Chrysin reduces heart endoplasmic reticulum stress-induced apoptosis by inhibiting PERK and Caspase 3-7 in high-fat diet-fed rats.

BACKGROUND: Chrysin (Chy) is a naturally occurring flavonoid found in fruits, vegetables, honey, propolis, and many plant extracts that has shown notable medicinal value. Chy exhibits diverse pharmacological properties, including anti-oxidative, anti-inflammatory, anti-apoptotic, anti-cholesteremic, and cardioprotective. However, the influence of Chy in mitigating high-fat diet (HFD)-induced ER stress of rat myocardium remains unknown. PURPOSE: The current work intended to determine the therapeutic potential of Chy against HFD-induced endoplasmic stress-mediated apoptosis. METHODS: To evaluate the therapeutic value of Chy in HFD-induced endoplasmic stress-mediated apoptosis in the myocardium; The male wistar rats were divided into different groups; control, HFD control, HFD fed followed by Chy-treated and HFD fed followed by atorvastatin (Atv) treated rats. RESULTS: When compared to the control group, the HFD-fed rats had significantly higher levels of marker enzymes such as CK-NAC and ALP, as well as lipid peroxidation and lipid profile (TC, TG, LDL, and VLDL). Chy therapy greatly reversed these marker enzymes and the lipid profile. qRT-PCR Studies showed that Chy supplementation considerably improved Nrf2 and its target genes. In addition, Chy lowered the expression of PERK, CHOP, ATF6, GRP78, and Caspase-3 genes in the heart tissue of HFD-fed rats. Immunohistochemistry results demonstrated that Chy substantially enhanced the Nrf2 and reduced PERK and Caspase3-7 protein expression in HFD-fed rats. CONCLUSION: The current study concluded that Chy may mediate the cardioprotective effect by activating Nrf2 and inhibiting PERK signaling pathway against ER stress-mediated apoptosis induced by HFD. Therefore, supplementation with Chy could serve as a promising therapeutic target against HFD-induced ER stress-mediated cardiac complication.

Fucoidan from Sargassum wightii reduces oxidative stress through upregulating Nrf2/HO-1 signaling pathway in alloxan-induced diabetic cardiomyopathy rats.

BACKGROUND: Diabetic cardiomyopathy (DCM) is a form of cardiac dysfunction caused by diabetes, increasing heart failure and death. Studies shown that hyperglycemia-induced oxidative stress significantly affects heart structure and functional changes during diabetic cardiomyopathy. Fucoidans are sulfated polysaccharide derived from naturally available seaweeds and reported for various biological functions such as antioxidant, anti-diabetic, and anti-inflammatory. However, the therapeutic potential of Indian seaweeds against DCM remains largely unexplored. Therefore, the current study aimed to work on the cardioprotective effect of extracted fucoidan from Sargassum wightii (SwF) in alloxan-induced DCM. METHODS AND RESULTS: Diabetes (DM) was induced with alloxan monohydrate (150 mg/kg-1) dissolved in Nacl (0.9%) overnight-fasted rats. Group III, IV rats were DM induced, followed by treated with SwF (150 mg/kg-1) and (300 mg/kg-1). Group V and VI were non-diabetic rats and received SwF (150 mg/kg-1) and (300 mg/kg-1). SwF reduced classical progressive DM complications such as hyperglycemia, polydipsia, polyphagia, and polyurea in alloxan-induced diabetic rats. Biochemical analysis showed that SwF decreased blood glucose, cardiac markers enzymes, and lipid peroxidation levels compared to diabetic rats. SwF administration significantly increased Nrf2, HO-1, SOD, Catalase, and NQO1 gene expression. In addition, SwF-treated rats showed reduced heart tissue damage with increased Nrf2 and HO-1 protein expression. CONCLUSION: The current research concludes that targeting oxidative stress with SwF provided an effective role in the prevention of DCM. Thus, fucoidan could be used to develop functional food ingredients for DCM.

Role of haptoglobin 2-2 genotype on disease progression and mortality among South Indian chronic kidney disease patients.

