Brent E. Wisse

The Inflammatory Syndrome: The Role of Adipose Tissue Cytokines in Metabolic Disorders Linked to Obesity

The metabolic effects of obesity have made this highly prevalent disease one of the most common risk factors for diabetes, hypertension, and atherosclerosis, the leading causes of end-stage renal failure. However, obesity per se, as defined by body mass index, is less predictive of the development of these diseases than is the presence of a constellation of obesity-related abnormalities now known as the metabolic syndrome. Recognition of this syndrome, which can readily be identified in clinical settings using defined threshold values for waist circumference, BP, fasting glucose, and dyslipidemia, allows for earlier intervention in these high-risk patients. Systemic insulin resistance has been implicated as one possible factor that links visceral obesity to adverse metabolic consequences; however, the mechanism whereby adipose tissue causes alterations in insulin sensitivity remains unclear. Infection and inflammation are commonly associated with insulin resistance, and visceral obesity is associated with a chronic, low-grade inflammatory state, suggesting that inflammation may be a potential mechanism whereby obesity leads to insulin resistance. Moreover, adipose tissue is now recognized as an immune organ that secretes numerous immunomodulatory factors and seems to be a significant source of inflammatory signals known to cause insulin resistance. Therefore, inflammation within white adipose tissue may be a crucial step contributing to the emergence of many of the pathologic features that characterize the metabolic syndrome and result in diabetes and atherosclerosis. This review describes the role of proinflammatory cytokines and hormones released by adipose tissue in generating the chronic inflammatory profile associated with visceral obesity.

Obesity is associated with hypothalamic injury in rodents and humans

Rodent models of obesity induced by consuming high-fat diet (HFD) are characterized by inflammation both in peripheral tissues and in hypothalamic areas critical for energy homeostasis. Here we report that unlike inflammation in peripheral tissues, which develops as a consequence of obesity, hypothalamic inflammatory signaling was evident in both rats and mice within 1 to 3 days of HFD onset, prior to substantial weight gain. Furthermore, both reactive gliosis and markers suggestive of neuron injury were evident in the hypothalamic arcuate nucleus of rats and mice within the first week of HFD feeding. Although these responses temporarily subsided, suggesting that neuroprotective mechanisms may initially limit the damage, with continued HFD feeding, inflammation and gliosis returned permanently to the mediobasal hypothalamus. Consistent with these data in rodents, we found evidence of increased gliosis in the mediobasal hypothalamus of obese humans, as assessed by MRI. These findings collectively suggest that, in both humans and rodent models, obesity is associated with neuronal injury in a brain area crucial for body weight control.

Vascular Inflammation, Insulin Resistance, and Reduced Nitric Oxide Production Precede the Onset of Peripheral Insulin Resistance

Objectives—Obesity causes inflammation and insulin resistance in the vasculature as well as in tissues involved in glucose metabolism such as liver, muscle, and adipose tissue. To investigate the relative susceptibility of vascular tissue to these effects, we determined the time course over which inflammation and insulin resistance develops in various tissues of mice with diet-induced obesity (DIO) and compared these tissue-based responses to changes in circulating inflammatory markers. Methods and Results—Adult male C57BL/6 mice were fed either a control low-fat diet (LF; 10% saturated fat) or a high-fat diet (HF, 60% saturated fat) for durations ranging between 1 to 14 weeks. Cellular inflammation and insulin resistance were assessed by measuring phospho-I&kgr;Bα and insulin-induced phosphorylation of Akt, respectively, in extracts of thoracic aorta, liver, skeletal muscle, and visceral fat. As expected, HF feeding induced rapid increases of body weight, fat mass, and fasting insulin levels compared to controls, each of which achieved statistical significance within 4 weeks. Whereas plasma markers of inflammation became elevated relatively late in the course of DIO (eg, serum amyloid A [SAA], by Week 14), levels of phospho-I&kgr;Bα in aortic lysates were elevated by 2-fold within the first week. The early onset of vascular inflammation was accompanied by biochemical evidence of both endothelial dysfunction (reduced nitric oxide production; induction of intracellular adhesion molecule-1 and vascular cell adhesion molecule-1) and insulin resistance (impaired insulin-induced phosphorylation of Akt and eNOS). Although inflammation and insulin resistance were also detected in skeletal muscle and liver of HF-fed animals, these responses were observed much later (between 4 and 8 weeks of HF feeding), and they were not detected in visceral adipose tissue until 14 weeks. Conclusions—During obesity induced by HF feeding, inflammation and insulin resistance develop in the vasculature well before these responses are detected in muscle, liver, or adipose tissue. This observation suggests that the vasculature is more susceptible than other tissues to the deleterious effects of nutrient overload.

