Acrp30 is a circulating protein synthesized in adipose tissue. A single injection in mice of purified recombinant Acrp30 leads to a 2-3-fold elevation in circulating Acrp30 levels, which triggers a transient decrease in basal glucose levels. Similar treatment in ob/ob, NOD (non-obese diabetic) or streptozotocin-treated mice transiently abolishes hyperglycemia. This effect on glucose is not associated with an increase in insulin levels. Moreover, in isolated hepatocytes, Acrp30 increases the ability of sub-physiological levels of insulin to suppress glucose production. We thus propose that Acrp30 is a potent insulin enhancer linking adipose tissue and whole-body glucose metabolism.

Adiponectin is an adipocyte-specific secretory protein that circulates in serum as a hexamer of relatively low molecular weight (LMW) and a larger multimeric structure of high molecular weight (HMW). Serum levels of the protein correlate with systemic insulin sensitivity. The full-length protein affects hepatic gluconeogenesis through improved insulin sensitivity, and a proteolytic fragment of adiponectin stimulates ␤ oxidation in muscle. Here, we show that the ratio, and not the absolute amounts, between these two oligomeric forms (HMW to LMW) is critical in determining insulin sensitivity. We define a new index, S A , that can be calculated as the ratio of HMW/(HMW ؉ LMW). db/db mice, despite similar total adiponectin levels, display decreased S A values compared with wild type littermates, as do type II diabetic patients compared with insulin-sensitive individuals. Furthermore, S A improves with peroxisome proliferator-activated receptor-␥ agonist treatment (thiazolidinedione; TZD) in mice and humans. We demonstrate that changes in S A in a number of type 2 diabetic cohorts serve as a quantitative indicator of improvements in insulin sensitivity obtained during TZD treatment, whereas changes in total serum adiponectin levels do not correlate well at the individual level. Acute alterations in S A (⌬S A ) are strongly correlated with improvements in hepatic insulin sensitivity and are less relevant as an indicator of improved muscle insulin sensitivity in response to TZD treatment, further underscoring the conclusions from previous clamp studies that suggested that the liver is the primary site of action for the full-length protein. These observations suggest that the HMW adiponectin complex is the active form of this protein, which we directly demonstrate in vivo by its ability to depress serum glucose levels in a dose-dependent manner.

Acrp30/adiponectin is an adipocyte-specific secretory protein that has recently been implicated as a mediator of systemic insulin sensitivity with liver and muscle as target organs. Acrp30 is found as two forms in serum, as a lower molecular weight trimer-dimer and a high molecular weight complex. Little is know about the regulation and significance of these Acrp30 complexes in serum and about the events that lead to the generation of the bioactive ligand. Here, we show that there is a profound sexual dimorphism of Acrp30 levels and complex distribution in serum. Female mice display significantly higher levels of the high molecular weight complex in serum than males. In both females and males, levels of the high molecular weight complex are significantly reduced in response to a systemic increase of insulin. The ratio of the two complexes is restored upon normalization of glucose levels. Structurally, we show that oligomer formation of Acrp30 critically depends on disulfide bond formation mediated by Cys-39. Mutation of Cys-39 results in trimers that are subject to proteolytic cleavage in the collagenous domain. Surprisingly, Acrp30(C39S) or wild-type Acrp30 treated with dithiothreitol are significantly more bioactive than the higher order oligomeric forms of the protein with respect to reduction of serum glucose levels. Furthermore, treatment of primary hepatocytes with trimeric and higher order forms of Acrp30 confirms that the increased bioactivity seen in vivo is reflected in an augmented potency to reduce glucose output in the presence of gluconeogenic stimuli. Combined, these results shed new light on the regulation of this complex protein and suggest a new model for in vivo activation of the protein, implicating a serum reductase activity.Adipose has been under appreciated as an endocrine tissue for decades because of the prevalent opinion that it served merely as storage for lipids. Recently, however, the importance of adipocytes to whole body energy homeostasis and metabolism has been underscored by several reports focusing on secreted products of adipocytes (1-4). There has been increased interest in adipose tissue as an endocrine organ, and several of these secreted proteins, termed adipokines, are currently undergoing extensive study regarding roles as divergent as feeding behavior to cardiovascular protection. For instance, leptin, the gene disrupted in ob/ob mice, has central roles in the hypothalamus, as well as peripheral effects in liver, muscle, and endothelial cells (5). Other adipose-secreted products, such as tumor necrosis factor ␣ and adipsin (complement factor D), have well established functions in innate immunity (6 -9). The recently identified adipokine resistin has been implicated as a modulator of insulin sensitivity and is also being studied for its effects on metabolism (4, 10).Acrp30 (also known as adiponectin, AdipoQ, and GBP28) is an adipokine exclusively synthesized and secreted by adipocytes (11-14). Acrp30 has recently been shown to influence glucose homeostasis and insulin se...

