Andrea M. Caricilli

Gut Microbiota Is a Key Modulator of Insulin Resistance in TLR 2 Knockout Mice

A genetic and pharmacological approach reveals novel insights into how changes in gut microbiota can subvert genetically predetermined phenotypes from lean to obese.

Inhibition of Hypothalamic Inflammation Reverses Diet-Induced Insulin Resistance in the Liver

Defective liver gluconeogenesis is the main mechanism leading to fasting hyperglycemia in type 2 diabetes, and, in concert with steatosis, it is the hallmark of hepatic insulin resistance. Experimental obesity results, at least in part, from hypothalamic inflammation, which leads to leptin resistance and defective regulation of energy homeostasis. Pharmacological or genetic disruption of hypothalamic inflammation restores leptin sensitivity and reduces adiposity. Here, we evaluate the effect of a hypothalamic anti-inflammatory approach to regulating hepatic responsiveness to insulin. Obese rodents were treated by intracerebroventricular injections, with immunoneutralizing antibodies against Toll-like receptor (TLR)4 or tumor necrosis factor (TNF)α, and insulin signal transduction, hepatic steatosis, and gluconeogenesis were evaluated. The inhibition of either TLR4 or TNFα reduced hypothalamic inflammation, which was accompanied by the reduction of hypothalamic resistance to leptin and improved insulin signal transduction in the liver. This was accompanied by reduced liver steatosis and reduced hepatic expression of markers of steatosis. Furthermore, the inhibition of hypothalamic inflammation restored defective liver glucose production. All these beneficial effects were abrogated by vagotomy. Thus, the inhibition of hypothalamic inflammation in obesity results in improved hepatic insulin signal transduction, leading to reduced steatosis and reduced gluconeogenesis. All these effects are mediated by parasympathetic signals delivered by the vagus nerve.

Inhibition of toll-like receptor 2 expression improves insulin sensitivity and signaling in muscle and white adipose tissue of mice fed a high-fat diet

The aims of the present study were to investigate the expression of toll-like receptor 2 (TLR2) in muscle and white adipose tissue (WAT) of diet-induced obesity (DIO) mice, and also the effects of its inhibition, with the use of TLR2 antisense oligonucleotide (ASON), on insulin sensitivity and signaling. The expression of TLR2 was increased in muscle and WAT of DIO mice, compared with those that received standard chow. Inhibition of TLR2 in DIO mice, by TLR2 ASON, improved insulin sensitivity and signaling in muscle and WAT. In addition, data show that the inhibition of TLR2 expression prevents the activation of IKBKB, MAPK8, and serine phosphorylation of IRS1 in DIO mice, suggesting that TLR2 is a key modulator of the crosstalk between inflammatory and metabolic pathways. We, therefore, suggest that a selective interference with TLR2 presents an attractive opportunity for the treatment of insulin resistance in obesity and type 2 diabetes.

Topical insulin accelerates wound healing in diabetes by enhancing the AKT and ERK pathways: a double-blind placebo-controlled clinical trial.

Background Wound healing is impaired in diabetes mellitus, but the mechanisms involved in this process are virtually unknown. Proteins belonging to the insulin signaling pathway respond to insulin in the skin of rats. Objective The purpose of this study was to investigate the regulation of the insulin signaling pathway in wound healing and skin repair of normal and diabetic rats, and, in parallel, the effect of a topical insulin cream on wound healing and on the activation of this pathway. Research Design and Methods We investigated insulin signaling by immunoblotting during wound healing of control and diabetic animals with or without topical insulin. Diabetic patients with ulcers were randomized to receive topical insulin or placebo in a prospective, double-blind and placebo-controlled, randomized clinical trial (NCT 01295177) of wound healing. Results and Conclusions Expression of IR, IRS-1, IRS-2, SHC, ERK, and AKT are increased in the tissue of healing wounds compared to intact skin, suggesting that the insulin signaling pathway may have an important role in this process. These pathways were attenuated in the wounded skin of diabetic rats, in parallel with an increase in the time of complete wound healing. Upon topical application of insulin cream, the wound healing time of diabetic animals was normalized, followed by a reversal of defective insulin signal transduction. In addition, the treatment also increased expression of other proteins, such as eNOS (also in bone marrow), VEGF, and SDF-1α in wounded skin. In diabetic patients, topical insulin cream markedly improved wound healing, representing an attractive and cost-free method for treating this devastating complication of diabetes. Trial Registration ClinicalTrials.gov NCT01295177

