Sanchita Gupta

Saroglitazar reduces obesity and associated inflammatory consequences in murine adipose tissue

Abstract Prevailing knowledge links chronic low‐grade inflammation in the adipose tissue to obesity and its associated metabolic complications. In this study, we evaluated immunometabolic effects of a recently launched dual peroxisome proliferator‐activated receptor (PPAR) &agr; & &ggr; agonist ‘Saroglitazar’ in a mouse model of diet‐induced obesity (DIO). Body composition analysis revealed that saroglitazar treatment promoted hepatic weight gain, while attenuated epididymal white adipose tissue (eWAT) mass in DIO. In the eWAT of saroglitazar treated mice, histological analysis showed reduced adipocyte hypertrophy and matrix deposition (picrosirius red staining). Immunological profiling of stromal vascular fraction isolated from eWAT showed decreased pro‐inflammatory cells (M1 macrophages, CD4 and CD8 T‐cells) and increased anti‐inflammatory M2 macrophages. Gene expression and western blot analysis suggested that saroglitazar promoted energy expenditure machinery and attenuated inflammatory as well as fibrotic markers in eWAT during DIO. In conclusion, for the first time we are reporting immunometabolic effects of dual PPAR&agr; & &ggr; agonist saroglitazar in DIO and insulin resistance (IR). Saroglitazar exerted its beneficial effects on adipose tissue by limiting, diet‐induced adipose tissue dysfunction, adipocyte hypertrophy, adipocyte cell damage and extracellular matrix deposition in obesity.

Ecliptal, a promising natural lead isolated from Eclipta alba modulates adipocyte function and ameliorates metabolic syndrome

ABSTRACT A swift increase has been observed in the number of individuals with metabolic syndrome worldwide. A number of natural compounds have been identified towards combating metabolic syndrome. Adding to this premise, here we report the pleiotropic activities of Ecliptal (EC); a natural compound isolated from the herb Eclipta alba. Administration of EC was shown to have prominent anti‐adipogenic effects in 3T3‐L1 and hMSC derived adipocytes. It was shown to activate Wnt‐pathway and alter AKT signaling. Additionally, it caused cell cycle arrest and inhibited mitotic clonal expansion. EC treatment augmented mitochondrial biogenesis as well as function as estimated by expression of PGC1&agr;, UCP‐1, mitochondrial complexes and estimation of oxygen consumption rate. EC also reduced LPS‐induced inflammation and tunicamycin induced ER stress. Further, EC enhanced insulin sensitivity by increasing AKT phosphorylation, inhibiting PKC&agr;/&bgr;II phosphorylation and reducing leptin/adiponectin ratio. Finally, EC administration in Syrian golden hamsters was shown to have potent anti‐dyslipidemic effects. Cumulatively, encompassing pleiotropic activities of EC, it could prove to be a potential drug candidate against obesity, insulin resistance and related metabolic syndrome. Graphical abstract Figure. No Caption available. HighlightsEcliptal shows prominent anti‐adipogenic effects.Ecliptal blocked mitotic clonal expansion.Ecliptal treatment augmented mitochondrial functions.Ecliptal exhibited anti‐inflammatory and anti‐ER stress activities.In‐vivo, Ecliptal was shown to have anti‐dyslipidemic effects too.

Chronic hyperinsulinemia induced miR-27b is linked to adipocyte insulin resistance by targeting insulin receptor

