Antibody blockade of activin type II receptors preserves skeletal muscle mass and enhances fat loss during GLP-1 receptor agonism.

OBJECTIVE: Glucagon-like peptide 1 (GLP-1) receptor agonists reduce food intake, producing remarkable weight loss in overweight and obese individuals. While much of this weight loss is fat mass, there is also a loss of lean mass, similar to other approaches that induce calorie deficit. Targeting signaling pathways that regulate skeletal muscle hypertrophy is a promising avenue to preserve lean mass and modulate body composition. Myostatin and Activin A are TGFß-like ligands that signal via the activin type II receptors (ActRII) to antagonize muscle growth. Pre-clinical and clinical studies demonstrate that ActRII blockade induces skeletal muscle hypertrophy and reduces fat mass. In this manuscript, we test the hypothesis that combined ActRII blockade and GLP-1 receptor agonism will preserve muscle mass, leading to improvements in skeletomuscular and metabolic function and enhanced fat loss. METHODS: In this study, we explore the therapeutic potential of bimagrumab, a monoclonal antibody against ActRII, to modify body composition alone and during weight loss induced by GLP-1 receptor agonist semaglutide in diet-induced obese mice. Mechanistically, we define the specific role of the anabolic kinase Akt in mediating the hypertrophic muscle effects of ActRII inhibition in vivo. RESULTS: Treatment of obese mice with bimagrumab induced a ∼10 % increase in lean mass while simultaneously decreasing fat mass. Daily treatment of obese mice with semaglutide potently decreased body weight; this included a significant decrease in both muscle and fat mass. Combination treatment with bimagrumab and semaglutide led to superior fat mass loss while simultaneously preserving lean mass despite reduced food intake. Treatment with both drugs was associated with improved metabolic outcomes, and increased lean mass was associated with improved exercise performance. Deletion of both Akt isoforms in skeletal muscle modestly reduced, but did not prevent, muscle hypertrophy driven by ActRII inhibition. CONCLUSIONS: Collectively, these data demonstrate that blockade of ActRII signaling improves body composition and metabolic parameters during calorie deficit driven by GLP-1 receptor agonism and demonstrate the existence of Akt-independent pathways supporting muscle hypertrophy in the absence of ActRII signaling.

Hybridization into a Bitopic Ligand Increased Muscarinic Receptor Activation for Isopilocarpine but Not for Pilocarpine Derivatives.

Pilocarpine (1), a secondary metabolite of several Pilocarpus species, is a therapeutically used partial agonist of muscarinic acetylcholine receptors (mAChRs). The available pharmacological data and structure-activity relationships do not provide comparable data for all five receptor subtypes. In this study, pilocarpine (1), its epimer isopilocarpine (2), racemic analogues pilosinine (3) and desmethyl pilosinine (4), and the respective hybrid ligands with a naphmethonium fragment (5-C6 to 8-C6) were synthesized and analyzed in mini-G nano-BRET assays at the five mAChRs. In line with earlier studies, pilocarpine was the most active compound among the orthosteric ligands 1-4. Computational docking of pilocarpine and isopilocarpine to the active M2 receptor suggests that the trans-configuration of isopilocarpine leads to a loss of the hydrogen bond from the lactone carbonyl to N6.52, explaining the lower activity of isopilocarpine. Hybrid formation of pilocarpine (1) and isopilocarpine (2) led to an inverted activity rank, with the trans-configured isopilocarpine hybrid (6-C6) being more active. The hydrogen bond of interest is formed by the isopilocarpine hybrid (6-C6) but not by the pilocarpine hybrid (5-C6). Hybridization thus leads to a modified binding mode of the orthosteric moiety, as the binding mode of the hybrid is dominated by the high-affinity allosteric moiety.

AKT controls protein synthesis and oxidative metabolism via combined mTORC1 and FOXO1 signalling to govern muscle physiology.