The Role of Gut Microbiota on Insulin Resistance

The development of obesity and insulin resistance has been extensively studied in the last decades, but the mechanisms underlying these alterations are still not completely understood. The gut microbiota has been identified as a potential contributor to metabolic diseases. It has been shown that obese individuals present different proportions of bacterial phyla compared with lean individuals, with an increase in Firmicutes and Actinobacteria and a decrease in Bacteroidetes. This alteration seems to interfere with intestinal permeability, increasing the absorption of lipopolysaccharide (LPS), which reaches circulation and initiates activation of Toll-like receptor (TLR) 4 and 2 and LPS receptor CD14, leading to increased activation of inflammatory pathways. With these activations, an impairment of the insulin signaling is observed, with decreased phosphorylation of the insulin receptor, insulin receptor substrate (IRS) and Akt, as well as increased inhibitory serine phosphorylation of IRS-1. Altered proportions of bacterial phyla have also been demonstrated to interfere with host’s biochemical pathways, increasing energy extraction and depot in adipose tissue. Therefore, understanding the mechanisms by which the alteration in the gut microbiota produces different signaling activations and phenotype changes may offer an interesting opportunity for the treatment of obesity and type 2 diabetes.

Targeted Disruption of Inducible Nitric Oxide Synthase Protects Against Aging, S -Nitrosation, and Insulin Resistance in Muscle of Male Mice

Accumulating evidence has demonstrated that S-nitrosation of proteins plays a critical role in several human diseases. Here, we explored the role of inducible nitric oxide synthase (iNOS) in the S-nitrosation of proteins involved in the early steps of the insulin-signaling pathway and insulin resistance in the skeletal muscle of aged mice. Aging increased iNOS expression and S-nitrosation of major proteins involved in insulin signaling, thereby reducing insulin sensitivity in skeletal muscle. Conversely, aged iNOS-null mice were protected from S-nitrosation–induced insulin resistance. Moreover, pharmacological treatment with an iNOS inhibitor and acute exercise reduced iNOS-induced S-nitrosation and increased insulin sensitivity in the muscle of aged animals. These findings indicate that the insulin resistance observed in aged mice is mainly mediated through the S-nitrosation of the insulin-signaling pathway.

Gut microbiota composition and its effects on obesity and insulin resistance.

Purpose of review Rising evidence suggest that variation in the gut microbiome at gene and species levels defines subsets of individuals who have increased risk of obesity-related metabolic disorders, including insulin resistance and type 2 diabetes, which is influenced by diet and genetic profile of the host. Our goal in this review is gathering the newest findings concerning gut microbiota composition and effects on hosts metabolism. Recent findings Dietary changes have been shown as the most prominent shaper of gut microbiota composition, reflecting major phenotypes, which can also be transmitted to other individuals, in spite of genetic variances. Gut microbiota composition has also been presented as diversity, which may have important implications in metabolite production and consequent interference with inflammatory activation, insulin resistance, and obesity. Summary Specific approaches made it possible to comprehend some of the interactions between certain bacterial strains and their host, and how their metabolites may interfere with hosts cell signaling, changing its metabolic profile. Herein, we discuss some of the mechanisms by which alterations in the gut microbiota composition may contribute to the pathophysiology of obesity and its related co-morbities.

