrate, which was in turn affecting

00:37:33.280 food intake. So the animal got fat that way. So the animal was effectively starving. Even though it

00:37:39.340 stored an unbelievable amount of energy, it was disproportionately taking those circulating

00:37:45.280 metabolic fuels and putting them into storage. So that if you buy this idea that something is

00:37:51.520 sensing energy availability, that would naturally drive the hyperphagia, right? Correct. And the other

00:37:58.240 model, the one more consistent, say, with the OB or the DB mouse, was the so-called push model,

00:38:05.340 which is that fat cells were being filled up because the brain itself had a... Was pushing.

00:38:11.140 Pushing the substrate. So these push-pull models were very much discussed in terms of what they said

00:38:19.720 about where the primary mechanisms might be for the control of body weight. Was it the adipose tissue

00:38:27.380 that was basically begging for more fuel and sucking it in and causing the animal to eat that way? Or was

00:38:33.920 there something going on primarily in the brain that was influencing the food intake and the

00:38:40.880 adipose tissue was being packed from the outside? And like all good problems in medicine,

00:38:46.060 there's examples of both that are quite elegant, right? I mean, that's sort of the challenge of

00:38:51.340 this is when you look at this in experimental models, you could find very elegant examples of

00:38:57.080 each, correct? You could find elegant examples that pointed in the direction of both of these.

00:39:03.320 It was interesting. I mean, it was the whole issue of lipoprotein lipase suggested that,

00:39:09.380 again, this model, the Zuckerrat, it was used to support this idea that the LPL was, as it was

00:39:15.980 referred to, was critical in this regard. But it was known that individuals who had total lipoprotein

00:39:23.380 lipase deficiency, and there are such individuals on a genetic basis, had very, very high levels of

00:39:30.460 circulating triglyceride. I mean, these can be life-threatening in terms of the levels that are

00:39:35.620 reached, but they had perfectly normal adipose tissue. So this suggested that maybe LPL wasn't

00:39:41.540 critical or absolutely necessary. Meaning you would expect those patients to have sky-high

00:39:46.680 triglycerides and be emaciated. Yes. And they weren't. And they aren't. So this was already sort

00:39:53.060 of a question about the lipoprotein lipase. And we know that those patients were completely

00:39:58.640 deficient in LPL? Yes. Does that mean they had no LPL on their muscle cells as well?

00:40:03.140 So the individuals who are totally deficient in LPL have no LPL on any cell type.

00:40:11.960 It's hard to imagine how they function. Does that mean they're completely dependent on glycolysis?

00:40:17.000 They are able to take up the fatty acids. They don't need the LPL to break down the triglyceride.

00:40:23.800 They do fine. I mean, in terms of being able to, there's nothing wrong with their manipulation or

00:40:28.840 movement of fatty acid. And that's probably why they have normal adipose tissue and normal muscle.

00:40:35.800 So it's interesting, the history of this, and just to mention this, we now know what the Zucker

00:40:42.560 mutation is. And one of the earliest experiments I did at Rockefeller, or one of the relatively early

00:40:49.480 experiments, is we tried to map the position of the Zucker obesity gene in the same way that we

00:40:58.360 were already trying to map where and ultimately clone the OB and DB gene for pretty obvious reasons

00:41:05.300 at this point, that we wanted to figure out what the signal and whether there was a receptor for this.

00:41:11.800 And as part of this work, a student who worked with me named Gary Truitt made a map of the position

00:41:19.480 of the Zucker gene, which we did by crossing the Zucker rats with another strain of rats so that we

00:41:25.760 could make what's referred to as a genetic map. And tell me how that works, because everything I've

00:41:30.960 ever done with genes came after we had the luxury of real-time PCR. And we just said, you know,

00:41:37.380 my generation had it so easy. You guys, like, when I hear stories about people doing positional gene

00:41:44.200 stuff, you know, 40, 50 years ago, I'm like, wow, that's incredible. So tell me a little technically,

00:41:50.780 like, what were you actually doing to positionally isolate that?

00:41:54.960 So the way these experiments are done, it's pretty much the same principle, is that you take,

00:42:01.480 in the case of rats or mice, which you have this luxury, you can cross one strain of,

00:42:07.380 mouse or rat, to another strain of mouse or rat, and then look for signposts of genetic variation

00:42:16.000 along the entire genome of the animal. And each strain has different sequences of DNA.

00:42:26.040 Major differences are in the so-called non-coding region. Some of the differences are in coding

00:42:31.840 regions. And back then, we used a technique called southern blotting to look for

00:42:37.080 basically putting down signposts on the DNA of an animal to figure out where the differences between

00:42:44.800 the strains resided. And by monitoring the obesity of the animal and knowing which strain was actually

00:42:54.420 carrying the gene that we didn't know what it was, but we know what strain it came from,

00:42:59.440 in this case the Zucker strain. We could actually mark the DNA of these animals that had been

00:43:06.880 interbred and figure out where the Zucker chromosome, the Zucker genetic markers, were segregating,

00:43:16.940 is the formal term for this, along with the level of obesity of the animal. So we would look for

00:43:22.160 correlation of region which has the Zucker genetic variation with the obesity of the animals. Because

00:43:31.280 if you do these crosses in the way that I described, you get some obese, you're going to get in some

00:43:36.840 non-obese animals. And then by relating where the sequences are corresponding to the strain from

00:43:44.520 which you know the obesity gene must have come, you can actually narrow the region of the genome down

00:43:51.080 to the area that must contain the gene. I mean...

00:43:54.520 And this was a qualitative assessment, not so much a quantitative assessment, right?

00:43:58.680 So the quantitative assessment is, is the animal obese or not obese? You absolutely have to get this

00:44:05.840 right because you're, the obese animal has, in this particular model, two copies of the mutant gene

00:44:12.720 that's producing the obesity. And if it's not obese, it either has one or zero copies. And you need to be

00:44:18.880 absolutely clear what the phenotype is, so to speak, of the animal is. And then if you put these markers

00:44:27.360 down, these signposts down, you can define them, an interval in which that gene must reside. So it's

00:44:37.080 actually, it's, it's in a sense, it's quantitative at the level of the phenotype.

00:44:42.540 So the best you could do is say this is on, you know, 17P or 17Q. That's, that's like the level of

00:44:49.160 resolution you can get out of this?

00:44:50.820 Yes.

00:44:51.560 What happened when they did a parabiosis with a Zucker rat? Did anyone do that?

00:44:55.180 There have been parabiosis experiments done with rats, but they've generally done these in a,

00:45:03.600 in a slightly different way from the way you asked the question. And those studies have again,

00:45:11.860 supported the idea that the Zucker rat may be overproducing, like the DB mouse, the product,

00:45:19.100 which is suppressive of food intake. And as a matter of fact, when we were trying to clone

00:45:25.120 these genes, I actually took blood out of Zucker rats and injected it into DB mice to see whether

00:45:33.280 or not I could slow down their food intake. Because at one time we were worried that we might not be

00:45:38.400 able to get the gene by mapping and would have to try to isolate the product out of the blood of an

00:45:43.300 animal. But the Zucker rat studies supported the idea that the same as the mice.

00:45:49.560 So we'll fast forward now into the late seventies, early eighties. You're working mostly now in the

00:45:55.800 lab. Your clinical responsibilities have shrunk significantly, right?

00:45:59.660 So at the time I was doing these experiments, I had some experiments still going on, no clinical

00:46:07.240 practice at all. But I was studying humans in the clinical research center at Rockefeller,

00:46:13.220 looking at the effects of weight loss and weight gain on energy metabolism in these individuals.

00:46:23.440 Again, beginning to try to understand what it is that's regulating body weight in a human. The idea

00:46:30.800 being that if you reduce the body weight of a human down by 10 or 20%, what happens to them

00:46:38.020 metabolically that might be consistent with some of these things that had been seen in mice,

00:46:44.260 that would suggest that again, in humans, there's regulatory pathways that are there in order to

00:46:52.340 regulate the amount of stored energy. So my time was basically split between doing these studies of

00:46:59.940 humans who were in the clinical research center for long periods of time, when we would study the

00:47:04.820 metabolic consequences of perturbing their body weight. The other was spent trying to identify

00:47:11.300 these genes or try to clone the genes. So how did that progress through the eighties as you were

00:47:18.180 working to try to identify specifically the OB OB gene? And again, the reason you were pretty confident

00:47:25.140 that it was a single gene was just based on the breeding pattern? Yes. The breeding pattern clearly

00:47:30.580 indicated that this was a single. Because it was sort of Mendelian. Yes, absolutely Mendelian.

00:47:35.940 It's a so-called autosomal recessive Mendelian, as is the DB mouse and the Zucker rat. So what we did

00:47:43.060 is what I basically described for the rats. Why was it harder for the mice? To clone the gene? Yes.

00:47:49.780 It was not harder. No, no. I think it was equally difficult. What I was describing is when we didn't

00:47:56.420 actually sort of finish that part of the conversation. We did make a map of where the

00:48:02.340 Zucker gene was located in the rat. But we didn't clone the gene out of the rat at that time. And

00:48:10.020 what that experiment showed is that that was interesting apropos this lipoprotein lipase

00:48:15.620 hypothesis is that the Zucker gene was in a part of the rat genome entirely separate from where the LPL

00:48:24.740 LPL gene was located. So from that experiment alone, we knew that it couldn't be LPL as the

00:48:34.660 causal mechanism of the Zucker rat. We didn't know what it was, but we could clearly identify what it

00:48:42.660 wasn't. Because if the gene is not in the location that is segregating with the phenotype, it's not the

00:48:51.060 gene. This was hard for people, I think, to understand or at least to accept at the time.

00:48:57.700 There still, I think, was not full appreciation of what an experiment like that represents,

00:49:05.300 which is basically an unequivocal falsifying experiment. That is, if it isn't in the place

00:49:12.180 where the gene must be, then it can't be the gene. I mean, it's a syllogism.

00:49:17.460 Yeah. And you don't get a lot of those in science.

00:49:20.420 No, you don't get those.

00:49:21.140 Especially in biology.

00:49:22.340 Right. And so this early on, this was one of the proofs that at least we knew one gene

00:49:27.780 that it wasn't.

00:49:28.580 It wasn't.

00:49:29.460 And then to sort of back then to your question, we used a very similar strategy,

00:49:35.140 an identical strategy with the mice. We crossed the mice to strains that were different from the

00:49:41.140 strain on which the mutation existed. And by doing that, we're able to make maps

00:49:45.620 of the region where the gene was located. And I was actually known for other reasons. Coleman

00:49:52.260 had figured it out. He knew which chromosome these genes were on. So we at least had that

00:50:00.260 as a sort of initial guide. But what the gene was or how it worked was obviously entirely unknown.

00:50:08.900 What was Coleman's training?

00:50:10.980 Coleman was trained as a biochemist at the University of Wisconsin, I think. And we actually

00:50:16.100 consulted with Dr. Coleman, Doug, very early on, as a matter of fact, very early on in the process,

00:50:22.020 about which strains of mice might be appropriate as so-called counter strains. That is,

00:50:27.380 mice that would be used to enable these maps to be made. And he was extremely helpful. I mean,

00:50:35.540 we used to come down, we would talk to him, go visit him at Jackson on at least one occasion,

00:50:42.020 met with him in Philadelphia.

