Intraperitoneal Glucose Tolerance Test Research Articles

GIP receptor agonism improves dyslipidemia and atherosclerosis independently of body weight loss in preclinical mouse model for cardio-metabolic disease

BackgroundAgonism at the receptor for the glucose-dependent insulinotropic polypeptide (GIPR) is a key component of the novel unimolecular GIPR:GLP-1R co-agonists, which are among the most promising drugs in clinical development for the treatment of obesity and type 2 diabetes. The therapeutic effect of chronic GIPR agonism to treat dyslipidemia and thus to reduce the cardiovascular disease risk independently of body weight loss has not been explored yet.MethodsAfter 8 weeks on western diet, LDL receptor knockout (LDLR-/-) male mice were treated with daily subcutaneous injections of long-acting acylated GIP analog (acyl-GIP; 10nmol/kg body weight) for 28 days. Body weight, food intake, whole-body composition were monitored throughout the study. Fasting blood glucose and intraperitoneal glucose tolerance test (ipGTT) were determined on day 21 of the study. Circulating lipid levels, lipoprotein profiles and atherosclerotic lesion size was assessed at the end of the study. Acyl-GIP effects on fat depots were determined by histology and transcriptomics.ResultsHerein we found that treatment with acyl-GIP reduced dyslipidemia and atherogenesis in male LDLR-/- mice. Acyl-GIP administration resulted in smaller adipocytes within the inguinal fat depot and RNAseq analysis of the latter revealed that acyl-GIP may improve dyslipidemia by directly modulating lipid metabolism in this fat depot.ConclusionsThis study identified an unanticipated efficacy of chronic GIPR agonism to improve dyslipidemia and cardiovascular disease independently of body weight loss, indicating that treatment with acyl-GIP may be a novel approach to alleviate cardiometabolic disease.

Nutritive/non-nutritive sweeteners and high fat diet contribute to dysregulation of sweet taste receptors and metabolic derangements in oral, intestinal and central nervous tissues.

Overconsumption of non-nutritive sweeteners is associated with obesity, whereas the underlying mechanisms remain controversial. This study aimed to investigate the effects of long-term consumption of nutritive or non-nutritive sweeteners with or without high fat diet on sweet taste receptor expression in nutrient-sensing tissues and energy regulation dependent on sweet-sensing. 50 Male Sprague-Dawley rats (140-160g) were assigned to 10 groups (n = 5/group). All received fructose at 2.5% or 10%, sucralose at 0.01% or 0.015% or water with a normal chow diet or high fat diet for 12weeks. Food and drink intake were monitored daily. Oral glucose tolerance test and intraperitoneal glucose tolerance test were performed at week 10 and 11 respectively. Serum was obtained for measurement of biochemical parameters. Tongue, duodenum, jejunum, ileum, colon and hypothalamus were rapidly removed to assess gene expression. Long-term consumption of sweeteners impaired glucose tolerance, increased calorie intake and body weight. A significant upregulation of sweet taste receptor expression was observed in all the four intestinal segments in groups fed 0.01% sucralose or 0.015% sucralose, most strikingly in the ileum, accompanied by elevated serum glucagon-like peptide-1 levels and up-regulated expression of sodium-dependent glucose cotransporter 1 and glucose transporter 2. A significant down-regulation in the tongue and hypothalamus was observed in groups fed 10% fructose or 0.015% sucralose, with alterations in hypothalamic appetite signals. The presence of high fat diet differentially modulates sweet taste perception in nutrient-sensing tissues. Long-term consumption of whether nutritive sweeteners or non-nutritive sweeteners combined with high fat diet contribute to dysregulation of sweet taste receptor expression in oral, intestinal and central nervous tissues.

Asprosin contributes to nonalcoholic fatty liver disease through regulating lipid accumulation and inflammatory response via AMPK signaling.

Nonalcoholic fatty liver disease (NAFLD) is a primary contributor to liver-related morbidity and mortality. Asprosin has been reported to be implicated in NAFLD. This work is to illuminate the effects of Asprosin on NAFLD and the possible downstream mechanism. The weight of NAFLD mice induced by a high-fat diet was detected. Quantitative reverse-transcription polymerase chain reaction (RT-qPCR) examined serum Asprosin expression. RT-qPCR and western blot analysis examined Asprosin expression in mice liver tissues. Intraperitoneal glucose tolerance test (IPGTT) and intraperitoneal insulin tolerance test (IPITT) were implemented. Biochemical kits tested liver enzyme levels in mice serum and liver tissues. Hematoxylin and eosin staining evaluated liver histology. Liver weight was also tested and oil red O staining estimated lipid accumulation. RT-qPCR and western blot analysis analyzed the expression of gluconeogenesis-, fatty acid biosynthesis-, fatty acid oxidation-, and inflammation-associated factors. Besides, western blot analysis examined the expression of AMP-activated protein kinase (AMPK)/p38 signaling-associated factors. In palmitic acid (PA)-treated mice hepatocytes, RT-qPCR and western blot analysis examined Asprosin expression. Lipid accumulation, gluconeogenesis, fatty acid biosynthesis, fatty acid oxidation, and inflammation were appraised again. Asprosin was overexpressed in the serum and liver tissues of NAFLD mice and PA-treated mice hepatocytes. Asprosin interference reduced mice body and liver weight, improved glucose tolerance and diminished liver injury in vivo. Asprosin knockdown alleviated lipid accumulation and inflammatory infiltration both in vitro and in vivo. Additionally, Asprosin absence activated AMPK/p38 signaling and AMPK inhibitor Compound C reversed the impacts of Asprosin on lipid accumulation and inflammatory response. Collectively, Asprosin inhibition suppressed lipid accumulation and inflammation to obstruct NAFLD through AMPK/p38 signaling.