BACKGROUND: Haptoglobin (HP), a plasma glycoprotein, binds to free hemoglobin and prevents the loss of iron and kidney damage. The variations of HP gene affect its enzyme activity, resulting in varied antioxidant, angiogenic and anti-inflammatory properties. HP 2-2 genotype showed 3.84 fold increased risk for the development of CKD in Taiwan population. With this background, the present work focused to conduct a prospective case-control study in South Indian population to evaluate whether the HP variants are associated to nondialysis (ND) (CKD stages 1-4) and ESRD (CKD stage 5) conditions. METHODS AND RESULTS: Totally 392 CKD patients (nondialysis, ND; n = 170, end-stage renal disease, ESRD; n = 222) and 202 healthy individuals were enrolled. The blood samples collected from the patients were used to determine biochemical parameters and HP genotyping. Gene frequency and biochemical parameters were statistically analyzed for disease association. Results showed that HP 2-2 genotypes were significantly associated with ND and ESRD disease development compared to controls. Higher HP2-2 genotype frequency showed an increased hazard ratio for overall disease progression among ND patients (hazard ratio = 3.86; 95% CI 1.88 to 7.93; P = 0.0002). Survival analysis also showed that non-HP2-2 patients have a statistically significantly decreased risk for mortality compared to patients with the HP2-2 genotype (ESRD patients hazard ratio = 4.05; P = 0.04). CONCLUSION: The present study confirms that HP2-2 polymorphism is statistically associated with the risk of CKD incidence, progression, and mortality among South Indians. Concluding our results, the HP2-2 genotype could be an independent predictor of all-cause mortality and disease progression in patients with CKD.

Ascorbic acid attenuates cadmium-induced myocardial hypertrophy and cardiomyocyte injury through Nrf2 signaling pathways comparable to resveratrol.

Chronic cadmium (Cd) exposure severely affects the structural integrity of the heart, leading to cardiovascular disease. This study investigates the protective role of ascorbic acid (AA) and resveratrol (Res) in cellular defense against Cd-induced cardiomyocyte damage and myocardial hypertrophy in H9c2 cardiomyocytes. Experimental results showed that AA and Res treatment significantly increased cell viability, reduced ROS production, attenuated lipid peroxidation, and increased antioxidant enzyme activity in Cd-induced H9c2 cells. AA and Res decreased the mitochondrial membrane permeability and protected the cells from Cd induced cardiomyocyte damage. This also suppressed the pathological hypertrophic response triggered by Cd, which increased the cell size of cardiomyocytes. Gene expression studies revealed that cells treated with AA and Res decreased the expression of hypertrophic genes ANP (two-fold), BNP (one-fold) and ß- MHC (two-fold) compared to Cd exposed cells. AA and Res promoted the nuclear translocation of Nrf2 and increased the expression of antioxidant genes (HO-1, NQO1, SOD and CAT) during Cd mediated myocardial hypertrophy. This study proves that AA and Res play a significant role in improving Nrf2 signaling, thereby reversing stress-induced injury, and facilitating the regression of myocardial hypertrophy.

Chrysin protects cardiac H9c2 cells against H2O2-induced endoplasmic reticulum stress by up-regulating the Nrf2/PERK pathway.

Oxidative and endoplasmic reticulum (ER) stress-mediated cardiac apoptosis is an essential pathological process in cardiovascular diseases (CVDs). Chrysin (Chy) is a natural flavonoid that exerts several health benefits, particularly anti-oxidative and anti-apoptotic effects. However, its protective effect against CVDs and its mechanism of action at a molecular level remains unclear. Therefore, the present study aimed to investigate the interaction of ER stress response protein with Chy by computational analysis and molecular action in H2O2-induced oxidative and ER stress in cardiomyoblast cells. H9c2 cells were pre-treated with 50 µM of Chy for 24 h and exposed to H2O2 for 1 h. Explore the Chy-mediated Nrf2 signalling on ER stress reduction, H9c2 cell lines were transfected with Nrf2 siRNA for 48 h and further treated with Chy for 24 h and subjected to H2O2 for 1 h. Chy pre-treatment increased the Nrf2-regulated gene expression, reduced the ER stress signalling genes such as CHOP and GRP78, and increased the PERK and AFT6 expression compared to H2O2-treated cells. Chy preincubation down-regulated the expression of PI3K, NF-κB, and caspase-3. Fluorescence staining revealed that Chy reduced intracellular ROS generation, ER stress, apoptosis, and increased MMP. This beneficial effect of Chy was abolished when silencing Nrf2 in H9c2 cells. Overall, the present study confirmed that Chy showed the cardioprotective effect by attenuating ER stress via the activation of Nrf2 signalling. Therefore, the study concluded that improving Nrf2 signalling by Chy supplementation could provide a promising therapeutic target in oxidative and ER stress-mediated CVDs complications.

Isolation and characterization of fucoidan from four brown algae and study of the cardioprotective effect of fucoidan from Sargassum wightii against high glucose-induced oxidative stress in H9c2 cardiomyoblast cells.