00:50:43.460 So he loved the collaboration.

00:50:45.540 He loved it. It was very encouraging. He always said that he wasn't sort of au courant with regard

00:50:53.300 to the tools of so-called molecular genetics. But in fact, I and Jeff Friedman, who was the

00:50:59.380 other sort of PI on this project, neither of us was expert in molecular genetics.

00:51:04.740 So we sort of had to train ourselves to do this, as did the students who worked with us on this project.

00:51:10.180 So when did Jeff enter the lab?

00:51:13.300 I got to know Jeff because he rotated as a resident, actually, at New York Hospital through

00:51:22.020 Jules' lab to work with another investigator there. So I got to know him when he rotated through.

00:51:28.260 But he and I started this project after he entered the Ph.D. program. He was a physician and then went

00:51:36.100 on to get a Ph.D. at Rockefeller in a different laboratory. But he and I started the project to

00:51:42.100 clone these genes together.

00:51:43.860 About what year was that?

00:51:45.700 1985 or six, I guess. And, you know, 86, probably.

00:51:51.380 What were the major steps that took place starting around the mid-'80s that were kind of like the

00:51:57.940 essential? I mean, I think one of the things that's just hard for people, I think, to understand is

00:52:03.060 how much you have to fail in science. I mean, it sounds simple, right? We want to clone a gene.

00:52:09.940 And yet, in that era, that was very hard to do.

00:52:14.820 Right. There were efforts underway at around the same time to clone the gene for Huntington's disease

00:52:22.100 and cystic fibrosis, muscular dystrophy, to name a few.

00:52:27.220 Other single genes.

00:52:28.340 Other single gene disorders. And for a number of reasons, genes that are on

00:52:33.940 sex chromosomes may be a little bit easier to handle in this regard. But the same tools were being used,

00:52:41.700 the so-called southern blotting, putting down markers across the genome, using this rather

00:52:46.580 tedious technique of having to put the DNA down on a blot of some kind and then exposing it with

00:52:54.420 radioactively tagged reagents to be able to find these individual markers. This was a

00:53:00.980 tedious process. One thing that helped with the mice is that there existed a chromosome

00:53:06.740 in mice. So these are called Robertsonian chromosomes in which you have two chromosomes

00:53:12.820 joined to each other. And there was a so-called Robertsonian chromosome that had both chromosome

00:53:18.260 4 and chromosome 6 of the mouse attached to each other. We knew based on Doug Coleman's work that

00:53:25.780 the OB gene was on chromosome 6 and the DB gene was on chromosome 4. And one of the students who came

00:53:32.740 to work on this project very early, a young man named Nate Bahari who came over from New York hospital

00:53:40.500 as a medical student and spent time on this project and then went on to get a PhD working on it, he took

00:53:47.860 glass needles, if you can believe this, and dissected out under a microscope the rough regions of the

00:53:56.100 chromosome 6 and 4 that we knew must have the gene in them and made from those needle dissected

00:54:05.060 chromosome fragments other markers that we could put down on the DNA from these mice that we had

00:54:13.460 intercrossed to make a finer and finer map of the region in which the gene must reside. And otherwise this was

00:54:21.700 identical to the Zucker strategy in which we very carefully looked at see whether a mouse was obese

00:54:29.300 or not, and again these are mentalizing phenotypes, but we had to be absolutely sure what the phenotype

00:54:36.340 of the mouse was and then we could use the DNA from the obese animal to make a finer and finer map around the

00:54:45.780 region where the gene must reside. I don't even understand how that would work. What resolution of a

00:54:50.180 microscope would you need to be tweaking chromosomes? Chromosomes aren't that small. I mean you can see

00:54:55.540 a chromosome quite easily with a high-powered just a light microscope. Yeah, light microscope. But

00:55:01.300 having the steady hands to be able to and the patience to be able to do this was quite an achievement.

00:55:08.420 I would say it was sine qua non without which this project wouldn't have been done at the time. We didn't

00:55:15.380 have the tools, some of the ones that you already mentioned, available. So we had to try to identify

00:55:21.540 additional signposts, if you will, in the region of the two genes that we were primarily interested in,

00:55:29.380 that is the OB and the DB gene. At the time you're basically thinking,

00:55:32.740 look, we're looking for a gene for a hormone, we're probably looking for a gene for a receptor,

00:55:36.660 so a ligand receptor. Did you have an expectation that those would be on the same chromosome or would

00:55:40.900 that be completely unnecessary? No, we knew that they absolutely weren't. No, no, I knew you knew

00:55:45.380 that they weren't, but would you expect that? No. For example, like in thyroid, is T4 and TSH or T4 and

00:55:54.980 TR, like you know the thyroid receptor, TSH and TSR, are they necessarily? No. No. There's no

00:56:01.220 relationship. No. Yeah. No. So the function doesn't require that the genes be in proximity to each other,

00:56:07.140 and if they are, it's almost always just a coincidence. Got it. So what was the kind of

00:56:13.300 critical breakthrough that led to the cloning of this gene? So what was done basically was to make

00:56:22.660 the map as fine as we could using the reagents that I mentioned to you and some others that we were able

00:56:29.620 to get hold of and using computer programs to put these data together in such a way as to be able to

00:56:37.780 generate as fine a map of the region around this gene as we could. And then what was done was to go

00:56:45.460 into the region that must contain the gene and begin to look for expressed, meaning that the DNA was

00:56:55.300 transcribed transcripts that were being read off of this part of the DNA. And by taking the transcripts

00:57:04.740 that came off and putting them against various organs from the OB or the DB mouse, we would look for

00:57:14.740 transcripts that were specifically altered in the animals that were mutant for the gene. So you would

00:57:23.220 expect, you might ordinarily expect maybe the level of the gene that we were looking for would be very low

00:57:29.300 in the organ that was affected. And again, even though we know now it was adipose tissue and there

00:57:35.380 were certainly reasons to think that might be where the OB gene was, when we did these experiments we

00:57:40.900 actually looked at every organ in the animal, brain, thyroid, muscle, adipose tissue, anything we could

00:57:48.580 get our hands on. We actually dissected out almost everything out of a mouse to look to see where the

00:57:56.180 deficiency or the excess might be. Sometimes gene transcripts are overexpressed, although they're not

00:58:02.500 functionally competent. And actually at a visit to the Jackson labs when Doug Coleman was retiring,

00:58:11.460 one of the investigators there asked me if I would like to have, or we would like to have

00:58:18.740 mice that appeared to have the OB mutation, but a different mouse. So the original mutation occurred

00:58:26.340 in the mouse that was identified at Jackson in the early 1950s. This was a new mutation that had arisen

00:58:33.380 and almost certainly wouldn't be identical to the one that had arisen earlier.

00:58:37.540 But it was similar in phenotypes? Absolutely identical in phenotypes. So I got those mice

00:58:43.220 and we took them to Rockefeller. What did you call them?

00:58:47.380 2Js. So the OB, the first mutation we referred to as 1J, that's just the nomenclature. And the

00:58:54.420 second one was OB 2J. And we took the OB organs out of the 1Js and the 2Js and ran these transcripts

00:59:03.700 against them using the technique called Northern blotting at the time.

00:59:07.620 Not to be confused with Western blotting.

00:59:09.540 Not to be confused with Western blotting. So Northern blotting is looking actually at RNA.

00:59:14.340 It turned out that the 2J was extremely valuable animal because those animals didn't express any

00:59:23.380 of what turned out to be the leptin transcript. Whereas the 1Js actually overexpressed it,

00:59:31.700 but it was functionally inadequate, functionally inactive. But the transcript was increased,

00:59:38.260 again, for reasons having to do with the molecular biology of how these systems work. But when we saw

00:59:43.780 that there was one animal that had a very low level and the other very high, this was a smoking gun,

00:59:50.740 so to speak. This was the evidence that that piece of the genome was actually a part of the OB gene.

00:59:58.340 So ultimately, where did this ligand, where did it show up?

01:00:03.380 The OB gene showed up almost solely, virtually solely, I mean, in adipose tissue. Fully consistent

01:00:11.140 with the idea that adipose tissue was producing something that was picked up in Doug's parabiosis experiments.

01:00:19.060 Which chromosome?

01:00:20.660 Chromosome 6 of the mouse, it's chromosome 7 of the human. We actually knew that because we could put the

01:00:30.100 orthologs of the mouse genes down on human DNA. And so just like we did with the Zucker rat,

01:00:37.460 we actually knew where the gene was.

01:00:41.380 Do mice have 23 pairs of chromosomes?

01:00:43.540 They have, yes, 23.

01:00:45.860 23, including the sex.

01:00:47.060 Yeah.

01:00:47.860 So what name was given to this?

01:00:50.580 I mean, it's referred to as the OB gene or the name that was given to the protein is leptin,

01:00:57.060 which was, the name was chosen because that suggested that whatever this was,

01:01:02.980 it had an effect to lighten the body weight of an animal.

01:01:06.580 So now you have it that the fat cell secretes leptin and the OB mouse can't make leptin, correct?

01:01:16.420 Correct.

01:01:17.460 And the DB mouse makes all the leptin in the world.

01:01:20.260 This wasn't known at the time because the gene, the first gene to be cloned was the OB gene.

01:01:27.140 And the fact that the DB gene was the receptor was actually learned in two ways.

01:01:34.260 One by taking the OB protein and putting it down on what's called an expression library

01:01:42.260 from part of the brain, the choroid plexus.

01:01:44.580 Wait, sorry, to interrupt one thing, Rudy, I apologize.

01:01:47.620 Did they ever do parabiosis between the OB and the DB?

01:01:50.500 Yes, that was done as well.

01:01:52.500 Okay.

01:01:53.060 Because that would, that should fix the OB.

01:01:56.180 It did.

01:01:57.060 But not the DB.

01:01:58.020 Correct.

01:01:58.420 Okay.

01:01:59.300 Which is another great and elegant example that one of them had the ligand deficiency,

01:02:03.860 the other had the receptor deficiency.

01:02:05.940 God, Mendelian genetics are freaking awesome when they work.

01:02:09.140 Yeah.

01:02:09.540 Well, so that was one of the reasons for using these animals is that you could use the genetics

01:02:14.020 to really help to narrow down the region where these genes had to be located.

01:02:19.220 If it were a polygenic disturbance, it's really, it's a whole different order of business.

01:02:24.020 So basically the DB gene was, or the protein that was nominated to be the DB gene was pulled

01:02:31.860 out of a library of what's called an expression library of choroid plexus by using the OB protein

01:02:38.980 as a probe to see what it stuck to.

01:02:42.020 And that gene that was pulled out of the choroid plexus in that way, we then took and mapped

01:02:51.860 into the DB crosses that we had and showed that that, if you will, nominated gene mapped

01:03:00.180 right in the right place for the DB gene.

01:03:03.860 Which was on chromosome four in the mouse.

01:03:06.100 Correct. And it also turned out that that gene, the DB gene, the leptin receptor, is the mutant

01:03:14.020 in the Zucker rat, which we showed by cloning the gene then out of the Zucker rat, knowing where it was.

01:03:21.700 So this was hailed as one of the most important insights and breakthroughs in all of obesity

01:03:27.460 research. This was what, 1994?

01:03:30.020 4. And then the DB gene a year or two later.

