The effects of changes in glutathione levels through exogenous agents on intracellular cysteine content and protein adduct formation in chronic alcohol-treated VL17A cells.

Alcohol-mediated liver injury is associated with changes in the level of the major cellular antioxidant glutathione (GSH). It is interesting to investigate if the changes in intracellular GSH level through exogenous agents affect the intracellular cysteine content and the protein adduct formation indicative of oxidative insult in chronic alcohol treated liver cells. In VL-17A cells treated with 2 mM N-acetyl cysteine (NAC) or 0.1 mM ursodeoxycholic acid (UDCA) plus 100 mM ethanol, an increase in cysteine concentration which was accompanied by decreases in hydroxynonenal (HNE) and glutathionylated protein adducts were observed. Pretreatment of 100 mM ethanol treated VL-17A cells with 0.4 mM buthionine sulfoximine (BSO) or 1 mM diethyl maleate (DEM) had opposite effects. Thus, altered GSH level through exogenous agents may either potentiate or ameliorate chronic alcohol-mediated protein adduct formation and change the cysteine level in chronic alcohol treated VL-17A cells. The gene expression of non-treated and ethanol-treated hepatocytes in 2 microarray datasets was also compared to locate differentially expressed genes involved in cysteine metabolism. The study demonstrates that increased protein adducts formation and changes in cysteine concentration occur under chronic alcohol condition in liver cells which may increase alcohol-mediated oxidative injury.

GSH protects against oxidative stress and toxicity in VL-17A cells exposed to high glucose.

PURPOSE: The deficiency of glutathione (GSH) has been linked to several diseases. The study investigated the role of GSH as a protective factor against hyperglycemia-mediated injury in VL-17A cells treated with 50 mM glucose. METHODS: The cell viability and different oxidative stress parameters including glyoxalase I activity were measured. RESULTS: GSH supplementation with 2 mM N-acetyl cysteine (NAC) or 0.1 mM ursodeoxycholic acid (UDCA) increased the viability, GSH level and the GSH-dependent glyoxalase I activity in 50 mM glucose-treated VL-17A cells. Further, pretreatment of 50 mM glucose-treated VL-17A cells with NAC or UDCA decreased oxidative stress (levels of reactive oxygen species and protein carbonylation), apoptosis (caspase 3 activity and annexin V-propidium iodide positive cells) and glutathionylated protein formation, a measure of oxidative stress. GSH depletion with 0.4 mM buthionine sulfoximine (BSO) or 1 mM diethyl maleate (DEM) potentiated the decrease in viability, glyoxalase I activity and increase in oxidative stress and apoptosis, with decreased GSH levels in 50 mM glucose-treated VL-17A cells. CONCLUSION: Thus, changes in GSH levels with exogenous agents such as NAC, UDCA, BSO or DEM modulate hyperglycemia-mediated injury in a cell model of VL-17A liver cells.

Inhibition of CYP2E1 leads to decreased advanced glycated end product formation in high glucose treated ADH and CYP2E1 over-expressing VL-17A cells.

BACKGROUND: In recent years, there has been a growing interest to explore the association between liver injury and diabetes. Advanced glycated end product (AGE) formation which characterizes diabetic complications is formed through hyperglycemia mediated oxidative stress and is itself a source for ROS. Further, in VL-17A cells over-expressing ADH and CYP2E1, greatly increased oxidative stress and decreased viability have been observed with high glucose exposure. METHODS: In VL-17A cells treated with high glucose and pretreated with the different inhibitors of ADH and CYP2E1, the changes in cell viability, oxidative stress parameters and formation of AGE, were studied. RESULTS: Inhibition of CYP2E1 with 10µM diallyl sulfide most effectively led to decreases in the oxidative stress and toxicity as compared with ADH inhibition with 2mM pyrazole or the combined inhibition of ADH and CYP2E1 with 5mM 4-methyl pyrazole. AGE formation was decreased in VL-17A cells when compared with HepG2 cells devoid of the enzymes. Further, AGE formation was decreased to the greatest extent with the inhibitor for CYP2E1 suggesting that high glucose inducible CYP2E1 and the consequent ROS aid AGE formation. CONCLUSIONS: Thus, CYP2E1 plays a pivotal role in the high glucose induced oxidative stress and toxicity in liver cells as observed through direct evidences obtained utilizing the different inhibitors for ADH and CYP2E1. GENERAL SIGNIFICANCE: The study demonstrates the role of CYP2E1 mediated oxidative stress in aggravating hyperglycemic insult and suggests that CYP2E1 may be a vital component of hyperglycemia mediated oxidative injury in liver.

Cost-effectiveness of response-adapted therapy (RAT) for advanced Hodgkin's Lymphoma compared with conventional treatment in India: a Markov-model based analysis.