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Caveolae organelles and caveolin-1 protein expression are most abundant in adipocytes and endothelial cells. Our initial report on mice lacking caveolin-1 (Cav-1) demonstrated a loss of caveolae and perturbations in endothelial cell function. More recently, however, observation of the Cav-1-deficient cohorts into old age revealed significantly lower body weights, as compared with wild-type controls. These results suggest that Cav-1 null mice may have problems with lipid metabolism and/or adipocyte functioning. To test this hypothesis directly, we placed a cohort of wild-type and Cav-1 null mice on a high fat diet. Interestingly, despite being hyperphagic, Cav-1 null mice show overt resistance to diet-induced obesity. As predicted, adipocytes from Cav-1 null null mice lack caveolae membranes. Early on, a lack of caveolin-1 selectively affects only the female mammary gland fat pad and results in a near complete ablation of the hypo-dermal fat layer. There are also indications of generalized adipose tissue pathology. With increasing age, a systemic decompensation in lipid accumulation occurs resulting in dramatically smaller fat pads, histologically reduced adipocyte cell diameter, and a poorly differentiated/hypercellular white adipose parenchyma. To gain mechanistic insights into this phenotype, we show that, although serum insulin, glucose, and cholesterol levels are entirely normal, Cav-1 null mice have severely elevated triglyceride and free fatty acid levels, especially in the postprandial state. However, this build-up of triglyceriderich chylomicrons/very low density lipoproteins is not due to perturbed lipoprotein lipase activity, a major culprit of isolated hypertriglyceridemia. The lean body phenotype and metabolic defects observed in Cav-1 null mice are consistent with the previously proposed functions of caveolin-1 and caveolae in adipocytes. Our results show for the first time a clear role for caveolins in systemic lipid homeostasis in vivo and place caveolin-1/ caveolae as major factors in hyperlipidemias and obesity.

Adiponectin or adipocyte complement-related protein of 30 kDa (Acrp30) is a circulating protein produced exclusively in adipocytes. Circulating Acrp30 levels have been associated with insulin sensitivity in adult mice and humans, yet the Acrp30 profile over the lifespan and its hormonal regulation in vivo have not been previously described. Hence, we set forth to determine whether hormonal and metabolic changes associated with sexual maturation, reproduction, aging, and calorie restriction affect Acrp30. In mice, Acrp30 levels increase during sexual maturation by 4-fold in males and 10-fold in females. Neonatal castration (CX) allows Acrp30 of adults to reach female levels. CX in adults does not lead to female Acrp30 levels unless glucocorticoid exposure is elevated simultaneously by implant. Ovariectomy of infant mice does not interfere with the pubertal rise of Acrp30. However, ovariectomy in adults increases Acrp30. Estrogen suppressed Acrp30 in mice and 3T3-L1 adipocytes. In parallel to changes in estrogen action, Acrp30 decreased in late gestation but increased in both calorie-restricted and old (anovulatory) mice. The reduction of Acrp30 in lactating dams is consistent with a suppressive effect of prolactin and a stimulating effect of bromocriptine. In summary, Acrp30 levels in serum are under complex hormonal control and may play a key role in determining systemic insulin sensitivity under the respective conditions. Diabetes 52:268 -276, 2003

Adiponectin is a plasma protein expressed exclusively in adipose tissue. Adiponectin levels are linked to insulin sensitivity, but a direct effect of chronically elevated adiponectin on improved insulin sensitivity has not yet been demonstrated. We identified a dominant mutation in the collagenous domain of adiponectin that elevated circulating adiponectin values in mice by 3-fold. Adiponectinemia raised lipid clearance and lipoprotein lipase activity, and suppressed insulin-mediated endogenous glucose production. The induction of adiponectin during puberty and the sexual dimorphism in adult adiponectin values were preserved in these transgenic animals. As a result of elevated adiponectin, serum PRL values and brown adipose mass both increased. The effects on carbohydrate and lipid metabolism were associated with elevated phosphorylation of 5'-AMP-activated protein kinase in liver and elevated expression of peroxisomal proliferator-activated receptor gamma2, caveolin-1, and mitochondrial markers in white adipose tissue. These studies strongly suggest that increasing endogenous adiponectin levels has direct effects on insulin sensitivity and may induce similar physiological responses as prolonged treatment with peroxisomal proliferator-activated receptor gamma agonists.