01:03:35.460 What was your expectation at the time? What type of hope did you hold out for

01:03:40.020 the discovery of the gene, but perhaps more importantly, the protein that it coded for?

01:03:45.620 So there were different schools of thought with regard to what this protein actually did. I think

01:03:51.860 one view was that if you saw this as a protein that suppresses food intake, which it definitely

01:03:59.940 does in an OB mouse. If you give it to an OB mouse, you can basically, quote, cure the animal.

01:04:06.980 And if you give it in very high doses to a mouse, it will suppress its food intake.

01:04:12.340 So one view was that this was a weight suppressing hormone. The other view, which was taken by

01:04:21.620 Jeff Flyer, who was at Harvard at the time, later became dean, still at Harvard.

01:04:28.020 Streamson Chua, who was working on this project or had worked on it, and myself, was that the

01:04:34.660 protein was actually more important in its deficiency state as a signal to the brain that you didn't have

01:04:42.020 enough energy to survive under circumstances of a fast or to have enough energy on board to successfully

01:04:48.660 complete a pregnancy. So one view was that very high levels would normally suppress body weight,

01:04:55.300 although it was hard to imagine at the time. Evolutionarily, that was not a natural state.

01:04:59.620 That's correct. So that's why we were wondering right from the beginning, why would nature invent

01:05:04.340 something that would... Right. It makes more sense that nature would say, here's a hormone that if

01:05:08.820 it's low, is a kick in the pants to eat. That's correct. Or, and maybe other things, right? Maybe

01:05:13.860 prevent you from getting pregnant. Yes. And that's, I think, turned out to be the correct model for how

01:05:19.220 this hormone, its primary mechanism of action is that it is there to tell the brain that you don't have

01:05:26.180 enough fat. And in an OB mouse, you don't make the hormone leptin, so the brain thinks you don't have

01:05:32.500 any fat. And in the DB mouse, it doesn't receive the signal from the perfectly adequate amounts,

01:05:39.220 actually high levels of leptin, so that the animal again thinks that it's starving.

01:05:44.740 Are there human conditions that mimic the OB and DB condition of the mice?

01:05:49.460 Yes. So there are humans. There are a handful of humans who have mutations of the OB gene and are obese

01:05:56.020 and have many of the phenotypic characteristics of a... And these are patients that you've cured with

01:06:01.700 this by giving... You give these patients leptin and you cure them.

01:06:04.100 Yes. They're curable by giving leptin. And there are a handful, again, of individuals with leptin

01:06:10.260 receptor mutations, which are comparable. How do you help those patients?

01:06:14.900 Those... The leptin won't help. You can't treat them with leptin. And there is no effective intervention

01:06:21.380 for those people at this time, although there are drugs that are being developed that are designed to

01:06:27.460 act downstream of some of the actions of leptin that might, if you could, rectify the activity

01:06:34.580 there. For example, at a receptor called the melanocortin-4 receptor, that would presumably

01:06:41.460 or possibly help to rectify the phenotype of those individuals. Although at present,

01:06:46.340 it's not a proven effective intervention. It might turn out to be.

01:06:50.180 I mean... It wasn't long after the discovery of leptin that... What drug company came in?

01:06:56.180 Amgen.

01:06:56.660 Amgen. So Amgen obviously bought the rights to do this, thinking this could be a blockbuster obesity

01:07:03.380 drug. It turned out not to be because unless you had a leptin deficiency, it didn't seem to help much,

01:07:09.300 right?

01:07:09.620 Correct. There was a major trial of the use of leptin as a therapeutic agent in obese individuals,

01:07:16.900 and neither the obese individuals or the lean individuals who were in the study as controls,

01:07:23.060 responded very much to raising blood levels as much as tenfold above where they were normally

01:07:31.220 located in those individuals. So I just did a blood draw. I've been doing a blood draw on myself

01:07:35.620 every seven days because I'm doing... You know me, Rudy. That's just the shit I do. So my leptin level

01:07:40.820 last Friday was two or less than two. The cutoff was two. So I have a low level of leptin. So if you

01:07:49.140 gave me enough leptin to raise my serum levels to levels that I rarely see clinically, like when I

01:07:55.220 check leptin levels on patients, which I always do, I always want to see leptin. I want to see

01:07:58.740 adiponectin. I want to see insulin. I want to see NIFA, FFA, etc. It's very unusual that I'll see a

01:08:05.540 leptin level, even in an overweight patient, more than about 40 or 50. So if you took me and you gave

01:08:12.020 me that much leptin, do you think it would regulate or depress my appetite in any way?

01:08:17.300 Well, first of all, your leptin is low now because you're not eating. So...

01:08:22.180 Oh, this is even... This is back when I was eating too.

01:08:24.260 Right. So leptin is very highly correlated with body fat. At any level of body fat, if you stop eating,

01:08:31.220 if you restrict calories, the level of leptin will drop by...

01:08:34.260 Within what period of time?

01:08:35.220 Within 12, 18 hours, it'll drop by 50%.

01:08:39.060 So if we check a leptin level on a patient in the morning following a 12-hour fast,

01:08:43.300 we can assume that that's about half what they're fed leptin level is?

01:08:46.660 Well, 12, it depends on what they've been eating. 12 may be a little short in that context. So

01:08:51.780 to be safe, I would say 24.

01:08:54.260 Okay.

01:08:54.620 But it'll start down within 12 hours. So a 12-hour fast will give you a leptin level,

01:08:59.180 which is not what it would be if you were to draw it when the patient is...

01:09:02.700 So let's say my fed level is 2 or 3 or 4. My fasting level is less than 2. If you injected

01:09:10.700 me with enough leptin such that, let's just say my level was now 100, the level that you would see

01:09:15.740 in the highest fed obese patient, what would that do to me?

01:09:20.060 Based on the studies that have been done, the so-called leptin trial, not very much.

01:09:25.420 So interesting.

01:09:26.780 You might have some, at least in some of these individuals, there was some reduction in body

01:09:33.340 weight, but not very significant.

01:09:35.500 Through the mechanism of reducing intake?

01:09:38.140 Yeah, primarily, apparently, yes. Although there weren't detailed formal studies of food

01:09:46.060 intake done in those experiments.

01:09:49.660 Now, going back to the DBs, who obviously have, in many ways, a worse problem. Earlier,

01:09:55.740 you alluded to Prader-Willi. What is the genetic defect in Prader-Willi, and what's the phenotype?

01:10:01.340 So Prader-Willi is a genetic disorder in which, rather than having a single gene affected, there's

01:10:09.100 a region of the genome which is, in most instances, not all, but the majority of instances, deleted.

01:10:15.660 So there's a region of chromosome 15 in which there is a large deletion that is a region that has normally

01:10:25.900 20 or so genes in it. And when deleted, when the paternal, the father's copy of this

01:10:35.180 interval is deleted, the individual becomes, has a very characteristic phenotype. The maternal,

01:10:42.940 the mother's copy, is not expressed. This is called an imprinted region of the genome, which again,

01:10:49.820 for technical reasons, the only one chromosome expresses the genes from that interval. And in the

01:10:57.900 Prader-Willi region, the maternal genes are silenced. So they're there, but they're not expressed.

01:11:03.820 And if you do... So the mom could be a silent carrier.

01:11:06.380 The mom could be a silent... Carrier is not the right term.

01:11:10.540 Almost all of these individuals are... All of these individuals are the result of

01:11:17.180 spontaneous deletions that occur after conception. So it's not... So it's sort of polygenic,

01:11:25.180 and you can have... So the one example that is in my world that I get is familial hypercholesterolemia.

01:11:32.620 It's a phenotypic description. There are at least 2,000 known mutations, distinct mutations,

01:11:39.740 that can produce the same phenotype. So in that case, no one really... I don't want to say nobody

01:11:45.420 cares, but it's a lot less interesting to try to map out each and every one of the genes that can

01:11:50.460 lead to familial hypercholesterolemia. It's more important to understand the phenotype and how to

01:11:54.940 treat it. I'm guessing Prader-Willi is not quite that diverse, but is it safe to say, if I'm

01:12:00.620 understanding you correctly, that you could have two patients with Prader-Willi that will have different

01:12:04.380 genes? There are rare instances of Prader-Willi in which the number of genes deleted, rather than

01:12:12.700 being that large interval that I mentioned, include only three genes, but they're still in the region

01:12:18.540 which is deleted in the large deletion patients. So... So they're necessary and sufficient genes.

01:12:25.020 Yes. We think that those... that smaller region probably has the necessary and sufficient genes,

01:12:31.900 although again, there's debate in the field about this. The children, though, who are affected with

01:12:38.060 this disorder have very characteristic phenotypes. They actually don't grow well. They don't feed well

01:12:43.420 early in life. They actually have failure to thrive. They don't... they're floppy and hard to feed and

01:12:52.300 not very responsive in terms of ingestive behaviors. And then, again, for reasons that are entirely

01:12:59.820 unknown at this point, between the ages of four, five, three years of age, they become hyperphagic. They,

01:13:08.220 again, the drive to eat becomes very, very strong. These are some of the most tragic stories I've ever

01:13:14.860 heard. These children who I've never taken... I'm not a pediatrician, so I've never taken care of them,

01:13:19.740 but I've talked to parents of children. They have to lock the refrigerators. I even had one parent tell

01:13:25.980 me that they actually have to lock the bathroom because the child is so hungry, so hyperphagic,

01:13:31.500 that he would go into the bathroom to try to eat out of the toilet. It's true that in the majority of

01:13:36.780 instances, these children require very close scrutiny of their access to food. And in many instances,

01:13:44.780 the parents or caretakers have to resort to locking up whatever food would otherwise be available to

01:13:53.260 the children. But I think it's important to point out that there are ways of managing this disorder,

01:14:00.220 which has other characteristics. There are many endocrine disturbances that go along with it,

01:14:05.980 including actually growth hormone deficiency. And administration of growth hormone to these

01:14:12.380 children and various other sort of interventions that require a great deal of parental attention,

01:14:19.900 no doubt about that. Some of the severity of the disease is manageable at a clinical level with very

01:14:26.460 high levels of scrutiny by the people who are taking care of them. So that not all children with

01:14:33.020 Prader-Willi will necessarily become hugely obese or suffer the consequences of severe obesity.

01:14:40.540 This is one of those things where I hope, I know there are folks out there trying to raise money to

01:14:44.860 do clinical trials in this because anecdotally, again, there are these very interesting stories.

01:14:49.820 The first I ever heard of this was probably four years ago. I was giving a talk. I don't even

01:14:54.140 remember where it was. It was in Chicago, but I can't even recall the audience. But afterwards,

01:14:58.300 a woman came up to me and I was giving a talk about, I don't know, longevity, like it wasn't an obesity

01:15:04.860 specific talk or anything like that. But a woman came up to me and introduced herself and said her

01:15:08.540 son had Prader-Willi and was going off the rails, et cetera. And at about the age of seven or eight,

01:15:14.140 she put him on a ketogenic diet. And it corrected the phenotype, including the cognitive developmental

01:15:22.220 part, which often accompanies this. She then, I guess, since that time had formed kind of a support

01:15:29.020 group and you had more and more of these parents that were putting their Prader-Willi kids onto

01:15:34.140 ketogenic diets. Again, it's very difficult to draw much of a conclusion when you don't have controlled

01:15:39.500 data and you have obviously the selection biases that go into these things. But it struck me as very

01:15:45.180 interesting. Do the Prader-Willi kids, are they hyperinsulinemic? They are hyperinsulinemic to the

01:15:52.860 level that would be anticipated based on their degree of adiposity. And in some research that

01:16:01.260 we've done very recently using some of the stem cell techniques, or at least using stem cells that I

01:16:07.260 alluded to much earlier, we've got some evidence that suggests that one of the major consequences of

01:16:17.740 deleting this region of the chromosome 15 by mechanisms that are somewhat complicated

01:16:26.060 influence the expression of an enzyme called proconvertase 1. And this enzyme is critical for

01:16:35.260 for the processing of many of the hormones and neuropeptides that are made in the body.