Cost effectiveness analysis of interim positron emission tomography (PET-2, done after 2 cycles of chemotherapy) based response adaptive therapy (RAT) approaches in advanced Hodgkin lymphoma (aHL) are not available from an Indian perspective. We used a five-year decision analytics model to assess the cost-effectiveness of the two RAT approaches [(escalation (RAT-1) or de-escalation (RAT-2)] compared with standard care (SOC) in aHL (mean age:35 years). Modelling data was derived from secondary sources and sensitivity analyses were performed to assess the robustness of the model. Net monetary benefit (NMB) gained from RAT2 in Indian rupees (INR) (INR 2,26,896) was higher than the RAT1 (INR 1,83,138) when compared with SOC. Proportion achieving the complete response after initial treatment (CR1) was the key determining factor for the RAT1/2 dominance over SOC. Despite higher initial input costs, response-adapted therapy of aHL was cost-effective by minimizing the cost incurred and disutility experienced during relapse and salvage.

Despite higher initial costs, response-adapted therapy based on the interim PET scan after 2 cycles of chemotherapy was more cost-effective when compared to standard therapy with 6 cycles of ABVD in patients with advanced Hodgkin's lymphoma. Among the RAT approaches, de-escalation (RAT-2) had better cost-effectiveness than the escalation approach (RAT-1).

Control status of hypertension in India: systematic review and meta-analysis.

BACKGROUND AND AIMS: Uncontrolled hypertension is a major risk factor for cardiovascular diseases (CVDs). The present study aimed to conduct a systematic review and meta-analysis to estimate the pooled prevalence of control status of hypertension in India. METHODS AND RESULTS: We carried out systematic search (PROSPERO No.: CRD42021239800) in PubMed and Embase published between April 2013 and March 2021 followed by meta-analysis with random-effects model. The pooled prevalence of controlled hypertension was estimated across geographic regions. The quality, publication bias and heterogeneity of the included studies were also assessed. We included 19 studies with 44 994 hypertensive population, among which 17 studies had low risk of bias. We found statistically significant heterogeneity ( P  ≤ 0.05) and absence of publication bias among the included studies. The pooled prevalence of control status among patients with hypertension was 15% (95% CI: 12-19%) and among those under treatment was 46% (95% CI: 40-52%). The control status among patients with hypertension was significantly higher in Southern India 23% (95% CI: 16-31%) followed by Western 13% (95% CI: 4-16%), Northern 12% (95% CI: 8-16%), and Eastern India 5% (95% CI: 4-5%). Except for Southern India, the control status was lower among the rural areas compared with urban areas. CONCLUSION: We report high prevalence of uncontrolled hypertension in India irrespective of treatment status, geographic regions and urban and rural settings. There is urgent need to improve control status of hypertension in the country.

Cost-Utility Analysis of Dabigatran and Warfarin for Stroke Prevention Among Patients With Nonvalvular Atrial Fibrillation in India.

OBJECTIVES: Dabigatran has a better safety profile and requires less monitoring, but is costlier than warfarin. This study evaluated the cost-utility of dabigatran relative to warfarin for preventing stroke in nonvalvular atrial fibrillation (NVAF) in India. METHODS: A Markov decision analysis model was developed to compare dabigatran (110 or 150 mg twice a day) to warfarin titrated to target prothrombin time in patients with NVAF at high risk of stroke. Model utilities and transition probabilities were based on literature and costs on market prices. Data on out-of-pocket expenses and income lost were taken from a nationally representative survey. We adopted a societal perspective and discounted both costs and outcomes at 3%. Ischemic stroke, intracranial bleed, other major bleeds, and death were outcomes of NVAF. The model projected the costs, life-years, and quality-adjusted life-years (QALYs) for each intervention over a lifetime. We used gross domestic product per capita of India (US dollars [US$]1889) as the cost-effectiveness threshold. Sensitivity analyses were conducted. RESULTS: Treatment with either dose of dabigatran was associated with gain in life-years and QALYs compared with warfarin. The discounted incremental cost-effectiveness ratios/QALYs for both doses of dabigatran (110 mg US$7519; 150 mg US$6634) were above the cost-effectiveness threshold, and the probability of being cost-effective at this threshold was low. Cost of dabigatran was an important factor in determining incremental cost-effectiveness ratio. Price reduction of 150 mg dose by 49% will make it cost-effective. CONCLUSIONS: Dabigatran is not cost-effective in the Indian societal context. Reducing the price of dabigatran 150 mg by half will make it cost-effective.

Cytochrome P450 2E1 and hyperglycemia-induced liver injury.

Cytochrome P450 2E1 (CYP2E1), a microsomal enzyme involved in xenobiotic metabolism and generation of oxidative stress, has been implicated in promoting liver injury. The review deals with the changes in various cellular pathways in liver linked with the changes in regulation of CYP2E1 under hyperglycemic conditions. Some of the hepatic abnormalities associated with hyperglycemia-mediated induction of CYP2E1 include increased oxidative stress, changes in mitochondrial structure and function, apoptosis, nitrosative stress, and increased ketone body accumulation. Thus, changes in regulation of CYP2E1 are associated with the injurious effects of hyperglycemia in liver.

Modulation of GSH with exogenous agents leads to changes in glyoxalase 1 enzyme activity in VL-17A cells exposed to chronic alcohol plus high glucose.