Oxidative stress plays a vital role in the initiation and progression of diabetic cardiomyopathy (DCM). Increased cardiac dysfunction and apoptosis in DCM are independent factors associated with hypertension or coronary artery disease. Fucoidan, a class of sulfated polysaccharides, is widely used as food supplements and reported to have various pharmacological properties. However, the pharmacological property of Indian seaweeds remains unexplored. The present study is focused on isolating and characterizing the fucoidan from four brown seaweeds such as Sargassum wightii (SwF), Sargassum swartzii (SsF), Sargassum polycystum (SpF), Turbinaria ornata (ToF), and aimed to investigate cardioprotective effect of fucoidan against High Glucose (HG) induced oxidative stress in H9c2 cells. The mild acid hydrolysis method was used to isolate crude fucoidan from four brown seaweeds purified by the FPLC system. The biochemical composition analysis showed that SwF had a high content of fucoidan and sulfate, followed by SsF, SpF, and ToF. Further, FTIR, XRD, NMR, and SEM analysis confirmed the isolated fucoidan structures. SwF showed higher DPPH activity compared to another isolated fucoidan. In vitro studies with SwF revealed significantly decreased cytotoxicity, prevented the loss of MMP, reduced lipid peroxidation, and increased cellular enzymatic and non-enzymatic activity. qRT-PCR results showed SwF significantly upregulated the Nrf2, HO-1, NQO1, and Bcl2 and down-regulated the Bax and Caspase-3 mRNA expression compared to HG-treated cells. In conclusion, SwF could be used to develop functional foods for diabetic-mediated CVD complications compared to another isolated fucoidan. PRACTICAL APPLICATIONS: Bioactive carbohydrates have gained significant interest among researchers to improve human health. The biomedical field showed great interest in seaweed research in managing various diseases. In particular, seaweeds contain many bioactive compounds because of their chemical and biological diversity. Despite the various beneficial effects of fucoidan in CVD, the therapeutic potential of Indian seaweeds remains largely unexplored. Hence, this study isolated fucoidan from four brown seaweeds and studied their bioactive properties. Results revealed that SwF showed higher free radical scavenging activity compared to another isolated fucoidan. Therefore, SwF was selected for the in vitro study. SwF increased the cytoprotection through increasing antioxidant levels against oxidative stress in H9c2 cells. Staining analysis showed SwF increased cellular protection via inhibiting ROS protection and increasing MMP. Overall, fucoidan from SwF could be developed as a functional food for CVD.

Chrysin reduces hypercholesterolemia-mediated atherosclerosis through modulating oxidative stress, microflora, and apoptosis in experimental rats.

Chrysin (Chy) is known for various biological proprieties such as inhibitory effects on inflammation, cancer, oxidative stress, aging, and atherosclerosis. However, the hypolipidemic activity of Chy and its mechanistic action remains unclear in cardiovascular diseases (CVD). In this study, we focused on the hypolipidemic proprieties of Chy in hypercholesterolemia-induced atherosclerosis. Male Wistar rats (150-220 g) were divided into four groups as follows: Group I control was fed with standard laboratory chow. Rats in Group II were fed a high-fat diet (HFD) for 60 days. After 60 days of HFD, Group III rats received Chy (100 mg/kg body weight); Group IV rats received Atorvastatin (Atv; 10 mg/kg body weight) for 30 days. Biochemical studies showed Chy, Atv treatment decreased the activities of liver marker enzymes and the levels of Reactive Oxygen Species (ROS) and lipid profile. Gene expression analysis on nuclear factor erythroid 2-related factor 2 (Nrf2) and its regulated genes were significantly reduced in the intestine and increased in the aorta by Chy and Atv. Gut microbial species such as Bacteroidetes, Lactobacillus, Enterococcus, and Clostridium leptum copy numbers were significantly increased by Chy and Atv treatment. In addition, Chy and Atv modulated the expression of inflammatory genes including TLR4, TNFα, NLRP3, and IL-17 in the aorta and intestine compared with hypercholesterolemic control rats. Chy and Atv effectively increased the caspase-3 mRNA expression in the intestine, but these decreased in the aorta. The present study concludes that by reducing oxidative stress and increasing gut microbial colonization, Chy may provide an effective therapeutic approach for the prevention of hypercholesterolemia-mediated atherosclerosis. PRACTICAL APPLICATIONS: Our study focused on a therapeutic model representing the clinical presentation of atherosclerosis in humans. Statins are commonly used in the treatment of cardiovascular complications, patients with hypercholesterolemia face difficulties in the continuation of statin therapy. The reason for statin discontinuation has been associated with toxicological effects. It is necessary to investigate the potentiality of the natural compound as an alternative medicine to statin with fewer side effects. The main theme of our study is to compare the therapeutic potential of Chy and Atv. Chy is a natural bioflavonoid that could be considered as an alternative medicinal compound to statins and to avoid toxicity problems associated with statins. Chy is a bioflavonoid present in Passiflora caerulea (blue passion flower), Oroxylum indicum (Indian trumpet flower), Pelargonium crispum, propolis, and honey. Consuming Chy-rich foods will reduce hypercholesterolemia-mediated cardiovascular complications. Overall, the present studies provided a key to developing bioactive compounds-based foods for CVD patients.

