Adipose tissue expresses tumor necrosis factor (TNF) and interleukin (IL)-6, which may cause obesity-related insulin resistance. We measured TNF and IL-6 expression in the adipose tissue of 50 lean and obese subjects without diabetes. Insulin sensitivity (S(I)) was determined by an intravenous glucose tolerance test with minimal-model analysis. When lean [body mass index (BMI) <25 kg/m(2)] and obese (BMI 30-40 kg/m(2)) subjects were compared, there was a 7.5-fold increase in TNF secretion (P < 0.05) from adipose tissue, and the TNF secretion was inversely related to S(I) (r = -0.42, P < 0.02). IL-6 was abundantly expressed by adipose tissue. In contrast to TNF, plasma (rather than adipose) IL-6 demonstrated the strongest relationship with obesity and insulin resistance. Plasma IL-6 was significantly higher in obese subjects and demonstrated a highly significant inverse relationship with S(I) (r = -0.71, P < 0.001). To separate the effects of BMI from S(I), subjects who were discordant for S(I) were matched for BMI, age, and gender. By use of this approach, subjects with low S(I) demonstrated a 3.0-fold increased level of TNF secretion from adipose tissue and a 2.3-fold higher plasma IL-6 level (P < 0.05) compared with matched subjects with a high S(I). Plasma IL-6 was significantly associated with plasma nonesterified fatty acid levels (r = 0.49, P < 0.002). Thus the local expression of TNF and plasma IL-6 are higher in subjects with obesity-related insulin resistance.

Adiponectin is a 29-kDa adipocyte protein that has been linked to the insulin resistance of obesity and lipodystrophy. To better understand the regulation of adiponectin expression, we measured plasma adiponectin and adipose tissue adiponectin mRNA levels in nondiabetic subjects with varying degrees of obesity and insulin resistance. Plasma adiponectin and adiponectin mRNA levels were highly correlated with each other (r ‫؍‬ 0.80, P < 0.001), and obese subjects expressed significantly lower levels of adiponectin. However, a significant sex difference in adiponectin expression was observed, especially in relatively lean subjects. When men and women with a BMI <30 kg/m 2 were compared, women had a twofold higher percent body fat, yet their plasma adiponectin levels were 65% higher (8.6 ؎ 1.1 and 14.2 ؎ 1.6 g/ml in men and women, respectively; P < 0.02). Plasma adiponectin had a strong association with insulin sensitivity index (S I ) (r ‫؍‬ 0.67, P < 0.0001, n ‫؍‬ 51) that was not affected by sex, but no relation with insulin secretion. To separate the effects of obesity (BMI) from S I , subjects who were discordant for S I were matched for BMI, age, and sex. Using this approach, insulin-sensitive subjects demonstrated a twofold higher plasma level of adiponectin (5.6 ؎ 0.6 and 11.2 ؎ 1.1 g/ml in insulin-resistant and insulin-sensitive subjects, respectively; P < 0.0005). Adiponectin expression was not related to plasma levels of leptin or interleukin-6. However, there was a significant inverse correlation between plasma adiponectin and tumor necrosis factor (TNF)-␣ mRNA expression (r ‫؍‬ ؊0.47, P < 0.005), and subjects with the highest levels of adiponectin mRNA expression secreted the lowest levels of TNF-␣ from their adipose tissue in vitro. Thus, adiponectin expression from adipose tissue is higher in lean subjects and women, and is associated with higher degrees of insulin sensitivity and lower TNF-␣ expression.

