COMMEMORATIVE LECTURE
PPARs and the complex journey to obesity
Ronald M. Evans
Howard Hughes Medical Institute, The Salk Institute for Biological Studies, San Diego, CA, USA(Received for publication on March 11, 2004)

Key words: PPARs, nuclear receptors, metabolic disease, obesity, diabetes

tors, we need to keep in mind that there probably is amatrix that connects the entire family, and thus there isa higher principle that is operating at the genetic level.
To brieﬂy summarize, nuclear receptors have acentral DNA binding domain (DBD) and a carboxyterminal ligand binding domain (LBD) (Fig. 1). The
DBD allows them to bind to and activate target genes,
thus deﬁning them as transcription factors. The LBD
modulates their activities, making them hormonedependent transcription factors. They act as genetic
switches that are controlled by ligands.

I would like to thank all of you for staying throughthis long day of lectures, as I get to have the last word. I
wish to thank Drs. Hayashi and Kizaki for their kind
remarks and for keeping everyone on time and to also
thank Dr. Miyashita, my co-recipient of this award
for staying until the end of these talks. We have had
two interesting sessions, one that has dealt with cognition and neuroscience, and another that has dealt withthe molecular biology of lipids and hormones. In somerespects, each of these can be thought of as a separate
science, which in fact they are. But from another perspective, all disciplines are linked and we have every
reason to believe that nuclear receptors and cognition
will one day intersect. Why? In part because nuclear
receptors are expressed in every region of the brain andhormones affect behavior in quite profound ways from
reproductive behavior to the development of the brain
itself. Nuclear receptors are widely expressed in neuralstem cells, differentiated neurons, and glial cells. Thyroid hormone, glucocorticoids, and retinoids are major
regulators of neural activity. Thus, at each of theselevels and from various perspectives, convergence between these various systems can be found.

Nuclear Receptor Subclasses
The receptors can be grouped into several classes asshown in Fig. 1. One class binds endocrine hormonessuch as the adrenal and gonadal steroids. These include
glucocorticoids and mineralocorticoids that regulate
sugar and salt levels in the body, as well as the sex steroids estrogen, progesterone, and androgen. Anotherclass comprises receptors for vitamin A derivatives,
thyroid hormone, and vitamin D. These compounds are
involved in activities that range from neural differentiation, treatment of leukemia, regulation of basal metabolism, and regulation of calcium absorption andbone mineralization. Together these receptor systems
play important roles in controlling organ physiology via
transcriptional mechanisms that differ from that of cellsurface receptors. Then there are the orphan receptors
which comprise the remainder, and by number thelargest part of the superfamily (Mangelsdorf and Evans,
1995).2 These are members that were identiﬁed ﬁrst byhomology as genes and their existence led us to search
for new ligands. When ligands are found for these orphan receptors, we take them out of the ‘‘orphanage’’
and adopt them. I will talk about some of these adopted
orphan receptors, speciﬁcally, the PPAR subfamily, in

The Nuclear Receptor Superfamily
Today I am going to talk about a segment of the nuclear receptor superfamily. At present in our genome,
based on sequence and what has been isolated, there
are 48 genes that encode a nuclear receptor (for review,
Mangelsdorf et al., 1995).1 These 48 receptors comprise
what we call a ‘‘superfamily’’, and this entity can bethought of as the combined functions of the individual
receptors. We can look at each receptor individually, or
take a more global view of their functions as a matrix or
network in which they are not really separate, but areinterconnected. While we can study individual recep-

Dr. Evans is the recipient of the 2003 Keio Medical Science Prize.
Presented at the Keio Medical Science Prize Symposium, December 4, 2003.
Reprint requests to: Dr. Ronald M. Evans, Howard Hughes Medical Institute, The Salk Institute for Biological Studies, Post Ofﬁce Box 85800,
San Diego, CA 92186-5800, USA53

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Evans RM: Nuclear receptors and obesity