Gallic acid attenuates cadmium mediated cardiac hypertrophic remodelling through upregulation of Nrf2 and PECAM-1signalling in rats.

Gallic acid (GA) is an abundant natural polyphenolic compound found in vegetable and fruits that reduces the cardiac disease risk factor. This study aims to evaluate GA's role on cadmium (Cd) induced cardiac remodelling in experimental rats. Male Wistar rats were exposed to Cd (15 ppm) in drinking water and administered with GA orally (15 mg/kg/d) for 60 days. The results showed that GA regulated the lipid profile and reduced the LDL to 57 % compared with Cd treated rats. GA inhibited cardiac marker enzymes activity of CK-NAC (to 72.7 %) and CK-MB (to 100.3 %). Moreover, GA attenuated lipid peroxidation and enhanced the cardiac glutathione S transferase (GST) activity (89.2 %), glutathione peroxidase (GPx) (87 %), superoxide dismutase (SOD) (88.4 %) and catalase (CAT) activity (86.5 %). Histopathological examination showed that GA impaired the ventricular hypertrophy and fibrotic proliferation induced by Cd in rats. The combination of GA + Cd, decreased the gene expression of ANP (1-fold), BNP (0.5-fold) and ß- MHC (0.9-fold). Furthermore, GA significantly reduced the expression of profibrotic (TGF-ß) and proinflammatory (MCP-1) gene in Cd intoxicated rats. GA upregulated the expression of Nrf2 (2-fold), HO-1 (3-fold), and PECAM-1 (0.6-fold), which augments the detoxifying enzyme activity and cellular immunity in Cd intoxicated rats. The increased protein expression of Nrf2, PECAM-1 and decreased AKT-1 levels confirmed the mechanical action of GA during the hypertrophic condition. Thus, our results suggest that GA could act as a potential therapeutic agent regulating Nrf2 and PECAM-1 signalling pathways, thereby ameliorating Cd-induced pathological cardiac remodelling.

Chrysin attenuates high-fat-diet-induced myocardial oxidative stress via upregulating eNOS and Nrf2 target genes in rats.

Hypercholesterolemia is one of the risk factors associated with increased morbidity and mortality in cardiovascular disorders. Chrysin (Chy) is reported to exhibit anti-inflammatory, anti-cancerous, anti-oxidative, anti-aging, and anti-atherogenic properties. In the present study, we aimed to investigate whether Chy would mediate the cardioprotective effect against hypercholesterolemia-triggered myocardial oxidative stress. Male Sprague Dawley rats were divided into different groups as control and fed with high-fat diet (HFD) followed by oral administration of Chy (100 mg/kg b.wt), atorvastatin (Atv) (10 mg/kg b.wt), and L-NAME (10 mg/kg b.wt) for 30 days. At the end of the experimental period, the rats were sacrificed and tissues were harvested. Biochemical results showed a significant increase of cardiac disease marker enzymes (ALT, AST, and CKMB), lipid peroxidation, and lipid profile (TC, TG, LDL, and VLDL) in HFD-fed rat tissues when compared to control, whereas oral administration of Chy significantly reduced the activities of these marker enzymes and controlled the lipid profile. qRT-PCR studies revealed that Chy administration significantly increased the expression of endothelial nitric oxide synthase (eNOS), and Nrf2 target genes such as SOD, catalase, and GCL3 in left ventricular heart tissue of HFD-challenged rats. Immunohistochemistry results also showed that Chy treatment increased myocardial protein expression of eNOS and Nrf2 in HFD-challenged rats. Concluding the results of the present study, the Chy could mediate the cardioprotective effect through the activation of eNOS and Nrf2 signaling against hypercholesterolemia-induced oxidative stress. Thus, the administration of Chy would provide a promising therapeutic strategy for the prevention of HFD-induced oxidative stress-mediated myocardial complications.

Renoprotective effect of tectorigenin glycosides isolated from Iris spuria L. (Zeal) against hyperoxaluria and hyperglycemia in NRK-49Fcells.