Sphingosine Kinase 2 Knockout Mice Resist HFD-Induced Obesity Through Increasing Energy Expenditure.

It has been reported that sphingosine kinase (SK) 2 plays a role in maintaining metabolism and glucose homeostasis. However, the mechanism remains uncertain. The present research aimed to further investigate the effect of SK2 knockout on high-fat diet (HFD)-induced obesity and metabolic regulation. Male SK2-/- and wild-type (WT) control mice were challenged with HFD for 8 weeks. Then, body composition, inguinal white adipose tissue (IWAT) histology, intraperitoneal glucose tolerance tests (IPGTT), and metabolic parameters were examined, and expression levels of uncoupling protein 1 (UCP1), a key molecular marker of thermogenesis, in IWAT were determined. After 8 weeks of HFD challenge, compared with WT mice, SK2-/- mice displayed decreased whole body, epididymal white adipose tissue (EWAT) and IWAT weights, reduced fat/lean body mass ratios and inguinal adipocytes size; also, SK2-/- mice exhibited improved intraperitoneal glucose tolerance. Next, elevated energy expenditure was observed in SK2-/- mice compared with WT mice; however, neither food intake nor physical activity showed obvious difference between SK2-/- and WT mice. Furthermore, we found that the expressions of UCP1 was markedly increased in IWAT from SK2-/- mice. SK2-/- mice may resist HFD-induced obesity through increasing energy expenditure by promoting thermogenesis in the beige adipose tissue.

Formononetin suppresses hyperglycaemia through activation of GLUT4-AMPK pathway

Background: Formononetin (FMN) is a flavonoid that has different pharmacological effects. Thus, the anti-diabetic effects of FMN has been investigated in a high-fat diet/Streptozotocin-(HFD/STZ)-induced diabetes mice model. Methods: Mice were fed with HFD followed by STZ. Diabetic mice were treated orally with FMN or metformin for 28 days before collecting plasma and soleus muscle for further analysis. Results: FMN reduced serum glucose (p>0.001) and increased serum insulin in diabetic group compared to the vehicle control. Additionally, FMN decreased homeostasis model assessment of insulin resistance (HOMA-IR). Fasting glucose level was also reduced with FMN during the intraperitoneal glucose tolerance test (IPGTT). GLUT4 and p-AMPK-α1 were upregulated following treatment with FMN. LDL, triglyceride, and cholesterol were reduced in diabetic mice treated with FMN. FMN reduced MDA, increased GSH levels , and reduced GSSG levels in diabetic mice. Conclusion: FMN could represent a promising therapeutic agent to treat T2D.

Schisandrin B, a potential GLP-1R agonist, exerts anti-diabetic effects by stimulating insulin secretion

Diabetes mellitus is a metabolic disease that is characterized by elevated blood sugar. Although glucagon-like peptide-1 receptor agonists (GLP-1RA) lower blood glucose in a glucose-dependent manner, most of them are macromolecule polypeptides. Macromolecular peptides are relatively expensive and inconvenient compared with small molecules. Therefore, this study sought to identify the small molecules binding to GLP-1R via cell membrane chromatography (CMC), confirm their agonistic activity, and further study its beneficial effects in a mouse model of type 2 diabetes mellitus (T2DM) induced by a combination of high-fat diet and streptozotocin. We used CMC, calcium imaging and molecular docking techniques to screen and identify the potential small molecule Schisandrin B (Sch B), which exhibits a strong binding effect to GLP-1R, from the small molecule library of traditional Chinese medicine. Through in-vitro experiments, we found that Sch B stimulated insulin secretion in β-TC-6 cells, while GLP-1R antagonist Exendin9-39, adenylate cyclase inhibitor SQ22536, and protein kinase A (PKA) inhibitor H89 could significantly inhibit the insulin secretion induced by Sch B. In vivo, Sch B significantly improved fasting blood glucose levels, intraperitoneal glucose tolerance test damage, and the status of pancreatic tissue damage, and reduced serum insulin levels, total cholesterol, triglyceride and low density lipoprotein in T2DM mice. These results indicate that Sch B alleviates T2DM by promoting insulin release through the GLP-1R/cAMP/PKA signaling pathway, suggesting that Sch B may be a potential GLP-1RA, which is expected to provide a new therapeutic strategy for the prevention and treatment of T2DM.

