Azadirachtin Attenuates Carcinogen Benzo(a) Pyrene-Induced DNA Damage, Cell Cycle Arrest, Apoptosis, Inflammatory, Metabolic, and Oxidative Stress in HepG2 Cells.

Azadirachtin (AZD), a limonoid from the versatile, tropical neem tree (Azadirachta indica), is well known for its many medicinal, and pharmacological effects. Its effects as an anti-oxidant, anti-inflammatory, and anti-cancer agent are well known. However, not many studies have explored the effects of AZD on toxicities induced by benzo(a)pyrene (B(a)P), a toxic component of cigarette smoke known to cause DNA damage and cell cycle arrest, leading to different kinds of cancer. In the present study, using HepG2 cells, we investigated the protective effects of Azadirachtin (AZD) against B(a)P-induced oxidative/nitrosative and metabolic stress and mitochondrial dysfunction. Treatment with 25 µM B(a)P for 24 h demonstrated an increased production of reactive oxygen species (ROS), followed by increased lipid peroxidation and DNA damage presumably, due to the increased metabolic activation of B(a)P by CYP 450 1A1/1A2 enzymes. We also observed intrinsic and extrinsic apoptosis, alterations in glutathione-dependent redox homeostasis, cell cycle arrest, and inflammation after B(a)P treatment. Cells treated with 25 µM AZD for 24 h showed decreased oxidative stress and apoptosis, partial protection from DNA damage, and an improvement in mitochondrial functions and bioenergetics. The improvement in antioxidant status, anti-inflammatory potential, and alterations in cell cycle regulatory markers qualify AZD as a potential therapeutic in combination with anti-cancer drugs.

Decoding numeracy and literacy in the human brain: insights from MEG and MVPA.

Numbers and letters are the fundamental building blocks of our everyday social interactions. Previous studies have focused on determining the cortical pathways shaped by numeracy and literacy in the human brain, partially supporting the hypothesis of distinct perceptual neural circuits involved in the visual processing of the two categories. In this study, we aim to investigate the temporal dynamics for number and letter processing. We present magnetoencephalography (MEG) data from two experiments (N = 25 each). In the first experiment, single numbers, letters, and their respective false fonts (false numbers and false letters) were presented, whereas, in the second experiment, numbers, letters, and their respective false fonts were presented as a string of characters. We used multivariate pattern analysis techniques (time-resolved decoding and temporal generalization), testing the strong hypothesis that the neural correlates supporting letter and number processing can be logistically classified as categorically separate. Our results show a very early dissociation (~ 100 ms) between numbers, and letters when compared to false fonts. Number processing can be dissociated with similar accuracy when presented as isolated items or strings of characters, while letter processing shows dissociable classification accuracy for single items compared to strings. These findings reinforce the evidence indicating that early visual processing can be differently shaped by the experience with numbers and letters; this dissociation is stronger for strings compared to single items, thus showing that combinatorial mechanisms for numbers and letters could be categorically distinguished and influence early visual processing.

Exercise alleviates diabetic complications by inhibiting oxidative stress-mediated signaling cascade and mitochondrial metabolic stress in GK diabetic rat tissues.

Type 2 diabetes, obesity (referred to as "diabesity"), and metabolic syndrome associated with increased insulin resistance and/or decreased insulin sensitivity have been implicated with increased oxidative stress and inflammation, mitochondrial dysfunction, and alterations in energy metabolism. The precise molecular mechanisms of these complications, however, remain to be clarified. Owing to the limitations and off-target side effects of antidiabetic drugs, exercise-induced control of hyperglycemia and increased insulin sensitivity is a preferred strategy to manage "diabesity" associated complications. In this study, we have investigated the effects of moderate exercise (1 h/day, 5 days a week for 60 days) on mitochondrial, metabolic, and oxidative stress-related changes in the liver and kidney of type 2 diabetic Goto-Kakizaki (GK) rats. Our previous study, using the same exercise regimen, demonstrated improved energy metabolism and mitochondrial function in the pancreas of GK diabetic rats. Our current study demonstrates exercise-induced inhibition of ROS production and NADPH oxidase enzyme activity, as well as lipid peroxidation and protein carbonylation in the liver and kidney of GK rats. Interestingly, glutathione (GSH) content and GSH-peroxidase and GSH reductase enzymes as well as superoxide dismutase (SOD) activities were profoundly altered in diabetic rat tissues. Exercise helped in restoring the altered GSH metabolism and antioxidant homeostasis. An increase in cytosolic glycolytic enzyme, hexokinase, and a decrease in mitochondrial Kreb's cycle enzyme was observed in GK diabetic rat tissues. Exercise helped restore the altered energy metabolism. A significant decrease in the activities of mitochondrial complexes and ATP content was also observed in the GK rats and exercise regulated the activities of the respiratory complexes and improved energy utilization. Activation of cytochrome P450s, CYP 2E1, and CYP 3A4 was observed in the tissues of GK rats, which recovered after exercise. Altered expression of redox-responsive proteins and translocation of transcription factor NFκB-p65, accompanied by activation of AMP-activated protein kinase (AMPK), SIRT-1, Glut-4, and PPAR-γ suggests the induction of antioxidant defense responses and increased energy metabolism in GK diabetic rats after exercise.