BACKGROUND: Skeletomuscular diseases result in significant muscle loss and decreased performance, paralleled by a loss in mitochondrial and oxidative capacity. Insulin and insulin-like growth factor-1 (IGF-1) are two potent anabolic hormones that activate a host of signalling intermediates including the serine/threonine kinase AKT to influence skeletal muscle physiology. Defective AKT signalling is associated with muscle pathology, including cachexia, sarcopenia, and disuse; however, the mechanistic underpinnings remain unresolved. METHODS: To elucidate the role of AKT signalling in muscle mass and physiology, we generated both congenital and inducible mouse models of skeletal muscle-specific AKT deficiency. To understand the downstream mechanisms mediating AKT's effects on muscle biology, we generated mice lacking AKT1/2 and FOXO1 (M-AKTFOXO1TKO and M-indAKTFOXO1TKO) to inhibit downstream FOXO1 signalling, AKT1/2 and TSC1 (M-AKTTSCTKO and M-indAKTTSCTKO) to activate mTORC1, and AKT1/2, FOXO1, and TSC1 (M-QKO and M-indQKO) to simultaneously activate mTORC1 and inhibit FOXO1 in AKT-deficient skeletal muscle. Muscle proteostasis and physiology were assessed using multiple assays including metabolic labelling, mitochondrial function, fibre typing, ex vivo physiology, and exercise performance. RESULTS: Here, we show that genetic ablation of skeletal muscle AKT signalling resulted in decreased muscle mass and a loss of oxidative metabolism and muscle performance. Specifically, deletion of muscle AKT activity during development or in adult mice resulted in a significant reduction in muscle growth by 30-40% (P  < 0.0001; n = 12-20) and 15% (P < 0.01 and P < 0.0001; n = 20-30), respectively. Interestingly, this reduction in muscle mass was primarily due to an ~40% reduction in protein synthesis in both M-AKTDKO and M-indAKTDKO muscles (P < 0.05 and P < 0.01; n = 12-20) without significant changes in proteolysis or autophagy. Moreover, a significant reduction in oxidative capacity was observed in both M-AKTDKO (P < 0.05, P < 0.01 and P < 0.001; n = 5-12) and M-indAKTDKO (P < 0.05 and P < 0.01; n = 4). Mechanistically, activation and inhibition of mTORC1/FOXO1, respectively, but neither alone, were sufficient to restore protein synthesis, muscle oxidative capacity, and muscle function in the absence of AKT in vivo. In a mouse model of disuse-induced muscle loss, simultaneous activation of mTORC1 and inhibition of FOXO1 preserved muscle mass following immobilization (~5-10% reduction in casted M-indFOXO1TSCDKO muscles vs. ~30-40% casted M-indControl muscles, P < 0.05 and P < 0.0001; n = 8-16). CONCLUSIONS: Collectively, this study provides novel insights into the AKT-dependent mechanisms that underlie muscle protein homeostasis, function, and metabolism in both normal physiology and disuse-induced muscle wasting.

Pregnane-Oximino-Alkyl-Amino-Ether Compound as a Novel Class of TGR5 Receptor Agonist Exhibiting Antidiabetic and Anti-Dyslipidemic Activities.

INTRODUCTION: The present study deals with the synthesis of pregnane-oximino-amino-alkyl-ethers and their evaluation for antidiabetic and anti-dyslipidemic activities in validated animal and cell culture models. METHODS: The effect on glucose tolerance was measured in sucrose-loaded rats; antidiabetic activity was evaluated in streptozotocin (STZ)-induced diabetic rats and genetically diabetic db/db mice; the anti-dyslipidemic effect was characterized in high-fructose, high-fat diet (HFD)-fed dyslipidemic hamsters. The effect on glucose production and glucose utilization was analyzed in HepG2 liver and L6 skeletal muscle cells, respectively. RESULTS: From the synthesized molecules, pregnane-oximino-amino-alkyl-ether (compound 14b) improved glucose clearance in sucrose-loaded rats and exerted antihyperglycemic activity on STZ-induced diabetic rats. Further evaluation in genetically diabetic db/db mice showed temporal decrease in blood glucose, and improvement in glucose tolerance and lipid parameters, associated with mild improvement in the serum insulin level. Moreover, compound 14b treatment displayed an anti-dyslipidemic effect characterized by significant improvement in altered lipid parameters of the high-fructose, HFD-fed dyslipidemic hamster model. In vitro analysis in the cellular system suggested that compound 14b decreased glucose production in liver cells and stimulated glucose utilization in skeletal muscle cells. These beneficial effects of compound 14b were associated with the activation of the G-protein-coupled bile acid receptor TGR5. CONCLUSION: Compound 14b exhibits antidiabetic and anti-dyslipidemic activities through activating the TGR5 receptor system and can be developed as a lead for the management of type II diabetes and related metabolic complications.