Fig. 1

Nuclear receptor superfamily.

more detail. I would like to mention two properties ofthese adopted orphans: one, they form heterodimers
with the retinoid X receptor (RXR), and two, they respond to dietary compounds such as lipids, that is,molecules that we take orally. This has opened up entire new areas of endocrine physiology.
‘‘Reverse Endocrinology’’
Next I will describe a strategy called ‘‘reverse endocrinology’’. Forward endocrinology is where you startwith the hormone and then search for the receptor. For
example, we used estrogen to ﬁnd the estrogen receptor. Reverse endocrinology consists of ﬁrst ﬁnding thereceptor and then looking for the hormones (Kliewer
et al., 1999).3 For the orphans, our approach has been
to search for ligands, which have turned out to be dietary lipids that include oxysterols and bile acids, bothof which are cholesterol derivatives, and fatty acids andxenobiotic lipids in drugs.
Because these orphan receptors are transcription
factors, we looked for their target genes and what we
found were genes that are involved in lipid detoxiﬁcation, transport and storage (Chawla et al., 2001).4
These include cytochrome P450s, the ABC cholesteroltransporters and cytosolic binding proteins that transport lipids. We then used genetics, transgenics orknockouts to characterize their various phenotypes andagain we observed problems with lipid homeostasis that
affected cholesterol, bile acid, fatty acid and drug metabolism. So the orphans appear to play a central role inbody physiology.

I will describe only brieﬂy the mechanism by which
the receptors work, as this has already been coveredin the earlier talks. The receptors are a relatively smallpart of the overall complex that we believe comprises
more than 75 or 80 proteins that are involved in transcriptional control (Collingwood et al., 1999).5 The retinoic acid receptor, in the absence of a hormone, is a
repressor and it collects a series of repressor proteins
like a magnet. Addition of the ligand reverses thepolarity of the magnet, causing the repressors to ﬂy
off and the coactivators to bind, resulting in chromatin
modiﬁcation to activate transcription. This is how thehormonal or physiological signals affect the recruitmentof co-factors that modify the chromatin in target genes.
Every cell has receptors and therefore can respond, buteach cell responds in its own unique way, and thus thesame hormone can have a different effect in a neuron
versus an epithelial skin cell versus a bone cell. In thisway each receptor has a genetic network and it is the
combined network that leads to the physiological effectin the body. Each of the 48 genes has its own network,so there is a very complicated set of pathways.
PPARs and Disease
Next I will focus on two of the PPAR receptor systems (Lee et al., 2003).6 They both respond to dietary
lipids and are involved in lipid metabolism. They bothform heterodimers with RXR and in response to theirligands, control the activity of a set of target genes.
That being said, these two very closely related receptors function in opposing ways in controlling lipid me-Keio J Med 2004; 53 (2): 53–58

Fig. 2

PPARs and heart disease.

tabolism, which I will talk about in more detail, particularly as it relates to obesity in obesity-related diseases
such as heart disease and insulin resistance.
How are the PPARs involved in heart disease? In
the California surfer view of this process, we start out
young, blond, slim and with exercise, we look good;then through the combination of genetics, a high-fat
diet, and little or no exercise as we get older, we evolveand ‘‘expand’’ into a more mature state (Fig. 2). Insteadof surﬁng in a bathing suit, we sit on the couch andsurf channels on television. This decrease in exercise
leads to an obesity-related problem called Syndrome X,
that includes insulin resistance, hypertension, hyperlipidemia and heart disease. Our society is facing the
problem that aiming the remote control for the television has often become the most exercise many ‘‘couch
potatoes’’ get.
The reality is that Syndrome X is an obesity-relateddisease (Haffner and Taegtmeyer, 2003).7 Sixty-ﬁve
percent of Americans are overweight, which is theleading risk factor for Syndrome X. The United States
is approximately three billion pounds overweight,
which is the basis for the obesity epidemic. We arecontinually getting heavier and this excess weight
carries a major health problem to society. The besttreatment for obesity is diet and exercise, but eventhough that advice is repeatedly given, it is generally
not taken, resulting in weight gain. There is a clear needto understand the nature of the problem and to use thisinsight to develop pharmacologic therapies. Developed
countries such as Japan and those in Europe are not far
behind. A recent study reported that Italy is the leading European nation in weight gain accompanied byan increased prevalence of Type II diabetes in theiradolescent population with spread into the childhoodgroup, similar to what is happening in the UnitedStates. It appears that the countries with highly-evolved