Liquiritigenin regulates insulin sensitivity and ameliorates inflammatory responses in the nonalcoholic fatty liver by activation PI3K/AKT pathway.

Nonalcoholic fatty liver disease (NAFLD) is a prevalent long-term disease in the world. Liquiritigenin (LQ) is protective against a variety of hepatotoxins. Herein, we report the potential mechanism of LQ on a high-fat diet (HFD) induced NAFLD. NAFLD mice model was established by HFD for 12 weeks, and LQ treatment for 1 week. Commercially available assay kits measure liver triglycerides (TG) and total cholesterol (TC) levels. Plasm TC, TG, high-density-lipoprotein (HDL-C), and low-density-lipoprotein cholesterol (LDL-C) levels were also monitored by biochemistry. Enzyme linked immunosorbent assay (ELISA) kits were performed to analyze the pro-inflammatory factors, and intraperitoneal glucose tolerance test (IPGTT), insulin tolerance test (IPITT), and serum insulin were also determined. GO and KEGG pathway enrichment analysis was employed to analyze the overlapping genes of LQ targets and NAFLD development-related targets. Western blot was performed on key proteins of the enriched signaling pathway. HFD mice showed significant increases in hepatic TG and TC, and plasm TC, TG, and LDL-C in blood lipids, while HDL-C significantly decreased, and LQ treatment reversed their levels (p < 0.05). LQ also alleviated HFD-induced elevated levels of IPGTT, IPITT, and homeostasis model assessment of insulin resistance (HOMA-IR). And serum levels of the pro-inflammatory factor were also suppressed by LQ. PI3K/AKT pathway was enriched by KEGG pathway enrichment, and its key proteins p-PI3K and p-AKT were elevated after LQ treatment (p < 0.05). We found for the first time that LQ improves lipid accumulation, alleviates insulin resistance, and suppresses inflammatory responses in NAFLD mice, which might be associated with the activation of the PI3K/AKT pathway.

Protection of pancreatic islets from oxidative cell death by a peripherally-active morphinan with increased drug safety

ObjectiveDextromethorphan (DXM) is a commonly used antitussive medication with positive effects in people with type 2 diabetes mellitus, since it increases glucose tolerance and protects pancreatic islets from cell death. However, its use as an antidiabetic medication is limited due to its central nervous side effects and potential use as a recreational drug. Therefore, we recently modified DXM chemically to reduce its blood–brain barrier (BBB) penetration and central side effects. However, our best compound interacted with the cardiac potassium channel hERG (human ether-à-go-go-related gene product) and the μ-opioid receptor (MOR). Thus, the goal of this study was to reduce the interaction of our compound with these targets, while maintaining its beneficial properties. MethodsReceptor and channel binding assays were conducted to evaluate the drug safety of our DXM derivative. Pancreatic islets were used to investigate the effect of the compound on insulin secretion and islet cell survival. Via liquor collection from the brain and a behavioral assay, we analyzed the BBB permeability. By performing intraperitoneal and oral glucose tolerance tests as well as pharmacokinetic analyses, the antidiabetic potential and elimination half-life were investigated, respectively. To analyze the islet cell-protective effect, we used fluorescence microscopy as well as flow cytometric analyses. ResultsHere, we report the design and synthesis of an optimized, orally available BBB-impermeable DXM derivative with lesser binding to hERG and MOR than previous ones. We also show that the new compound substantially enhances glucose-stimulated insulin secretion (GSIS) from mouse and human islets and glucose tolerance in mice as well as protects pancreatic islets from cell death induced by reactive oxygen species and that it amplifies the effects of tirzepatide on GSIS and islet cell viability. ConclusionsWe succeeded to design and synthesize a novel morphinan derivative that is BBB-impermeable, glucose-lowering and islet cell-protective and has good drug safety despite its morphinan and imidazole structures.

