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(The FASEB Journal. 2001;15:312-321.)
© 2001 FASEB

Diseases of liporegulation: new perspective on obesity and related disorders

ROGER H. UNGER*,{dagger}1 and LELIO ORCI{ddagger}

* Gifford Laboratories, Touchstone Center for Diabetes Research, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas 75390-8854, USA;
{dagger} Veterans Affairs North Texas Health Care System, Dallas, Texas, USA; and
{ddagger} Department of Morphology, University of Geneva Medical School, Geneva, Switzerland

1Correspondence: Gifford Laboratories, Touchstone Center for Diabetes Research, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75390-8854, USA. E-mail: runger{at}mednet.swmed.edu


   ABSTRACT
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
Obesity-related diseases now threaten to reach epidemic proportions in the United States. Here we review in a rodent model of genetic obesity, the fa/fa Zucker diabetic fatty (ZDF) rat, the mechanisms involved in the most common complications of diet-induced human obesity, i.e., noninsulin-dependent diabetes mellitus, and myocardial dysfunction. In ZDF rats, hyperphagia leads to hyperinsulinemia, which up-regulates transcription factors that stimulate lipogenesis. This causes ectopic deposition of triacylglycerol in nonadipocytes, providing fatty acid (FA) substrate for damaging pathways of nonoxidative metabolism, such as ceramide synthesis. In ß cells and myocardium, the resulting functional impairment and apoptosis cause diabetes and cardiomyopathy. Interventions that lower ectopic lipid accumulation or block nonoxidative metabolism of FA and ceramide formation completely prevent these complications. Given the evidence for a similar etiology for the complications of human obesity, it would be appropriate to develop strategies to avert the predicted epidemic of lipotoxic disorders.—Unger, R. H., Orci, L. Diseases of lipid overflow: new perspective on obesity and related disorders.


Key Words: lipotoxicity • lipoapoptosis • complications of obesity • noninsulin-dependent diabetes • cardiomyopathy


   INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
THROUGHOUT THE 2.5 MILLION YEAR history of human development the principal threat to survival has been recurrent famine. The evolution of adipocytes provided a means for coping with the cycles of undernutrition by enabling the preloading of calories for subsequent use (1) . During the 20th century, however, an unprecedented change in the pattern of caloric availability took place in many Western countries. Recurrent undernutrition was replaced by unending overnutrition, the consequences of which were greatly amplified by the permanent state of underexertion imposed by sedentary occupations and immobilizing technologies of modern life. Since the compensatory mechanisms that buffer the metabolic consequences of short-term overnutrition were incapable of compensating for such chronic changes in caloric balance, diseases of overnutrition became increasingly prevalent.

The first such disease to be identified was coronary artery disease (2) . Scientific advances of the past three decades have elucidated its mechanisms (3) and have reduced its health consequences (4 , 5) . A second group of disorders, the complications of obesity, now threatens to replace it as a major health problem in Westernized societies (6) . These complications, which include dyslipidemia, insulin resistance, noninsulin-dependent diabetes mellitus (NIDDM), and heart disease, are often referred to collectively as ‘metabolic syndrome X’, ‘insulin resistance syndrome’, or ‘Reaven syndrome’ (6 7 8) . Given the inexorable increase in the prevalence of obesity, these complications will constitute an ever-increasing cause of morbidity, mortality, and health care costs in the United States and elsewhere (6) .

A similar array of complications in genetically obese Zucker diabetic fatty (ZDF) rats has been referred to as ‘lipotoxicity’ (9 10 11 12 13) . This presumably monogenic disorder is far more suitable for intensive study of obesity-related disease mechanisms than is the environmentally induced, polygenic human counterpart, because the complications appear by the 14th week of age in ~100% of male homozygotes, provided that their diet contains at least 6% of the calories as fat. The lipotoxicity is attributed to products of excessive non-ß-oxidative metabolism of FA excess in skeletal muscle, pancreatic islets, and myocardium (10 11 12) . High levels of these metabolic products are believed (13) to cause the common complications of obesity, insulin resistance, cardiovascular disease, and diabetes by disrupting cell function and ultimately by promoting programmed cell death (‘lipoapoptosis’) (11 , 12) . These complications are completely preventable in rodents by means of currently available pharmacologic interventions that increase oxidative and reduce the nonoxidative metabolism of FA (14 , 15) .

