Weikang Cai

Insulin Action in Brain Regulates Systemic Metabolism and Brain Function

Insulin receptors, as well as IGF-1 receptors and their postreceptor signaling partners, are distributed throughout the brain. Insulin acts on these receptors to modulate peripheral metabolism, including regulation of appetite, reproductive function, body temperature, white fat mass, hepatic glucose output, and response to hypoglycemia. Insulin signaling also modulates neurotransmitter channel activity, brain cholesterol synthesis, and mitochondrial function. Disruption of insulin action in the brain leads to impairment of neuronal function and synaptogenesis. In addition, insulin signaling modulates phosphorylation of tau protein, an early component in the development of Alzheimer disease. Thus, alterations in insulin action in the brain can contribute to metabolic syndrome, and the development of mood disorders and neurodegenerative diseases.

Insulin resistance in brain alters dopamine turnover and causes behavioral disorders

Significance Both types 1 and 2 diabetes are associated with increased risks of age-related decay in cognitive function and mood disorders, especially depression. Insulin action has been shown to regulate neuronal signaling and plasticity. Here we investigate whether brain-specific knockout of insulin receptor (NIRKO) in mice causes behavioral changes and how these are mechanistically linked. We find that NIRKO mice exhibit age-related anxiety and depressive-like behavior. This is due to altered mitochondrial function, aberrant monoamine oxidase (MAO) expression, and increased dopamine turnover in the mesolimbic system, and can be reversed by treatment with Mao inhibitors. Thus, brain insulin resistance alters dopamine turnover and induces anxiety and depressive-like behaviors. These findings demonstrate a potential molecular link between central insulin resistance and behavioral disorders. Diabetes and insulin resistance are associated with altered brain imaging, depression, and increased rates of age-related cognitive impairment. Here we demonstrate that mice with a brain-specific knockout of the insulin receptor (NIRKO mice) exhibit brain mitochondrial dysfunction with reduced mitochondrial oxidative activity, increased levels of reactive oxygen species, and increased levels of lipid and protein oxidation in the striatum and nucleus accumbens. NIRKO mice also exhibit increased levels of monoamine oxidase A and B (MAO A and B) leading to increased dopamine turnover in these areas. Studies in cultured neurons and glia cells indicate that these changes in MAO A and B are a direct consequence of loss of insulin signaling. As a result, NIRKO mice develop age-related anxiety and depressive-like behaviors that can be reversed by treatment with MAO inhibitors, as well as the tricyclic antidepressant imipramine, which inhibits MAO activity and reduces oxidative stress. Thus, insulin resistance in brain induces mitochondrial and dopaminergic dysfunction leading to anxiety and depressive-like behaviors, demonstrating a potential molecular link between central insulin resistance and behavioral disorders.

Insulin and IGF-1 receptors regulate FoxO-mediated signaling in muscle proteostasis

Diabetes strongly impacts protein metabolism, particularly in skeletal muscle. Insulin and IGF-1 enhance muscle protein synthesis through their receptors, but the relative roles of each in muscle proteostasis have not been fully elucidated. Using mice with muscle-specific deletion of the insulin receptor (M-IR-/- mice), the IGF-1 receptor (M-IGF1R-/- mice), or both (MIGIRKO mice), we assessed the relative contributions of IR and IGF1R signaling to muscle proteostasis. In differentiated muscle, IR expression predominated over IGF1R expression, and correspondingly, M-IR-/- mice displayed a moderate reduction in muscle mass whereas M-IGF1R-/- mice did not. However, these receptors serve complementary roles, such that double-knockout MIGIRKO mice displayed a marked reduction in muscle mass that was linked to increases in proteasomal and autophagy-lysosomal degradation, accompanied by a high-protein-turnover state. Combined muscle-specific deletion of FoxO1, FoxO3, and FoxO4 in MIGIRKO mice reversed increased autophagy and completely rescued muscle mass without changing proteasomal activity. These data indicate that signaling via IR is more important than IGF1R in controlling proteostasis in differentiated muscle. Nonetheless, the overlap of IR and IGF1R signaling is critical to the regulation of muscle protein turnover, and this regulation depends on suppression of FoxO-regulated, autophagy-mediated protein degradation.

Domain-dependent effects of insulin and IGF-1 receptors on signalling and gene expression

Despite a high degree of homology, insulin receptor (IR) and IGF-1 receptor (IGF1R) mediate distinct cellular and physiological functions. Here, we demonstrate how domain differences between IR and IGF1R contribute to the distinct functions of these receptors using chimeric and site-mutated receptors. Receptors with the intracellular domain of IGF1R show increased activation of Shc and Gab-1 and more potent regulation of genes involved in proliferation, corresponding to their higher mitogenic activity. Conversely, receptors with the intracellular domain of IR display higher IRS-1 phosphorylation, stronger regulation of genes in metabolic pathways and more dramatic glycolytic responses to hormonal stimulation. Strikingly, replacement of leucine973 in the juxtamembrane region of IR to phenylalanine, which is present in IGF1R, mimics many of these signalling and gene expression responses. Overall, we show that the distinct activities of the closely related IR and IGF1R are mediated by their intracellular juxtamembrane region and substrate binding to this region.