To examine the role of adipose-resident macrophages in insulin resistance, we examined the gene expression of CD68, a macrophage marker, along with macrophage chemoattractant protein-1 (MCP-1) in human subcutaneous adipose tissue using real-time RT-PCR. Both CD68 and MCP-1 mRNAs were expressed in human adipose tissue, primarily in the stromal vascular fraction. When measured in the adipose tissue from subjects with normal glucose tolerance, covering a wide range of BMI (21-51 kg/m 2 ) and insulin sensitivity (S I ) (0.6 -8.0 ؋ 10 ؊4 min ؊1 ⅐ U -1 ⅐ ml -1 ), CD68 mRNA abundance, which correlated with the number of CD68-positive cells by immunohistochemistry, tended to increase with BMI but was not statistically significant. However, there was a significant inverse relation between CD68 mRNA and S I (r ‫؍‬ ؊0.55, P ‫؍‬ 0.02). In addition, there was a strong positive relationship among adipose tissue CD68 mRNA, tumor necrosis factor-␣ (TNF-␣) secretion in vitro (r ‫؍‬ 0.79, P < 0.005), and plasma interleukin-6 (r ‫؍‬ 0.67, P < 0.005). To determine whether improving S I in subjects with impaired glucose tolerance (IGT) was associated with decreased CD68 expression, IGT subjects were treated for 10 weeks with pioglitazone or metformin. Pioglitazone increased S I by 60% and in the same subjects reduced both CD68 and MCP-1 mRNAs by >50%.

Acyl-coenzyme A:diacylglycerol transferase (DGAT), fatty acid synthetase (FAS), and LPL are three enzymes important in adipose tissue triglyceride accumulation. To study the relationship of DGAT1, FAS, and LPL with insulin, we examined adipose mRNA expression of these genes in subjects with a wide range of insulin sensitivity (S I ). DGAT1 and FAS (but not LPL) expression were strongly correlated with S I . In addition, the expression of DGAT1 and FAS (but not LPL) were higher in normal glucose-tolerant subjects compared with subjects with impaired glucose tolerance (IGT) (P , 0.005). To study the effects of insulin sensitizers, subjects with IGT were treated with pioglitazone or metformin for 10 weeks, and lipogenic enzymes were measured in adipose tissue. After pioglitazone treatment, DGAT1 expression was increased by 33 6 10% (P , 0.05) and FAS expression increased by 63 6 8% (P , 0.05); however, LPL expression was not altered. DGAT1, FAS, and LPL mRNA expression were not significantly changed after metformin treatment. The treatment of mice with rosiglitazone also resulted in an increase in adipose expression of DGAT1 by 2-to 3-fold, as did the treatment of 3T3 F442A adipocytes in vitro with thiazolidinediones.

Obesity-related insulin resistance may be caused by adipokines such as IL-6, which is known to be elevated with the insulin resistance syndrome. A previous study reported that IL-6 knockout mice (IL-6−/−) developed maturity onset obesity, with disturbed carbohydrate and lipid metabolism, and increased leptin levels. Because IL-6 is associated with insulin resistance, one might have expected IL-6−/− mice to be more insulin sensitive. We examined body weights of growing and older IL-6−/− mice and found them to be similar to wild-type (IL-6+/+) mice. Dual-energy X-ray absorptiometry analysis at 3 and 14 mo revealed no differences in body composition. There were no differences in fasting blood insulin and glucose or in triglycerides. To further characterize these mice, we fed 11-mo-old IL-6−/− and IL-6+/+ mice a high- (HF)- or low-fat diet for 14 wk, followed by insulin (ITT) and glucose tolerance tests (GTT). An ITT showed insulin resistance in the HF animals but no difference due to genotype. In the GTT, IL-6−/− mice demonstrated elevated postinjection glucose levels by 60% compared with IL-6+/+ but only in the HF group. Although IL-6−/− mice gained weight and white adipose tissue (WAT) with the HF diet, they gained less weight than the IL-6+/+ mice. Total lipoprotein lipase activity in WAT, muscle, and postheparin plasma was unchanged in the IL-6 −/− mice compared with IL-6+/+ mice. There were no differences in plasma leptin or TNF-α due to genotype. Plasma adiponectin was ∼53% higher (71.7 ± 14.1 μg/ml) in IL-6−/− mice than in IL-6+/+ mice but only in the HF group. Thus these data show that IL-6−/− mice do not demonstrate obesity, fasting hyperglycemia, or abnormal lipid metabolism, although HF IL-6−/− mice demonstrate elevated glucose after a GTT.

