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.

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.

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.

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.

Gut microbial degradation of organophosphate insecticides-induces glucose intolerance via gluconeogenesis.

BACKGROUND: Organophosphates are the most frequently and largely applied insecticide in the world due to their biodegradable nature. Gut microbes were shown to degrade organophosphates and cause intestinal dysfunction. The diabetogenic nature of organophosphates was recently reported but the underlying molecular mechanism is unclear. We aimed to understand the role of gut microbiota in organophosphate-induced hyperglycemia and to unravel the molecular mechanism behind this process. RESULTS: Here we demonstrate a high prevalence of diabetes among people directly exposed to organophosphates in rural India (n = 3080). Correlation and linear regression analysis reveal a strong association between plasma organophosphate residues and HbA1c but no association with acetylcholine esterase was noticed. Chronic treatment of mice with organophosphate for 180 days confirms the induction of glucose intolerance with no significant change in acetylcholine esterase. Further fecal transplantation and culture transplantation experiments confirm the involvement of gut microbiota in organophosphate-induced glucose intolerance. Intestinal metatranscriptomic and host metabolomic analyses reveal that gut microbial organophosphate degradation produces short chain fatty acids like acetic acid, which induces gluconeogenesis and thereby accounts for glucose intolerance. Plasma organophosphate residues are positively correlated with fecal esterase activity and acetate level of human diabetes. CONCLUSION: Collectively, our results implicate gluconeogenesis as the key mechanism behind organophosphate-induced hyperglycemia, mediated by the organophosphate-degrading potential of gut microbiota. This study reveals the gut microbiome-mediated diabetogenic nature of organophosphates and hence that the usage of these insecticides should be reconsidered.

Regression of oxidative stress by targeting eNOS and Nrf2/ARE signaling: a guided drug target for cardiovascular diseases.

Cardiovascular diseases (CVDs) are the major health concern and the leading cause of death. Imbalance between free radicals and anti-oxidant defence is associated with cellular dysfunctions leading to the pathophysiology of various diseases including cardiac and vascular diseases. The stress responsive transcription factor NF-E2-related factor 2/antioxidant response element (Nrf2/ARE) regulates the expression of many detoxifying genes. Nitric oxide (NO) produced by endothelial nitric oxide synthase (eNOS) is an important regulator of vascular function. Involvement of NO in modulating Nrf2 signaling is well established. Thus, it is apparent that increasing NO bioavailability and antioxidant status in vascular and myocardial tissue can be considered as a potential strategy to prevent the onset of vascular dysfunction and CVDs and is therefore of therapeutical interest. Based on the marked protective effect of Nrf2/ARE signalling and intriguing links between antioxidant mechanism and endothelial derived NO, the aim of the present review is to compile conclusive evidence for the involvement of NO-Nrf2/ARE axis in the regulation of cardiovascular function. This review also discusses on improving eNOS and Nrf2 signalling by Nrf2 activators which holds promise for countering cardiac and vascular disorders.

Recombinant Lactobacillus plantarum expressing and secreting heterologous oxalate decarboxylase prevents renal calcium oxalate stone deposition in experimental rats.

BACKGROUND: Calcium oxalate (CaOx) is the major constituent of about 75% of all urinary stone and the secondary hyperoxaluria is a primary risk factor. Current treatment options for the patients with hyperoxaluria and CaOx stone diseases are limited. Oxalate degrading bacteria might have beneficial effects on urinary oxalate excretion resulting from decreased intestinal oxalate concentration and absorption. Thus, the aim of the present study is to examine the in vivo oxalate degrading ability of genetically engineered Lactobacillus plantarum (L. plantarum) that constitutively expressing and secreting heterologous oxalate decarboxylase (OxdC) for prevention of CaOx stone formation in rats. The recombinants strain of L. plantarum that constitutively secreting (WCFS1OxdC) and non-secreting (NC8OxdC) OxdC has been developed by using expression vector pSIP401. The in vivo oxalate degradation ability for this recombinants strain was carried out in a male wistar albino rats. The group I control; groups II, III, IV and V rats were fed with 5% potassium oxalate diet and 14th day onwards group II, III, IV and V were received esophageal gavage of L. plantarum WCFS1, WCFS1OxdC and NC8OxdC respectively for 2-week period. The urinary and serum biochemistry and histopathology of the kidney were carried out. The experimental data were analyzed using one-way ANOVA followed by Duncan's multiple-range test. RESULTS: Recombinants L. plantarum constitutively express and secretes the functional OxdC and could degrade the oxalate up to 70-77% under in vitro. The recombinant bacterial treated rats in groups IV and V showed significant reduction of urinary oxalate, calcium, uric acid, creatinine and serum uric acid, BUN/creatinine ratio compared to group II and III rats (P < 0.05). Oxalate levels in kidney homogenate of groups IV and V were showed significant reduction than group II and III rats (P < 0.05). Microscopic observations revealed a high score (4+) of CaOx crystal in kidneys of groups II and III, whereas no crystal in group IV and a lower score (1+) in group V. CONCLUSION: The present results indicate that artificial colonization of recombinant strain, WCFS1OxdC and NC8OxdC, capable of reduce urinary oxalate excretion and CaOx crystal deposition by increased intestinal oxalate degradation.