Endothelial insulin receptors differentially control insulin signaling kinetics in peripheral tissues and brain of mice

Significance Circulating hormones must cross the vascular endothelium to elicit their actions in target tissues via either transcytosis or paracellular diffusion. Insulin receptors on endothelial cells are believed to mediate transcytosis of circulating insulin, but how this affects insulin action in vivo is unknown. Here, we demonstrate that knockout of insulin receptors on endothelial cells delays the kinetics of activation of insulin signaling in skeletal muscle, fat, and several regions of the brain but not in liver or olfactory bulb. This alters the kinetics of insulin action in vivo and induces tissue-specific insulin resistance leading to dysregulated glucose and body weight homeostasis. Insulin receptors (IRs) on endothelial cells may have a role in the regulation of transport of circulating insulin to its target tissues; however, how this impacts on insulin action in vivo is unclear. Using mice with endothelial-specific inactivation of the IR gene (EndoIRKO), we find that in response to systemic insulin stimulation, loss of endothelial IRs caused delayed onset of insulin signaling in skeletal muscle, brown fat, hypothalamus, hippocampus, and prefrontal cortex but not in liver or olfactory bulb. At the level of the brain, the delay of insulin signaling was associated with decreased levels of hypothalamic proopiomelanocortin, leading to increased food intake and obesity accompanied with hyperinsulinemia and hyperleptinemia. The loss of endothelial IRs also resulted in a delay in the acute hypoglycemic effect of systemic insulin administration and impaired glucose tolerance. In high-fat diet-treated mice, knockout of the endothelial IRs accelerated development of systemic insulin resistance but not food intake and obesity. Thus, IRs on endothelial cells have an important role in transendothelial insulin delivery in vivo which differentially regulates the kinetics of insulin signaling and insulin action in peripheral target tissues and different brain regions. Loss of this function predisposes animals to systemic insulin resistance, overeating, and obesity.

Viral insulin-like peptides activate human insulin and IGF-1 receptor signaling: A paradigm shift for host–microbe interactions

Significance Although there has been tremendous progress in understanding hormone action and its relationship to human physiology and disease, there has been no comprehensive approach to search the viral genome for the presence of human-like hormones. Here, using a bioinformatics approach, we have identified 16 different human peptide hormones/growth factors, including four insulin/insulin growth factor (IGF)1-like peptides (VILPs) that have homologous sequences in viruses. When these VILPs were chemically synthesized, the resulting peptides could bind to human and murine insulin and IGF1 receptors, stimulate postreceptor signaling, increase glucose uptake, and activate proliferation of cells. Injection of VILPs into mice can significantly lower the blood glucose. Thus, VILPs are members of the insulin superfamily and first characterized viral hormones. Viruses are the most abundant biological entities and carry a wide variety of genetic material, including the ability to encode host-like proteins. Here we show that viruses carry sequences with significant homology to several human peptide hormones including insulin, insulin-like growth factors (IGF)-1 and -2, FGF-19 and -21, endothelin-1, inhibin, adiponectin, and resistin. Among the strongest homologies were those for four viral insulin/IGF-1–like peptides (VILPs), each encoded by a different member of the family Iridoviridae. VILPs show up to 50% homology to human insulin/IGF-1, contain all critical cysteine residues, and are predicted to form similar 3D structures. Chemically synthesized VILPs can bind to human and murine IGF-1/insulin receptors and stimulate receptor autophosphorylation and downstream signaling. VILPs can also increase glucose uptake in adipocytes and stimulate the proliferation of fibroblasts, and injection of VILPs into mice significantly lowers blood glucose. Transfection of mouse hepatocytes with DNA encoding a VILP also stimulates insulin/IGF-1 signaling and DNA synthesis. Human microbiome studies reveal the presence of these Iridoviridae in blood and fecal samples. Thus, VILPs are members of the insulin/IGF superfamily with the ability to be active on human and rodent cells, raising the possibility for a potential role of VILPs in human disease. Furthermore, since only 2% of viruses have been sequenced, this study raises the potential for discovery of other viral hormones which, along with known virally encoded growth factors, may modify human health and disease.

Insulin regulates astrocyte gliotransmission and modulates behavior

&NA; Complications of diabetes affect tissues throughout the body, including the central nervous system. Epidemiological studies show that diabetic patients have an increased risk of depression, anxiety, age‐related cognitive decline, and Alzheimers disease. Mice lacking insulin receptor (IR) in the brain or on hypothalamic neurons display an array of metabolic abnormalities; however, the role of insulin action on astrocytes and neurobehaviors remains less well studied. Here, we demonstrate that astrocytes are a direct insulin target in the brain and that knockout of IR on astrocytes causes increased anxiety‐ and depressive‐like behaviors in mice. This can be reproduced in part by deletion of IR on astrocytes in the nucleus accumbens. At a molecular level, loss of insulin signaling in astrocytes impaired tyrosine phosphorylation of Munc18c. This led to decreased exocytosis of ATP from astrocytes, resulting in decreased purinergic signaling on dopaminergic neurons. These reductions contributed to decreased dopamine release from brain slices. Central administration of ATP analogs could reverse depressive‐like behaviors in mice with astrocyte IR knockout. Thus, astrocytic insulin signaling plays an important role in dopaminergic signaling, providing a potential mechanism by which astrocytic insulin action may contribute to increased rates of depression in people with diabetes, obesity, and other insulin‐resistant states.