Defect in insulin signaling leads to the development of insulin resistance followed by type 2 diabetes. Exploiting our previously developed physiological chronic hyperinsulinemia (CI)-mediated insulin resistance (IR) model, we wanted to understand how miRNAs contribute to the development of IR. Amongst the identified and validate miRNAs, the expression of miR-27b was found to be highly upregulated during CI-induced IR in 3T3-L1 adipocytes. We also validated the expression of miR-27b in CI-induced IR in human mesenchymal stem cell (hMSC)-derived adipocytes and in vivo high fat diet (HFD)-induced IR mice model. Bioinformatics target prediction softwares and luciferase reporter assay identified insulin receptor (INSR) as one of a prime target of miR-27b. Lentiviral mediated overexpression of miR-27b impairs insulin signaling by modulating INSR expression that in turn led to decreased glucose uptake in both 3T3-L1 and hMSC-derived adipocytes. Conversely, inhibition of miR-27b reversed CI-mediated suppression of target protein INSR and improved phosphorylation of Akt, a nodal protein of insulin signaling that is impaired by CI treatment. Lentiviral mediated overexpression of miR-27b in in vivo C57BL/6 mice impaired whole body glucose tolerance and adipose tissue insulin sensitivity. Furthermore, inhibition of miR-27b in HFD-induced insulin resistance mice model improved glucose tolerance and adipose tissue insulin sensitivity by increasing the expression of its target gene INSR in eWAT. Thus, our results indicate that miR-27b functions as a prime modulator of CI-induced IR via regulating the expression of INSR.Key messagesmiR-27b is upregulated in different in vitro and in vivo models of insulin resistance.miR-27b directly suppresses the expression of INSR by targeting 3’UTR of INSR.Modulation of miR-27b expression regulates insulin sensitivity by targeting INSR.

Ethyl acetate fraction of Eclipta alba: a potential phytopharmaceutical targeting adipocyte differentiation

Natural products have always fascinated mankind for their miraculous properties. Eclipta alba (E. alba), a medicinal herb has long been used in traditional medicine for curing several pathologies. It has been shown to have anti-diabetic effect as well as hepato-protective activity. Here, in order to address metabolic derangements, the study was designed to evaluate the efficacy of E. alba and its fractions in adipogenesis inhibition and dyslipidemia. Of the crude extract and fractions screened, ethyl acetate fraction of E. alba inhibited adipocyte differentiation in 3T3-L1 pre-adipocytes and hMSC derived adipocytes. It inhibited mitotic clonal expansion and caused cell cycle arrest in G1 and S phase as suggested by western blot analysis and flow cytometry. It was also shown to have lipolytic effects. Oral administration of ethyl acetate fraction of E. alba to hamsters unveiled its anti-adipogenic as well as anti-dyslipidemic activity in-vivo. Mass spectrometry analysis of ethyl acetate fraction confirmed the presence of several bioactive components, projecting it as an effective phytopharmaceutical agent. In conclusion, ethyl acetate fraction of E. alba possesses potent anti-adipogenic as well as anti-dyslipidemic activity and could be projected as an herbal formulation towards obesity.

Chronic hyperinsulinemia promotes meta-inflammation and extracellular matrix deposition in adipose tissue: Implications of nitric oxide

Various imperative studies support the notion that hyperinsulinemia (HI) itself serves as the common link between adipose tissue inflammation (ATI) and metabolic syndrome. However, the contribution of HI mediated ATI and its metabolic consequences are yet to be explored. We induced chronic HI per se in mice by administration of exogenous insulin for 8 weeks through mini-osmotic pumps. For the reduction of circulating insulin in response to excess calorie intake, we have partially ablated β-cells by using streptozotocin (STZ) in the diet-induced obesity (DIO) and genetic mice models (db/db). Flow cytometry analysis was performed for the quantification of immune cells in stromal vascular fraction (SVF) isolated from epididymal white adipose tissue (eWAT). Our studies demonstrated that chronic HI augmented ATI in terms of elevated pro-inflammatory cells (M1 macrophages and NK-cells) and suppressed anti-inflammatory cells (M2 macrophages, eosinophils and regulatory T-cells). These results were correlated with altered obesity-associated metabolic phenotype. Partial reduction of circulating insulin level attenuated excess calorie-induced ATI and improved insulin sensitivity. Mechanistically, an imbalance in M1 and M2 macrophage proportions in eWAT promoted iNOS (inducible nitric oxide synthase): arginase-1 imbalance that resulted into extracellular matrix (ECM) deposition and insulin resistance (IR) development. However, iNOS-/- mice were protected from HI-induced M1:M2 macrophage imbalance, ECM deposition and IR in adipose tissue. Overall, we conclude that chronic HI per se contributed in ATI and iNOS corroborated ECM deposition.