1727-P: Mitochondrial-Encoded Peptide MOTS-c Prevents Senescence-Induced Diabetes

A polymorphism MOTS-c, a mitochondrial-derived miniprotein, was reported to be associated with longevity in humans. Previous reports showed that MOTS-c regulates senescence in cells, enhances physical capacity in old mice and in humans. However, it is still unknown how MOTS-c is able to intervene the cellular senescence, what tissue or cell-type MOTS-c targets, and whether the regulation of aging by MOTS-c can lead to the prevention of aging- or senescence-induced disorders. Here, using S961 (or senescence)-induced mouse model, we show that MOTS-c is capable of targeting pancreas and is able to prevent senescence-induced diabetes. To test how MOTS-c is associated with β cell senescence, we firstly assess whether β cell senescence is associated with MOTS-c expression using pancreatic β cells isolated from 12-week-old mice. After isolation, pancreatic β cells were treated with H2O2 in concentration-dependent manner. Treatment of H2O2 lowered MOTS-c expression in concentration-dependent manner, but increased other senescence-related markers such as β-gal expression. Furthermore, MOTS-c treatment decreased H2O2-induced senescence-associated β-gal expression and SASPs (i.e., IL-1). To understand the inhibitory effect of MOTS-c against β cell senescence in vivo, we have transplanted S961-osmotic pump in 28-week-old mice with or without MOTS-c for 2 weeks. Mice treated with MOTS-c showed lowered random glucose level (310 mg/dL versus 180 mg/dL) and decreased level of pancreatic β-gal expression compared to S961-treated mice. Also, an i.p. glucose tolerance test (IPGTT) in 30-week-old mice revealed that MOTS-c treatment significantly improved glucose clearance. As a result, we showed that senescence/S961-induced diabetes were inhibited by MOTS-c treatment. Disclosure B.Kong: None. S.Hong: None. Y.Cho: Consultant; LG Chem. Funding Ministry of Health &amp; Welfare, Republic of Korea (HI14C1277)

1538-P: Brain TGFß Regulates Glucose Tolerance in Male Mice with Metabolic Syndrome

Hypothalamic inflammation contributes to insulin resistance and obesity. Transforming growth factor beta (TGFβ) is among the inflammatory mediators inducing hypothalamic inflammation. In this study, we investigated the role of brain TGFβ on metabolic function. We hypothesized that TGFβ in the brain promotes insulin resistance and reduces glucose tolerance in diabetic and obese mice. C57BL/J male mice were fed either hypercaloric diet (HCD, 52% fat) or regular diet (RD) from weaning. At 3 months age, intraperitoneal glucose tolerance test (GTT), insulin tolerance test (ITT) and weight confirmed the acquisition of metabolic syndrome. Hypothalamic protein expression was assessed for TGFβ receptor 2, phosphorylated SMAD3, SMAD6 and SMAD7, as downstream signal transducer proteins for TGFβ. Compared to RD male mice (n=6), mice on HCD (n=6) showed glucose intolerance (AUC; 60587 ±2460 vs. 30802±1923, p&amp;lt;0.0001), insulin intolerance (AUC; Mean±SEM; 19025836.0 vs.11436±1087, p=0.02) and weight gain (33.67±2.028 vs. 26.89±0.4231 g). HCD mice also exhibited increased protein levels of TGFβR2 (normalized to total protein, Fold Change (FC)±SEM to RD group; 3.54±0.3, p&amp;lt;0.0001), pSMAD3 (FC 2±0.5, p=0.04) while inhibitors for TGFβ signaling such as SMAD6 and SMAD7, were downregulated (FC 0.7±0.1, p=0.03 and 0.8±0.04, p=0.03, respectively). To understand the role of brain TGFβ on metabolic regulation, 3 months HCD mice were subjected to intracerebroventricular (ICV) infusion with either TGFβ neutralizing antibody (nAb, n=4) or IgG control antibody (n=4) using subcutaneous osmotic pumps connected to ICV cannula for 2 weeks. GTT was measured after 2-week infusion. Treatment with TGFβ nAb resulted in lower fasting blood glucose compared to before treatment (166.1 ±6.878 vs. 229.9 ±3.923 mg/dL) and at 120 min time point (229.9 ±28.69 vs. 382.2 ±24.33 mg/dL). In conclusion, brain TGFβ plays a key role in regulating glucose metabolism probably through inflammation-induced insulin resistance. Disclosure M.Elgazzaz: None. K.Miao: None. A.R.Nuzzo: None. L.Restivo: None. K.Moles: None. E.D.Lazartigues: None. Funding American Diabetes Association (1-19-IBS-291 to E.D.L.)

767-P: TERN-601, a Novel Oral GLP-1R Agonist, Suppresses Food Intake and Improves Glucose Tolerance in Transgenic Mice Expressing Human GLP-1 Receptor