Identification of Body Fat Mass as a Major Determinant of Metabolic Rate in Mice

OBJECTIVE Analysis of energy expenditure (EE) in mice is essential to obesity research. Since EE varies with body mass, comparisons between lean and obese mice are confounded unless EE is normalized to account for body mass differences. We 1) assessed the validity of ratio-based EE normalization involving division of EE by either total body mass (TBM) or lean body mass (LBM), 2) compared the independent contributions of LBM and fat mass (FM) to EE, and 3) investigated whether leptin contributes to the link between FM and EE. RESEARCH DESIGN AND METHODS We used regression modeling of calorimetry and body composition data in 137 mice to estimate the independent contributions of LBM and FM to EE. Subcutaneous administration of leptin or vehicle to 28 obese ob/ob mice and 32 fasting wild-type mice was used to determine if FM affects EE via a leptin-dependent mechanism. RESULTS Division of EE by either TBM or LBM is confounded by body mass variation. The contribution of FM to EE is comparable to that of LBM in normal mice (expressed per gram of tissue) but is absent in leptin-deficient ob/ob mice. When leptin is administered at physiological doses, the plasma leptin concentration supplants FM as an independent determinant of EE in both ob/ob mice and normal mice rendered leptin-deficient by fasting. CONCLUSIONS The contribution of FM to EE is substantially greater than predicted from the metabolic cost of adipose tissue per se, and the mechanism underlying this effect is leptin dependent. Regression-based approaches that account for variation in both FM and LBM are recommended for normalization of EE in mice.

Hypothalamic inflammation and energy homeostasis: resolving the paradox.

Determining the effect of hypothalamic inflammatory signals on energy balance presents a paradox. On the one hand, a large body of work has identified inflammatory signaling in the hypothalamus as an essential mediator of the sickness response--the anorexia, cachexia, fever, inactivity, lethargy, anhedonia and adipsia that are triggered by systemic inflammatory stimuli and promote negative energy balance. On the other hand, numerous recent studies implicate inflammatory activation within the hypothalamus as a key factor whereby high-fat diets--and saturated fats in particular--cause central leptin and insulin resistance and thereby promote the defense of elevated body weight. This paradox will likely remain unresolved until several issues have been addressed. Firstly, the hypothalamus--unlike many peripheral inflamed tissues--is an extremely heterogeneous tissue comprised of astrocytes, oligodendrocytes, microglia, endothelial cells, ependymal cells as well as numerous neuronal subgroups. Determining exactly which cells activate defined inflammatory signals in response to a particular stimulus--i.e. sepsis vs. nutrient excess--may yield critical clues. Secondly, for the sake of simplicity many studies evaluate inflammation as an on/off phenomenon. More realistically, inflammatory signaling occurs as a cascade or cycle that changes and progresses over time. Accordingly, even within the same cell type, the low-grade, chronic signal induced by nutrient excess may invoke a different cascade of signals than a strong, acute signal such as sepsis. In addition, because tolerance can develop to certain inflammatory mediators, physiological outcomes may not correlate with early biochemical markers. Lastly, the neuroanatomical location, magnitude, and duration of the inflammatory stimulus can undoubtedly influence the net CNS response. Rigorously evaluating the progression of the inflammatory signaling cascade within specific hypothalamic cell types is a key next step towards resolving the paradox surrounding the effect of inflammatory signaling on energy homeostasis.