Naringenin confers protection against oxidative stress through upregulation of Nrf2 target genes in cardiomyoblast cells

Cardiovascular diseases are the major health concern and the leading cause of death. Numerous studies have shown that oxidative stress stimuli have been incriminated in the pathogenesis of both acute and chronic heart disease. Though it is well known that bioflavonoids protect cells against reactive oxygen species (ROS)-induced damage, the molecular mechanisms involved are uncertain. Understanding the possible intracellular signaling pathways triggered by flavonoids will help to overcome the cardiac diseases resulting from oxidative stress. In the present study, we investigated whether naringenin (NGN) supplementation would improve the antioxidant defence under oxidative stress through the activation of Nrf2 signaling in cultured cardiomyoblast. NGN pretreatment significantly reduced stress-mediated apoptotic cell death and lipid peroxidation and showed increased level of reduced glutathione in H2O2-treated cardiomyoblast. In addition, NGN inhibited the production of NO and trigged the synthesis of antioxidant marker enzymes. Gene expression studies revealed that NGN upregulated the transcription of Akt and downregulated NF-êB andCaspase 3 genes. Notably, transcription of Nrf2 and its target genes was also upregulated. Taken together, the present study revealed that NGN elicits potent cytoprotective effect against oxidative stress by regulating Nrf2 and its target genes. In conclusion, the present work suggests that improving Nrf2 signaling by NGN supplementation would be a rational approach to facilitate ROS detoxification by augmenting both expression and activity of phase II detoxification enzymes for the alleviation of cardiac complications

Naringenin confers protection against oxidative stress through upregulation of Nrf2 target genes in cardiomyoblast cells.

Cardiovascular diseases are the major health concern and the leading cause of death. Numerous studies have shown that oxidative stress stimuli have been incriminated in the pathogenesis of both acute and chronic heart disease. Though it is well known that bioflavonoids protect cells against reactive oxygen species (ROS)-induced damage, the molecular mechanisms involved are uncertain. Understanding the possible intracellular signaling pathways triggered by flavonoids will help to overcome the cardiac diseases resulting from oxidative stress. In the present study, we investigated whether naringenin (NGN) supplementation would improve the antioxidant defence under oxidative stress through the activation of Nrf2 signaling in cultured cardiomyoblast. NGN pretreatment significantly reduced stress-mediated apoptotic cell death and lipid peroxidation and showed increased level of reduced glutathione in H2O2-treated cardiomyoblast. In addition, NGN inhibited the production of NO and trigged the synthesis of antioxidant marker enzymes. Gene expression studies revealed that NGN upregulated the transcription of Akt and downregulated NF-κB and Caspase 3 genes. Notably, transcription of Nrf2 and its target genes was also upregulated. Taken together, the present study revealed that NGN elicits potent cytoprotective effect against oxidative stress by regulating Nrf2 and its target genes. In conclusion, the present work suggests that improving Nrf2 signaling by NGN supplementation would be a rational approach to facilitate ROS detoxification by augmenting both expression and activity of phase II detoxification enzymes for the alleviation of cardiac complications.

L-Arginine ameliorates cardiac left ventricular oxidative stress by upregulating eNOS and Nrf2 target genes in alloxan-induced hyperglycemic rats.

Hyperglycemia is independently related with excessive morbidity and mortality in cardiovascular disorders. L-Arginine-nitric oxide (NO) pathway and the involvement of NO in modulating nuclear factor-E2-related factor-2 (Nrf2) signaling were well established. In the present study we investigated, whether L-arginine supplementation would improve the myocardial antioxidant defense under hyperglycemia through activation of Nrf2 signaling. Diabetes was induced by alloxan monohydrate (90 mg kg(-1) body weight) in rats. Both non-diabetic and diabetic group of rats were divided into three subgroups and they were administered either with L-arginine (2.25%) or L-NAME (0.01%) in drinking water for 12 days. Results showed that L-arginine treatment reduced the metabolic disturbances in diabetic rats. Antioxidant enzymes and glutathione levels were found to be increased in heart left ventricles, thereby reduction of lipid peroxidation by L-arginine treatment. Heart histopathological analysis further validates the reversal of typical diabetic characteristics consisting of alterations in myofibers and myofibrillary degeneration. qRT-PCR studies revealed that L-arginine treatment upregulated the transcription of Akt and downregulated NF-κB. Notably, transcription of eNOS and Nrf2 target genes was also upregulated, which were accompanied by enhanced expression of Nrf2 in left ventricular tissue from diabetic and control rats. Under these findings, we suggest that targeting of eNOS and Nrf2 signaling by L-arginine supplementation could be used as a potential treatment method to alleviate the late diabetic complications.