Synthesis,and In Vitro and In Silico α-Glucosidase Inhibitory Studies of 5-Chloro-2-Aryl Benzo[d]thiazoles

Twenty-five derivatives of 5-chloro-2-aryl benzo[d]thiazole (1–25) were synthesized and evaluated for their α-glucosidase (S. cerevisiae EC 3.2.1.20) inhibitory activity in vitro. Among them eight compounds showed potent activity with IC₅₀ values between 22.1 ± 0.9 and 136.2 ± 5.7 μM, when compared with standard acarbose (IC₅₀ = 840 ± 1.73 μM). The most potent compounds 4, 9, and 10 showed IC₅₀ values in the range of 22.1 ± 0.9 to 25.6 ± 1.5 μM. Compounds 2, 5, 11, and 19 showed IC₅₀ values within the range of 40.2 ± 0.5 to 60.9 ± 2.0 μM. Compounds 1 and 3 were also found to be good inhibitors with IC₅₀ values 136.2 ± 5.7 and 104.8 ± 9.9 μM, respectively. Their activities were compared with α-glucosidase inhibitor drug acarbose (standard) (IC₅₀ = 840 ± 1.73 μM). The remaining compounds were inactive. Structure-activity relationships (SAR) have also been established. Kinetics studies indicated compounds 2, 3, 10, 19, and 25 to be non-competitive, while 1, 5, 9, and 11 as competitive inhibitors of α-glucosidase enzyme. All the active compounds (1–5, 9–11, and 19) were also found to be non-cytotoxic, in comparison to the standard drug i.e., doxorubicin (IC₅₀ = 0.80 ± 0.12 μM) in MTT assay. Furthermore, molecular interactions of active compounds with the enzyme binding sites were predicted through molecular modeling studies.     

Changes in glucose metabolism from discrete regions of rat brain and its relationship to reproductive failure during experimental diabetes

This study reports the effects of alloxan induced diabetes on glucose metabolism enzymes viz. Hexokinase, Lactate dehydrogenase, and Glucose-6-phosphate dehydrogenase from discrete brain regions. Enzymes activity was assayed from hypothalamic areas such as medial preoptic area and median eminence-arcuate region which have gonadotropin releasing hormone cell bodies and their terminals, respectively and other brain regions like septum, amygdala, hippocampus, and thalamus. In all the areas studied, induction of diabetes resulted in a significant decrease in particulate bound HK activity, whereas soluble HK, LDH and G6PDH activity showed increase at 3, 8, 15 and 28 days intervals. Insulin treatment of diabetic rats led to recovery in enzyme activity. Blood glucose levels increased significantly after induction of diabetes and recovery was seen after insulin treatment. The present results suggest that altered cerebral glucose metabolism may also be responsible for reproductive failure observed in diabetic rats. (Mol Cell Biochem141: 97–102, 1994)     

Personalized microbiome-driven effects of non-nutritive sweeteners on human glucose tolerance

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Diabetes Inhibits Cerebral Ischemia-Induced Astrocyte Activation - an Observation in the Cingulate Cortex

The objective of this study was to study the effect of diabetic hyperglycemia on astrocytes after forebrain ischemia. Streptozotocin (STZ)-injected hyperglycemic and vehicle-injected normoglycemic rats were subjected to 15 minutes of forebrain ischemia. The brains were harvested in sham-operated controls and in animals with 1 and 6 h of recirculation following ischemia. Brain damage was accessed by haematoxylin and eosin (H&E) staining, cleaved caspase-3 immunohistochemistry and TdT-mediated-dUTP nick end labeling (TUNEL). Anti-GFAP antibody was employed to study astrocytes. The results showed that the 15-minute ischemia caused neuronal death after 1 and 6 h of reperfusion as revealed by increased numbers of karyopyknotic cells, edema, TUNEL-positive and active caspase-3-positive cells. Ischemia also activated astrocytes in the cingulated cortex as reflected by astrocyte stomata hypertrophy, elongated dendrites and increases in the number of dendrites, and immunoreactivity of GFAP. Diabetic hyperglycemia further enhanced neuronal death and suppressed ischemia-induced astrocyte activation. Further, diabetes-damaged astrocytes have increased withdrawal of the astrocyte end-foot from the cerebral blood vessel wall. It is concluded that diabetes-induced suppression and damages to astrocytes may contribute to its detrimental effects on recovery from cerebral ischemia.     