Oxidative stress has been identified as an underlying factor in the development of insulin resistance, ß-cell dysfunction, impaired glucose tolerance and type 2 diabetes mellitus and it also play major role in kidney stone formation. The present study is aimed to elucidate the in vitro nephroprotective activity of two isoflavonoid glycosides, tectorigenin 7-O-ß-D-glucosyl-(1→6)-ß-D-glucoside (1) and tectorigenin 7-O-ß-D-glucosyl-4'-O-ß-D-glucoside (2) isolated from the n-BuOH fraction of Iris spuria L. (Zeal) rhizome MeOH extract against oxalate and high glucose-induced oxidative stress in NRK-49F cells. The results revealed that compounds 1 and 2 significantly increased the antioxidant enzyme activities and decreased MDA levels in both oxalate and high glucose stress. Treatment with these phytochemicals effectively down-regulated expression of crystal modulator genes and pro-fibrotic genes in oxalate and high glucose-mediated stress respectively. This study indicates cytoprotective, antioxidant, anti-urolithic and anti-diabetic effects of compounds 1 and 2 against oxalate and high glucose stress.[Figure: see text].

Loss of ERAD bridging factor UBX2 modulates lipid metabolism and leads to ER stress-associated apoptosis during cadmium toxicity in Saccharomyces cerevisiae.

The endoplasmic reticulum (ER) stress potentially activates the unfolded protein response (UPR) and ER-associated protein degradation (ERAD) as quality-control mechanisms. During ERAD process, the ERAD adaptor protein Ubx2 serves as a bridging factor and transports the misfolded proteins from the ER to the cytosol for subsequent ubiquitylation and proteasomal degradation. Cadmium (Cd) is a toxic metal that initiates ER stress and has an impact on lipid homeostasis and this study focuses on the synergistic impact of Cd exposure and ERAD (using ubx2∆ strain). With Cd exposure in ubx2∆ strain, we observed stunted growth and induction of ER stress. The ER stress was confirmed by measuring the expression of UPR marker (Kar2p), and mRNA expression of ER stress-responsive genes (HAC1, IRE1, ERO1, and PDI1), heat shock responsive genes (HSP104 and HSP60), ERAD pathway genes (DOA10, CDC48, HRD1, and YOS9), and proteasome regulators (UBI14, and RPN4). We also observed aberrant membrane morphology with DiOC6 staining, and interrupted mitochondria with mitotracker dye using microscopic analysis. The cell's inability to relieve stress through adaptive response results in apoptosis and was assessed using acridine orange (AO)-ethidium bromide (EtBr) staining. In ubx2∆ strain, there was reduction in triacylglycerol (TAG) and lipid droplets (LDs), and increase in the phospholipids. The mRNA expression of lipid metabolic genes (LRO1, DGA1, ARE1, ARE2, and OLE1) supported the lipid pattern observed. Collectively, our data suggest that in Saccharomyces cerevisiae, the Cd exposure on ubx2∆ strain induced cellular stress and has an impact on ERAD, UPR, and LD homeostasis.

GSTM1-null allele predicts rapid disease progression in nondialysis patients and mortality among South Indian ESRD patients.

Chronic kidney disease (CKD) is one of the main causes of early death in humans worldwide. Glutathione S-Transferases (GSTs) are involved in a series of xenobiotics metabolism and free radical scavenging. The previous studies elucidated the interlink between GST variants and to the development of various diseases. The present case-control study performed to ascertain whether GST polymorphisms are associated with the incidence and advancement of CKD. From the Southern part of India, a total of 392 CKD patients (nondialysis, ND; n = 170, end-stage renal disease, ESRD; n = 222) and 202 healthy individuals were enrolled. Patients were followed-up for 70 months. Serum biochemical parameters were recorded, and the extraction of DNA was done from the patient's blood samples. To genotype study participants, multiplex PCR for GSTM1/T1 was performed. Statistical analysis was carried out to analyze the relationship between gene frequency and sonographic grading, as well as biochemical parameters for disease development. The GSTM1-null genotype showed threefold increased risk (OR = 2.9304; 95% CI 1.8959 to 4.5296; P < 0.0001) to CKD development and twofold increased risk (OR = 1.8379; 95% CI 1.1937 to 2.8299; P = 0.0057) to ESRD progression. During the mean follow-up of 41 months study, multivariate Cox regression analysis revealed that GSTM1-null genotype has 4 times increased the risk for all-cause rapid disease progression to ESRD among ND patients and 3.85-fold increased risk for death among ESRD patients. Survival analysis revealed that patients with GSTM1-present allele showed a significantly diminished risk of mortality compared to patients bearing the GSTM1-null allele among ESRD patients with a hazard ratio of 4.6242 (P < 0.0001). Thus, present data confirm that GSTM1-null genotype increased the risk for all-cause rapid disease progression to ESRD among ND patients. Based on our results, GSTM1-null genotype could be considered as a significant predictor for causing mortality among CKD patients when compared to all other variables.