Gluthathione (GSH) is a major cellular antioxidant. The present study utilizing VL-17A cells exposed to chronic alcohol plus high glucose investigated the changes in oxidative stress, toxicity, and glyoxalase 1 activity as a detoxification pathway due to changes in GSH level through GSH supplementation with N-acetyl cysteine (NAC) or ursodeoxycholic acid (UDCA) and its depletion through buthionine sulfoximine (BSO) or diethyl maleate (DEM). Glyoxalase 1 plays an important role in detoxification of methylglyoxal which is formed as a precursor of advanced glycated end products formed due to high glucose mediated oxidative stress. Significant changes in glyoxalase 1 activity utilizing methylglyoxal or glyoxal as substrates occurred with NAC or UDCA or BSO or DEM supplementation in chronic alcohol plus high glucose treated VL-17A cells. NAC or UDCA administration in chronic alcohol plus high glucose treated VL-17A cells increased viability and decreased ROS levels, lipid peroxidation and 3-nitrotyrosine adduct formation. Similarly, GSH depletion with BSO or DEM had an opposite effect on the parameters in chronic alcohol plus high glucose treated VL-17A cells. In conclusion, modulation of GSH with NAC or UDCA or BSO or DEM leads to significant changes in oxidative stress, glyoxalase 1 enzyme activity and toxicity in chronic alcohol plus high glucose treated VL-17A cells.

Increased oxidative stress and toxicity in ADH and CYP2E1 overexpressing human hepatoma VL-17A cells exposed to high glucose.

High glucose mediated oxidative stress and cell death is a well documented phenomenon. Using VL-17A cells which are HepG2 cells over-expressing alcohol dehydrogenase (ADH) and cytochrome P450 2E1 (CYP2E1) and control HepG2 cells, the association of ADH and CYP2E1 with high glucose mediated oxidative stress and toxicity in liver cells was investigated. Cell viability was measured and apoptosis or necrosis was determined through caspase-3 activity, Annexin V-propidium iodide staining and detecting decreases in mitochondrial membrane potential. Reactive oxygen species, lipid peroxidation and the formation of advanced glycated-end products were assessed. The levels of several antioxidants which included glutathione, glutathione peroxidase, catalase and superoxide dismutase were altered in high glucose treated VL-17A cells. Greater toxicity was observed in VL-17A cells exposed to high glucose when compared to HepG2 cells. Oxidative stress parameters were greatly increased in high glucose exposed VL-17A cells and apoptotic cell death was observed. Inhibition of CYP2E1 or caspase 3 or addition of the antioxidant trolox led to significant decreases in high glucose mediated oxidative stress and toxicity. Thus, the over-expression of ADH and CYP2E1 in liver cells is associated with increased high glucose mediated oxidative stress and toxicity.

Bio-informatics based analysis of genes implicated in alcohol mediated liver injury.

Alcohol induced liver injury has been studied extensively. Using literature search and bioinformatics tools, the present study characterizes the genes involved in alcohol induced liver injury. The cellular and metabolic processes in which genes involved in alcohol induced liver injury are implicated are also discussed. The genes related to alcohol induced liver injury are also involved in affecting certain molecular functions and metabolism of drugs, besides being associated with diseases. In conclusion, the changes in regulation of genes implicated in alcohol induced liver injury apart from causing alcohol mediated hepatic dysfunction may affect other vital processes in the body.

Elevated glutathione level does not protect against chronic alcohol mediated apoptosis in recombinant human hepatoma cell line VL-17A over-expressing alcohol metabolizing enzymes--alcohol dehydrogenase and Cytochrome P450 2E1.

Chronic consumption of alcohol leads to liver injury. Ethanol-inducible Cytochrome P450 2E1 (CYP2E1) plays a critical role in alcohol mediated oxidative stress due to its ability to metabolize ethanol. In the present study, using the recombinant human hepatoma cell line VL-17A that over-expresses the alcohol metabolizing enzymes-alcohol dehydrogenase (ADH) and CYP2E1; and control HepG2 cells, the mechanism and mode of cell death due to chronic ethanol exposure were studied. Untreated VL-17A cells exhibited apoptosis and oxidative stress when compared with untreated HepG2 cells. Chronic alcohol exposure, i.e., 100 mM ethanol treatment for 72 h caused a significant decrease in viability (47%) in VL-17A cells but not in HepG2 cells. Chronic ethanol mediated cell death in VL-17A cells was predominantly apoptotic, with increased oxidative stress as the underlying mechanism. Chronic ethanol exposure of VL-17A cells resulted in 1.1- to 2.5-fold increased levels of ADH and CYP2E1. Interestingly, the level of the antioxidant GSH was found to be 3-fold upregulated in VL-17A cells treated with ethanol, which may be a metabolic adaptation to the persistent and overwhelming oxidative stress. In conclusion, the increased GSH level may not be sufficient enough to protect VL-17A cells from chronic alcohol mediated oxidative stress and resultant apoptosis.