Synthetic peptide agonists of the GLP-1 receptor (GLP-1R) are approved for the treatment of diabetes and obesity based on their ability to lower blood glucose and induce weight loss. TERN-601 is a novel, oral small molecule GLP-1R agonist in preclinical development. In vitro, TERN-601 potently stimulated (EC50 = 2.9 ± 0.8 nM) intracellular cAMP generation, a downstream consequence of GLP-1R activation, in CHO-K1 cells stably expressing human GLP-1R. In vivo, single oral doses of TERN-601 (10, 30, or 60 mg/kg) led to significant dose-dependent suppression of food intake (2- to 24-hours post dose) in transgenic C57BL/6J mice (n = 10 per group) expressing human GLP-1R (hGLP-1R). In an intraperitoneal glucose tolerance test (IPGTT) on overnight fasted hGLP-1R mice (n = 7 per group), single oral doses of TERN-601 (0.3, 1, and 3 mg/kg) administered 30 minutes prior to IP glucose injection significantly lowered blood glucose area under the curve (AUC0-120min) with comparable efficacy to liraglutide (0.3 mg/kg). TERN-601 is a potent, orally bioavailable small molecule agonist of human GLP-1R in preclinical development for the treatment of chronic obesity. In transgenic mice expressing human GLP-1R, single oral doses of TERN-601 significantly suppressed food intake and improved glucose tolerance. Together these data support the continued development of TERN-601. Disclosure C.Jones: Employee; Terns Pharmaceuticals. K.P.Quinn: None. C.H.Nelson: Employee; Terns Pharmaceuticals, Gilead Sciences, Inc., Stock/Shareholder; Terns Pharmaceuticals, Gilead Sciences, Inc. O.Osborn: Employee; Terns, Stock/Shareholder; ClicBio. G.Luehr: None. J.R.Jasper: Advisory Panel; Rubedo Life Sciences, Employee; Terns Pharmaceuticals, Rubedo Life Sciences.

1587-P: The NADPH Oxidase 4 Regulates Anti-inflammatory Signals to Modulate Lipid Depot Distribution between Adipose Tissue and Liver

Excessive white adipose tissue (WAT) expansion and lipid load is associated with the onset of insulin resistance and Type 2 diabetes. In addition, lipid storage may “overflow” into the liver triggering the apparition of nonalcoholic fatty liver disease (NAFLD). Two major components linking white adipose tissue (WAT) dysfunction and NAFLD to insulin resistance are the deregulation of cellular redox homeostasis and the onset of chronic, low-grade inflammation. NADPH oxidases (NOX-es) produce oxidant molecules that control redox homeostasis and cytokine signaling in inflammatory cells. In particular, NOX4 is essential for the signal transduction of the transcription factor, Stat6 that mediates anti-inflammatory actions of IL-4 and IL-13. In order to elucidate the in vivo interaction between NOX4 and Stat6 and to assess their relevance in the onset of “obese” WAT and NAFLD, we created mice with double knock-out for NOX4 and Stat6 (DKO) and compared their metabolic phenotype to littermates with single NOX4 deletion (NOX4KO) upon a 13-week high-fat diet feeding (60 % of calorie from fat). DKO and NOX4KO mice displayed similar food intake. By contrast, DKO mice displayed less body weight gain than NOX4KO mice due to lower energy efficiency leading to reduced WAT accumulation. DKO mice shifted lipid deposition away from the WAT and enhanced lipid storage in the liver. DKO mice displayed similar fed and starved glucose levels with no alterations in glucose tolerance assessed by intra-peritoneal glucose tolerance tests. Gene expression analysis confirmed a less inflammatory state of both WAT and liver of DKO mice. Our results demonstrate that anti-inflammatory signaling is modulated by NOX4-derived signals in vivo in mice, and that NOX4 and IL-4/IL-13 interaction is a critical component in the regulation surplus lipid storage distribution between WAT and the liver. Disclosure I. Szanto: None. Funding Insuleman Foundation (ISZ-1); Foundation for the Innovation in Research in Cancer and Biological Sciences

Replenishable prevascularized cell encapsulation devices increase graft survival and function in the subcutaneous space.

Beta cell replacement therapy (BCRT) for patients with type 1 diabetes (T1D) improves blood glucose regulation by replenishing the endogenous beta cells destroyed by autoimmune attack. Several limitations, including immune isolation, prevent this therapy from reaching its full potential. Cell encapsulation devices used for BCRT provide a protective physical barrier for insulin-producing beta cells, thereby protecting transplanted cells from immune attack. However, poor device engraftment posttransplantation leads to nutrient deprivation and hypoxia, causing metabolic strain on transplanted beta cells. Prevascularization of encapsulation devices at the transplantation site can help establish a host vascular network around the implant, increasing solute transport to the encapsulated cells. Here, we present a replenishable prevascularized implantation methodology (RPVIM) that allows for the vascular integration of replenishable encapsulation devices in the subcutaneous space. Empty encapsulation devices were vascularized for 14 days, after which insulin-producing cells were inserted without disrupting the surrounding vasculature. The RPVIM devices were compared with nonprevascularized devices (Standard Implantation Methodology [SIM]) and previously established prevascularized devices (Standard Prevascularization Implantation Methodology [SPVIM]). Results show that over 75% of RPVIM devices containing stem cell-derived insulin-producing beta cell clusters showed a signal after 28 days of implantation in subcutaneous space. Notably, not only was the percent of RPVIM devices showing signal significantly greater than SIM and SPVIM devices, but the intraperitoneal glucose tolerance tests and histological analyses showed that encapsulated stem-cell derived insulin-producing beta cell clusters retained their function in the RPVIM devices, which is crucial for the successful management of T1D.