Temporal Profile of Astrocytes and Changes of Oligodendrocyte-Based Myelin Following Middle Cerebral Artery Occlusion in Diabetic and Non-diabetic Rats

The long-term impacts of cerebral ischemia and diabetic ischemia on astrocytes and oligodendrocytes have not been defined. The objective of this study is to define profile of astrocyte and changes of myelin in diabetic and non-diabetic rats subjected to focal ischemia.Focal cerebral ischemia of 30-min duration was induced in streptozotocin-induced diabetic and vehicle-injected normoglycemic rats. The brains were harvested for immunohistochemistry of glial fibrillary acidic protein (GFAP) and 2'', 3''-cyclic nucleotide 3''-phosphodiesterase (CNPase) at various reperfusion endpoints ranging from 30 min up to 28 days. The results showed that activate astrocytes were observed after 30 min and peaked at 3 h to 1 day after reperfusion in ischemic penumbra, and peaked at 7 days of reperfusion in ischemic core. Diabetes inhibited the activation of astrocytes in ischemic hemisphere. Demyelination occurred after 30 min of reperfusion in ischemic core and peaked at 1 day. Diabetes caused more severe demyelination compared with non-diabetic rats. Remyelination started at 7 days and completed at 14 and 28 days in ischemic region. Diabetes inhibited the remyelination processes. It is concluded that ischemia activates astrocytes and induces demyelination. Diabetes inhibits the activation of astrocytes, exacerbates the demyelination and delays the remyelination processes. These may contribute to the detrimental effects of hyperglycemia on ischemic brain damage.     

A Zebrafish Model of Diabetes Mellitus and Metabolic Memory

Diabetes mellitus currently affects 346 million individuals and this is projected to increase to 400 million by 2030. Evidence from both the laboratory and large scale clinical trials has revealed that diabetic complications progress unimpeded via the phenomenon of metabolic memory even when glycemic control is pharmaceutically achieved. Gene expression can be stably altered through epigenetic changes which not only allow cells and organisms to quickly respond to changing environmental stimuli but also confer the ability of the cell to "memorize" these encounters once the stimulus is removed. As such, the roles that these mechanisms play in the metabolic memory phenomenon are currently being examined.We have recently reported the development of a zebrafish model of type I diabetes mellitus and characterized this model to show that diabetic zebrafish not only display the known secondary complications including the changes associated with diabetic retinopathy, diabetic nephropathy and impaired wound healing but also exhibit impaired caudal fin regeneration. This model is unique in that the zebrafish is capable to regenerate its damaged pancreas and restore a euglycemic state similar to what would be expected in post-transplant human patients. Moreover, multiple rounds of caudal fin amputation allow for the separation and study of pure epigenetic effects in an in vivo system without potential complicating factors from the previous diabetic state. Although euglycemia is achieved following pancreatic regeneration, the diabetic secondary complication of fin regeneration and skin wound healing persists indefinitely. In the case of impaired fin regeneration, this pathology is retained even after multiple rounds of fin regeneration in the daughter fin tissues. These observations point to an underlying epigenetic process existing in the metabolic memory state. Here we present the methods needed to successfully generate the diabetic and metabolic memory groups of fish and discuss the advantages of this model.     

Gestational diabetes mellitus epigenetically affects genes predominantly involved in metabolic diseases

Offspring exposed to gestational diabetes mellitus (GDM) have an increased risk for chronic diseases, and one promising mechanism for fetal metabolic programming is epigenetics. Therefore, we postulated that GDM exposure impacts the offspring’s methylome and used an epigenomic approach to explore this hypothesis. Placenta and cord blood samples were obtained from 44 newborns, including 30 exposed to GDM. Women were recruited at first trimester of pregnancy and followed until delivery. GDM was assessed after a 75-g oral glucose tolerance test at 24–28 weeks of pregnancy. DNA methylation was measured at > 485,000 CpG sites (Infinium HumanMethylation450 BeadChips). Ingenuity Pathway Analysis was conducted to identify metabolic pathways epigenetically affected by GDM. Our results showed that 3,271 and 3,758 genes in placenta and cord blood, respectively, were potentially differentially methylated between samples exposed or not to GDM (p-values down to 1 × 10⁻⁰⁶; none reached the genome-wide significance levels), with more than 25% (n = 1,029) being common to both tissues. Mean DNA methylation differences between groups were 5.7 ± 3.2% and 3.4 ± 1.9% for placenta and cord blood, respectively. These genes were likely involved in the metabolic diseases pathway (up to 115 genes (11%), p-values for pathways = 1.9 × 10⁻¹³ < p < 4.0 × 10⁻⁰³; including diabetes mellitus p = 4.3 × 10⁻¹¹). Among the differentially methylated genes, 326 in placenta and 117 in cord blood were also associated with newborn weight. Our results therefore suggest that GDM has epigenetic effects on genes preferentially involved in the metabolic diseases pathway, with consequences on fetal growth and development, and provide supportive evidence that DNA methylation is involved in fetal metabolic programming.     