Fructose-induced AGEs-RAGE signaling in skeletal muscle contributes to impairment of glucose homeostasis.

Increased fructose intake has been linked to the development of dyslipidemia, obesity and impaired glucose tolerance. Due to its specific metabolic fate, fructose impairs normal lipid and carbohydrate metabolism and facilitates the non-enzymatic glycation reaction leading to enhanced accumulation of advanced glycation end products (AGEs). However, the formation of fructose-AGEs under in vivo setup and its tissue specific accumulation is less explored. Here, we investigated the impact of high fructose on AGEs accumulation in skeletal muscle and its causal role in impaired glucose homeostasis. In L6 rat skeletal muscle cells, chronic exposure to fructose induced AGEs accumulation and the cellular level of the receptor for AGEs (RAGE) and the effect was prevented by pharmacological inhibition of glycation. Under in vivo settings, Sprague Dawley rats exposed to 20% fructose in drinking water for 16 weeks, displayed increased fasting glycemia, impaired glucose tolerance, decreased skeletal muscle Akt (Ser-473) phosphorylation, and enhanced triglyceride levels in serum, liver and gastrocnemius muscle. We also observed a high level of AGEs in serum and gastrocnemius muscle of fructose-supplemented animals, associated with methylglyoxal accumulation and up regulated expression of RAGE in gastrocnemius muscle. Treatment with aminoguanidine inhibited fructose-induced AGEs accumulation and normalized the expression of RAGE and Dolichyl-Diphosphooligosaccharide-Protein Glycosyltransferase (DDOST) in gastrocnemius muscle. Inhibition of AGEs-RAGE axis counteracted fructose-mediated glucose intolerance without affecting energy metabolism. These data reveal diet-derived AGEs accumulation in skeletal muscle and the implication of tissue specific AGEs in metabolic derangement, that may open new perspectives in pathogenic mechanisms and management of metabolic diseases.

MondoA drives muscle lipid accumulation and insulin resistance.

Obesity-related insulin resistance is associated with intramyocellular lipid accumulation in skeletal muscle. We hypothesized that in contrast to current dogma, this linkage is related to an upstream mechanism that coordinately regulates both processes. We demonstrate that the muscle-enriched transcription factor MondoA is glucose/fructose responsive in human skeletal myotubes and directs the transcription of genes in cellular metabolic pathways involved in diversion of energy substrate from a catabolic fate into nutrient storage pathways including fatty acid desaturation and elongation, triacylglyeride (TAG) biosynthesis, glycogen storage, and hexosamine biosynthesis. MondoA also reduces myocyte glucose uptake by suppressing insulin signaling. Mice with muscle-specific MondoA deficiency were partially protected from insulin resistance and muscle TAG accumulation in the context of diet-induced obesity. These results identify MondoA as a nutrient-regulated transcription factor that under normal physiological conditions serves a dynamic checkpoint function to prevent excess energy substrate flux into muscle catabolic pathways when myocyte nutrient balance is positive. However, in conditions of chronic caloric excess, this mechanism becomes persistently activated leading to progressive myocyte lipid storage and insulin resistance.

Correction: Airway epithelial cells prime plasmacytoid dendritic cells to respond to pathogens via secretion of growth factors.

The original version of this Article contained an error in the spelling of the author Hannah Nguyen, which was incorrectly given as Hannah Ngyuen. This has now been corrected in both the PDF and HTML versions of the Article.

Airway epithelial cells prime plasmacytoid dendritic cells to respond to pathogens via secretion of growth factors.

