Obesity alters adipose tissue metabolic and endocrine function and leads to an increased release of fatty acids, hormones, and proinflammatory molecules that contribute to obesity associated complications. To further characterize the changes that occur in adipose tissue with increasing adiposity, we profiled transcript expression in perigonadal adipose tissue from groups of mice in which adiposity varied due to sex, diet, and the obesity-related mutations agouti (Ay) and obese (Lepob). We found that the expression of 1,304 transcripts correlated significantly with body mass. Of the 100 most significantly correlated genes, 30% encoded proteins that are characteristic of macrophages and are positively correlated with body mass. Immunohistochemical analysis of perigonadal, perirenal, mesenteric, and subcutaneous adipose tissue revealed that the percentage of cells expressing the macrophage marker F4/80 (F4/80+) was significantly and positively correlated with both adipocyte size and body mass. Similar relationships were found in human subcutaneous adipose tissue stained for the macrophage antigen CD68. Bone marrow transplant studies and quantitation of macrophage number in adipose tissue from macrophage-deficient (Csf1op/op) mice suggest that these F4/80+ cells are CSF-1 dependent, bone marrow-derived adipose tissue macrophages. Expression analysis of macrophage and nonmacrophage cell populations isolated from adipose tissue demonstrates that adipose tissue macrophages are responsible for almost all adipose tissue TNF-alpha expression and significant amounts of iNOS and IL-6 expression. Adipose tissue macrophage numbers increase in obesity and participate in inflammatory pathways that are activated in adipose tissues of obese individuals.

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Obesity is associated with macrophage accumulation in adipose tissue

Abundant data link hypercholesterolaemia to atherogenesis. However, only recently have we appreciated that inflammatory mechanisms couple dyslipidaemia to atheroma formation. Leukocyte recruitment and expression of pro-inflammatory cytokines characterize early atherogenesis, and malfunction of inflammatory mediators mutes atheroma formation in mice. Moreover, inflammatory pathways promote thrombosis, a late and dreaded complication of atherosclerosis responsible for myocardial infarctions and most strokes. The new appreciation of the role of inflammation in atherosclerosis provides a mechanistic framework for understanding the clinical benefits of lipid-lowering therapies. Identifying the triggers for inflammation and unravelling the details of inflammatory pathways may eventually furnish new therapeutic targets.

Inflammation in atherosclerosis

Adiponectin is an adipocyte-derived hormone. Recent genome-wide scans have mapped a susceptibility locus for type 2 diabetes and metabolic syndrome to chromosome 3q27, where the gene encoding adiponectin is located. Here we show that decreased expression of adiponectin correlates with insulin resistance in mouse models of altered insulin sensitivity. Adiponectin decreases insulin resistance by decreasing triglyceride content in muscle and liver in obese mice. This effect results from increased expression of molecules involved in both fatty-acid combustion and energy dissipation in muscle. Moreover, insulin resistance in lipoatrophic mice was completely reversed by the combination of physiological doses of adiponectin and leptin, but only partially by either adiponectin or leptin alone. We conclude that decreased adiponectin is implicated in the development of insulin resistance in mouse models of both obesity and lipoatrophy. These data also indicate that the replenishment of adiponectin might provide a novel treatment modality for insulin resistance and type 2 diabetes.

The fat-derived hormone adiponectin reverses insulin resistance associated with both lipoatrophy and obesity

Obesity is a principal causative factor in the development of metabolic syndrome. Here we report that increased oxidative stress in accumulated fat is an important pathogenic mechanism of obesity-associated metabolic syndrome. Fat accumulation correlated with systemic oxidative stress in humans and mice. Production of ROS increased selectively in adipose tissue of obese mice, accompanied by augmented expression of NADPH oxidase and decreased expression of antioxidative enzymes. In cultured adipocytes, elevated levels of fatty acids increased oxidative stress via NADPH oxidase activation, and oxidative stress caused dysregulated production of adipocytokines (fat-derived hormones), including adiponectin, plasminogen activator inhibitor-1, IL-6, and monocyte chemotactic protein-1. Finally, in obese mice, treatment with NADPH oxidase inhibitor reduced ROS production in adipose tissue, attenuated the dysregulation of adipocytokines, and improved diabetes, hyperlipidemia, and hepatic steatosis. Collectively, our results suggest that increased oxidative stress in accumulated fat is an early instigator of metabolic syndrome and that the redox state in adipose tissue is a potentially useful therapeutic target for obesity-associated metabolic syndrome.