The possibilities that the complications of human obesity are mechanistically similar to those of obese ZDF rats and can be blocked by the same interventions prompted this review of rodent lipotoxicity, its prevention, and its relevance to human disease.


   NORMAL FATTY ACID (FA) HOMEOSTASIS
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
Compensatory up-regulation of FA oxidation
Normally, FA delivery to nonadipose tissues is tightly coupled to their need for fuel. Plasma free FA levels rise during exercise and fasting to meet the metabolic requirements, leaving little or no unoxidized FA in these cells (Fig. 1A ). During chronic overnutrition, however, FA influx into tissues may exceed FA usage, in which case compensatory up-regulation of FA oxidation is required to maintain intracellular FA homeostasis (Fig. 1B ). The excess FA themselves probably provide the signals for such metabolic adjustments by serving as ligands for peroxisome proliferator-activated receptors (PPAR) (16 17 18 19) .



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Figure 1. Normal and abnormal fatty acid (FA) homeostasis in nonadipose tissues. A) Caloric equilibrium: FA supplied and FA used in nonadipose tissues are equal, leaving no unoxidized FA. B) Compensated caloric excess. During chronic caloric excess, the FA supply to nonadipose tissues may exceed their oxidative needs; however, up-regulation of PPAR{alpha}, the transcription factor for enzymes of FA oxidation (CPT-1 and ACO) and uncoupling protein (UCP-2), will promote compensatory oxidation of the surplus FA, and the unneeded energy will be dissipated as heat. This compensatory system requires leptin and a normal leptin receptor (OB-R). C) Uncompensated caloric excess. In the absence of leptin or its receptor OB-R, hyperphagia results in a caloric intake in excess of caloric requirements. FA storage occurs not only in adipocytes, resulting in obesity, but also in nonadipocytes, causing lipotoxicity. With PPAR{alpha} expression reduced, surplus FA influx presumably binds to the high levels of PPAR{gamma}, which up-regulates the lipogenic enzymes ACC and FAS. This causes ectopic accumulation of TG, increased nonoxidative FA metabolism, and lipoapoptosis (see Fig. 2 ).

PPAR{alpha} is a transcription factor that up-regulates the oxidative enzymes (Fig. 1B ) carnitine palmitoyl transferase-1 (CPT-1) and acyl CoA oxidase (ACO) (18 19) . Normally it is expressed in nonadipose tissues at relatively high levels compared to PPAR{gamma}, the lipogenic transcription factor. It has been suggested that surplus FA may up-regulate PPAR{alpha} and the oxidative machinery, and thus prevent the overaccumulation of unoxidized lipids (20) . The supplemental energy thereby generated is presumably dissipated as heat (21) , as inferred from leptin-mediated up-regulation of uncoupling protein 2 (22 , 23) and from earlier studies of the thermogenic effects of nutrients (24) .

In most normal nonadipose tissue, PPAR-{gamma} and the enzymes of fatty acid synthesis are expressed at low levels compared to adipocytes (25) , suggesting that they have relatively low lipogenic capacities, an important factor in normal intracellular homeostasis.


   ABNORMAL FA HOMEOSTASIS
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
Sources of FA excess
The putative liporegulatory system described above is leptin-requiring, i.e., it exists only in tissues of normally leptinized organisms. In leptin-unresponsive obese ZDF rats homozygous for the fa mutation in the leptin receptor (OB-R) (26 , 27) , FA-mediated up-regulation of oxidative enzymes does not occur (20) . The surplus FA may therefore enter pathways of lipogenesis and nonoxidative metabolism (10 , 12) such as lipid peroxidation and ceramide-mediated apoptosis (Fig. 1C ). While this is, in part, the consequence of failure of FA to up-regulate expression of PPAR{alpha} and the enzymes of ß-oxidation (10) , the major cause of the excess lipid deposition in nonadipocytes is the high rate of lipogenesis derived from both elevated plasma free FA and triacylglycerol (TG) levels and from increased intracellular FA synthesis (10) .

