Transthyretin Antisense Oligonucleotides Lower Circulating RBP4 Levels and Improve Insulin Sensitivity in Obese Mice.

Circulating transthyretin (TTR) is a critical determinant of plasma retinol-binding protein 4 (RBP4) levels. Elevated RBP4 levels cause insulin resistance, and the lowering of RBP4 levels improves glucose homeostasis. Since lowering TTR levels increases renal clearance of RBP4, we determined whether decreasing TTR levels with antisense oligonucleotides (ASOs) improves glucose metabolism and insulin sensitivity in obesity. TTR-ASO treatment of mice with genetic or diet-induced obesity resulted in an 80-95% decrease in circulating levels of TTR and RBP4. Treatment with TTR-ASOs, but not control ASOs, decreased insulin levels by 30-60% and improved insulin sensitivity in ob/ob mice and high-fat diet-fed mice as early as after 2 weeks of treatment. The reduced insulin levels were sustained for up to 9 weeks of treatment and were associated with reduced adipose tissue inflammation. Body weight was not changed. TTR-ASO treatment decreased LDL cholesterol in high-fat diet-fed mice. The glucose infusion rate during a hyperinsulinemic-euglycemic clamp was increased by 50% in high-fat diet-fed mice treated with TTR-ASOs, demonstrating improved insulin sensitivity. This was also demonstrated by 20% greater inhibition of hepatic glucose production, a 45-60% increase of glucose uptake into skeletal and cardiac muscle, and a twofold increase in insulin signaling in muscle. These data show that decreasing circulating TTR levels or altering TTR-RBP4 binding could be a potential therapeutic approach for the treatment of type 2 diabetes.

Downregulation of STRA6 in adipocytes and adipose stromovascular fraction in obesity and effects of adipocyte-specific STRA6 knockdown in vivo.

To investigate the mechanisms by which elevated retinol-binding protein 4 (RBP4) causes insulin resistance, we studied the role of the high-affinity receptor for RBP4, STRA6 (stimulated by retinoic acid), in insulin resistance and obesity. In high-fat-diet-fed and ob/ob mice, STRA6 expression was decreased 70 to 95% in perigonadal adipocytes and both perigonadal and subcutaneous adipose stromovascular cells. To determine whether downregulation of STRA6 in adipocytes contributes to insulin resistance, we generated adipose-Stra6(-/-) mice. Adipose-Stra6(-/-) mice fed chow had decreased body weight, fat mass, leptin levels, insulin levels, and adipocyte number and increased expression of brown fat-selective markers in white adipose tissue. When fed a high-fat diet, these mice had a mild improvement in insulin sensitivity at an age when adiposity was unchanged. STRA6 has been implicated in retinol uptake, but retinol uptake and the expression of retinoid homeostatic genes (encoding retinoic acid receptor ß [RARß], CYP26A1, and lecithin retinol acyltransferase) were not altered in adipocytes from adipose-Stra6(-/-) mice, indicating that retinoid homeostasis was maintained with STRA6 knockdown. Thus, STRA6 reduction in adipocytes in adipose-Stra6(-/-) mice fed chow resulted in leanness, which may contribute to their increased insulin sensitivity. However, in wild-type mice with high-fat-diet-induced obesity and in ob/ob mice, the marked downregulation of STRA6 in adipocytes and adipose stromovascular cells does not compensate for obesity-associated insulin resistance.

The role of skeletal muscle insulin resistance in the pathogenesis of the metabolic syndrome.

We examined the hypothesis that insulin resistance in skeletal muscle promotes the development of atherogenic dyslipidemia, associated with the metabolic syndrome, by altering the distribution pattern of postprandial energy storage. Following ingestion of two high carbohydrate mixed meals, net muscle glycogen synthesis was reduced by approximately 60% in young, lean, insulin-resistant subjects compared with a similar cohort of age-weight-body mass index-activity-matched, insulin-sensitive, control subjects. In contrast, hepatic de novo lipogenesis and hepatic triglyceride synthesis were both increased by >2-fold in the insulin-resistant subjects. These changes were associated with a 60% increase in plasma triglyceride concentrations and an approximately 20% reduction in plasma high-density lipoprotein concentrations but no differences in plasma concentrations of TNF-alpha, IL-6, adiponectin, resistin, retinol binding protein-4, or intraabdominal fat volume. These data demonstrate that insulin resistance in skeletal muscle, due to decreased muscle glycogen synthesis, can promote atherogenic dyslipidemia by changing the pattern of ingested carbohydrate away from skeletal muscle glycogen synthesis into hepatic de novo lipogenesis, resulting in an increase in plasma triglyceride concentrations and a reduction in plasma high-density lipoprotein concentrations. Furthermore, insulin resistance in these subjects was independent of changes in the plasma concentrations of TNF-alpha, IL-6, high-molecular-weight adiponectin, resistin, retinol binding protein-4, or intraabdominal obesity, suggesting that these factors do not play a primary role in causing insulin resistance in the early stages of the metabolic syndrome.

Beta3 integrin deficiency promotes atherosclerosis and pulmonary inflammation in high-fat-fed, hyperlipidemic mice.

Hyperlipidemia promotes the chronic inflammatory disease atherosclerosis through poorly understood mechanisms. Atherogenic lipoproteins activate platelets, but it is unknown whether platelets contribute to early inflammatory atherosclerotic lesions. To address the role of platelet aggregation in diet-induced vascular disease, we studied beta3 integrin-deficient mice (lacking platelet integrin alphaIIbbeta3 and the widely expressed nonplatelet integrin alphavbeta3) in two models of atherosclerosis, apolipoprotein E (apoE)-null and low-density lipoprotein receptor (LDLR)-null mice. Unexpectedly, a high-fat, Western-type (but not a low-fat) diet caused death in two-thirds of the beta3-/-apoE-/- and half of the beta3-/-LDLR-/- mice due to noninfectious pneumonitis. In animals from both models surviving high-fat feeding, pneumonitis was absent, but aortic atherosclerosis was 2- to 6-fold greater in beta3-/- compared with beta+/+ littermates. Expression of CD36, CD40L, and CD40 was increased in lungs of beta3-/-LDLR-/- mice. Each was also increased in smooth muscle cells cultured from beta3-deficient mice and suppressed by retroviral reconstitution of beta3. These data show that the platelet defect caused by alphaIIbbeta3 deficiency does not impair atherosclerotic lesion initiation. They also suggest that alphavbeta3 has a suppressive effect on inflammation, the loss of which induces atherogenic mediators that are amplified by diet-induced hyperlipidemia.