01:16:42.060 Virtually all hormones and neuropeptides are made as pro-hormones, that is, precursor hormones

01:16:47.900 that are then processed by proconvertase 1. And that is what produces or generates the mature

01:16:57.420 form of the hormone or neuropeptide. And we think, actually, that the phenotype,

01:17:03.260 many of the phenotypes of the Prader-Willi patients may, in fact, be due to a deficiency or under-activity

01:17:11.420 of proconvertase 1, which is not in the interval where this deletion occurs, but is influenced downstream

01:17:18.860 by one or another of the genes in that interval. And it's relevant in the question of insulin, because

01:17:27.020 insulin is processed by proconvertase. So, some of the-

01:17:30.780 That's where the C-peptide gets cleaved off the pre-hormone.

01:17:33.340 That's correct. So, some of the hyperinsulinemia of the Prader-Willi might be pro-insulin.

01:17:41.660 Proinsulin has biological activity, but much lower than native insulin. And so, if this turns out to

01:17:49.660 be the case, then it might be possible to rectify some of the Prader-Willi phenotypes by

01:17:57.100 by being able to increase the activity of proconvertase 1, pharmacologically or by other means.

01:18:04.460 So, there's all this incredible work, all this incredible molecular biology and molecular

01:18:08.780 genetics that's identified leptin deficiency, leptoreceptor deficiencies, the cluster of defects

01:18:14.620 that lead to Prader-Willi. And yet, the question is, how much do these things tell us about the more

01:18:21.980 broad condition of obesity that afflicts, I don't even know the stats, I don't really keep track

01:18:27.740 of this stuff anymore, but I don't know what, a third of Americans are probably obese now?

01:18:32.060 Mm-hmm.

01:18:33.580 So, I'll tell you, an interesting perspective on that question is, and I mentioned it earlier,

01:18:40.620 while we were doing the efforts to clone these genes, I was studying patients in the clinical

01:18:47.100 research center with Jules Hirsch, Mike Rosenbaum, long-standing associates. And what we were doing

01:18:53.740 is looking at the metabolic consequences of weight reduction and showing that if you reduce the body

01:18:58.940 weight of a human by 10%, 20%, you get a reduction in energy expenditure, which is greater than what you

01:19:06.220 would predict from the loss of just body size or that they're...

01:19:10.700 In other words, their metabolic rate didn't reduce just to support the new reduced weight,

01:19:15.980 it reduced even further, which would almost try to return them to a heavier weight.

01:19:20.700 Right. So, they had a disproportionate reduction in energy expenditure, and some of the data

01:19:27.900 suggested that the major change in energy expenditure was not in resting metabolic rate,

01:19:34.460 but in the energy cost of low levels of physical activity, of muscle work, so to speak, but at very

01:19:41.820 low levels of activity. And we went on to show that, as you would expect, based on a lot of what we've

01:19:51.180 been talking about, the leptin levels were low in these individuals, low but proportional to the

01:19:56.460 reduced amount of body fat. So, one hypothesis that we had is that this reduction in energy expenditure

01:20:02.380 was, in fact, a reflection of the fact that the body was sensing the reduction in low in leptin and

01:20:10.060 interpreting that as a starvation state. That is, for that individual, that new lower body weight

01:20:16.860 represented a threat, if you will, to survival or reproduction. And we've done experiments

01:20:23.180 subsequently in which we've injected leptin into those individuals. None of them has a genetic

01:20:28.620 lesion of leptin gene. So, they're just weight-reduced.

01:20:30.700 They're just weight-reduced normal individuals. We've put leptin back into them by injection to

01:20:37.260 raise the blood level back to where it was before they lost their body weight. So, these are very low

01:20:42.540 doses of leptin. And that intervention will restore their energy expenditure back to where it was before

01:20:51.500 they lost the body weight, even though they're now still maintaining the lower body weight.

01:20:57.340 So, put this in some quantitative terms for me. That infusion of leptin gave them how many more

01:21:04.700 kcal per day in energy expenditure, all things equal between movement, activity, et cetera?

01:21:10.780 You're talking about 100 kcal a day? Like, what kind of a delta are you producing?

01:21:14.140 Two or 300, because these are big people to start with.

01:21:17.900 That's huge. And that's a huge delta.

01:21:19.260 Right. And we could show that that effect is primarily being conveyed through skeletal muscle.

01:21:25.180 So, what happens is that the muscle becomes less efficient, if you want to think about it that way,

01:21:32.060 after the leptin administration.

01:21:33.820 So, there are two ways that we broadly in the research setting measure energy expenditure,

01:21:39.340 indirect calorimetry and doubly labeled water. Can you spend a second explaining each of those?

01:21:44.700 So, indirect calorimetry is really classical physics, if you will, in which the rate of oxygen consumption

01:21:54.060 and the rate of carbon dioxide are measured in a number of ways. The most frequent is to put a mask or

01:22:02.460 hood over the head of the individual in whom the measurement is made and simply use gas sensors and flow

01:22:09.340 meters to quantify the amount of oxygen consumed and CO2 released. And from that information,

01:22:17.100 one can quite easily calculate how many kcals of energy are being expended to correlate, if you will,

01:22:25.980 to account for the amount of oxygen consumed and the CO2 produced. And by looking at the ratio of

01:22:32.220 those two, one can even get a indication of the kind of fuel that's being burned to support the

01:22:40.940 metabolic rate.

01:22:41.980 So, the V, and I'll just spit it out, so folks can, if they're, we'll link to all of this stuff,

01:22:47.180 but the RQ or the respiratory quotient is the ratio of CO2 produced or VCO2 divided by

01:22:54.620 the amount of oxygen consumed or the VO2. And that'll vary sort of between, you know,

01:22:59.500 typically about 0.7 and 1, right? Correct.

01:23:02.140 You can see it outside of those ranges. The lower it is, the more they're partitioning and drawing

01:23:07.420 from fat. The higher it is, the more they're drawing from glycogen. Correct.

01:23:11.980 And if I still remember my Weir coefficients, energy expenditure is something to the effect

01:23:16.460 of 3.94 times the VO2 plus 1.11 times the VCO2. Well, your memory is better than mine, right?

01:23:23.980 I can't recite you the Weir equation off the top of my head.

01:23:26.780 You know, it's really funny. About five years ago, I asked Kevin Hall to send me

01:23:31.100 the derivation of it, and it was so great to get this page, two pages of differential equations,

01:23:38.620 so I could go through and actually see how the Weir coefficients were described.

01:23:43.100 Which, of course, then led to the inevitable question that you and I have talked about in

01:23:45.980 the past, which is, do those coefficients hold in the presence of radical dietary change,

01:23:52.700 such as, you know, a very, very low-fat, high-carbohydrate diet, or the opposite.

01:23:57.660 So then, if that's how indirect calorimetry works, and of course, by the way, these things happen in

01:24:02.700 metabolic chambers. That's the ultimate way to do it, right? Where you can put a person into a room,

01:24:06.380 and the room itself has all of these sensors embedded in it.

01:24:09.980 Right. And that's exactly the same principle, but there, the individual's not restricted in

01:24:14.780 terms of their motion, other than in the room that they're in. So you can get exactly the same

01:24:20.380 information, but over a much longer period of time. I mean, you can do it for days.

01:24:24.060 There was a point when I wanted to build one of these at my home. I was so obsessed with knowing

01:24:29.820 every... I wanted to know, like, my VO2 and VCO2 almost every minute of the day. And then I found

01:24:35.820 out it would cost at least five million to build one and realized I couldn't build one. But I thought,

01:24:39.420 boy, if I ever make enough money to build one of those, that's a great fun... That's like,

01:24:45.100 that's fun for the whole family, if you really stop to think about it, you know? I mean, who wouldn't

01:24:49.020 want to go sit in the metabolic chamber on the weekend, just kicking it, watching football?

01:24:53.660 We just built one of these actually at Columbia, and it's quite a... You're right. It's not only

01:24:59.900 expensive, but requires a great deal of attention to things like leaks and airflow and...

01:25:06.700 You know, I broke one once.

01:25:09.180 Doing what?

01:25:10.860 So you'll recall, I used to spend time in these chambers, and I guess I exceeded the level of

01:25:15.980 CO2 production that it was sort of calibrated for because I was on my bicycle, and I was riding

01:25:20.540 pretty vigorously, and the CO2 sensor broke. And it's one of those things where you don't get a big

01:25:25.500 red flag. It was just after the fact that data didn't make sense. And this is where you appreciate

01:25:29.500 what you're saying, which is the engineering in this thing is amazing. And so you got to pour through,

01:25:34.460 you know, the longest spreadsheets in the world to start to look for where the mistakes are.

01:25:38.780 But it's really beautiful stuff. And there's not many of these in the country. Like, there's...

01:25:43.260 Obviously, Columbia has some. NIH has some. TRI has some. Pennington has some. Baylor has some.

01:25:50.460 I mean, there are not many.

01:25:52.780 Like you said, they're tricky to construct and require high level of attention to their

01:25:58.700 maintenance and operation. It's not just like going down in your basement and measuring your RQ.

01:26:05.340 So how does doubly labeled water work, which I also have had the privilege of ingesting?

01:26:10.540 So doubly labeled water is a technique which uses two isotopes of water, which is H2O.

01:26:20.220 In one, the oxygen is a heavier version of the normal or the more prevalent oxygen molecule. And the

01:26:29.660 other, the hydrogen is a heavy atom as opposed to the more prevalent kind of hydrogen in the environment.

01:26:39.740 And what you do is you can give these as a mixture. You have to know exactly how much of each one you've

01:26:45.820 given. But you can give this as a drink, basically, of what amounts to heavy isotope labeled water.

01:26:53.500 And then look at the relative concentration of these two isotopes. You can do it in blood. You can do it in

01:27:02.540 urine. We often do it in both. And the difference in the rate of excretion of the so-called O18 water, the heavy isotope of water,

01:27:13.180 which is oxygen is borne out both in urine, but also in the expired air of an individual as carbon dioxide.

01:27:23.020 And the H2O, which comes out by non-respiratory means, you can actually get a measure of how much CO2

01:27:32.940 the individual is producing. And you can use that and the nature of the diet that the individual is on

01:27:42.540 to back-calculate from what would be the RQ back to the oxygen consumption.

01:27:48.220 So you would use the FQ, basically.

01:27:51.000 Yes, correct.

01:27:51.380 So the FQ is if you stuck the person's food into a bomb calorimeter, burned it, that would produce the RQ.