Effect of cadmium on essential metals and their impact on lipid metabolism in Saccharomyces cerevisiae.

Cadmium (Cd) is a toxic heavy metal that induces irregularity in numerous lipid metabolic pathways. Saccharomyces cerevisiae, a model to study lipid metabolism, has been used to establish the molecular basis of cellular responses to Cd toxicity in relation to essential minerals and lipid homeostasis. Multiple pathways sense these environmental stresses and trigger the mineral imbalances specifically calcium (Ca) and zinc (Zn). This review is aimed to elucidate the role of Cd toxicity in yeast, in three different perspectives: (1) elucidate stress response and its adaptation to Cd, (2) understand the physiological role of a macromolecule such as lipids, and (3) study the stress rescue mechanism. Here, we explored the impact of Cd interference on the essential minerals such as Zn and Ca and their influence on endoplasmic reticulum stress and lipid metabolism. Cd toxicity contributes to lipid droplet synthesis by activating OLE1 that is essential to alleviate lipotoxicity. In this review, we expanded our current findings about the effect of Cd on lipid metabolism of budding yeast.

Intramuscular Immunization of Streptococcus pyogenes SF370 protein extract and identification of multiple virulence factors through proteomic profiling in RHD induced Balb/c mice.

Group A streptococcus (GAS) and autoimmunity are associated with heart related mitral valve damage, in adults. In this study Balb/c mice were intramuscularly immunized with S. pyogenes SF370 for 4 weeks. Prior to euthanization, physiological parameters like body weight and electrical signalling of the heart were recorded. After euthanization, the heart tissue homogenate was prepared and proteomic alterations were studied using SDS-PAGE and 2D electrophoresis. The expression levels of inflammatory genes like TNFα, IFNγ and TGF-ß were quantified using real time PCR. Insilico analysis was performed to identify the functions of hypothetical proteins and virulence factors involved in the induction of rheumatic carditis. The results showed a reduction in body weight, ulceration, inflammation, cardiac lesions and prolonged PR interval in mice immunized with S. pyogenes SF370, as a result of RHD. The heart related proteins like α-actinin, fatty acid binding protein-heart, myosin light chain 3, hemoglobin subunit alpha, myoglobin regulatory light chain 2, (ventricular/cardiac muscle isoform), myosin-6, troponin-1 were found to be up-regulated when compared with the control. The functional annotation of S. pyogenes (SF370) was carried out by retrieving 1696 identified proteins and 653 hypothetical protein sequences in NCBI genome database. The conserved domain was identified for 505 proteins. The pfam database documented that the super families of 279 sequences and 40 signal peptides enabled the classification of proteins in different categories like biological (20%), cellular (22%) and molecular functions (36%). Putative transcription repair coupling factor and putative lysine aminopeptidase N terminal are the two virulence factors identified by VICMPRED in S. pyogenes SF370. The two identified virulence factors are involved in altering the mice heart proteome and thereby controlling the streptococcus pyogenes infection. Thus, the results of the present study reveals the role of immunogenic proteins in induction of rheumatic carditis and to elucidate the molecular mechanisms leading to autoimmune reactions in Balb/c mice.

The -866G/A polymorphism in the promoter of the UCP2 gene is associated with risk for type 2 diabetes and with decreased insulin levels.

BACKGROUND AND AIM: Oxidative stress and impaired insulin secretion is an underlying major risk factor for the development of type 2 diabetes (T2D). Uncoupling protein-2 (UCP2) is involved in the regulation of reactive oxygen species production, insulin secretion, and lipid metabolism. Based on this we aimed to find an association of UCP2 (G-866A) polymorphism with the risk of T2D in South Indian population. METHODS: A total of 318 T2D patients and 312 controls were enrolled in this study. All the study subjects were genotyped for UCP2 (G-866A) polymorphism using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Fasting blood glucose, HbA1c, serum lipid profile, systolic and diastolic blood pressure were measured by standard biochemical methods. Fasting serum insulin level was measured by ELISA. RESULTS: In UCP2 (G-866A) polymorphism, the distribution of GA (46%) and AA (14%) genotypes were significantly higher in T2D patients than the healthy controls. The frequency of GA and AA genotypes have high risk towards the development of T2D with an Odds Ratio (OR) of 1.55 (P = 0.01) and 2.04 (P = 0.01) respectively. Moreover, SNP-866 G>A allele was found to be significantly associated with T2D (OR = 1.48, P = 0.001, 95% CI = 1.16-1.88). Further, the UCP2 AA genotype showed significantly decreased level of insulin by the reduction in pancreatic ß-cell function in T2D patients. CONCLUSION: UCP2 (G-866A) polymorphism may play a crucial role in the pathogenesis of insulin secretion thus leads to the development of T2D.