Plasmacytoid dendritic cells (PDCs) are critical for defense against respiratory viruses because of their propensity to secrete high levels of type I interferons (IFN). The functions of PDCs in the lung can be influenced by airway epithelial cells. We examined the effect of human primary bronchial epithelial cells (PBECs) on PDC functions by performing RNA-sequencing of PDCs after co-culture with air liquid interface differentiated PBECs. Functional analysis revealed that PDCs co-cultured with PBECs displayed upregulation of type I IFN production and response genes. Upregulated transcripts included those encoding cytosolic sensors of DNA, ZBP-1,IRF-3, and NFkB as well as genes involved in amplification of the IFN response, such as IFNAR1, JAK/STAT, ISG15. In keeping with the RNA-seq data, we observe increased secretion of type I IFN and other cytokines in response to influenza in PDCs co-cultured with PBECs. The PDCs also primed Th1 responses in T cells. The enhanced response of PDCs co-cultured with PBECs was due to the action of growth factors, GMCSF, GCSF, and VEGF, which were secreted by PBECs on differentiation. These data highlight possible mechanisms to enhance the production of type-I IFN in the airways, which is critical for host defense against respiratory infections.

Comparative evaluation of the effect of different disinfecting agents on bond strength of composite resin to dentin using two-step self-etch and etch and rinse bonding systems: An in-vitro study.

OBJECTIVE: This study was carried out to examine the effect of the application of different disinfecting agents on the shear bond strength (SBS) of an etch-and-rinse system. MATERIALS AND METHODS: A total of 120 dentin surfaces of human molars were taken. Sixty samples were taken for immediate (testing was done after 24 h) and delayed (testing was done after 6 months) each. Further, they were divided into three subgroups (n = 20) according to the disinfectant used: Group I: control (no disinfectant); Group II: chlorhexidine based; and Group III: Aloe barbadensis miller (Aloe vera) based. Dentine bonding agent was applied, and resin composite build-ups were done for SBS testing. Samples were examined under scanning electron microscope for observing the gap formation for dentin-restoration interface. SBS results were statistically analyzed. RESULTS: The results of the study showed that both chlorhexidine and aloe vera had improved bond strength as compared to the control group both for immediate and delayed SBSs. CONCLUSION: Thus, aloe vera can be used as an alternative to chlorhexidine for cavity disinfection.

Evaluation of antibacterial efficacy of Chitosan, Chlorhexidine, Propolis and Sodium hypochlorite on Enterococcus faecalis biofilm : An in vitro study.

BACKGROUND: Long term successful root canal treatment requires effective debridement and disinfection of root canal system. Persistent periradicular lesions are usually associated with Enterococccus faecalis. Prompt research for natural alternatives for irrigation is mainly due to the constant increase in antibiotic resistant strains and side effects caused by synthetic drugs. Sodium hypochlorite; the gold standard for irrigation has many disadvantages. Therefore, the present study was aimed to explore newer irrigants probably be as more effective and at the same time would be less irritating to the tissues than NaOCl. MATERIAL AND METHODS: Ninety extracted human mandibular premolars were biomechanically prepared, vertically sectioned, placed in tissue culture wells exposing the root canal surface to E. faecalis to form a biofilm. At the end of 3rd week, all groups were irrigated with 3 ml of test solutions and control for 10 minutes. The samples were then scraped with a scalpel, inoculated on tryptone soy agar plates and incubated for 24 hours at 37ºC. The plates were then subjected to digital colony counter and evaluated for E. faecalisgrowth. The growth was statistically analysed by ANOVA & Post Hoc Tukey tests. RESULTS: Chitosan + Chlorhexidine, NaOCl and Chlorhexidine showed no statistically significant difference, whereas all the other inter­group differences were statistically significant (P<0.05). CONCLUSIONS: Chitosan + Chlorhexidine, Chlorhexidine and Propolis were found to be as efficacious as sodium hypochlorite. The use of natural alternatives as root canal irrigation solutions might prove to be advantageous considering several unfavorable properties of NaOCl. Key words:Antibacterial efficacy, Chitosan, Enterococcus faecalis, Root canal irrigation.

Antibacterial Effect of Azadirachta indica (Neem) or Curcuma longa (Turmeric) against Enterococcus faecalis Compared with That of 5% Sodium Hypochlorite or 2% Chlorhexidine in vitro.