01:27:58.140 So the limitation of this is a couple things. One, the patient's got to be weight stable, right? This

01:28:05.180 starts to fall flat when they're losing or gaining tremendous amounts of weight. And two, you have to

01:28:10.680 have a pretty darn good idea of what they're eating. So each of these methods, ingenious and completely

01:28:16.100 distinct, both have huge limitations. The former, of course, requires an artificial environment. They

01:28:22.820 don't get to be free living. And you can't really study them for that long. But the latter, where they

01:28:28.600 can be free living and you can study them for, you know, two weeks at a time with a single ingestion

01:28:33.680 of the doubly labeled water, you better make sure that they're not gaining and losing weight. And you

01:28:38.380 better make sure you know exactly what they're eating. And in an ideal world, it better be the same

01:28:42.440 thing almost every day, right?

01:28:44.060 And in general, because of the issues that you just mentioned, I think most investigators in

01:28:52.620 this field prefer, at least in terms of its nominal accuracy, the room calorimeter over

01:29:00.740 the doubly labeled water. But the doubly labeled water does have the virtue, as you pointed out,

01:29:06.580 of being applicable over much longer periods of time so that you can get integrated idea of

01:29:13.540 how much energy an individual is expending over literally days or even weeks. What's sometimes

01:29:20.800 done is to re-dose the individual at some point in the experiment and simply continue it.

01:29:27.600 So when I did this, I did several days in several chambers and then separately did the LW.

01:29:34.500 I was kind of amazed at how closely they overlapped. Of course, it was hard to replicate my chamber

01:29:40.520 activity in the real world because there's so much more, you know, movement outside of the chamber,

01:29:46.500 but they were surprisingly in line, which is interesting because at the time I was on a

01:29:50.900 ketogenic diet, which there's a lot of things that could be changing there. So I want to go back to

01:29:57.080 something that we kind of got into a little bit, but it's, to me, it's one of the most fascinating

01:30:00.940 discussions slash debates, which is the idea of appetite being regulated centrally versus peripherally.

01:30:10.360 And you and I have had some incredible dinners having these discussions. And I've always found

01:30:17.260 your perspectives to be interesting because I think few people are more qualified to talk about

01:30:23.480 that in that they know so much about both. I mean, in many ways, leptin at its inception was really

01:30:31.040 viewed as a peripheral way to regulate appetite. So let me explain what I mean by that for the listener.

01:30:36.500 When you're hungry, are you, is this all being driven by what your brain is saying?

01:30:42.260 Is it possible that somewhere outside of the brain, which we refer to as the periphery, your liver,

01:30:47.500 your fat cells, your muscles, the level of blood glucose that's circulating, is there some other

01:30:52.720 signal? You know, it's kind of interesting that here we are in the year 2018 and it's still not

01:30:59.200 entirely clear where appetite is regulated. So how has your thinking on this topic evolved?

01:31:06.500 through your own immense knowledge of both the central and peripheral components of appetite.

01:31:13.760 You want to take a sip of that Topo Chico before you try to answer that? For the listener,

01:31:17.840 I've introduced Rudy to Topo Chico today. So he's, one sip into it, he declared it the finest bottled

01:31:23.000 water. I would describe the system, you know, from high altitude as the following, that there are

01:31:29.580 CNS, central nervous system mechanisms. They're not all in the hypothalamus, but many of them are. And

01:31:37.600 many of those that aren't do interact with the hypothalamus. And that that can be regarded,

01:31:43.960 that organ or set of cells can be regarded as the integrating center for basically the reception of

01:31:54.200 signals that are of relevance to how an organism ought to respond to its environment in terms of

01:32:01.380 food intake and also for control of some of the aspects of energy expenditure that are not necessarily

01:32:10.880 under voluntary control, such as autonomic nervous system. And what's coming into that system

01:32:17.440 is, I think, broader than we had originally anticipated. So clearly, leptin is an example of

01:32:26.280 a hormone that's secreted from adipose tissue. But there are many other peptides, actually,

01:32:32.080 or hormones that come out of adipose tissue. There are neural signals that are now being identified.

01:32:40.120 That is, nerve-borne signals that look like they're coming out of and going into adipose tissue that may be

01:32:46.820 playing a much more important role than we had sort of previously appreciated. The gastrointestinal

01:32:53.220 tract, which for obvious reasons is a very relevant aspect of the system, originally was, I think,

01:33:01.680 regarded pretty much the way that adipose tissue was, as a tube that absorbed your calories and didn't

01:33:08.160 really participate actively in the regulation of that aspect of biology. Clearly, the advent of

01:33:16.820 various surgeries for control of body weight have indicated that that sort of premise is

01:33:23.620 incorrect. That is, that there are very powerful influences of the gastrointestinal tract, which are

01:33:29.020 brought out by some of the surgeries that are done. And it's now very clear that the gastrointestinal

01:33:35.380 tract, at many levels, is communicating with the brain, both by products that it secretes and by direct

01:33:44.320 neural input, probably primarily, but not entirely via the vagus nerve into the brainstem and then

01:33:51.420 further up into the brain. So just, you know, taking that as a sort of very broad picture,

01:34:00.180 it's much clearer now that there is no sort of single definitive part of the biology of an individual

01:34:10.500 which is dictating this very complex system. It's really a product of the interaction of these

01:34:16.300 peripheral signals, as you referred to them, including metabolites, such as glucose and free

01:34:22.400 fatty acids and so forth, and the central nervous system, not just the hypothalamus, but areas like

01:34:29.660 the brainstem, the amygdala, the frontal cortex. There's many other areas of the brain that interact with

01:34:35.320 these signals to influence both the behaviors and the unconscious, if you will, the vegetative

01:34:42.680 responses that ultimately go into the regulation of body weight. So I think it's obviously a very

01:34:50.980 complex and overdetermined system. And it's that way because it is absolutely critical to survival.

01:34:59.440 So it's not surprising in a way that it's as complex as it is. But I think what it clearly does is it

01:35:06.680 introduces all sorts of possibilities for mischief with regard to body weight regulation, starting in

01:35:14.940 the frontal cortex where decisions are made, at least with regard to some aspects of this, all the way down

01:35:21.180 into the colon where you've got large numbers of bacteria that are producing things, metabolites and

01:35:28.340 molecules themselves that, again, we're only just beginning to understand. So we're not only talking

01:35:35.140 about a regulatory system that has all of these elements, but a bunch of organisms literally living

01:35:40.900 within us that are influencing it in ways that I don't think we've really fully understood. The

01:35:46.940 miracle is actually when you think about all the ways that it could go wrong, that it doesn't go wrong

01:35:53.640 anymore than it actually does. And it looks to me like the major problem in terms of the obesity that

01:36:03.180 you referred to earlier that's increasing has to do a great deal with the environment and what people

01:36:09.740 are being exposed to that's very novel with regard to the system. I mean, this system was not designed

01:36:16.840 for the regulation of metabolism and body weight in an environment in which, with a cell phone, you can

01:36:24.200 bring literally as many calories as you want into your living room without getting up off your behind.

01:36:32.940 So we're dealing with an organism which has this very complex and beautiful system designed to regulate

01:36:39.420 various aspects of the biology, which is now in an environment which is entirely, at least in many

01:36:46.440 regards, novel to the system. And it fails in the sense that there's a lot more obesity than there was

01:36:55.240 prior to the environment that we've been able to create. But it's not totally dysregulated. I think people

01:37:04.400 are just now regulating their body weights at somewhat higher levels than they were in other

01:37:10.700 circumstances.

01:37:12.480 Again, I don't follow the statistics. It's not something I'm paying huge attention to. But

01:37:16.840 are the rates of obesity plateauing or are they still increasing? Or is the curve concave up or concave down?

01:37:24.140 So several years ago, it looked like the rates were either slowing down or level. Now it looks like maybe

01:37:32.000 they're continuing to inch up, but not as rapidly as they were, say, a decade ago. So there's been

01:37:38.040 some relenting of the rate of increase, excuse me, in obesity in both adults and children. But it

01:37:45.960 looks like it's still rising, although at a lower rate.

01:37:50.420 Do you have concern that there are epigenetic implications of this, that 10,000 years ago,

01:37:57.880 the prevalence of obesity, again, largely probably driven by the environment that made it pretty

01:38:04.800 hard to become obese? Not just in terms of food availability, in terms of absolute calories,

01:38:09.940 but presumably the types of calories that were available. I mean, I think my recollection is that

01:38:14.720 10,000 years ago, nothing came in a package, right?

01:38:17.600 As far as we know.

01:38:18.660 So we didn't have Pop-Tarts, we didn't have breakfast cereal, we didn't have orange juice,

01:38:26.800 we didn't have French fries and whatever else are the culprit potentials. So if Johnny got a set of

01:38:34.160 genes that were slightly suboptimal, Johnny Jr. probably got some of those suboptimal genes, but

01:38:41.460 there was nothing being imprinted on the genome that was being transmitted to generations. And

01:38:48.620 I mean, what do you think about this idea that we could be reaching kind of a dangerous point

01:38:54.600 in the evolution of our species where children, maybe during critical windows of development,

01:39:00.800 if exposed to, you know, again, in my opinion, it's probably a lot of the sort of really highly

01:39:06.660 refined crap, high amounts of fructose that we're seeing with the NAFLD, that this is setting them up

01:39:12.480 for a real problem later in life. It's going to make it a lot harder, not just for them to lose

01:39:17.260 weight one day, but also for their offspring. Is there any evidence of that?

01:39:21.940 I think there is evidence that's, I'm not so sure I would use the term imprinting. That has

01:39:29.020 sort of specific biological meaning and implications. I mean, we could talk about that in a minute, but

01:39:35.800 if what you're saying is, are there perhaps critical periods of development in a child in which if the

01:39:47.180 child is caused to become more obese than he or she might be in some other environment, that that will

01:39:54.960 leave a mark on the regulatory system that will then cause that individual to want to sustain or

01:40:01.900 maintain a higher body weight? I think the answer is probably, or at least possibly yes. That is that

01:40:09.760 there may be critical periods in human development. Certainly, I think the data are also available to

01:40:19.520 some extent in animal studies that suggest that if you manipulate body weight or some of these systems

01:40:27.060 at critical periods of development, you end up with an animal which is more likely to be obese as an

01:40:34.940 adult than if you hadn't done that. And exactly what the molecular and neuroanatomical or other

01:40:43.580 physiological consequences of that manipulation are that lead to the maintenance of a higher body weight,

01:40:51.780 this is obviously a critical area of study. It's actually something that people in my lab are quite

01:40:58.940 interested in. That is whether we can impose by dietary or other manipulations a higher body weight

01:41:06.500 or a higher sustained body weight in an animal by early manipulations, either by diet or endocrine

01:41:13.960 manipulations. And I think the answer is probably yes. But what is the consequence of that in terms of the

01:41:23.120 regulatory elements that we were talking about? We don't yet know. Another aspect or another perspective

01:41:31.520 on this is the following question. If you manipulate the metabolism of a pregnant animal or human,

01:41:40.540 what are the consequences of that on the developing brain or physiology of the fetus?

01:41:48.200 I mean, there's evidence now that a mother who is hyperinsulinemic or has gestational diabetes is

01:41:54.700 increasing the child's risk of obesity. Is that not the case? There are data that suggests that-

01:41:59.860 It's registry data, so it's hard to know. Right. But that obesity in a pregnant woman does confer

01:42:08.020 by, again, by mechanisms that we don't understand, higher risk of obesity in her offspring.