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Increased oxidative stress in obesity and its impact on metabolic syndrome

Adiponectin (Ad) is a hormone secreted by adipocytes that regulates energy homeostasis and glucose and lipid metabolism. However, the signaling pathways that mediate the metabolic effects of Ad remain poorly identified. Here we show that phosphorylation and activation of the 5'-AMP-activated protein kinase (AMPK) are stimulated with globular and full-length Ad in skeletal muscle and only with full-length Ad in the liver. In parallel with its activation of AMPK, Ad stimulates phosphorylation of acetyl coenzyme A carboxylase (ACC), fatty-acid oxidation, glucose uptake and lactate production in myocytes, phosphorylation of ACC and reduction of molecules involved in gluconeogenesis in the liver, and reduction of glucose levels in vivo. Blocking AMPK activation by dominant-negative mutant inhibits each of these effects, indicating that stimulation of glucose utilization and fatty-acid oxidation by Ad occurs through activation of AMPK. Our data may provide a novel paradigm that an adipocyte-derived antidiabetic hormone, Ad, activates AMPK, thereby directly regulating glucose metabolism and insulin sensitivity in vitro and in vivo.

Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase

Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity.

IL-18 is a novel cytokine that induces interferon (IFN)-gamma secretion and plays an important role in antitumor immunity. In the present study, we constructed plasmid vectors encoding the murine mature IL-18 cDNA linked with the Igkappa leader sequence and the pro-IL-18 cDNA to estimate the efficacy of the mature IL- 18 vector and to evaluate IL-18--producing tumor cells as a tumor vaccine. Colon 26 cells were transfected with the abovementioned vectors or with vector alone (mock). Reverse transcription-polymerase chain reaction analysis showed increased expression of murine IL-18 cDNA in both mature IL-18 and pro-IL-18 transfectants in comparison to that in mock transfected cells. The ability of the culture supernatants of mature IL-18 transfectants to induce IFN-gamma secretion was extremely high (40-140 pg/10(6) cells) in comparison to that of pro-IL-18 transfectants (4-18 pg/10(6) cells). When injected into syngeneic BALB/c mice, the growth of mature IL-18 transfectants, but not pro-IL-18 transfectants, was significantly less than that in mock transfected cells ( P< .01, by ANOVA and analysis of covariance). In addition, injection of colon 26 or Meth-A cells into mice immunized with a mature IL-18 transfectant revealed acquired immunity. Depletion of natural killer cells did not affect the growth of transfectants. However, the growth inhibitory effects were partially abrogated following treatment with anti-CD4+ and anti-CD8+ antibodies. These data suggest that the rejection of mature IL-18/colon 26 cells was mediated through T-cell activation. Gene therapy using mature IL-18 transfectants containing a plasmid vector and the Igkappa leader sequence may be a useful tumor vaccine.

Successful immunogene therapy using colon cancer cells (colon 26) transfected with plasmid vector containing mature interleukin-18 cDNA and the Igκ leader sequence

It remains controversial whether or not a correlation exists between serum leptin levels and insulin resistance, and, if such a correlation does exist, whether it is independent of adiposity. To investigate the possible existence of an independent correlation, we have assessed serum leptin levels and insulin resistance in Japanese diabetic and non-diabetic subjects by means of Homeostatic Model Assessment (HOMA-R). Sixty-four Japanese patients with Type 2 diabetes mellitus (DM) (33 men and 31 women) and 53 sex-, age-, and body mass index (BMI)-matched non-diabetic adults (29 men and 24 women) were enrolled. The fasting plasma level of glucose (FPG) and the fasting serum levels of immunoreactive insulin (FIRI) and leptin were determined. Multiple linear regression analysis demonstrated that, in both male diabetic and male non-diabetic subjects, HOMA-R and BMI were independently correlated with serum leptin levels. In females, BMI, but not HOMA-R, was correlated to the serum levels of leptin in both groups. There was no statistically significant difference in the partial regression coefficients between male diabetic and male non-diabetic subjects. These results suggest that the correlation of HOMA-R to the serum levels of leptin in females is dependent on BMI. In males, the relationship between serum leptin levels and the insulin resistance was not affected by the extent of glucose intolerance.

Yasuichi Ohmoto

Papers

Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity.

Successful immunogene therapy using colon cancer cells (colon 26) transfected with plasmid vector containing mature interleukin-18 cDNA and the Igκ leader sequence

Gender-Related Difference in Relationship between Insulin Resistance and Serum Leptin Level in Japanese Type 2 Diabetic and Non-Diabetic Subjects.