Identification of promoter PcadR, in silico characterization of cadmium resistant gene cadR and molecular cloning of promoter PcadR from Pseudomonas aeruginosa BC15.

Cadmium (Cd) remediation in Pseudomonas aeruginosa is achieved through the function of two vital genes, cadA and cadR, that code for P-type ATPase (CadA) and transcription regulatory protein (CadR), respectively. Although numerous studies are available on these metal-sensing and regulatory proteins, the promoter of these genes, metal sensing and binding ability, are poorly understood. The present work is aimed at the characterization of the CadR protein, identification of the PcadR promoter and protein-promoter-metal binding affinity using bioinformatics and to validate the results by cloning the PcadR promoter in Escherichia coli DH5α. The promoter regions and its curvature were identified and analysed using PePPER software (University of Groningen, The Netherland) and the Bendit program (Version: v.1.0), respectively. Using Phyre, the three-dimensional structure of CadR was modelled, and the structure was validated by Ramachandran plots. The DNA-binding domain was present in the N-terminal region of CadR. A dimeric interface was observed in helix-turn-helix and metal ion-binding sites at the C-terminal. Docking studies showed higher affinity of Cd to both CadR (Atomic contact energy = -15.04 kcal/Mol) and PcadR (Atomic contact energy = -40.18 kcal/Mol) when compared to other metal ions. CadR with PcadR showed the highest binding affinity (Atomic contact energy= -250.40 kcal/Mol) when compared with PcadA. In vitro studies using green fluorescent protein tagged with PcadR (gfp-PcadR) cloned in E. coli-expressed gfp protein in a concentration-dependent manner upon Cd exposure. Based on our in silico studies and in vitro molecular cloning analysis, we conclude that PcadR and CadR are active only in the presence of Cd. The CadR protein has the highest binding affinity with PcadR. As it became apparent that the cadR gene regulates the PcadR activity in the presence of Cd with high specificity, and the cadR and PcadR can be used as a biological tool for development of a microbial biosensor.

Chronic intake of 4-Methylimidazole induces Hyperinsulinemia and Hypoglycaemia via Pancreatic Beta Cell Hyperplasia and Glucose Dyshomeostasis.

Caramel colours are the preferential food colouring agent globally, reaches wide age groups through eatables. Colas, a sweetened carbonated drink are most common caramel coloured beverage and its consumption is linked with diabetes, obesity, pancreatic cancer and other endocrine disorders. A major by-product produced during caramelization is 4-methylimidazole (4-MEI) that is detected in noteworthy concentrations in colas and other beverages. Previous studies revealed the neurotoxic and carcinogenic potential of 4-MEI in animals at higher doses but the effect of 4-MEI at theoretical maximum daily intake dose on glucose homeostasis is unexplored. Here, mice treated with 4-MEI (32 µg/kg bodyweight/day) for seven weeks exhibited severe hypoglycaemia and hyperinsulinemia mediated by hyperplasia of pancreatic beta cells and induces metabolic alterations. On combinatorial treatment, 4-MEI suppressed the glucogenic potential of non-artificial sweeteners and promotes lipogenesis. Furthermore, increased levels of C-peptide, LDL-cholesterol and triglycerides were observed in the humans with regular intake of 4-MEI containing beverages. In summary, 4-MEI induced pancreatic beta cell hyperplasia and leads to disruption of glucose and lipid homeostasis. This study suggests the need for further assessment and reconsideration of the wide usage of 4-MEI containing caramels as food additives.

Association of paraoxonase-1 gene polymorphisms with insulin resistance in South Indian population.