01:42:16.240 Although I'd love to see it stratified by hyperinsulinemic versus non-hyperinsulinemic obesity.

01:42:21.740 Because I feel like I remember reading that it also seems to be dependent on the gestational stage.

01:42:28.280 In other words, like when the dorsal and ventral buds of the pancreas are forming,

01:42:32.640 which I can't even remember when that is, maybe week seven, eight, nine, something like that.

01:42:36.060 Those are very critical windows, right? That's when islet cell formation is taking place and

01:42:41.560 nutrient exposure or insulin exposure during that period of time could have a more profound effect

01:42:46.520 than say late in the third trimester where perhaps it's not as effective. Again, very, I mean, I'm just,

01:42:52.680 I don't follow this literature, so I don't want to sound like a complete moron. And the last time I

01:42:58.080 really paid attention to this stuff was probably four years ago. And it was, again, largely registry-based

01:43:02.880 data, so very difficult to know cause versus effect.

01:43:07.500 Again, you know, the literature, as you sort of implied, is wobbly on this point. But I'll just

01:43:14.900 tell you about a couple of experiments, actually, that we've done. One to answer, to address this

01:43:20.200 question of hyperinsulinemia in a dam, in a pregnant animal, and its effects on the fetus.

01:43:27.520 We were able to do an experiment in which we were interested in separating the issue of

01:43:34.140 high body weight and adiposity from the effect of insulin itself. And so we actually used a genetic

01:43:42.480 model of hyperinsulinemia and were able to make the dams hyperinsulinemic without being obese.

01:43:48.300 So these are perfectly normal body weight dams that were very hyperinsulinemic. And the pups that

01:43:55.720 they were carrying, some of those pups had no abnormality of any insulin-related genes. So they

01:44:02.620 were what we would call wild-type pups being incubated in a hyperinsulinemic dam. And we were very

01:44:08.980 interested to see what the consequences would be on the metabolism and body weight of these animals.

01:44:14.200 And very interestingly, these animals showed transient elevations of body weight and body fat

01:44:24.700 when they were in what I guess we would describe as adolescent period of a mouse, which tended to

01:44:31.980 revert back to normal over time. You could definitely see an effect in the body weight and metabolism of

01:44:39.560 the mouse, but it appeared to be transient. And to be clear, the mother was euglycemic?

01:44:46.040 The mother was euglycemic and hyperinsulinemic. That's a tough needle to thread, my friend.

01:44:51.040 Yes. So again, it's the power of genetics. So we could do that. So these kind of experiments,

01:44:55.760 we also looked at the brains of these animals, and there were some very subtle differences in the

01:45:00.480 way the hypothalamic cells, the numbers of different types of hypothalamic cells, which suggested

01:45:05.940 in sort of support of the inference of your implication of your question, there are consequences

01:45:14.180 of these metabolic states of the dam that can influence the subsequent development of the pup,

01:45:21.320 even if the pup doesn't have the genetic etiology that led to the derangement, if you want to call

01:45:27.980 it that, in the dam. We've more recently been doing some experiments along these lines looking at

01:45:33.500 leptin in a similar way, and again, can see evidence of influence, long-term influence of

01:45:40.060 manipulations of leptin in this way, which again suggests that these transient periods of exposure

01:45:46.640 at various levels or various periods of development can have long-term consequences for organisms that

01:45:54.840 don't themselves have any primary abnormality of these genes or their functions.

01:46:01.720 So I think this probably supports the notion that there are things going on in the environment

01:46:08.080 that are not reversible once the environment is necessarily reversed. That is, the exposure to

01:46:15.040 the environment may change the biology in such a way that even when the insult, if you want to call

01:46:21.780 it that, is removed, the biology is permanently impacted.

01:46:27.120 We've spent a long time talking about low-carbohydrate diets. You were part of one of the

01:46:33.620 studies that was funded by NUSI, and while the study that you were a major part of was not a diet

01:46:40.320 study, so I wasn't really looking at diets, it used diets as a tool to ask questions. Clinically,

01:46:46.240 we know that low-carbohydrate diets are very effective. I think it would be very difficult for

01:46:50.940 people to dispute that at this point in time. The interesting question, if you're rudely liable,

01:46:57.580 is why? What do you think? Why do these diets seem so effective for so many people? Let's not even

01:47:04.700 get into the fancy stuff about the diabetes reversal and stuff like that, where I think

01:47:08.560 the results are especially pronounced, but just in terms of obesity reduction, why do you think these

01:47:13.400 things work? I guess my view of this is that to the extent that diet composition has an effect on

01:47:21.460 body weight, which clearly it can, that this is the result of the interaction of the components of

01:47:33.000 the diet with aspects of the regulatory control of food intake. I guess the way I describe this is

01:47:41.220 the sort of hedonic aspects of the food that's being ingested will influence the drive to ingest that

01:47:51.680 particular food. I think individuals differ in terms of their sensitivity or susceptibility to diets of

01:47:58.360 different composition, and there clearly are some individuals who, when fed or offered or impose

01:48:07.360 diets that have pretty radical compositional differences, will respond very well. In other

01:48:15.620 words, there are people who respond well to high-carb diets, there are people who respond well to low-carb

01:48:20.740 diets. I think that it's primarily some aspect of the hedonics of these diets, if you want to call it

01:48:29.720 that, that's driving the favorable response. I know, and I'm sure you and I have discussed this many, many

01:48:37.260 times, there are individuals who view the diet composition as having, in addition to what effects

01:48:44.160 I just described, also an impact on energy homeostasis itself, meaning that the composition of the diet can

01:48:51.900 influence the degree of caloric expenditure per unit of calories ingested. I'm not a sort of strong

01:49:01.820 proponent of that view. I guess I try to remain agnostic about it, but my bias, if you want to call

01:49:09.340 it that, from my experience is that that's right. Diet composition can have an effect on body weight. It

01:49:15.840 doesn't do so by effects primarily, or maybe at all, on energy expenditure per se, but can have pretty

01:49:24.540 striking effects on an individual's desire to eat those calories delivered in that form.

01:49:31.580 And so, there's two ways to think about that, right? Again, if you leave the energy expenditure side

01:49:36.140 away, on the intake side, is it the case that a low-carbohydrate diet, as you say, it's purely a hedonic

01:49:43.020 issue. People just, when they're on these diets, they're just not as hungry because, I don't know,

01:49:47.280 the food is not as diverse or not as palatable. An alternative viewpoint is they eat a lot less

01:49:52.200 because they're eating themselves more. In other words, they're, you know, the way I try to explain

01:49:56.220 this to my patients when we're talking about nutrition is, you want to differentiate between

01:50:00.960 exogenous and endogenous calories. So, you know, most people walk around, if they're weight stable,

01:50:06.640 purely subsiding on exogenous calories. But if you want to lose weight, by definition,

01:50:11.340 you must start to consume endogenous calories. You have to start eating yourself.

01:50:16.480 And this gets back to the question of the regulation of body weight peripherally. If you

01:50:23.620 are eating yourself, can the CNS figure that out and say, hey, we've got all this lipolysis going on.

01:50:30.620 We're liberating all of this fat out of a fat cell. That's energy. Fantastic. I will program the

01:50:38.180 system. You need less exogenously. So those are two kind of very different ideas, right? The one is

01:50:43.680 just, hey, this food is boring and doesn't taste great. I'm just going to eat less of it. Personally,

01:50:48.540 I don't find that theory as appealing because having worked with so many patients that do this,

01:50:53.000 if that were the case, I'd suspect they'd be more hungry, but yet they don't seem to be very hungry,

01:50:58.320 which is, I think for most people, the only thing that's intolerable over long periods of time is

01:51:04.540 hunger. You know, you look back at the Ancel Keys starvation experiments and those guys weren't

01:51:09.700 really like, you know, what they were eating, like what, maybe 1600 calories a day, but they were

01:51:13.900 losing their minds, right? I mean, these guys lost their minds on 1600 calories per day. So

01:51:20.820 starvation is clearly an unacceptable state in the long run. I got to be honest with you. I'm glad I

01:51:27.080 don't work on this problem. I'm much more interested in just trying to figure out this other easy

01:51:31.100 problem of longevity. But this obesity thing is a real bear. Because at the same time, as much

01:51:37.460 success as I've seen for people on low carbohydrate diets, I also know that they don't work for

01:51:41.360 everybody. And therein lies the challenge of why? Why would something, I mean, again, it's a naive

01:51:50.520 question in the sense that you've just spent the last two hours explaining some of the most complex

01:51:55.480 biology of obesity. Why should everyone respond to the same treatment? But it does. It really frustrates

01:52:04.020 me that there are certain patients in which no matter what one does or no matter what they do,

01:52:09.580 you know, I think patients that try really, really hard and can't lose weight, I find that to be a very

01:52:14.420 frustrating clinical problem. So I guess maybe I should rephrase the way I answered your question.

01:52:21.460 And that is to say that I believe that the composition of the diet can influence aspects of

01:52:29.100 ingestive behavior. I emphasize the hedonic aspect, but I'm certainly open to the possibility

01:52:37.560 that if you feed a diet of, you know, divergent composition, you will definitely have some influence.

01:52:46.540 I mean, this is well documented on the substrates that are in the circulation as a result of feeding

01:52:52.440 that diet. Those substrates themselves, metabolites, you were mentioning fatty acid, but many other

01:52:59.020 things change, may themselves have a effect on the drive to eat. I'm sympathetic to that idea.