Banga A, Unal R, Tripathi P, Pokrovskaya I, Owens RJ, Kern PA, Ranganathan G. Adiponectin translation is increased by the PPAR␥ agonists pioglitazone and -3 fatty acids.

The perilipins are highly phosphorylated adipocyte proteins that are localized at the surface of the lipid droplet. With activation by protein kinase A, perilipins translocate away from the lipid droplet and allow hormone-sensitive lipase to hydrolyze the adipocyte triglycerides to release nonesterified fatty acids (NEFA). Because of the potential importance of adipocyte lipolysis to obesity and insulin resistance, we measured perilipin protein and mRNA levels in nondiabetic subjects with varying degrees of insulin resistance. By Northern and Western blotting, we could detect perilipin A, but not perilipin B. Perilipin A protein and mRNA levels were quantitated and were highly correlated with each other. There was a significant positive relationship between perilipin expression and obesity (r ‫؍‬ 0.55; P < 0.01, perilipin mRNA vs. percent body fat). However, there was no significant relationship between perilipin expression and blood NEFA, nor was there a significant relationship between perilipin expression and insulin resistance, using the insulin sensitivity index derived from the iv glucose tolerance test with minimal modeling. In addition, there was no significant relationship between perilipin and adipocyte or systemic inflammatory markers, such as TNF␣, IL-6, and adiponectin. Thus, perilipin was elevated in obese subjects, perhaps as a compensatory mechanism to limit basal lipolysis. However, there was no relationship between perilipin and insulin resistance. (J Clin Endocrinol Metab 89: [1352][1353][1354][1355][1356][1357][1358] 2004)

A cDNA library, prepared from poly(A)+ RNA isolated from quiescent AKR-2B cells 4 hr after stimulation with epidermal growth factor in the presence of cycloheximide, was screened to identify RNA transcripts whose abundance is specifically increased as a primary response to growth stimulation. Approximately 40% of the inducible clones detected by this procedure corresponded to either cytoskeletal 13-or y-actin genes. One nonactin clone, designated c99, was found to be derived from an 8.5-kilobase RNA whose abundance began to increase as early as 30 min after stimulation. DNA sequencing established the identity of this RNA as fibronectin. Several additional mitogens were then tested and found to efficiently induce fibronectin mRNA. These included fetal calf serum, platelet-derived growth factor, and transforming growth factor type P. For at least one inducer, fetal calf serum, the increase in mRNA was preceded by an increase in fibronectin gene transcription. This increase was rapid, reaching maximal levels within 10 min, and was accompanied by near-coordinate increases in both c-fos and 13-actin transcription. These results indicate that fibronectin is a member of a class of "early-response" genes, typified by c-fos and including j3-actin, whose rapid expression may be important in mediating cellular responses to peptide growth factors.Peptide growth factors govern a wide range of cellular activity including embryonic development and differentiation, maintenance of cell-type homeostasis, and wound healing. Genetic and biochemical evidence indicates that cellular responses to growth factors are mediated, in part, through the regulation of specific gene expression. Thus, one approach to understanding how growth factors influence cell behavior is to clone and identify responsive genes. Estimates derived from such endeavors are inexact (1) but suggest that growth factors exert their influence through the action of perhaps 50-100 genes.In studies of cell growth control, particular attention has been focused on genes expressed soon after growth-factor stimulation of quiescent cells in culture. Such genes presumably encode proteins important to growth stimulation and have been shown to include the c-fos and c-myc protooncogenes (2, 3). Interestingly, enhanced expression of the cytoskeletal actin genes is also a prominent and rapid response to mitogenic stimuli in several cell types (4-7). This contrasts with the traditional view of the cytoskeletal actins as passive structural proteins but is broadly consistent with other data suggesting an important role for actin microfilaments in mediating cellular responses to growth factors (8) and in neoplastic transformation (9).Although the exact role of actin in cell growth control is not clear, the degree of anchorage of cells to the extracellular matrix has long been known to influence both cell division and cell differentiation (reviewed in ref. 10). Among the major mediators of cell adhesion are transmembrane glycoproteins that link actin microfilaments on the cytop...