PPAR{gamma}, a lipogenic transcription factor normally expressed at high levels in adipocytes, is also expressed at high levels in nonadipose tissues of fa/fa rats (20) . PPAR{gamma} up-regulates the lipogenic enzymes (17 18 19) , acetyl CoA carboxylase (ACC), and fatty acid synthase (FAS), which catalyze FA synthesis, and glycerol-phosphate acyl transferase, which catalyzes FA esterification. The high PPAR{gamma} in nonadipocytes such as islets, in turn, is accompanied by increased expression of the adipocyte determination and differentiation factor 1 (ADD-1)/sterol regulatory element binding protein (SREBP-1) (28 , 29) in the affected nonadipose tissues of the ZDF rat. SREBP-1 is a strong candidate for the proximal transcription factor that increases lipogenic capacity, because it can be up-regulated by hyperinsulinemia induced by overnutrition (30 , 31) . Figure 2 depicts a putative concept for diet-induced hypertrophy and hyperplasia of adipocytes induced by overnutrition and ultimate ectopic lipogenesis in nonadipocytes.



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Figure 2. Mechanisms of increased nonoxidative metabolism in tissues. A plausible explanation for the abnormal FA homeostasis and accumulation of TG in nonadipose tissues is depicted schematically in Fig. 1C . Overnutrition causes secondary hyperinsulinemia, which up-regulates adipocyte SREPB-1 (ADD-1) (27 , 28) , leading to hypertrophy of adipocytes. Recruitment of new adipocytes by insulin-like growth factor 1 (29) and other adipocyte products leads to recruitment of new adipocytes and adipocyte hyperplasia. This is reflected by increased plasma levels of FFA, ICF-1, and leptin. At some point leptin-mediated protection of nonadipose tissues (Fig. 1B ) wanes ({emptyset}). SREBP-1c and lipogenic enzymes increase and TG (yellow dots) accumulates in excess in skeletal muscle, ß cells, myocardium, and other nonadipose tissues.

Although the abnormalities depicted in Fig. 1C have so far been identified only in pancreatic ß cells (11 , 33) and myocardium (34) , they could also be a cause of other complications of obesity (Table 1 ).


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Table 1. Clinical states in which abnormal leptin-dependent FA homeostasis may increase


   MECHANISMS OF LIPOTOXICITY
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
The products of non-ß-oxidative FA metabolism that can injure cells include TG (35) , ceramide (12 , 36 , 37) , and products of lipid peroxidation (38) (Fig. 1C ). FA-induced damage through direct detergent action on membranes, through omega oxidation or through inadequate stearoyl CoA desaturase activity, have not yet been explored.

Triacylglycerol excess (steatosis)
TG are the most abundant products of excessive non-ß-oxidative metabolism of FA, but they are probably relatively inert and therefore not toxic in a direct sense. Marked TG excess in a nonadipocyte could theoretically interfere directly with certain cell functions, such as muscular contraction. For example, the high incidence of gallstones in obese humans could conceivably be the result of bile stasis due to impaired contractility of the gall bladder secondary to steatosis of its wall, but this question has not been addressed.

Excess TG may cause fibrosis in nonadipocytes. Hepatic fibrosis, a common finding in alcohol-induced hepatic steatosis, also occurs in obese nonalcoholic patients with excessive hepatic TG deposition (35) . Abnormal fibrosis also develops in the fat-laden islets (14) and heart (34) of ZDF rats. Although the mechanism of the poststeatotic fibrosis is unknown, transforming growth factor ß is suspected (39 , 40) .