55

technology centers, penetration of computer-based systems and wide spread cell phone use appear to bemoving in this direction. These ‘‘advances’’ are eachassociated with changes such as reduced exercise andincreased calorie consumption.
We believe that the PPARs play a central and very
important role in the development and treatment
of Syndrome X (Desvergne and Wahli, 1999).8 There
are three PPAR isoforms. I will not discuss much here
about PPAR alpha (PPARa), which is expressed athigh levels in the liver and in the heart, and is involved
in beta oxidation of fatty acids. It is the target of a classof prescription drugs called the ﬁbrates, cloﬁbrate andciproﬁbrate. PPAR gamma (PPARg) as we have heard
in Dr. Kadowaki’s talk, is required for the formation of
the adipocyte, and it is the target for a class of drugs
called thiazolidinediones (TZDs) ‘‘insulin sensitizers’’.
Sales from TZDs now comprise a US$4 billion market.
The ﬁrst of the insulin sensitizers was developed inJapan but subsequently because of liver problems wasremoved from the market. However, the TZDs still
comprise a widely used class of drugs and there is muchinterest in its molecular target PPARg.
Involvement of PPARs in Heart Disease
PPAR delta (PPARd) is the receptor that we least
understand and this is the primary one upon which I
will focus. PPARg and PPARd are both involved in theformation of heart disease. The heart, just like the restof the body, is sensitive to a high-fat diet, poor exercise
and genetics. The ﬁrst inﬁltration of fatty streaks often
develops in childhood and continues in adolescence. It
can actually develop in the fetus, if the mother eats avery high-fat diet, and the fetal heart develops atherogenic lesions that persist throughout life. A lesion is
populated by macrophages which invade the arterial
wall. When these macrophages accumulate lipid, they
become foamy and are thus called ‘‘foam cells’’. Uponbuilding up in the intima of the artery, they initiallyform a fatty streak which can subsequently progress
into atherosclerosis. Among ﬁfty percent of all Americans and in most Western countries, heart attack is the
major cause of death. These cardiac macrophages arevery rich in PPARg and PPARd and both receptors canplay a beneﬁcial role in reversing and preventing heartdisease, so we think there may be a natural basis tounderstanding their medical functions.
PPARg and the Cholesterol Pathway
I will not describe the data that led to the model
shown in Fig. 3, as you all know how hard it is to build agenetic pathway (Lee et al., 2003a).9 But in this pathway, what we show is that low-density lipoproteins

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Evans RM: Nuclear receptors and obesity

Fig. 3 PPARs are lipid sensors regulating macrophage inﬂammationand lipid homeostasis.