Aegeline inspired synthesis of novel β3-AR agonist improves insulin sensitivity in vitro and in vivo models of insulin resistance

BACKGROUND AND PURPOSE In our drug discovery program of natural product, earlier we have reported Aegeline that is N-acylated-1-amino-2- alcohol, which was isolated from the leaves of Aeglemarmelos showed anti-hyperlipidemic activity for which the QSAR studies predicted the compound to be the β3-AR agonist, but the mechanism of its action was not elucidated. In our present study, we have evaluated the β3-AR activity of novel N-acyl-1-amino-3-arylopropanol synthetic mimics of aegeline and its beneficial effect in insulin resistance. In this study, we have proposed the novel pharmacophore model using reported molecules for antihyperlipidemic activity. The reported pharmacophore features were also compared with the newly developed pharmacophore model for the observed biological activity. EXPERIMENTAL APPROACH Based on 3D pharmacophore modeling of known β3AR agonist, we screened 20 synthetic derivatives of Aegeline from the literature. From these, the top scoring compound 10C was used for further studies. The in-slico result was further validated in HEK293T cells co-trransfected with human β3-AR and CRE-Luciferase reporter plasmid for β3-AR activity.The most active compound was selected and β3-AR activity was further validated in white and brown adipocytes differentiated from human mesenchymal stem cells (hMSCs). Insulin resistance model developed in hMSC derived adipocytes was used to study the insulin sensitizing property. 8 week HFD fed C57BL6 mice was given 50 mg/Kg of the selected compound and metabolic phenotyping was done to evaluate its anti-diabetic effect. RESULTS As predicted by in-silico 3D pharmacophore modeling, the compound 10C was found to be the most active and specific β3-AR agonist with EC50 value of 447 nM. The compound 10C activated β3AR pathway, induced lipolysis, fatty acid oxidation and increased oxygen consumption rate (OCR) in human adipocytes. Compound 10C induced expression of brown adipocytes specific markers and reverted chronic insulin induced insulin resistance in white adipocytes. The compound 10C also improved insulin sensitivity and glucose tolerance in 8 week HFD fed C57BL6 mice. CONCLUSION This study enlightens the use of in vitro insulin resistance model close to human physiology to elucidates the insulin sensitizing activity of the compound 10C and edifies the use of β3AR agonist as therapeutic interventions for insulin resistance and type 2 diabetes.

Temporal immmunometabolic profiling of adipose tissue in HFD-induced obesity: manifestations of mast cells in fibrosis and senescence

Background/Objectives:Chronic low-grade inflammation/meta-inflammation in adipose tissue leads to obesity-associated metabolic complications. Despite growing understanding, the roles of immune cell subsets, their interrelationship, and chronological events leading to progression of obesity-associated insulin resistance (IR) remains unclear.Methods:We carried out temporal immunometabolic profiling of adipose tissue from C57BL/6 mice fed a high-fat diet (HFD) for 4, 8, 12, 16, and 20 weeks. We used clodronate sodium liposomes (CLODs) to deplete macrophages and disodium cromoglycate sodium liposomes (DSCGs) to stabilize mast cells.Results:In the temporal HFD settings, mice showed progressive glucose intolerance, insulin resistance, and adipose tissue senescence. Histochemistry analysis of epididymal white adipose tissue (eWAT) using picro-sirius red and Masson’s trichrome staining showed extensive collagen deposition in the 16th and 20th weeks. Flow cytometry analysis of the stromal vascular fraction (SVF) from eWAT revealed T-cell subsets as early-phase components and pro-inflammatory macrophages, as well as mast cells as the later phase components during obesity progression. In our therapeutic strategies, macrophage depletion by CLOD and mast stabilization by DSCG attenuated obesity, adipose tissue fibrosis, and improved whole-body glucose homeostasis. In addition, mast cell stabilization also attenuated senescence (p53 and X-gal staining) in eWAT, signifying the role of mast cells over macrophages during obesity.Conclusion:New-generation mast cell stabilizers can be exploited for the treatment of obesity-associated metabolic complications.