Ceramide excess
An expanded reservoir of TG within nonadipocytes is a potential source of excess FA that may enter the lipotoxic pathways of de novo ceramide synthesis. Ceramide accumulation in ß cells, long implicated in apoptotic pathway of autoimmune destruction of ß cells and attributed to increased sphingomyelin breakdown (41) , can also be formed directly via de novo synthesis from FA (12) (Fig. 1C ). This pathway appears to play a central role in FA-induced apoptosis (11 , 12) (Fig. 3A ). The islets of obese fa/fa ZDF rats exhibit an increase in de novo synthesis of [3H]-ceramide from its precursors, [3H]-palmitate and [3H]-serine, compared to +/+ controls (Fig. 3B ). Moreover, the expression of serine palmitoyl transferase, which catalyzes the first step in ceramide synthesis, condensation of serine and palmitoyl CoA (42) (Fig. 3A ), is increased in such islets (12) (Fig. 3C ). Ceramide increases the expression of inducible nitric oxide synthase (iNOS) through activation of nuclear factor {kappa}B (43) , and thereby augments the production of nitric oxide (NO) (44) . NO forms potent oxidants, such as peroxynitrite (45 , 46) , that cause the apoptosis (Fig. 3B ).



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Figure 3. Role of de novo ceramide synthesis in lipotoxicity. A) Simplified scheme showing fatty acid-induced ceramide synthesis from condensation of palmitoyl CoA and serine. Elevated ceramide is believed to be the cause of FA-induced apoptosis of pancreatic ß cells. This pathway is distinct from cytokine-induced breakdown of sphingomyelin, the mechanism proposed for apoptosis of autoimmune diabetes. The validity of the fatty acid-induced pathway is supported by the fact that ceramide formation, iNOS up-regulation, and apoptosis can all be reduced or blocked by L-cycloserine (CS) and by fumonisin B1 (FB-1), by reducing islet lipid content (14) , and by the experiments in panel B. B) Comparison of de novo [3H]-ceramide formation from [3H]-serine or [3H]-palmitate in islets isolated from normal lean ZDF rats (+/+) and obese diabetic ZDF rats (fa/fa). C) The ratio of serine palmitoyl transferase mRNA to ß-actin mRNA in islets isolated from normal lean ZDF rats (+/+) or obese ZDF rats (fa/fa). At 7 wk of age the obese rats are prediabetic; at 14 wk of age they are diabetic. D) Apoptosis, as evidenced by DNA laddering, in the pancreatic islets of obese prediabetic ZDF (fa/fa) rats cultured in the absence or presence of 1 mM fatty acids (FA) and its relationship to leptin action and Bcl2 expression. Prior to culture, the islets were treated with either AdCMV-ß-gal (as a control) or AdCMV-OB-Rb to cause transgenic overexpression of a normal leptin receptor. In the AdCMV-ß-gal-treated islets cultured in 1 mM FA, the already high level of DNA laddering, an index of apoptosis, rose by twofold. Expression of the antiapoptotic factor Bcl2 declined; the addition of leptin did not prevent these changes. By contrast, in the AdCMV-leptin-treated islets in which the normal leptin receptor is overexpressed, FA still increased DNA laddering and reduced Bcl2 expression, but 20 ng/ml of recombinant leptin completely blocked both the apoptotic effect of FA and the FA-induced down-regulation of Bcl2. (Reprinted with permission of Proceedings of the National Academy of Sciences U.S.A.)

Type 1 and 2 diabetes share a distal portion of the apoptotic pathway
Obesity-related Type 2 diabetes of ZDF rats and autoimmune Type 1 diabetes appear to share the apoptotic pathway mediated by ceramide and iNOS (Fig. 3A ). However, in lipotoxic islet disease only those ß cells with the highest fat content succumb to lipoapoptosis; this leaves enough functioning ß cells to maintain insulin independence, but not sufficient insulin-producing capacity to compensate for the insulin resistance and prevent Type 2 diabetes. In contrast, in cytokine-mediated apoptosis of autoimmunity, the destruction of ß cells is indiscriminate and usually total, causing insulin-dependent or Type 1 diabetes.