(LDL), the so-called bad cholesterol, becomes evenworse when it becomes oxidized. Macrophages try toclear out oxidized LDL (oxLDL) and as they do, theligands for PPARg in this particle activate certain genes
such as CD36, a receptor for oxidized LDL. The presence of oxidized LDL creates a cycle that allows theclearance of this toxic lipid particle from the blood into
the macrophage. PPARg also activates another orphan
receptor, an oxysterol receptor (LXR), whose lipid
ligands are also present in the oxLDL particle. LXR
when activated in turn triggers the reverse cholesteroltransporter ABCA1, causing the release of HDL fromthe macrophage. The net result is that the bad cholesterol is processed in the macrophage and released as
good cholesterol. Thus the macrophage functions as aliving machine that tries to purge the body of bad cholesterol and produce good cholesterol. But if you havetoo much oxidized LDL, the body is unable to keep
up and stores most of it. Through pharmacology, we
can try to stimulate the export arm, using chemicals toaccelerate HDL release and clear out the artery. Thatmodel is currently being tested in patients. So we believe that PPARg and LXR will play central roles in the
future of the treatment of coronary artery disease by
non-invasive methods through an orally-active pill.
PPARd and Clearance of Excess Cholesterol
What about PPARd? PPARd does not respond to
LDL, but it does respond to VLDL and is a VLDL
sensor (Lee et al., 2003b).6 What is VLDL? VLDL
stands for very low density lipoprotein, which is produced by the liver and is a triglyceride-rich particle. We
take in dietary cholesterol and other types of fats andfatty acids which are packaged chylomicrons and delivered to the liver where they are repackaged to form
LDL and VLDL. These particles comprise the forward
arm of cholesterol transport and this is how lipids
are delivered through the circulation, to the cells inthe body. As triglycerides are stripped from VLDL, it
becomes converted to the cholesterol-rich particle,
LDL. Excess cholesterol is efﬂuxed back to the liver inthe form of HDL. This is the reverse arm of cholesteroltransport and is often considered a measure of goodcholesterol. In the liver, cholesterol is metabolized into
bile acids (BAs) and delivered to the intestine for excretion. While most (> 95%) of all bile acid is resorbed,
a small amount (< 5%) is lost in the stool. Because BAs
circulate nearly 15 times a day, this is one of the very
few mechanisms by which cholesterol can be removed
from the body. The problem of overeating cholesterol isthat much of it remains in the body, which over a lifetime causes many problems as excess cholesterol is very
toxic.
Function of PPARd in Fat
I will talk about the function of PPARd in two tissues, adipose and muscle. These are the two key tissues
for the metabolism of fats. When we think about metabolism, there are two components that are relevant:fat storage and fat burning. What are the controls forthese processes? The two receptors that we have heard
about – PPARg for fat storage and PPARd for fat
burning – are required for the formation of the adipose
cell and for storing fat after the cell is formed. Because
they are required both to generate and to maintain thefat cells, they are a major target for drug development.
I will describe a series of experiments on the role
PPARd in fat metabolism that were published earlier
this year (Wang et al., 2003).10 We expressed a form of
PPARd in transgenic mice using a fat-speciﬁc promoter
called aP-2. This form of PPARd has a VP-tag on it that
activates the receptor without a ligand. In this way, it
can be selectively activated in fat cells. It causes a net
reduction in the fat pad, compared to the wild-type fat
pad. The fat cells form and they accumulate lipid, butwhat we observed is increased metabolism in the cells
that causes fat burning and therefore a reduction in
total fat content. This has very important implications
for body physiology and I will describe a few examplesof what happens in these transgenic mice.
First of all, the mice with the activated PPARd havegreatly reduced serum triglycerides – about a 40% decrease, as well as a 20% decrease in serum free fatty
acids. This signiﬁcant reduction, which is what we try toachieve with drugs, we were able to do genetically. If
we put these animals on a high-fat diet – one that corresponds to the Western diet, which is about 30% fat,Keio J Med 2004; 53 (2): 53–58