Leptin Activates a Novel CNS Mechanism for Insulin-Independent Normalization of Severe Diabetic Hyperglycemia

The brain has emerged as a target for the insulin-sensitizing effects of several hormonal and nutrient-related signals. The current studies were undertaken to investigate mechanisms whereby leptin lowers circulating blood glucose levels independently of insulin. After extending previous evidence that leptin infusion directly into the lateral cerebral ventricle ameliorates hyperglycemia in rats with streptozotocin-induced uncontrolled diabetes mellitus, we showed that the underlying mechanism is independent of changes of food intake, urinary glucose excretion, or recovery of pancreatic β-cells. Instead, leptin action in the brain potently suppresses hepatic glucose production while increasing tissue glucose uptake despite persistent, severe insulin deficiency. This leptin action is distinct from its previously reported effect to increase insulin sensitivity in the liver and offers compelling evidence that the brain has the capacity to normalize diabetic hyperglycemia in the presence of sufficient amounts of central nervous system leptin.

Hypothalamic Inflammation: Marker or Mechanism of Obesity Pathogenesis?

Obesity and its associated metabolic and cardiovascular disorders are among the most challenging health problems confronting developed countries. Not only is obesity remarkably common, affecting more than one-third of U.S. adults, and very challenging to treat, but it is also tightly linked to type 2 diabetes and related metabolic disorders. A major obstacle to effective obesity treatment is that lost weight tends to be regained over time (1). Although the mechanisms underlying recovery of lost weight are incompletely understood, a large literature suggests that body fat stores are subject to homeostatic regulation, and that this process occurs in obese as well as normal-weight individuals. From this perspective, obesity can be viewed as a disorder in which the biologically defended level of body fat mass is increased. Recent findings implicate inflammation in key hypothalamic areas for body weight control in this process. In this review, we present an overview of energy homeostasis—the biological process that underlies the control of body fat mass—and describe evidence that defects in this regulatory system contribute to obesity pathogenesis. We then address molecular characteristics of hypothalamic inflammation and their implications for obesity pathogenesis (detailed more extensively in refs. 2,3), followed by evidence linking high-fat diet (HFD) feeding to neuropathological alteration of key hypothalamic areas controlling energy balance. We conclude by considering how cell-cell interactions may contribute to this deleterious hypothalamic response and the implications of these interactions for obesity pathogenesis. ### Homeostatic response to weight loss. Following a period of caloric restriction, lost weight is gradually but inexorably recovered in most individuals. This effect involves the capacity of the brain to sense the reduction of body energy stores and activate responses to compensate for this deficit. In simple terms, voluntary weight loss triggers increases of both appetite and energy efficiency, such that both sides of the energy balance equation shift …

Leptin Deficiency Causes Insulin Resistance Induced by Uncontrolled Diabetes

OBJECTIVE Depletion of body fat stores during uncontrolled, insulin-deficient diabetes (uDM) results in markedly reduced plasma leptin levels. This study investigated the role of leptin deficiency in the genesis of severe insulin resistance and related metabolic and neuroendocrine derangements induced by uDM. RESEARCH DESIGN AND METHODS Adult male Wistar rats remained nondiabetic or were injected with the β-cell toxin, streptozotocin (STZ) to induce uDM and subsequently underwent subcutaneous implantation of an osmotic minipump containing either vehicle or leptin at a dose (150 μg/kg/day) designed to replace leptin at nondiabetic plasma levels. To control for leptin effects on food intake, another group of STZ-injected animals were pair fed to the intake of those receiving leptin. Food intake, body weight, and blood glucose levels were measured daily, with body composition and indirect calorimetry performed on day 11, and an insulin tolerance test to measure insulin sensitivity performed on day 16. Plasma hormone and substrate levels, hepatic gluconeogenic gene expression, and measures of tissue insulin signal transduction were also measured. RESULTS Physiologic leptin replacement prevented insulin resistance in uDM via a mechanism unrelated to changes in food intake or body weight. This effect was associated with reduced total body fat and hepatic triglyceride content, preservation of lean mass, and improved insulin signal transduction via the insulin receptor substrate–phosphatidylinositol-3-hydroxy kinase pathway in the liver, but not in skeletal muscle or adipose tissue. Although physiologic leptin replacement lowered blood glucose levels only slightly, it fully normalized elevated plasma glucagon and corticosterone levels and reversed the increased hepatic expression of gluconeogenic enzymes characteristic of rats with uDM. CONCLUSIONS We conclude that leptin deficiency plays a key role in the pathogenesis of severe insulin resistance and related endocrine disorders in uDM. Treatment of diabetes in humans may benefit from correction of leptin deficiency as well as insulin deficiency.