   GENETIC AND NONGENETIC CAUSES OF LIPOTOXICITY (TABLE 1)
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
Congenital generalized lipodystrophy (CGL)
CGL is a rare genetic disorder characterized by absence of adipocytes and deficiency of their secretory products. The leptin deficiency (47) causes hyperphagia (48) , whereas the lack of leptin-mediated protection against ectopic lipid accumulation, coupled with a lack of adipocytes in which to store excess TG, results in severe lipotoxicity in nonadipocytes. The clinical manifestations of the disorder include a voracious appetite, hypertriglyceridemia, severe insulin resistance, diabetes, hepatic steatosis, and cardiac disorders such as hypertrophic cardiomyopathy appearing in early adulthood (48 , 49) . In mouse models of CGL, leptin administration reduces the insulin resistance and hepatic steatosis (50) , as does transplantation of adipocytes (51) .

Based on this evidence of lipotoxicity in rodent CGL, it would seem appropriate in human CGL patients to maintain equality between FA influx and FA utilization by reducing their caloric intake and/or increasing their FA utilization. Exercise and/or antisteatotic treatment with leptin (50) , or possibly troglitazone (15) , might also be helpful in protecting against lipotoxicity.

Lack of leptin action
Loss-of-function mutations in the genes encoding leptin (52) or its receptor (26 , 27) block compensatory ß-oxidation of excess FA and enhance lipogenesis in nonadipose tissues, thereby predisposing to lipotoxicity whenever influx of FA exceeds intrinsic oxidative needs. Because leptin-mediated regulation of hypothalamic appetite centers is absent in such rodents, their caloric intake is high and they lack leptin-mediated protection against the ectopic deposition of the increased FA and its consequences (Fig. 1C ). Although such mutations are extremely rare in humans, they have been reported (53 , 54) .

Diet-induced obesity
The clinical course of the lipotoxicity complicating diet-induced obesity is as different from the genetic obesity of monogenic leptin-resistant ZDF fa/fa rats as is the course of coronary artery disease of diet-induced hypercholesterolemia from that of monogenic familial hypercholesterolemia. Diet-induced lipotoxicity develops far more gradually, at widely varying rates, and at a later age than genetic lipotoxicity. This is because normally the protective action of the increasing hyperleptinemia (Fig. 1B ) will at first maintain normal FA homeostasis and minimize ectopic TG deposition. Only later in the course of obesity, for reasons that are not yet fully understood, does severe leptin resistance appear. The causes of postreceptor leptin resistance are under study (55) . It is possible that aging itself contributes to the leptin resistance and complications that appear after decades of uncomplicated dietary obesity.

Aging
The increased morbidity and mortality of old age have been attributed to increased oxidative stress and lipid peroxidation in various tissues (56 57 58 59) . Caloric restriction and exercise are reputed to extend life by reducing non-ß-oxidative metabolism of lipids (60 61 62) , as leptin might be expected to do. It is therefore noteworthy that as normal rats grow to maturity, they become increasingly leptin resistant (63) . In elderly rats the effectiveness of leptin in reducing the TG content of nonadipose tissues is less than 10% of young controls (64) , raising the possibility that leptin resistance contributes to the phenotype of old age.

Impaired cardiac function and sarcopenia play central roles in the ‘dwindling’ of advanced age (65) . These and other abnormalities could be the consequence of an age-related lipotoxicity in which leptin-mediated protective ß-oxidation is lost, permitting lipids to shift into nonadipose tissues. Human ß cells have, in fact, been reported to undergo an age-related accumulation of neutral lipids (66) , perhaps explaining the decline in ß cell function in the elderly.


   LIPOTOXIC DISORDERS: DO THEY OCCUR IN HUMANS?
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
Lipotoxic heart disease
In rodents
‘Lipid cardiomyopathy’ was identified more than 30 years ago in the hypertrophied heart of gold thioglucose obese mice (67) , and the deleterious effect of FA on myocytes has subsequently been well documented (68 , 69) . In ZDF fa/fa rats, evidence of increased non-ß-oxidative FA metabolism in the myocardium is reflected by elevations in myocardial TG and ceramide content (34) (Fig. 5A ) and by increased myocardial oxidative stress (38) . Myocardial expression of iNOS expression is high (34) , and the increased nitric oxide is believed to interact with superoxide to induce apoptosis (45) , as reflected by the increase in DNA laddering (34) (Fig. 4B ). There is echocardiographic evidence of reduced myocardial contractility (Fig. 4C ) attributed to loss of functioning myocytes through apoptosis (34) . These changes can be completely prevented by treatment with troglitazone, which reduces the myocardial TG and ceramide content and preserves the contractile function of the heart (34) .