The purpose of this study was to compare the antibacterial properties of Azadirachta indica (neem) or Curcuma longa (turmeric) against Enterococcus faecalis with those of 5% sodium hypochlorite or 2% chlorhexidine as root canal irrigants in vitro. The activity of neem, chlorhexidine, sodium hypochlorite, or turmeric against E. faecalis was measured on agar plates using the agar diffusion method. The tube dilution method was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the irrigants used. Chlorhexidine or neem exhibited the greatest antibacterial activity when used as endodontic irrigants against E. faecalis, followed by sodium hypochlorite. No statistically significant difference was observed between neem, sodium hypochlorite, or chlorhexidine. The MIC of neem was 1: 128, which was similar to that of chlorhexidine. The MBC for each of these irrigants was 1: 16. Neem yielded antibacterial activity equivalent to 2% chlorhexidine or sodium hypochlorite against E. faecalis, suggesting that it offers a promising alternative to the other root canal irrigants tested.

Synthesis of substituted 2H-benzo[e]indazole-9-carboxylate as a potent antihyperglycemic agent that may act through IRS-1, Akt and GSK-3ß pathways.

Based on high throughput screening of our chemical library, we identified two 4,5-dihydro-2H-benzo[e]indazole derivatives (5d and 5g), which displayed a significant effect on glucose uptake in L6 skeletal muscle cells. Based on these lead molecules, a series of benzo[e]indazole derivatives were prepared. Among all the synthesized dihydro-2H-benzo[e]indazoles, 8-(methylthio)-2-phenyl-6-p-tolyl-4,5-dihydro-2H-benzo[e]indazole-9-carboxylate (5e) showed significant glucose uptake stimulation in L6 skeletal muscle cells, even better than lead compounds. Additionally, 5e decreased glucagon-induced glucose release in HepG2 hepatoma cells. The 2H-benzo[e]indazole 5e exerted an antihyperglycemic effect in normal, sucrose challenged streptozotocin-induced diabetic rats and type 2 diabetic db/db mice. Treatment with 5e at a dose of 30 mg kg-1 in db/db mice caused a significant decrease in triglyceride and total cholesterol levels and increased the HDL-C level in a significant manner. The mechanistic studies revealed that the 2H-benzo[e]indazole 5e significantly stimulated insulin-induced signaling at the level of IRS-1, Akt and GSK-3ß in L6 skeletal muscle cells, possibly by inhibiting protein tyrosine phosphatase-1B. This new 2H-benzo[e]indazole derivative has potential for the treatment of diabetes with improved lipid profile.

Comparative evaluation of the effect of chlorhexidine and Aloe barbadensis Miller (Aloe vera) on dentin stabilization using shear bond testing.

INTRODUCTION: The main objective of adhesive dentistry is to create an effective, durable union between the tooth structure and restorative material. However, degradation of adhesive dentine interface remains largely responsible for the relatively short lifetime of tooth colored resin restoration. AIM: The aim of the study is to compare the dentin collagen stabilization property of Chlorhexidine (CHX) and Aloe barbadensis Miller using shear bond strength testing. MATERIALS AND METHODS: Occlusal reduction was done in sixty extracted human mandibular molars to expose the mid coronal dentin and divided into three groups n = 20. Following the surface pretreatment (Group 1 = control, Group 2 = CHX, Group 3 = Aloevera), dentine bonding agent and composite resin were applied and cured. The specimens were then subjected to shear bond strength testing. RESULTS: From the results analyzed, it was noted that there was statistically significant difference between the groups Control and CHX and Control and A. barbadensis Miller (P < 0.05), specifically the values of Control < CHX and Control < A. barbadensis Miller (P < 0.05). However, there was no statistically significant difference between CHX and A. barbadensis Miller (P > 0.05). Hence, the following result for the shear bond strengths to dentin was obtained: Control < CHX ≈ A. barbadensis Miller. CONCLUSION: CHX and A. barbadensis Miller, as pretreatment agents of acid demineralized dentin collagen, has no adverse effect on the immediate shear bond strength of a two-step etch and rinse adhesive to dentin.

Endodontic Management of Maxillary First Molar with Five Root Canals, Including Two Distobuccal Root Canals: A Case Report.