01:53:07.720 And exactly in any given individual who does or doesn't respond to a specific diet, which of the

01:53:14.380 mechanisms, you know, whether it's occurring at the level of the taste of the diet or some of the

01:53:21.960 metabolic consequences of the diet, I would say to the extent that somebody is successful in

01:53:27.840 losing weight on that diet, A, they have to be in negative energy balance. That is, there's no way that

01:53:33.860 they can eat the number of calories that they're expending and lose weight. So they need to be in

01:53:39.960 negative energy balance. And I mean, that's just a given. And then the mechanism by which the diet

01:53:46.460 composition influences the success of their ability to sustain a hypocaloric state, which as you pointed

01:53:53.400 out, is uncomfortable. I'm open to the possibility that it could be at any level of the sort of neural

01:54:01.920 circuitry that regulates food intake. It could be very proximal in terms of taste aspects, but it could

01:54:10.040 also be some of the mechanisms that you mentioned. I think that's perfectly reasonable. How that works,

01:54:18.100 as you pointed out, is a tough nut to figure out. And I think, like you said, there are individuals who

01:54:26.380 respond very well to diets of quite divergent composition, but they have to be taking less

01:54:33.240 calories than they are spending. Right. And that's an easy thing to do mathematically. It's a very hard

01:54:39.760 thing to do clinically because, again, I think patients can tolerate a lot, but I don't think

01:54:45.500 people can tolerate hunger for very long. I think to ask somebody to be constantly hungry

01:54:51.020 is a losing proposition. And that's why I believe clinically low carbohydrate diets seem to be more

01:54:59.000 satiating. And again, whether they're satiating because of something in their composition or whether

01:55:05.420 they're just satiating because you end up eating more of yourself. I mean, I've seen lots of patients

01:55:10.000 who, and this has even been published, who on 16, 1700 calories a day of a ketogenic diet seem

01:55:16.520 completely fine. And yet Ancel Keys' patients at 1,400, 1,500, 1,600, 700 calories of basically

01:55:22.640 very high carbohydrate diet, very low fat diets, lost their minds. What did you believe 10 years

01:55:29.860 ago that you don't believe today to be true? I think over the past decade, the field of the

01:55:37.280 sort of neuroscience of body weight regulation or ingested behavior and energy expenditure has expanded

01:55:44.040 the role of other elements in the central nervous system beyond what began as our very intense

01:55:52.680 focus on the hypothalamus. Partly that's historical and for reasons having to do with the tractability

01:55:58.920 of some of the cell types and mechanisms in the hypothalamus. But I would say the biggest change

01:56:04.760 in my thinking about this has been that these supra-hypothalamic aspects of the central nervous

01:56:12.660 system, again, starting with the cortex and working down, have a much stronger, I think,

01:56:19.500 ultimate impact on this regulatory system than the way I perceived it a decade ago. And what this does

01:56:27.000 is it opens up, I think, a lot of very interesting aspects of the biology that maybe we weren't either

01:56:34.560 capable of or didn't focus on adequately, part of which actually is in response to the point you were

01:56:40.840 just raising. That is, what is the effect of diet composition on hunger and so forth? It's almost

01:56:47.480 certainly an aspect of this sort of view of the broader central nervous system impact on ingested

01:56:55.700 behavior than just the hypothalamus. You know, my mentor, Jules Hirsch, I think, who unfortunately

01:57:02.820 passed away. He passed away about four years ago? Yeah, three years ago. He would agree with this. I

01:57:08.080 mean, he always, I think, was suspicious of focusing, you know, slavishly, if you will, on the hypothalamus

01:57:16.760 as the center for all of this. He always, I think, had a much broader perspective on some of the

01:57:23.180 psycho-affective aspects, as he would describe them, of ingestive behavior. And I think he would see the

01:57:30.160 sort of flowering of broader view of some of these regulatory aspects of food intake to other parts

01:57:37.600 of the central nervous system. He would say, I told you so. And he was right. I think he's looking

01:57:43.360 down at you saying that. Yeah, he's probably saying a lot of other stuff too. If we could wave a magic

01:57:50.200 wand and not only were resources unlimited, but more importantly, any of the technical considerations

01:57:56.600 that make this type of study challenging were just ameliorated, what's the dream experiment you

01:58:02.740 would want to do? If you got one more shot at this, you could spend the rest of your career

01:58:07.260 doing a dream experiment with infinite resources, infinite subject participation. Screw the ethics.

01:58:14.800 There's no IRB. You've got a bunch of clones out of Westworld. You can do whatever the hell you want.

01:58:19.420 What experiment do you want to do? Once to take your final kick at this problem.

01:58:26.600 There is a gene that was identified now 10, 12 years ago.

01:58:33.580 FTO?

01:58:34.360 Yes.

01:58:34.880 You tested me for this.

01:58:36.260 Yes.

01:58:37.600 I was negative.

01:58:39.200 Right. It was identified by Mark McCarthy and some other investigators in Great Britain

01:58:44.060 as sending the strongest genetic signal for obesity or elevated body fat, not severe effect,

01:58:52.980 but the strongest statistical signal ever detected in humans. It remains the same.

01:58:59.880 The prevalence of the variants that are associated with higher body fat are very high in the general

01:59:06.180 population. So there are a lot of these variants of this gene that are contributing to

01:59:13.500 human obesity, not of the severity, certainly, of the OB or the monocortin-4 receptor.

01:59:21.480 There are various schools of thought about how this gene is doing it. The interesting thing is

01:59:26.480 that the variation in the gene that is conferring the risk of obesity is variation in what we call

01:59:34.260 non-coding part of the genome. So as people will know, a small fraction of the genome,

01:59:41.020 2% or 3% actually encodes the proteins that are made that we've been talking about, actually.

01:59:48.620 The rest of it is regulatory parts and elements that we don't understand very well.

01:59:54.340 The variants that are implicated for the FTO gene actually occur in the non-coding region.

02:00:00.880 We know exactly where they are. It's not a mystery where they are, but they're in non-coding. They're in

02:00:07.180 the first intron, so to speak, of this gene. The mechanism by which this very, very common variant

02:00:14.820 leads to subtle increases in body fatness in a large number of individuals. To me, this is a very

02:00:23.700 intriguing question. There are some that view the effect as being conveyed through what's called

02:00:30.800 browning of adipose tissue, increasing the energy expenditure of adipose tissue. There are others

02:00:37.220 like myself who think it's affecting central nervous system circuits that affect food intake,

02:00:43.680 but the mechanism, at least in terms of the central nervous system, is really unknown. The primary

02:00:50.780 phenotype that's seen in these individuals who do or don't have what we call the risk variance is food

02:00:57.300 intake. I mean, if you measure their food intake very carefully in a lab, you can see that the

02:01:02.820 individuals with the variants will eat slightly more. They actually may have a slight preference

02:01:08.080 for fattier, higher fat foods than individuals who don't, but what exactly is going on in the central

02:01:16.320 nervous system is not clear. They have actually, in one study, pretty characteristic functional magnetic

02:01:24.100 resonance imaging differences among individuals who do or don't have this variant, but the mechanism

02:01:31.540 is unknown. And this is something which has intrigued me now for eight or ten years. That's the experiment I

02:01:38.740 would love to do. I mean, we're actually trying to do it. Tell me specifically what the actual

02:01:42.700 experiment is, but before you do that, I want to make sure I'm, and this is going to be an oversimplified

02:01:47.400 analogy, but what you're basically saying is there's a genetic marker effectively that creates or

02:01:53.820 identifies those who are predisposed in a high nutrient environment to overeat. So even though our

02:02:01.240 ancestors would have had these genes, the reason, or nevermind our ancestors, even though a hundred years

02:02:06.220 ago or 50 years ago, this gene still existed, probably at the same prevalence, you didn't have the food

02:02:11.660 environment to pour fuel on that fire. Now, just to push back a little bit, isn't that sort of like

02:02:18.180 saying, we've got all these people getting lung cancer. It only started when they started smoking

02:02:23.560 and, you know, only one in 10 of these smokers are getting lung cancer. There's a gene that might be

02:02:29.160 predisposing them to lung cancer. So you're, you're basically saying, I want to look at the problem of

02:02:34.680 why are certain people susceptible to an environmental trigger versus what's the environmental trigger?

02:02:41.620 In the case of smoking, it was so obvious. It was the goddamn cigarettes. In the case of obesity,

02:02:46.820 it's a little more complicated.

02:02:48.100 Correct.

02:02:48.580 So what's the experiment you would do?

02:02:50.520 So the experiment that I would do, and actually we're trying to do these now, is to actually look

02:02:57.160 in animals in whom the variants of the genes that we think are affected by this, because even though

02:03:06.680 the intronic region is not encoding anything, we think it's affecting genes that do encode something.

02:03:14.040 And we think we know at least what one of those genes is. And what I would like to see is what the

02:03:19.540 consequences of these variants are on the functional activity of the system that we think is mediating the

02:03:30.700 effects of these genes. Because I, again, for reasons that are quite technical, I think that one of the

02:03:38.620 major impacts of this gene or these variants in this gene is actually on the structure of the nervous

02:03:46.540 system. So I think their impact is occurring early in the development of the brain. And that what it's

02:03:54.540 doing is changing the circuitry slightly in a very maybe subtle way, probably subtle way, that favors

02:04:02.860 the increased food intake that you just described. And the implication of this is very relevant to some

02:04:13.640 of the things we were just talking about earlier. That is, can the environment impose permanent

02:04:20.680 signatures on the feeding circuits on the feeding circuits, so to speak. And this is, I think, a prime

02:04:27.120 candidate in that regard. And the virtue of the FTO study that we've been talking about is that we know

02:04:34.680 at least enough about the genetics and some of the mechanisms that we can design the experiments primarily,

02:04:43.560 but not totally, but not totally in mice to be able to look precisely at where the changes are occurring.

02:04:51.480 We now just coming online are the tools to look at the circuits as they are acting physiologically. We can

02:04:59.800 activate these circuits in the brain of an animal by using various neurophysiological and neurogenetic

02:05:07.720 techniques that will, I think, permit us to see where the structural consequences of these variants

02:05:15.400 reside. And my guess is that that would give us an insight into the sort of prevalent genetic

02:05:25.160 susceptibility or one of the prevalent genetic susceptibility.

02:05:27.560 What would we do with that information? Let's assume you had all of that information. How does that

02:05:32.440 take obesity from 35% to 5%? Well, you're asking somebody who is not

02:05:38.760 an epidemiologist. And I, you know, some of us do what we know how to do.

02:05:43.320 You're a hammer, you look for a nail.

02:05:45.000 You're a hammer, you look for a nail. So I'm a hammer looking for the nail of whether or not

02:05:49.560 I can show a gene which alters the structure of the central nervous system in a developing organism

02:05:58.120 in such a way as to make it susceptible to the environment. The animals that we have studied

02:06:03.960 where we've manipulated this gene definitely are sensitive to the environment in the sense that they

02:06:12.280 will choose or eat more of a high-fat diet than a normal or, you know, a regular diet.

02:06:19.000 How many such SNPs are there in humans?

02:06:21.400 There are at least a half a dozen SNPs in the first intron.

02:06:26.760 In the first intron alone.

02:06:28.040 And, but that's where they are.

02:06:29.400 That's where they all are.

02:06:30.120 That's where they all are.

02:06:31.000 And what is the prevalence in, to the best of our knowledge,

02:06:35.320 of people today who have at least one of these SNPs in that first intron?

02:06:40.520 60%.

02:06:42.280 So 60% of the population would be at least somewhat susceptible to obesity.

02:06:47.960 Yeah. Which is, you know, sort of fits with what we see because the effect size of this genetic

02:06:55.000 variation is not on or off. It just widens, I think, the, or increases the susceptibility

02:07:03.080 to whatever it is in the environment that's doing this.

02:07:05.800 Are there pharmacologic things that could be done to combat this?

02:07:09.240 Well, we don't know what it is, so we don't know.

02:07:11.320 Because we don't know what the, we don't know what the approach,

02:07:13.000 it's so much more complicated because it's non-coding.

02:07:15.080 Correct. But that's what fascinates me about it. And I think it's producing

02:07:19.880 something in the central nervous system which is structural in consequence, at least in part.

02:07:25.000 And that's why I'm fascinated by it. Because what this means is that if you have this

02:07:30.600 predisposition, it may change the nature of your central nervous system structurally in such a way

02:07:37.640 as to make you either more or less.

02:07:39.560 So if you take just hypothetically an animal that's born with these introns,

02:07:44.760 with this pattern of introns, and you could magically use CRISPR and you could delete them

02:07:49.320 and edit them and make them a wild type again, do you fix the phenotype? Because if not, that would

02:07:55.160 certainly point to something early on in neurologic development, right?

02:07:59.800 Yes, you do. So we can do that kind of thing.

02:08:02.760 You've done that experiment?

02:08:03.640 Yeah. And we've also done experiment.