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Figure 5. Oil red O staining for lipids in human and rat heart and rat skeletal muscle showing similarity of excess lipid deposition in obesity. Frozen sections. A) Upper panels: cardiac muscle samples from (left) an untreated obese ZDF fa/fa rat or (right) from a troglitazone (TGZ)-treated obese ZDF fa/fa rat. Lower panels: skeletal muscle samples from (left) an untreated fa/fa rat, or (right) from a TGZ-treated fa/fa rat. B) Autopsy material from human cardiac muscle from (left) a 57-year-old obese male [body mass index (BMI) = 42] or (right) a 67-year-old lean male (BMI = 28). There is accumulation of Oil red O-stained lipids in samples from both the obese human, presumably diet-induced, and the untreated fa/fa ZDF rat with monogenic obesity.



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Figure 4. A) Cardiac triacylglycerol (TG) and ceramide content 14-wk-old ZDF (fa/fa) rats and wild-type controls (+/+). There is a statistically significant increase in both TG and ceramide content. Troglitazone (TGZ) treatment for 6 wk significantly reduces TG and ceramide content toward normal. *P<0.01. B) DNA laddering, an index of apoptosis, is increased in heart of obese fa/fa ZDF rats at age 14 wk. The increase is entirely blocked by treatment of the rats for 6 wk with the antisteatotic agent troglitazone (TGZ) beginning at 7 wk of age. Data are expressed as fold change from the 7-wk-old baseline in fa/fa rats, which is ~10-fold that of lean +/+ rats at that age. C) Left panel: echocardiographic images from a normal lean (+/+) rat at age 20 wk and an obese fa/fa age-matched ZDF rat. The dotted lines represent the cavity area for end-diastole (left) and end-systole (right). Both the end-diastolic and end-systolic areas are much larger in the fa/fa rat, indicating a dilated ventricle with poor systolic contraction. Right panel: quantification of circumferential fractional shortening, showing functional loss in 20-wk-old fa/fa prevented by treatment with the antisteatotic drug, troglitazone (TGZ).

In humans
Dilated cardiomyopathy, sudden death, restrictive cardiomyopathy, and congestive heart failure attributed to lipid-related abnormalities have all been observed in obese humans (70 71 72 73 74 75 76) . In fact, obesity in asymptomatic young women is associated with echocardiographic evidence of diastolic dysfunction (76) . Left ventricular dilatation, eccentric hypertrophy, and diminished compensatory reserve, sometimes leading to overt congestive failure, have been reported in obesity (76) . It appears that left ventricular abnormalities are greater in visceral-type than in subcutaneous-type obesity (75) , in keeping with the greater metabolic activity of the former.

Despite the foregoing evidence that FA and ceramide may damage cardiomyocytes, cardiac lipotoxicity is not currently recognized as a clinical entity in the United States. When congestive failure, chest pain, or arrhythmias occur in obese patients, they are usually attributed to other coexisting diseases, such as coronary artery disease and/or hypertension, rather than to lipotoxic dysfunction of myocytes. Nonetheless, a reduction in the number of cardiac myocytes through apoptosis is now a recognized cause of heart failure (77) , and lipotoxicity could well be an important contributory cause of apoptosis. Figure 5B provides the first preliminary morphological evidence that lipid excess can occur in the myocardium of obese humans, as it does in rodents. A human counterpart of the lipotoxic heart disease of obese rodents is therefore a very real possibility.

Lipotoxic diabetes
In rodents
As mentioned, diabetes occurs before the age of 14 wk in ~100% of male ZDF fa/fa rats, provided that 6% or more of their caloric intake is derived from fat. Two weeks before the onset of hyperglycemia (9) , plasma levels of FFA and TG increase dramatically. Islet TG content, which at first rises gradually, increases abruptly 1 wk before the onset of hyperglycemia (9) . Ceramide also increases at this time. The chronologic relationships of islet TG and ceramide content to the islet morphology and the clinical course of the disorder are depicted in Fig. 6 .