their lipid levels increased sharply. However, in thetransgenic animals, there is a 55% reduction in triglycerides compared to the wild-type mice, demonstratingthat there is resistance to the elevation even though
they eat the same amount of fat. The same is true forthe free fatty acids. There is a resistance – of about
35% reduction. Cholesterol, on the other hand, is notchanged. We believe this is because PPARd controls
the triglyceride component of fat but does not inﬂuencecholesterol levels itself. How does it do this? PPARd is
switched on in brown and white adipose tissue, which
activates a set of target genes that produce a lean phenotype and a depletion of serum lipids. These animals
weigh less than wild-type animals. In the brown adipose
tissue of wild-type animals on a high-fat diet, the excess
fat is stored in vacuoles. But the transgenic mouseburns the excess fat and the brown adipose tissue isfound to have a reduced lipid content.
We also examined what happens in leptin receptordefective (ob/ob) animals that are genetically prone towards obesity. They get fat by excessive eating, andtheir fat cells contain an excessive amount of fat. If wegive them a drug that is speciﬁc for PPARd, the drugcauses increased metabolism and fat burning and normalizes the appearance of the brown adipose tissue.
This demonstrates that we have an orally active drugthat mimics the effect of the activated transgene byincreasing fat burning.
To show what happens in the absence of PPARd
function, we knocked out PPARd completely in someanimals. These mice, because they lack PPARd, cannot
upregulate fat burning. As a result, in a three-week experiment on a high fat diet, they nearly doubled theirbody weight, which is a tremendous amount of weightgain. In the wild-type animals, upregulation of fat
burning occurs during the early phase of the experiment, protecting the animals from acute weight gain.
On the other hand, the knockout animals gain weight
immediately and quite dramatically.
PPARd Increases Type I Muscle Fiber
The last topic that I want to mention is the consequence of PPARd activation in muscle. These results
were quite surprising. I am going to use the example of
two Olympic runners: Frank Shorter, a long-distancerunner who has more of a type I (slow-twitch) muscle
ﬁber and the sprinter Maurice ‘‘Mo’’ Green, the only
human to have run the 100-meter dash in 9.79 seconds
without steroids (Ben Johnson did it with steroids) who
have type II (fast-twitch) muscle. At birth, we have acertain ratio of type I to type II muscle. It is a genetically determined ratio for each individual, but as you
become obese, you lose type I ﬁbers. If you train byrunning, you gain more type I ﬁber, so there is an57

adaptive switch. These two types of muscle ﬁbers arevery different. They have different amounts of mitochondria, different troponins, different amounts ofmyoglobin and myosins and require different amountsof energy to maintain.
We wanted to ask what happens if we express
PPARd in the muscle (Wang et al., submitted).11 Would
it affect muscle metabolism? To our surprise, the transgenic muscle that expresses PPARd from a myosinheavy chain gene promoter was much redder (increased
levels of type I muscle) than the wild-type. This wasquite clearly observed in the gastrocnemius muscle,
which normally has a mixed amount of type I and typeII ﬁbers. Thus, the wild-type animal had a typical mixture of type I and II ﬁbers, and the transgenic had theslow-twitch, Frank Shorter type I muscle. This was
conﬁrmed by ﬁber type-speciﬁc ATPase staining. There
are different types of troponins that are expressed inthis muscle that confer on it what we would call a longdistance running phenotype.
Increased Running Endurance in PPARd
Transgenic Mice
An experiment that is rarely done in science is atrue gain-of-function experiment. Most of what we do isknock genes out; we lose function, we lose gonadal differentiation or we lose some feature of hippocampal
function like LTP or LTD. Rarely do we have experiments where we have a net gain-of-function. If I wereto ask, ‘‘How would you improve vision?’’ or ‘‘How
would you improve motor function?’’, can you do that
simply by making more retinal cells, simply by making
more motor neurons? Those are questions that are very
difﬁcult to answer. We now had the opportunity to ask
the question, ‘‘What happens if you make more type I
muscle ﬁber?’’ In this genetically-engineered strain of
mice, the question was, do these new slow-twitch ﬁbers
become innervated normally by the motor neurons anddo they have the right type of response to affect thephysiology of these animals?
We addressed this question by putting the mice on atreadmill to see whether the engineered mouse could
actually run longer than its wild-type littermates. We
placed both wild-type and transgenic mice on treadmills, and monitored various parameters. A platform
was situated at the back of the treadmill where themice, after running to exhaustion, will be deposited.
When both hind and fore paws are on the platform, theexperiment is terminated.
The wild-type animals were able to run for a little
less than 90 minutes, and the transgenics ran for about
150 minutes, about 60 minutes longer, which represented a tremendous increase in time on the treadmill.
Since we increased the speed of the treadmill over time,