Multiple canals in the root are part of the normal morphology of the tooth. A canal may sometimes be overlooked, however, and this may lead to failure of treatment. The first step in successful endodontic treatment, therefore, is gaining access to the pulp chamber and locating all the canals. In order to achieve this goal, practitioners need to be familiar with all possible variations in root canal morphology, and should thoroughly explore roots to ensure that all canals are identified, debrided, and obturated. Here, we report the diagnosis, treatment planning, and endodontic management of a maxillary first molar with five root canals, including two distobuccal root canals, in a 22-year-old woman.

Radix Entomolaris: A Report of Two Cases.

A comprehensive knowledge of the anatomy of the root canal is a basic prerequisite for the success of endodontic treatment. Mandibular molars may have an additional root located lingually (radix entomolaris) or buccally (radix paramolaris). An awareness of the potential for and understanding of unusual external and internal root canal mor-phology contributes to the successful outcome of root canal treatment. Here, we report two cases of radix entomolaris to increase awareness and understanding of this unusual morphology so that procedural errors during endodontic therapy might better be avoided.

Fructose induces mitochondrial dysfunction and triggers apoptosis in skeletal muscle cells by provoking oxidative stress.

Mitochondrial dysfunction in skeletal muscle has been implicated in the development of insulin resistance, a major characteristic of type 2 diabetes. There is evidence that oxidative stress results from the increased production of reactive oxygen species and reactive nitrogen species leads to mitochondrial dysfunction, tissue damage, insulin resistance, and other complications observed in type 2 diabetes. It has been suggested that intake of high fructose contributes to insulin resistance and other metabolic disturbances. However, there is limited information about the direct effect of fructose on the mitochondrial function of skeletal muscle, the major metabolic determinant of whole body insulin activity. Here, we assessed the effect of fructose exposure on mitochondria-mediated mechanisms in skeletal muscle cells. Exposure of L6 myotubes to high fructose stimulated the production of mitochondrial reactive oxygen species and nitric oxide (NO), and the expression of inducible NO synthase. Fructose-induced oxidative stress was associated with increased translocation of nuclear factor erythroid 2-related factor-2 to the nucleus, decreases in mitochondrial DNA content and mitochondrial dysfunctions, as evidenced by decreased activities of citrate synthase and mitochondrial dehydrogenases, loss of mitochondrial membrane potential, decreased activity of the mitochondrial respiratory complexes, and impaired mitochondrial energy metabolism. Furthermore, positive Annexin-propidium iodide staining and altered expression of Bcl-2 family members and caspases in L6 myotubes indicated that the cells progressively became apoptotic upon fructose exposure. Taken together, these findings suggest that exposure of skeletal muscle cells to fructose induced oxidative stress that decreased mitochondrial DNA content and triggered mitochondrial dysfunction, which caused apoptosis.

Free fatty acid induced impairment of insulin signaling is prevented by the diastereomeric mixture of calophyllic acid and isocalophyllic acid in skeletal muscle cells.

Elevated fatty acid levels play a pathogenic role in the development of insulin resistance, associated with type 2 diabetes. Interventions with ability to ameliorate fatty acid-induced insulin resistance might be useful for the management of diabetes. Here, we explored the effect of the diastereomeric mixture of calophyllic acid and isocalophyllic acid (F015) on palmitate-induced insulin resistance in skeletal muscle cells. An incubation of L6 myotubes with palmitate inhibited insulin-stimulated glucose uptake and translocation of GLUT4 to cell surface. Addition of F015 strongly prevented these inhibitions. Furthermore, F015 effectively inhibited the ability of palmitate to reduce insulin-stimulated phosphorylation of IRS-1, AKT and GSK-3ß in L6 myotubes. F015 presented a strong inhibition on palmitate-induced production of reactive oxygen species and associated inflammation, as the activation JNK, ERK1/2 and p38 MAPK were greatly reduced. F015 also inhibited inflammation-stimulated IRS-1 serine phosphorylation and restored insulin-stimulated IRS-1 tyrosine phosphorylation in presence of palmitate, resulted in enhanced insulin sensitivity. Results suggest that F015 inhibits palmitate-induced, reactive oxygen species-associated MAPK kinase activation and restored insulin sensitivity through regulating IRS-1 function. All these indicate F015 to be a potentially therapeutic candidate for insulin resistance and type 2 diabetes.