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Figure 6. A) Schematic representation of the relative changes in insulin requirements and insulin production as ZDF rats progress from preobesity to overt diabetes. B) Longitudinal course of plasma free FA levels (mM) and islet TG levels (µg/islet) and ceramide content in preobese, obese prediabetic, and obese diabetic ZDF fa/fa rats. Lipid abnormalities precede by ~2 wk the onset of diabetes (stippled zone), consistent with a lipotoxic mechanism. C) Immunocytochemical staining for insulin of representative islets obtained from ZDF fa/fa rats in the preobese, obese prediabetic, and obese diabetic phase of the illness (upper panels). Islets are morphometrically normal in the preobese phase, but ß cells have increased ~fourfold in the obese, compensated prediabetic phase, and decline sharply when islet TG content rises further. Oil red O staining for lipid accumulation in skeletal muscle appears under the 1) preobese islet (far left) and 2) under the obese diabetic islet (far right).

In the preobese state, the islet morphology and the TG content appear to be perfectly normal. During the prediabetic stage, as obesity increases, islet TG content rises ~10-fold, in association with a ~4-fold increase in the ß cell mass and insulin production. Since these same changes can be induced in vitro by culturing normal islets in FA (78) , it is assumed they are mediated by the excess of FA. The increasing hyperinsulinemia parallels the rising insulin requirements, thus keeping blood glucose levels within a normal range. The insulin resistance has been attributed to lipid excess in skeletal muscle (79 80 81 82) , and this is readily apparent on Oil Red O staining (14) (Fig. 5A ).

By the age of 10 wk the obesity and insulin requirement have progressed further, islet TG may be ~50x normal, and ß cell function has declined. The ceramide content has risen significantly, accompanied by a ~10-fold increase in DNA laddering (11) . The ß cell mass now begins to decrease and insulin production is further reduced, essentially wiping out the earlier gain of ß cells. Insulin production can no longer meet the increased production required to compensate for the insulin resistance and diabetes is now present (Fig. 6) . Figure 7 displays the morphological changes that characterize the ß cells of obese ZDF rats at the time of their failure.



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Figure 7. Upper panels: Immunofluorescent staining for insulin of an islet of Langerhans in an untreated ZDF fa/fa rat (left) or in a troglitazone (TGZ) -treated ZDF fa/fa rat (right). Note the reduction and irregular distribution of the insulin cells in the untreated islet. Middle panels: Immunofluorescent staining for glucose transporter-2 (Glut-2) of an islet in an untreated fa/fa rat (left) or in a TGZ-treated fa/fa rat (right). Note the marked reduction of Glut-2 staining on the insulin cells in the untreated fa/fa rat. Left upper and middle panels and right upper and middle panels are consecutive serial sections. Lower panels: aldehyde-fuchsin and Masson’s trichrome staining of islets in untreated, or TGZ-treated fa/fa rats to compare fibrosis. The purple-stained insulin cells are severely reduced in number in the islet of the untreated fa/fa rat (left), accompanied by a marked accumulation of collagen, which is stained in blue-green. The bars represent 50 µm.

In humans
Conclusive evidence that lipotoxic diabetes occurs in humans is lacking because of the unavailability of islets and other tissues for longitudinal measurements of TG. However, fat accumulation in human islets was first reported in 1902 (83) and found to be greater in diabetic islets than in nondiabetic controls (84) . As mentioned, a recent study of human islets has demonstrated increased neutral lipids in pancreatic sections and ß cells of older humans (66) . Furthermore, treatment with the lipopenic drug troglitazone (85) or with caloric restriction (86) , measures shown to lower the islet lipid content in rats (14 , 51) , improves glucose tolerance in both humans (84) and obese rats (14 , 86) . These findings are consistent with a common cause of disease in the two species.