METABOLIC DYSFUNCTIONS CAUSED BY EARLY FRUCTOSE INGESTION ARE PREVENTED IN CONCURRENTLY EXERCISED ADULT MALE RATS

This study evaluated whether fructose ingestion at adolescence programs the development ofmetabolic syndrome (MetS) in adult rats and whether moderate-intensity combined exercisecan attenuate fructose-induced MetS compounds. Pubertal rats were trained on a treadmill(55-65% VO 2max ), 3 times/week, 44 min/session, for one month. They were allocated into fourgroups: sedentary control (SC), trained control (TC), sedentary fructose (SF), and trainedfructose (TF). Groups F ingested 10% fructose. Food intake (AI) and body weight (BW) weremeasured weekly. At 60 and 120 days of age, metabolic parameters were evaluated. Fructoseintake mainly affected animals in adult life, and these changes were related to glucoseintolerance (p=0.001), periepididymal fat (p&amp;lt;0.03), and increase in total cholesterol (TC) (p&amp;lt;0.001), triglycerides (TAGs) (p&amp;lt;0.0001) and insulin (p&amp;lt;0.0001) levels. Early physicalexercise was able to decrease periepididymal (p&amp;lt;0.003) and retroperitoneal (p&amp;lt;0.0002)adipose tissue, TC levels (p&amp;lt;0.0001) and glucose tolerance (p=0.21). Rats that receivedcombined treatments during adolescence showed low glycemia (p=0.003) and insulinemia(p&amp;lt;0.0001) during the intraperitoneal glucose tolerance test (ipGTT). HOMA-IR (p=0.02) andTYG (p=0.0003) scores also improved in these rats. We conclude that early ingestion of 10%fructose was effective in programming adult rats to MetS, and simultaneous moderateexercise attenuated these aggressions, causing changes in glycemic homeostasis and lipidmetabolism.

Deficiency of S100 calcium binding protein A9 attenuates vascular dysfunction in aged mice

BackgroundS100 calcium-binding protein A9 (S100A9) is a danger-associated molecular pattern molecule that mediates the inflammatory response. Inflammation is essential in aging-related cardiovascular diseases. However, less is known regarding the role of S100A9 in vascular aging. MethodsS100A9 null mice were used to investigate the role of S100A9 in aging-related pathologies. Artery rings were used to measure the functional characteristics of vascular with a pressurized myograph. Telomere length, Sirtuin activity, oxidative stress, and endothelial nitric oxide synthetase (eNOS) activity were used to elevate vascular senescence. Intraperitoneal glucose tolerance (IPGTT) and insulin sensitivity test (IST) were employed to investigate the effects of S100A9 on insulin resistance. Inflammation response was reflected by the concentration of inflammatory cytokines. The Toll-like receptor 4 (TLR4) and receptor for advanced glycation end products (RAGE) inhibitors were used to identify the downstream molecular mechanisms of S100A9 in aging-induced senescence in endothelial cells. ResultsS100A9 expression in vascular increased with aging in mice and humans. Deficiency of S100A9 alleviated vascular senescence in aged mice, as evidenced by increased telomere length, Sirtuin activity, and eNOS activity. Meanwhile, S100A9 knockout improved endothelium-dependent vasodilatation and endothelial continuity in aged mice. Moreover, the increased insulin resistance, oxidative stress, and inflammation were mitigated by S100A9 deletion in aged mice. In vitro, S100A9 induced senescence in endothelial cells, and that effect was blunted by TLR4 but not RAGE inhibitors. ConclusionThe present study suggested that S100A9 may contribute to aging-related pathologies and endothelial dysfunction via the TLR4 pathway. Therefore, targeting S100A9/TLR4 signaling pathway may represent a crucial therapeutic strategy to prevent age-related cardiovascular diseases.