Topiramate Treatment Improves Hypothalamic Insulin and Leptin Signaling and Action and Reduces Obesity in Mice

Topiramate (TPM) treatment has been shown to reduce adiposity in humans and rodents. The reduction in adiposity is related to decreased food intake and increased energy expenditure. However, the molecular mechanisms through which TPM induces weight loss are contradictory and remain to be clarified. Whether TPM treatment alters hypothalamic insulin, or leptin signaling and action, is not well established. Thus, we investigate herein whether short-term TPM treatment alters energy balance by affecting insulin and leptin signaling, action, or neuropeptide expression in the hypothalamus of mice fed with a high-fat diet. As expected, short-term treatment with TPM diminished adiposity in obese mice mainly due to reduced food intake. TPM increased anorexigenic signaling by enhancing the leptin-induced leptin receptor/Janus kinase 2/signal transducer and activator of transcription 3 pathway and the insulin-induced insulin receptor substrate/Akt/forkhead box O1 pathway in parallel to reduced phosphatase protein expression in the hypothalamus of obese mice. These effects were independent of body weight. TPM also raised anorexigenic neuropeptides such as POMC, TRH, and CRH mRNA levels in obese mice. In addition, TPM increased the activation of the hypothalamic MAPK/ERK pathway induced by leptin, accompanied by an increase in peroxisome proliferator-activated receptor-coactivator α and uncoupling protein 1 protein levels in brown adipose tissue. Furthermore, TPM increased AMP-activated protein kinase and acetyl-coenzyme A carboxylase phosphorylation in peripheral tissues, which may help improve energy metabolism in these tissues. Together, these results provide novel insights into the molecular mechanisms through which TPM treatment reduces adiposity.

Tub Has a Key Role in Insulin and Leptin Signaling and Action In Vivo in Hypothalamic Nuclei

Mutation of tub gene in mice induces obesity, suggesting that tub could be an important regulator of energy balance. In the current study, we investigated whether insulin, leptin, and obesity can modulate Tub in vivo in hypothalamic nuclei, and we investigated possible consequences on energy balance, neuropeptide expression, and hepatic glucose metabolism. Food intake, metabolic characteristics, signaling proteins, and neuropeptide expression were measured in response to fasting and refeeding, intracerebroventricular insulin and leptin, and Tub antisense oligonucleotide (ASO). Tub tyrosine phosphorylation (Tub-p-tyr) is modulated by nutritional status. Tub is a substrate of insulin receptor tyrosine kinase (IRTK) and leptin receptor (LEPR)–Janus kinase 2 (JAK2) in hypothalamic nuclei. After leptin or insulin stimulation, Tub translocates to the nucleus. Inhibition of Tub expression in hypothalamus by ASO increased food intake, fasting blood glucose, and hepatic glucose output, decreased O2 consumption, and blunted the effect of insulin or leptin on proopiomelanocortin, thyroid-releasing hormone, melanin-concentrating hormone, and orexin expression. In hypothalamus of mice administered a high-fat diet, there is a reduction in leptin and insulin-induced Tub-p-tyr and nuclear translocation, which is reversed by reducing protein tyrosine phosphatase 1B expression. These results indicate that Tub has a key role in the control of insulin and leptin effects on food intake, and the modulation of Tub may contribute to insulin and leptin resistance in DIO mice.

Modulation of hypothalamic PTP1B in the TNF-α-induced insulin and leptin resistance

We have associated functional and molecular studies of insulin and leptin to investigate the effect of TNF‐α on central insulin and leptin signaling in rats pre‐treated with PTP1B‐ASO. The icv infusion of TNF‐α‐induced an increase in PTP1B protein expression and activity, and attenuated insulin and leptin sensitivity and signaling in the hypothalamus. However, TNF‐α was able to completely blunt the leptin and insulin effect in rats treated with PTP1B‐ASO, suggesting that TNF‐α does not require PTP1B to fully attenuate the leptin and insulin effects. In addition, our data also show that other mechanisms of insulin and leptin resistance are activated in the hypothalamus by TNF‐α.