TRPV1 neurons regulate β-cell function in a sex-dependent manner

There is emerging evidence to support an important role for the transient receptor potential vanilloid type 1 (TRPV1) sensory innervation in glucose homeostasis. However, it remains unknown whether the glucoregulatory action of these afferent neurons is sex-biased and whether it is pancreatic β-cell-mediated. Objective We investigated in male and female mice whether denervation of whole-body or pancreas-projecting TRPV1 sensory neurons regulates adult functional β-cell mass and alters systemic glucose homeostasis. Methods We used a combination of pharmacological and surgical approaches to ablate whole-body or pancreatic TRPV1 sensory neurons and assessed islet β-cell function and mass, aspects of glucose and insulin homeostasis, and energy expenditure. Results Capsaicin-induced chemodenervation of whole-body TRPV1 sensory neurons improved glucose clearance and enhanced glucose-stimulated insulin secretion without alterations in β-cell proliferation and mass, systemic insulin sensitivity, body composition, and energy expenditure. Similarly, denervation of intrapancreatic TRPV1 afferents by pancreas intraductal injection of capsaicin or surgical removal of the dorsal root ganglia projecting into the pancreas lowered post-absorptive glucose levels and increased insulin release upon glucose stimulation. The beneficial effects of TRPV1 sensory denervation on glucose tolerance and β-cell function were observed in male but not female mice. Conclusion Collectively, these findings suggest that TRPV1 neurons regulate glucose homeostasis, at least partly, through direct modulation of glucose-induced insulin secretion and that this regulation operates in a sex-dependent manner.

IGFBP2 Is Increased after Gastric Bypass in Humans—Potential Impact on Hepatic Metabolism

To identify molecular mechanisms contributing to postoperative metabolic improvements after Roux-en-Y gastric bypass (RYGB), we performed comprehensive proteomic analysis of fasting human plasma samples obtained from the SLIMM-T2D longitudinal clinical trial, which randomized obese individuals with T2D to RYGB surgery or a one-year intensive medical diabetes and weight management (DWM) program, and followed them for 3 years. Somalogic proteomic analysis was performed on plasma collected in the fasting state at baseline and during longitudinal follow-up, and proteins differentially abundant in RYGB vs. DWM at each time point were identified. The protein with highest magnitude of differential abundance at 3 years was insulin-like growth factor binding protein 2 (IGFBP2), upregulated by 2.2-fold in RYGB vs. DWM (p=4.37E-06); differences were confirmed by ELISA. IGFBP2 inversely correlated at 3 years with BMI (r=-0.74, p=5.31E-05) and HbA 1c (r=-0.68, p=0.00004). Notably, IGFBP2 levels were increased at the first postoperative study visit (10% weight loss) following RYGB or at similar weight loss for the DWM group, suggesting an early weight-independent contribution. IGFBP2 is predominantly expressed in liver and upregulated after RYGB in mice (2.8-fold at 8 weeks, GSE68812). To identify IGFBP2-dependent mechanisms contributing to improved hepatic metabolism, we overexpressed mouse IGFBP2 or control vector in mouse AML12 hepatocytes, achieving a 6-fold increase in IGFPB2 protein. qPCR revealed 15-30% downregulation of lipogenic genes (e.g., Fasn, Srebp1) with a 2-fold increase in expression of genes regulating fatty acid oxidation (e.g., Ppara, Ppargc1a). Analysis of metabolism (Seahorse flux analyzer) revealed increased fatty acid oxidation (3-fold, p In conclusion, increased IGFBP2 expression may contribute to improved hepatic lipid oxidative metabolism, and thus induce systemic metabolic improvement after RYGB surgery. Disclosure Y. Yuchi: None. W. Cai: None. T. Takagi: None. H. Pan: None. J. Dreyfuss: None. K. Foster: None. A.H. Vernon: None. D.C. Simonson: Advisory Panel; Self; GI Windows, Inc.. Stock/Shareholder; Self; GI Windows, Inc.. Stock/Shareholder; Spouse/Partner; Phase V Technologies, Inc. A. Goldfine: Employee; Self; Novartis AG. A. Hoeflich: None. M.E. Patti: Research Support; Self; Janssen Pharmaceuticals, Inc.. Other Relationship; Self; Xeris Pharmaceuticals, Inc.. Research Support; Self; Ethicon US, LLC., Coviden, MedImmune. Other Relationship; Self; Novo Nordisk Inc., XOMA Corporation, AstraZeneca, Nestle. Research Support; Self; Dexcom, Inc.. Consultant; Self; Eiger BioPharmaceuticals.