Chronic supplementation of branched chain amino acids and its impact on glucose and hepatic metabolism during insulin sensitivity

Dysfunctional interactions between branched chain amino acids (BCAAs) and facets of glucose and lipid metabolism are central features of insulin resistant tissues. However, it is unclear whether insulin resistance is a prerequisite for the biochemical relationships between BCAAs and macronutrient networks. We hypothesize that BCAAs will enhance glucose and lipid oxidation, lower gluconeogenesis and decrease hepatic de novo lipogenesis to optimize insulin sensitivity. Male mice (C57BL/6NJ) were placed on either a low-fat (LF; 10% fat Kcal) or LF supplemented with 1.5X BCAAs (LB) diet for a period of 12-32 wks. Oxygen consumption and energy expenditure were determined by indirect calorimetry after 12 and 24-wks on the diets. After 32-wks on the diets and a 4-hr fast, LF and LB mice underwent intraperitoneal glucose tolerance test (IPGTT) with the 2g/kg glucose containing 50% [ 13 C 6 ]glucose. Blood glucose was measured every 15-min and blood was collected every 30-min for 2 hours. Enrichment of 13 C from glucose into pyruvate and other metabolic intermediates was determined by gas-chromatography and mass-spectrometry (GC-MS). Following 1-wk of recovery, these LF and LB mice were randomly split into fed and overnight fasted groups. [ 13 C 3 ] pyruvate (20 mg) was injected intraperitoneally to determine the rate of incorporation of pyruvate into glucose via. gluconeogenesis, utilizing GC-MS. Hepatic gene expression in fed and fasted livers were profiled using qPCR. Supplementation of BCAAs for 24-wks increased energy expenditure and oxygen consumption in the LB mice vs. their LF counterparts (p≤0.05). Additionally, IPGTT revealed improved insulin sensitivity in LB mice vs. their LF counterparts (p&lt;0.05). Dilution of [ 13 C 6 ]glucose was lower in the LB plasma by the 90-min time point of the glucose tolerance test (% enrichment; LB; 16.42±0.74 vs. LF; 18.06±0.54, p=0.09), while the LB blood glucose levels during IPGTT remained lower, indicative of higher rates of glucose oxidation in LB mice. Further, following the pyruvate tolerance test, incorporation of [ 13 C 3 ]pyruvate into glucose tended to be higher in the fasted BCAA supplemented group (% enrichment; LB; 11.79±0.23 vs. LF; 10.58±0.53, p=0.07). Basal levels of tricarboxylic acid (TCA) cycle intermediates in plasma were significantly different between LF vs. LB mice (p&lt;0.05). Fed-to-fasted fold induction of genes involved in lipid oxidation ( Lcad, P para) were higher in the LB livers vs. their LF counterparts (p≤0.05). On the contrary, fed-to-fasted suppression of lipogenic gene expression (A cc, F as) was greater in the LB livers vs. their LF counterparts (p&lt;0.05). In summary, our results indicate that chronic BCAA supplementation to insulin sensitive mice positively impacts aspects of glucose and lipid metabolism. The ability of BCAAs to modulate macronutrient metabolism could have beneficial effects in alleviating metabolic dysfunction during early stages of insulin resistance and fatty liver disease. National Institute of Health R01DK112865 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 592: Marfan Syndrome Mice Fed On High Fat Diet Have An Altered Metabolism

Introduction: Marfan syndrome (MFS) is a genetic disorder, caused by different mutations in the fibrillin-1 gene ( FBN1 ) and affects the connective tissue. Patients suffer from a wide range of clinical symptoms at the cardiovascular, skeletal and ocular level. However, MFS patients are also often characterized as very slender. Body weight gain in these patients can be a challenge, and the reason for this is still not fully understood. Objective: To determine if individuals with MFS have an altered metabolism. Methods: 9-11 weeks old male Fbn1 C1041G/+ (MFS) mice and wild-type littermates were given either a chow, high cholesterol diet (HCD) or a high fat diet (HFD), for 12 weeks. These mice were then housed individually in a metabolic cage for 1 week before euthanasia was performed. Results: HFD resulted in higher body weights compared to chow diet, and the HFD groups became insulin resistant as measured by the intraperitoneal glucose tolerance test. However, body weight gain was not different in this MFS mouse model, as compared to WT mice, regardless of diet. The HFD groups had increased fat weight compared to chow groups. Interestingly, MFS mice had an enhanced heart weight compared to WT mice, independent of the diet. As expected, aortic root diameters were enlarged in MFS compared to WT mice. However, in the ascending aorta there was a significant increase in diameter in the WT HFD as compared to WT chow, indicating that enhanced lipid levels promote aortic growth. Metabolic cage data showed that the HFD groups had a lower respiratory exchange ratio (RER), indicating that HFD groups used fat as a predominant source of fuel and that chow groups used primarily carbohydrates as an energy source. MFS mice had overall lower RER compared to WT mice, revealing that MFS mice use more lipids under any condition, most likely to manage the significantly elevated activity in the MFS mice. Conclusion: Taken together, MFS mice seem metabolically different.

Mustn1 ablation in skeletal muscle results in increased glucose tolerance concomitant with upregulated GLUT expression in male mice.