Identification of a physiological role for leptin in the regulation of ambulatory activity and wheel running in mice

Mechanisms regulating spontaneous physical activity remain poorly characterized despite evidence of influential genetic and acquired factors. We evaluated ambulatory activity and wheel running in leptin-deficient ob/ob mice and in wild-type mice rendered hypoleptinemic by fasting in both the presence and absence of subcutaneous leptin administration. In ob/ob mice, leptin treatment to plasma levels characteristic of wild-type mice acutely increased both ambulatory activity (by 4,000 ± 200 beam breaks/dark cycle, P < 0.05) and total energy expenditure (TEE; by 0.11 ± 0.01 kcal/h during the dark cycle, P < 0.05) in a dose-dependent manner and acutely increased wheel running (+350%, P < 0.05). Fasting potently increased ambulatory activity and wheel running in wild-type mice (AA: +25%, P < 0.05; wheel running: +80%, P < 0.05), and the effect of fasting was more pronounced in ob/ob mice (AA: +400%, P < 0.05; wheel running: +1,600%, P < 0.05). However, unlike what occurred in ad libitum-fed ob/ob mice, physiological leptin replacement attenuated or prevented fasting-induced increases of ambulatory activity and wheel running in both wild-type and ob/ob mice. Thus, plasma leptin is a physiological regulator of spontaneous physical activity, but the nature of leptins effect on activity is dependent on food availability.

Forebrain melanocortin signaling enhances the hindbrain satiety response to CCK-8

Melanocortin 4 receptors (MC4R) are hypothesized to mediate the central nervous system actions of leptin to enhance the satiety effects of cholecystokinin (CCK). To further elucidate this mechanism, we confirmed that peripheral administration of CCK-8 is less effective in producing this effect in MC4R-deficient mice (MC4R(-/-)). Whereas intraperitoneal (ip) CCK-8 at 0.75 nmol/kg lean body mass (lbm) suppressed food intake in wild-type mice, CCK-8 doses of 7.5 nmol/kg lbm were required to attenuate food intake in MC4R(-/-) mice. To determine whether melanocortin signaling in the hypothalamic paraventricular nucleus (PVN) participates in regulating this CCK satiety response, we administered the MC3/MC4R antagonist, SHU9119, into the PVN of rats before ip CCK-8 administration. PVN administration of SHU9119 attenuated the ability of CCK-8 to reduce 30-min food intake by 20%. To determine whether MC4R are expressed by PVN neurons that project directly to hindbrain nuclei involved in the satiety response to ip CCK-8, the retrograde tracer fluorescent cholera toxin subunit B was injected into the nucleus tractus solitarius (NTS) of the hindbrain. After 4 days, labeled PVN neurons were collected by laser capture microdissection and found to express MC4R mRNA by quantitative RT-PCR analysis. These data provide evidence for a neuroanatomical link between hypothalamic melanocortin signaling in the PVN and NTS neurons that regulate food intake. These findings highlight the contribution of melanocortin signaling in the PVN toward regulating the satiety effects of CCK-8 while acknowledging that melanocortin-dependent pathways in other brain regions and/or melanocortin-independent mechanisms are also important in this mechanism.