4-Hydroxyisoleucine ameliorates fatty acid-induced insulin resistance and inflammatory response in skeletal muscle cells.

The 4-hydroxyisoleucine (4-HIL), an unusual amino acid isolated from the seeds of Trigonella foenum-graecum was investigated for its metabolic effects to ameliorate free fatty acid-induced insulin resistance in skeletal muscle cells. An incubation of L6 myotubes with palmitate inhibited insulin stimulated-glucose uptake and -translocation of glucose transporter 4 (GLUT4) to the cell surface. Addition of 4-HIL strongly prevented this inhibition. We then examined the insulin signaling pathway, where 4-HIL effectively inhibited the ability of palmitate to reduce insulin-stimulated phosphorylation of insulin receptor substrate-1 (IRS-1), protein kinase B (PKB/AKT), AKT substrate of 160 kD (AS160) and glycogen synthase kinase 3ß (GSK-3ß) in L6 myotubes. Moreover, 4-HIL presented strong inhibition on palmitate-induced production of reactive oxygen species (ROS) and associated inflammation, as the activation of NF-κB, JNK1/2, ERK1/2 and p38 MAPK was greatly reduced. 4-HIL also inhibited inflammation-stimulated IRS-1 serine phosphorylation and restored insulin-stimulated IRS-1 tyrosine phosphorylation in the presence of palmitate, leading to enhanced insulin sensitivity. These findings suggested that 4-HIL could inhibit palmitate-induced, ROS-associated inflammation and restored insulin sensitivity through regulating IRS-1 function.

Synthesis of novel triterpene and N-allylated/N-alkylated niacin hybrids as α-glucosidase inhibitors.

Diabetes mellitus is a metabolic disorder characterized by chronic hyperglycemia. α-Glucosidase (EC 3.2.1.20) inhibitors interfere with enzymatic action to slow down the liberation of d-glucose from oligosaccharides and disaccharides, resulting in delayed glucose absorption and decreased postprandial plasma glucose levels. In continuation of our drug discovery program on antidiabetic agents, we synthesized novel N-allylated/N-alkylated niacin and α-amyrin (4-9) and lupeol (12-16) hybrids and tested for their α-glucosidase inhibiting activity. Compounds 4-9 showed better activity profile than the marketed α-glucosidase inhibitor i.e. acarbose. Compound 4 possess the highest inhibitory action with IC50 of 5 µM. Kinetic and CD studies revealed that 4 inhibited the α-glucosidase in a noncompetitive manner and caused conformational changes in secondary structure of the enzyme protein.

Diastereomeric mixture of calophyllic acid and isocalophyllic acid stimulates glucose uptake in skeletal muscle cells: involvement of PI-3-kinase- and ERK1/2-dependent pathways.

The diastereomeric mixture of calophyllic acid and isocalophyllic acid (F015) isolated from the leaves of Calophyllum inophyllum was investigated for the metabolic effect on glucose transport in skeletal muscle cells. In L6 myotubes, F015 dose-dependently stimulated glucose uptake by increasing translocation of glucose transporter4 (GLUT4) to plasma membrane without affecting their gene expression. The effects on glucose uptake were additive to insulin. Inhibitors analyses revealed that F015-induced glucose uptake was dependent on the activation of phosphatidylinositol-3-kinase (PI-3-K) and extracellular signal-regulated kinases 1 and 2 (ERK1/2), while independent to the activation of 5'AMP-activated kinase (AMPK). F015 significantly increased the phosphorylation of AKT, AS160 and ERK1/2, account for the augmented glucose transport capacity in L6 myotubes. Furthermore, F015 improved glucose tolerance and enhanced insulin sensitivity in skeletal muscle of dexamethasone-induced insulin resistant mice. Our findings demonstrate that F015 activates glucose uptake in skeletal muscle cells through PI-3-K- and EKR1/2-dependent mechanisms and can be a potential lead for the management of diabetes and obesity.