Glucose homeostasis is closely regulated to maintain energy requirements of vital organs and skeletal muscle plays a crucial role in this process. Mustn1 is expressed during embryonic and postnatal skeletal muscle development and its function has been implicated in myogenic differentiation and myofusion. Whether Mustn1 plays a role in glucose homeostasis in anyway remains largely unknown. As such, we deleted Mustn1 in skeletal muscle using a conditional knockout (KO) mouse approach. KO mice did not reveal any specific gross phenotypic alterations in skeletal muscle. However, intraperitoneal glucose tolerance testing (IPGTT) revealed that 2-month-old male KO mice had significantly lower glycemia than their littermate wild type (WT) controls. These findings coincided with mRNA changes in genes known to be involved in glucose metabolism, tolerance, and insulin sensitivity; 2-month-old male KO mice had significantly higher expression of GLUT1 and GLUT10 transporters, MUP-1 while OSTN expression was lower. These differences in glycemia and gene expression were statistically insignificant after 4 months. Identical experiments in female KO and WT control mice did not indicate any differences at any age. Our results suggest a link between Mustn1 expression and glucose homeostasis during a restricted period of skeletal muscle development/maturation. While this is an observational study, Mustn1's relationship to glucose homeostasis appears to be more complex with a possible connection to other key proteins such as GLUTs, MUP-1, and OSTN. Additionally, our data indicate temporal and sex differences. Lastly, our findings strengthen the notion that Mustn1 plays a role in the metabolic capacity of skeletal muscle.

Increased methylglyoxal formation in plasma and tissues during a glucose tolerance test is derived from exogenous glucose.

The dicarbonyl compound methylglyoxal (MGO) is a major precursor in the formation of advanced glycation endproducts (AGEs). MGO and AGEs are increased in subjects with diabetes and are associated with fatal and non-fatal cardiovascular disease. Previously we have shown that plasma MGO concentrations rapidly increase in the postprandial phase, with a higher increase in individuals with type 2 diabetes. In current study, we investigated whether postprandial MGO formation in plasma and tissues originates from exogenous glucose and whether the increased plasma MGO concentration leads to a fast formation of MGO-derived AGEs. We performed a stable isotope labelled oral glucose tolerance test (OGTT) in 12 healthy males with universally labelled D(+)13C glucose. Analysis of plasma labelled 13C3 MGO and glucose levels at eleven time-points during the OGTT revealed that the newly formed MGO during OGTT is completely derived from exogenous glucose. Moreover, a fast formation of protein-bound MGO-derived AGEs during the OGTT was observed. In accordance, ex vivo incubation of MGO with plasma or albumin showed a rapid decrease of MGO and a fast increase of MGO-derived AGEs. In an intraperitoneal glucose tolerance test in C57BL/6J mice, we confirmed that the formation of postprandial MGO is derived from exogenous glucose in plasma and also showed in tissues that MGO is increased and this is also from exogenous glucose.Collectively, increased formation of MGO during a glucose tolerance test arises from exogenous glucose both in plasma and in tissues, and this leads to a fast formation of MGO-derived AGEs.

PSTPIP2 alleviates obesity associated adipose tissue inflammation and insulin resistance in diabetes mice through promoting M2 macrophage polarization via activation of PPARγ

BackgroundProline-serine-threonine phosphatase-interacting protein 2 (PSTPIP2) plays a role in inflammatory disease. In diabetes, very little is known about PSTPIP2 until now. Hence, this study aimed to determine PSTPIP2 functional role in diabetes. MethodsDiabetes mouse model was constructed by feeding high fat diet (HFD). Intraperitoneal glucose tolerance test and intraperitoneal insulin tolerance test were examined the glucose and insulin tolerance. The expression of genes and proteins was detected by quantitative real time PCR, immunohistochemistry and western blotting. The pathological changes of epididymal adipose tissues were examined by hematoxylin-eosin staining. RAW264.7 macrophages were treated with GW9662 (PPARγ antagonist). Flow cytometry examined the proportion of M1/M2 macrophages. ResultsHFD enhanced the body weight, glucose and insulin tolerance, and inhibited PSTPIP2 expression in mice. PSTPIP2 overexpression alleviated glucose and insulin tolerance, reduced inflammation and macrophage accumulation in the epididymal adipose tissues of diabetic mice. The expression of iNOS and TNF-α was increased, the expression of IL-10 and Arg-1 was decreased in diabetic mice, which was abrogated by PSTPIP2 overexpression. In vitro, PSTPIP2 overexpression reduced the proportions of iNOS-positive cells and enhanced the proportions of CD206-positive cells in RAW264.7 cells. PPARγ and p-STAT6 were up-regulated, STAT6 was down-regulated in RAW264.7 cells. GW9662 impaired PSTPIP2 overexpression-mediated up-regulation of Arg-1, YM-1 and FIZZ1 in RAW264.7 cells. ConclusionPSTPIP2 alleviates obesity associated adipose tissue inflammation and insulin resistance in diabetic mice through promoting M2 macrophage polarization via activation of PPARγ, suggesting that PSTPIP2 is a prospective target for diabetes treatment.