   PREVENTION OF LIPOTOXICITY AND LIPOAPOPTOSIS
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
The dysfunctional and apoptotic consequences of lipotoxicity can be blocked at multiple sites (Fig. 3A ). Restoration of leptin action, reduction of the lipid excess, blockade of ceramide formation, and inhibition of iNOS are all effective (11 , 12 , 43) . Blocking the ectopic deposition of lipids by the daily administration of troglitazone to prediabetic fa/fa ZDF rats completely prevents both the diabetes (14) and the heart disease (33) that otherwise occur in 100% of untreated animals. Troglitazone appears to work by reducing lipogenesis in nonadipose tissues such as the islets (14) and heart (34) , while increasing lipogenesis in adipocytes. By lowering plasma FFA, it not only reduces substrate overload to tissues but also diminishes putative FA-mediated signals to nuclear receptors (87). Moreover, it may improve insulin sensitivity by minimizing activation of a serine/threonine kinase cascade postulated to block insulin receptor substrate association with phosphotidylinositol 3-kinase, a key event in insulin action (88). By confining lipogenesis to the adipocytes, the excessive apoptosis of ß-cells (14) and cardiac myocytes is prevented (34) . The iNOS inhibitors aminoguanidine and nicotinamide similarly prevent the diabetes (44) . Their efficacy in preventing lipotoxic heart disease has not been studied.

Profound alterations in 85% of mitochondria have been observed in ß cells of untreated rats (14); these are completely prevented by troglitazone therapy, as is the accompanying fibrosis and deformation of islets (Fig. 7C ). Loss of the ß cell glucose transporter GLUT-2 (Fig. 4D ) and of glucose-responsive insulin secretion (data not shown) is also prevented (14) . Similarly, the echocardiographic evidence of impaired myocardial contractility and the increased apoptosis are completely prevented by troglitazone treatment, in conjunction with a reduction of myocardial TG and ceramide content and iNOS expression (34) (Fig. 3A ).


   TRANSLATING 20th CENTURY SCIENCE INTO 21st CENTURY MEDICINE
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 
Here we have attempted to link important molecular discoveries of the last century [leptin (52) , ADD-1/SREBP-1 (29 30 31) , PPAR{alpha}, and {gamma} (87) ] to an impending health problem of the present century, the tissue disease and clinical consequences resulting from abnormal FA homeostasis caused by dietary excess. Although absolute proof is lacking, circumstantial evidence suggests that the findings described in rats may be relevant to human disease. The inexorable increase in the prevalence of human obesity and the decrease in its age of onset predicts a rising incidence of lipotoxic complications that could approach the 50% level in Pima Indians (88) . Since pharmacologic strategies that effectively prevent lipotoxic complications in rodents now exist, a concerted effort to minimize the impact of lipotoxic diseases in humans is indicated until a means of controlling obesity becomes available.

Note added in proof: The efficacy of troglitazone in improving the metabolic control of patients with lipodystrophy and diabetes. Ariuglu, E. et al. (2000) Ann. Int. Med. 133, 203–274.


   ACKNOWLEDGMENTS
 
We thank Susan Kennedy for outstanding secretarial work, Kay McCorkle, Nadine Dupont, Gérard Negro, and P.-Alain Rüttimann for excellent assistance with the illustrations, and R. Sanders Williams for critical review of the manuscript. This work supported by the Department of Veterans Affairs Institutional Support, the National Institutes of Health (NIH) (DK02700–37), the NIH/Juvenile Diabetes Foundation Diabetes Interdisciplinary Research Program, the Novo-Nordisk Corporation, and the Swiss National Science Foundation (L.O.).

Received for publication August 23, 2000. Accepted for publication September 15, 2000.


   REFERENCES
TOP
ABSTRACT
INTRODUCTION
NORMAL FATTY ACID (FA)...
ABNORMAL FA HOMEOSTASIS
MECHANISMS OF LIPOTOXICITY
GENETIC AND NONGENETIC CAUSES...
LIPOTOXIC DISORDERS: DO THEY...
PREVENTION OF LIPOTOXICITY AND...
TRANSLATING 20th CENTURY SCIENCE...
REFERENCES
 

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