Suppression of autophagy is protective in high glucose-induced cardiomyocyte injury

Hyperglycemia is linked to increased heart failure among diabetic patients. However, the mechanisms that mediate hyperglycemia-induced cardiac damage remain poorly understood. Autophagy is a cellular degradation pathway that plays important roles in cellular homeostasis. Autophagic activity is altered in the diabetic heart, but its functional role has been unclear. In this study, we determined if mimicking hyperglycemia in cultured cardiomyocytes from neonatal rats and adult mice could affect autophagic activity and myocyte viability. High glucose (17 or 30 mM) reduced autophagic flux compared with normal glucose (5.5 mM) as indicated by the difference in protein levels of LC3-II (microtubule-associated protein 1 light chain 3 form II) or the changes of punctate fluorescence patterns of GFP-LC3 and mRFP-LC3 in the absence and presence of the lysosomal inhibitor bafilomycin A₁. Unexpectedly, the inhibited autophagy turned out to be an adaptive response that functioned to limit high glucose cardiotoxicity. Indeed, suppression of autophagy by 3-methyladenine or short hairpin RNA-mediated silencing of the Becn1 or Atg7 gene attenuated high glucose-induced cardiomyocyte death. Conversely, upregulation of autophagy with rapamycin or overexpression of Becn1 or Atg7 predisposed cardiomyocytes to high glucose toxicity. Mechanistically, the high glucose-induced inhibition of autophagy was mediated at least partly by increased mTOR signaling that likely inactivated ULK1 through phosphorylation at serine 467. Together, these findings demonstrate that high glucose inhibits autophagy, which is a beneficial adaptive response that protects cardiomyocytes against high glucose toxicity. Future studies are warranted to determine if autophagy plays a similar role in diabetic heart in vivo.     

Diabetic liver injury from streptozotocin is regulated through the caspase-8 homolog cFLIP involving activation of JNK2 and intrahepatic immunocompetent cells

The endemic occurrence of obesity and the associated risk factors that constitute the metabolic syndrome have been predicted to lead to a dramatic increase in chronic liver disease. Non-alcoholic steatohepatitis (NASH) has become the most frequent liver disease in countries with a high prevalence of obesity. In addition, hepatic steatosis and insulin resistance have been implicated in disease progression of other liver diseases, including chronic viral hepatitis and hepatocellular carcinoma. The molecular mechanisms underlying the link between insulin signaling and hepatocellular injury are only partly understood. We have explored the role of the antiapoptotic caspase-8 homolog cellular FLICE-inhibitory protein (cFLIP) on liver cell survival in a diabetic model with hypoinsulinemic diabetes in order to delineate the role of insulin signaling on hepatocellular survival. cFLIP regulates cellular injury from apoptosis signaling pathways, and loss of cFLIP was previously shown to promote injury from activated TNF and CD95/Apo-1 receptors. In mice lacking cFLIP in hepatocytes (flip^−/−), loss of insulin following streptozotocin treatment resulted in caspase- and c-Jun N-terminal kinase (JNK)-dependent liver injury after 21 days. Substitution of insulin, inhibition of JNK using the SP600125 compound in vivo or genetic deletion of the mitogen-activated protein kinase (MAPK)9 (JNK2) in all tissues abolished the injurious effect. Strikingly, the difference in injury between wild-type and cFLIP-deficient mice occurred only in vivo and was accompanied by liver-infiltrating inflammatory cells with a trend toward increased amounts of NK1.1-positive cells and secretion of proinflammatory cytokines. Transfer of bone marrow from rag-1-deficient mice that are depleted from B and T lymphocytes prevented liver injury in flip^−/− mice. These findings support a direct role of insulin on cellular survival by alternating the activation of injurious MAPK, caspases and the recruitment of inflammatory cells to the liver. Thus, increasing resistance to insulin signaling pathways in hepatocytes appears to be an important factor in the initiation and progression of chronic liver disease.     

Ultrasonic synthesis of tyramine derivatives as novel inhibitors of α-glucosidase in vitro

Tyramine derivatives 3–27 were synthesized by using conventional and environmental friendly ultrasonic techniques. These derivatives were then evaluated for the first time for their α-glucosidase (Sources: Saccharomyces cerevisiae and mammalian rat-intestinal acetone powder) inhibitory activity by using in vitro mechanism-based biochemical assays. Compounds 7, 14, 20, 21 and 26 were found to be more active (IC₅₀?=?49.7?±?0.4, 318.8?±?3.7, 23.5?±?0.9, 302.0?±?7.3 and 230.7?±?4.0?μM, respectively) than the standard drug, acarbose (IC₅₀?=?840.0?±?1.73?μM (observed) and 780?±?0.028?μM (reported)) against α-glucosidase obtained from Saccharomyces cerevisiae. Kinetic studies were carried out on the most active members of the series in order to determine their mode of inhibition and dissociation constants. Compounds 7, 20 and 26 were found to be the competitive inhibitors of α-glucosidase. These compounds were also screened for their protein antiglycation, and dipeptidyl peptidase-IV (DPP-IV) inhibitory activities. Only compounds 20, 22 and 27 showed weak antiglycation activity with IC₅₀ values 505.27?±?5.95, 581.87?±?5.50 and 440.58?±?2.74?μM, respectively. All the compounds were found to be inactive against DDP-IV enzyme. Inhibition of α-glucosidase, DPP-IV enzymes and glycation of proteins are valid targets for the discovery of antidiabetic drugs. Cytotoxicity of compounds 3–27 was also evaluated by using mouse fibroblast 3T3 cell lines. All the compounds were found to be noncytotoxic. The current study describes the synthesis α-glucosidase inhibitory activity of derivatives, based on a natural product tyramine template. The compounds reported here may serve as the starting point for the design and development of novel α-glucosidase inhibitors as antidiabetic agents.