BACKGROUND AND AIM: Insulin resistance plays a crucial role in the pathogenesis of type 2 diabetes and cardiovascular diseases. Recently, paraoxonase-1(PON1) is reported to have an ability to reduce insulin resistance by promoting glucose transporter-4 (GLUT-4) expression in vitro. Single nucleotide polymorphism (SNP) in PON1 is associated with variability in enzyme activity and concentration. Based on this we aimed to investigate the association of PON1 (Q192R and L55M) polymorphisms with the risk of developing insulin resistance in adult South Indian population. METHODS: Two hundred and eighty seven (287) Type 2 diabetes patients and 293 healthy controls were enrolled in this study. All the study subjects were genotyped for PON1 (Q192R and L55M) missense polymorphisms using polymerase chain reaction-restriction fragment length polymorphism (PCRRFLP) method. Fasting serum insulin level was measured by ELISA. RESULTS: The distribution of QR/RR and LM/MM genotypes were significantly higher in type 2 diabetes patients compared with healthy controls. Moreover, the R and M alleles were significantly associated with type 2 diabetes with an Odds Ratio of 1.68 (P < 0.005) and 2.24 (P < 0.005) respectively. SNP 192 Q > R genotypes were found to be significantly associated with higher BMI, cholesterol, triglycerides, LDL, fasting serum insulin and HOMA-IR. Further, the mutant allele or genotypes of PON1 L55M were associated with higher BMI, triglycerides, VLDL, fasting serum insulin and HOMA-IR among adult type 2 diabetes patients. CONCLUSION: PON1 (Q192R and L55M) polymorphisms may play a crucial role in pathogenesis and susceptibility of insulin resistance thus leads to the development of type 2 diabetes in South Indian population.

Parthenolide attenuates 7,12-dimethylbenz[a]anthracene induced hamster buccal pouch carcinogenesis.

Over the decades, the survival rates for oral cancer have not improved despite development in novel diagnostic and therapeutic strategies. Therefore, the present study is aimed at investigating the chemopreventive potential of parthenolide in DMBA-induced hamster buccal pouch carcinogenesis. The hamsters were divided into 4 groups (n = 6/group). Group I was treated as control. Groups II and III were painted with a solution of 0.5% DMBA three times per week for 14 weeks on the left buccal pouches. In addition, group III were orally administrated with parthenolide 2 mg/kg b.w on days alternate to the DMBA application. Group IV received only parthenolide. At the end of 14th week all hamsters were sacrificed. Buccal tissues from all hamsters were evaluated for histopathology. Biochemical studies were carried out using plasma, liver, and buccal mucosa of control and experimental hamsters. Gene and protein expression studies of apoptotic markers p53, Bcl-2, and Bax were performed. The results showed 100% tumor formation and marked alterations in histopathology, status of detoxification enzymes, lipid peroxidation, and antioxidant profile in group II hamsters. Oral administration of parthenolide completely prevented tumor formation and significantly reduced the severity of histopathological changes in group III hamsters. The status of detoxification enzymes, lipid peroxidation, and antioxidants were significantly restored in parthenolide treated group compared to group II hamsters. The apoptotic gene p53 and antiapoptotic gene Bcl-2 were significantly down regulated; whereas, pro-apoptotic gene Bax was up regulated in group III hamsters compared to group II. The results of the present study suggest that parthenolide have potent chemopreventive, antioxidant, and apoptotic effect in DMBA-induced oral carcinogenesis.

Molecular analysis of oxalate-induced endoplasmic reticulum stress mediated apoptosis in the pathogenesis of kidney stone disease.

Oxalate, a non-essential end product of metabolism, causes hyperoxaluria and eventually calcium oxalate (CaOx) stone disease. Kidney cells exposed to oxalate stress results in generation of reactive oxygen species (ROS) and progression of stone formation. Perturbations in endoplasmic reticulum (ER) result in accumulation of misfolded proteins and Ca2+ ions homeostasis imbalance and serve as a common pathway for various diseases, including kidney disorders. ER stress induces up-regulation of pro-survival protein glucose-regulated protein 78 (GRP78) and pro-apoptotic signaling protein C/EBP homologous protein (CHOP). Since the association of oxalate toxicity and ER stress on renal cell damage is uncertain, the present study is an attempt to elucidate the interaction of GRP78 with oxalate by computational analysis and study the role of ER stress in oxalate-mediated apoptosis in vitro and in vivo. Molecular docking results showed that GRP78-oxalate/CaOx interaction takes place. Oxalate stress significantly up-regulated expression of ER stress markers GRP78 and CHOP both in vitro and in vivo. Exposure of oxalate increased ROS generation and altered antioxidant enzyme activities. N-Acetyl cysteine treatment significantly ameliorated oxalate-mediated oxidative stress and moderately attenuated ER stress marker expression. The result indicates oxalate toxicity initiated oxidative stress-induced ER stress and also activating ER stress mediated apoptosis directly. In addition, the up-regulation of transforming growth factor ß-1 revealed oxalate may induce kidney fibrosis through ER stress-mediated mechanisms. The present study provide insights into the pathogenic role of oxidative and ER stress by oxalate exposure in the formation of calcium oxalate stone.