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Evans RM: Nuclear receptors and obesity

the transgenics actually ran about twice the distance ofthe controls. From this we calculated that normal animals could run for about 900 meters before they were
exhausted, whereas the transgenics could continue forover 1.8 kilometers, a dramatic difference. This result
was very exciting because it indicates that we havefound the ﬁrst transcriptional pathway to alter muscle
ﬁber-type formation in an animal that has a true functional consequence. We believe that this has many
consequences besides just running ability. For example,
these transgenic animals are apparently resistant to ahigh-fat diet; although they put on some weight on ahigh-fat diet, it is nowhere near the amount that thewild-type animals gain. Over the same period of time,
the wild-type mice gained almost 20 grams more than
the transgenics fed the same diet. In addition, the wildtype animals’ fat cells were greatly enlarged relative tothe transgenics.
We showed that we can mimic this effect with a drug.
We can take wild-type mice and give them a PPARdspeciﬁc drug, conferring on them resistance to weightgain. We thus have an orally active molecule thatcan prevent weight gain on a high-fat diet. What is themechanism behind this? We looked at the oxygen consumption and respiration of these animals. We put theminto metabolic cages and found that at every time point,
the animals treated with the drug had an increased
respiratory coefﬁcient. What this means is that the
drug causes increased metabolic activity and as result,increased fat burning.
Concluding Remarks
I wish to conclude the presentation by saying that
we have two very interesting receptors, PPARg and
PPARd, that control distinct genetic pathways. We believe that activation of both of these will improve Syndrome X. PPARg is a known drug target for insulin
sensitizing drugs such as Actos and Avandia. We are
currently developing drugs for PPARd, and there will
likely be announcements of the ﬁrst clinical trials ofthese drugs in humans soon. If the results of these drugs
in people are similar to what we have seen in mice, theyshould produce resistance to weight gain, a lowering oftriglycerides and possibly increased insulin sensitivity.
We therefore have a PPAR-mediated metabolic
switch that is controlled in part by diet, in part by circadian rhythm, in part by exercise and by certain classesof drugs. It involves two receptors, PPARg and PPARd,that activate a set of target genes. Interestingly, al-though they are expressed in the same cells, they activate different target genes, although a few, like ADRP,a lipid storage gene, are the same. Most of the target
genes are very different between these two closely related receptors. Nature appears to have used similarity
in receptor structure to control two different, opposing
pathways in terms of lipid metabolism – one controllingenergy storage, the other controlling energy expenditure. I am very excited about the potential of drugs forthese receptors.
This work was done by members of my lab and I
want to thank three of the key people who did most of
the work I talked about today, Yongxu Wang, Chi-Hao
Lee and Ruth Yu. Some of it started with Ajay Chawla
who is now at Stanford, Heonjoong Kang, a former
chemist in my lab who produces the chemical compounds that we use for these studies. I thank all of myformer students in the context of the award, my postdocs, technicians, staff and collaborators for all of their
help that helped lead to the Keio Medical Prize Symposium today. Thank you all very much for being here.
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The nuclear receptor superfamily: the second decade. Cell 1995;
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2. Mangelsdorf DJ, Evans RM: The RXR heterodimers and orphan
receptors. Cell 1995; 83: 841–850
3. Kliewer SA, Lehmann JM, Willson TM: Orphan nuclear receptors: shifting endocrinology into reverse. Science 1999; 284: 757–
760
4. Chawla A, Repa JJ, Evans RM, Mangelsdorf DJ: Nuclear
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coactivators, corepressors and chromatin remodeling in the control of transcription. J Mol Endocrinol 1999; 23: 255–275
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Fiber Type and Running Endurance by PPAR delta. (submitted)