02:08:06.040 And sorry, it did or did not fix the phenotype?

02:08:07.960 It fixes it.

02:08:08.680 Okay, so that means that it has an influence that transcends development, right?

02:08:13.320 That it's part of development. I mean, because the fix occurs at the level of the fertilization of the...

02:08:20.600 Oh, no, you've never fixed it post.

02:08:23.320 Not fixed it post.

02:08:24.600 Okay, okay.

02:08:25.080 Right. So, Rudy, you're never going to stop working, are you?

02:08:28.120 All right. Well, if I got to figure this out, I'll have to stay at it for a little bit longer.

02:08:32.760 There are a lot of other things I want to talk with you about. Maybe we'll just spend a couple

02:08:36.040 minutes. There's something else I want to talk about, which is the most complicated topic ever,

02:08:41.720 insulin resistance. What the hell does insulin resistance mean? How can an insulin resistant

02:08:48.280 person be fat when, by definition, insulin needs to tell the adipocyte to take up the free fatty

02:08:55.320 acid as triglyceride? So does insulin resistance mean insulin resistance of the muscle, insulin

02:09:01.960 resistance of the liver, but insulin sensitivity of the adipocyte? What does that mean?

02:09:07.320 The metabolic disposition of glucose, for example, is obviously, or maybe not so obviously,

02:09:15.080 it's not primarily via adipose tissue. I mean, glucose, when it gets into the circulation and is

02:09:21.240 removed or taken up. It's mostly in the muscle. Yes. It's mostly in the muscle. So, to the extent

02:09:27.560 that insulin resistance is functionally denominated by elevated blood glucose.

02:09:36.120 It would seem to be the muscle is the primary defect. It would seem to be the muscle, which is the

02:09:40.520 primary defect. Now, the mechanism of the insulin resistance of muscle, I mean, this is an area that

02:09:48.280 is not one that I work in primarily, but there appears to be that what's going on at the sort of

02:09:57.080 distal end of the insulin resistance path is that the glucose transporters are not

02:10:04.280 getting to the surface. They're not responding to insulin the way that they do in a non-insulin

02:10:13.320 resistant person. And when the insulin transporters, the glucose transporters don't get to the muscle,

02:10:20.680 get to the surface of the muscle cell, they can't transport glucose. So at a molecular level,

02:10:26.600 depending on exactly whose model of this you look at, there's something wrong with the transmission of

02:10:33.160 the signal from the insulin receptor. Into the cell to send the glute transporter up.

02:10:38.680 Right. And exactly what that defect is, again, depends on who you talk to or what mechanism is

02:10:47.880 invoked. That appears to be, at least on one level, the sort of primary problem.

02:10:52.680 So what's happening in the liver? When we do an oral glucose tolerance test,

02:10:56.760 the reason we do it for such a short period of time, relatively speaking, aside from who wants to

02:11:00.840 keep a patient in the lab forever is, you pretty much eliminate de novo glycogenesis or you eliminate

02:11:06.760 DNL, right? So you're basically looking at uptake by muscle, but there's some being taken up by the

02:11:12.600 liver. And I don't know that I understand fully, aside from capacity, right? The liver has a relatively

02:11:19.240 small capacity for glycogen relative to the muscles. What else determines that partitioning of how much

02:11:25.240 glucose in the bloodstream is going into the muscle versus the liver? And then ultimately,

02:11:30.760 anything that exceeds those capacities will then undergo DNL and the de novo glycogenic pathway will,

02:11:36.280 you know, produce addition free fatty acid. And again, what does it mean to be insulin resistant in

02:11:41.400 the liver? So it's interesting. You know, the liver, if you want to look at it this way,

02:11:45.400 there are sort of two major metabolic things going on. There are a lot of things going on in the liver,

02:11:50.040 but as far as what we're talking about, one is the ability to make lipid, to synthesize lipid,

02:11:56.600 and the other is the ability to make or take up glycogen, to release or take up glycogen. They are

02:12:03.640 differentially sensitive to insulin suppression or enhancement, if you want to look at it that way.

02:12:11.640 And what happens, I mean, just again, at a biochemical level is you can get insulin resistant in the liver

02:12:19.080 to the suppression of gluconeogenesis while you maintain sensitivity to the effect of insulin on

02:12:27.240 the lipids.

02:12:27.960 To normal lipogenesis pathway.

02:12:29.400 Yes.

02:12:29.560 That's the worst outcome.

02:12:30.760 Yes. But that's what happens. I mean, that's the state, I think, in many individuals who have

02:12:39.080 issues with regard to insulin homeostasis, if you want to describe it that way.

02:12:43.560 Yeah. So for the listener, what we're basically saying is the doomsday scenario would be

02:12:47.800 your muscles become resistant to the signaling of insulin and they don't put these glute transporters

02:12:53.240 up there and you don't bring glucose into your muscle.

02:12:55.240 Yeah. But why that happens actually, in other words, what is it about becoming obese that is

02:13:01.720 actually doing something to skeletal muscle?

02:13:06.200 What if it's the reverse?

02:13:07.240 Well, it may be. I mean, again, this is an area of somewhat hot debate, but one possibility,

02:13:13.320 I guess, is that there's something that happens to adipose tissue when you store

02:13:19.160 whatever for that individual is an excess amount of lipid that's producing another molecule or some

02:13:26.600 other mechanism by which muscle is affected with regard to its insulin sensitivity.

02:13:33.960 The other possibility, which I think, again, many people think is at least partly relevant,

02:13:39.720 is that when you increase the amount of lipid in your adipose tissue, you also increase the amount

02:13:46.440 of lipid in your muscle, the fat that's in your muscle, actually, the marbling of your own muscle,

02:13:53.240 and that that itself has an effect on glucose homeostasis. So there's a whole...

02:14:00.520 What does Gerald Schumann think of this? Doesn't he have an idea that it starts in the liver?

02:14:05.400 Isn't that his view?

02:14:07.640 So Gerald Schumann has done a lot of work on the mechanism by which insulin affects

02:14:15.400 glucose transport in muscle. I don't want to speak for him, I mean, in terms of what he

02:14:20.440 thinks about this, but he certainly is one of the people who I think has advanced the

02:14:26.600 field in terms of understanding why, for example, ambient fatty acids seems to suppress

02:14:34.200 glucose uptake in muscle. This was originally described by Randall.

02:14:38.440 And just to be clear, that's not Randall the patient.

02:14:40.920 That's correct. Not that Randall.

02:14:42.280 You didn't know shit about.

02:14:43.320 Right. It's spelled differently also.

02:14:46.040 So I think what Gerald Schulman has done is to advance our understanding of

02:14:50.280 a very, I think, compelling mechanism by which high levels of ambient fatty acid themselves

02:14:55.640 might suppress insulin signaling. Some of that fatty acid could, in fact, come from

02:15:02.440 lipid that's stored in the muscle as opposed to lipid or fatty acids that are in the circulation.

02:15:08.040 But there are these two different phenotypes, right? When you look at Cahill's 40-day fasted

02:15:12.760 subjects, their free fatty acids were through the roof. Their insulin was very low. Their glucose

02:15:17.880 was very low. Their ketones were very high. It's interesting to know, would you call them

02:15:22.440 insulin-sensitive or insulin-resistant, right? Because probably if you challenge them with

02:15:27.080 glucose immediately, they would have hyperglycemia. Physiologically, their muscles probably would,

02:15:32.280 in the short moment, say, hey, I don't want this stuff. Save every gram of that for the brain.

02:15:38.520 But presumably, if you refed them carbs for three days, they'd probably be as insulin-sensitive as

02:15:42.920 they'd ever been. Right. So I mean, this is one of the reasons why I think you're right. But I think

02:15:49.000 maybe what Shulman or Randall would say is that the reason for the lack of response to the transport

02:15:56.920 of glucose is that the system has now been downset by virtue of the high ambient free fatty acids,

02:16:03.560 and it may take some time to... Interesting. So you're thinking that could be the

02:16:07.320 short-term signal. Correct. And that takes time. That's why, again, I don't do this for a living.

02:16:12.600 I mean, you would know this. But why people are encouraged not to restrict their carbohydrates

02:16:18.600 prior to having a glucose tolerance test, for example. Right.

02:16:22.040 Because you can manipulate the system in the way that you just mentioned.

02:16:26.440 Rudy, I could sit here and talk about this stuff all night long, but

02:16:29.560 your wife will probably be pissed off that I kept you here all night. So maybe not. Maybe she's

02:16:34.120 actually enjoying the fact that you're right. Yeah, yeah, yeah. This has been super interesting.

02:16:39.000 You're really one of the most thoughtful people I've ever spoken to on this topic. And I think

02:16:43.640 there are a lot of areas where we don't necessarily see eye to eye clinically, which again, that sounds

02:16:48.520 like a much more harsh statement than it is. I just think we look at the world a little differently on

02:16:52.360 some level, but... Yeah, I'm a hammer.

02:16:54.120 You're a hammer looking for a nail, and I'm kind of a general contractor looking for everything in sight.

02:16:59.640 I mean, I think it's interesting. I think your FTO example is a great example. I mean,

02:17:03.800 if I were tackling this problem, and I'm glad I'm not, I'd be wanting to understand the

02:17:08.600 environmental trigger. Is it the cigarette that's causing lung cancer? You're trying to solve a

02:17:13.160 different problem, which is, why is it that only some people get lung cancer when they smoke?

02:17:18.520 And those people, before the adage of the invention of cigarettes, would have never got lung cancer.

02:17:23.720 But now that these cigarettes are here, probably can't do much about it. Let's figure out who they are.

02:17:28.200 And not just for that reason. I mean, just as an intellectual question, I find it very intriguing.

02:17:34.760 But knowledge of whatever would lead a person who smokes not to get lung cancer,

02:17:41.640 it would be very nice to know what that is, that whatever that biology is, for many,

02:17:47.240 many reasons, not just related necessarily to lung cancer. And I see the issue of the

02:17:53.880 consequences of a genetic variant, very prevalent in the population, which predisposes some individuals

02:18:00.520 to the environmental consequences and others not. If we knew what that mechanism is,

02:18:05.640 is there's no reason to believe that at some point we might not be able to manipulate it,

02:18:10.920 not only in service of those who have the genetic variant, but others who might benefit from

02:18:17.720 knowledge of the circuitry.

02:18:19.480 Yeah. I mean, that makes a lot of sense. And you've given me some great pearls over the years.

02:18:25.160 I remember you, I think it was one night at dinner when you, me, Mike Rosenbaum were playing patty

02:18:31.880 cakes and it somehow came up that in your 10 to 20% weight reduced subjects,

02:18:37.080 a little bit of T3 could overcome some of the deficit. And I remember having this eureka moment

02:18:42.120 because I was like, wait a minute, all these patients I see that lose weight have a reverse

02:18:46.360 T3 spike. That would exactly explain why giving them T3 rather than T4 would address the problem.

02:18:53.480 You give a patient with high reverse T3, T4, you're just, you're making them worse,

02:18:58.040 but you give them T3, you can bypass it. So I'm incredibly grateful to you,

02:19:03.240 to your work and for your friendship. So thank you very much. Thank you.

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The Peter Attia Drive - December 17, 2018

#33 - Rudy Leibel, M.D.： Finding the obesity gene and discovering leptin

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