Alterations in Inflammatory Cytokines and Redox Homeostasis in LPS-Induced Pancreatic Beta-Cell Toxicity and Mitochondrial Stress: Protection by Azadirachtin.

Inflammation and redox imbalance are hallmarks of cancer, diabetes, and other degenerative disorders. Pathophysiological response to these disorders leads to oxidative stress and mitochondrial dysfunction by alterations and reprogramming in cellular signaling and metabolism. Pancreatic beta cells are very sensitive to the inflammatory and altered nutrient signals and hence play a crucial role in diabetes and cancer. In this study, we treated insulin-secreting pancreatic beta cells, Rin-5F, with the bacterial endotoxin, LPS (1 µg/ml) to induce an inflammatory response in vitro and then treated the cells with a known anti-inflammatory, anticancer and antioxidant phytochemical, azadirachtin (AZD, 25 µM for 24 h). Our results demonstrated lipid peroxidation and nitric oxide production causing increased nitro/oxidative stress and alterations in the activities of anti-oxidant enzymes, superoxide dismutase and catalase after LPS treatment. Pro-inflammatory responses caused by translocation of nuclear factor kappa B and release of inflammatory cytokines were also observed. These changes were accompanied by GSH-dependent redox imbalance and alterations in mitochondrial membrane potential and respiratory complexes enzyme activities leading to mitochondrial respiratory dysfunction, reduced ATP synthesis, and intrinsic caspase-9 mediated apoptosis. Caspase-9 was activated due to alterations in Bcl-2 and Bax proteins and release of cytochrome c into the cytosol. The activities of oxidative stress-sensitive mitochondrial matrix enzymes, aconitase, and glutamate dehydrogenase were also inhibited. Treatment with AZD showed beneficial effects on the recovery of antioxidant enzymes, inflammatory responses, and mitochondrial functions. GSH-dependent redox homeostasis also recovered after the treatment with AZD. This study may help in better understanding the etiology and pathogenesis of inflammation-induced disorders in pancreatic beta cells to better manage therapeutic strategies.

Diabetes Mellitus Alters the Immuno-Expression of Neuronal Nitric Oxide Synthase in the Rat Pancreas.

Nitric oxide is generated from nitric oxide synthase following hyperglycemia-induced oxidative stress during the course of diabetes mellitus (DM). We examined the temporal immuno-expression of neuronal nitric oxide synthase (nNOS) in the pancreas of diabetic and non-diabetic rats using immunohistochemical, immunofluorescence and western blot techniques 12 h, 24 h, 1 week, 2 weeks, 1, 8 and 15 months after induction of DM. nNOS co-localized with pancreatic beta cells but disappears 12 h after the onset of DM. In contrast, the nNOS content of pancreatic nerves increased significantly (p < 0.001) 24 h after the induction of DM, and decreased sharply thereafter. However, nNOS-positive ganglion cells were observed even 15 months post-diabetes. ROS increased by more than 100% two months after the onset of DM compared to non-diabetic control but was significantly (p < 0.000001) reduced at 9 months after the induction of DM. The pancreatic content of GSH increased significantly (p < 0.02) after 9 months of DM. Although, TBARS content was significantly (p < 0.009; p < 0.002) lower in aged (9 months) non-diabetic and DM rats, TBARS rate was markedly (p < 0.02) higher 9 months after the induction of DM when compared to younger age group. In conclusion, nNOS is present in pancreatic beta cell, but disappears 12 h after the onset of diabetes. In contrast, the tissue level of nNOS of pancreatic nerves increased in the first week of diabetes, followed by a sharp reduction. nNOS may play important roles in the metabolism of pancreatic beta cell.

Alterations in Energy Metabolism, Mitochondrial Function and Redox Homeostasis in GK Diabetic Rat Tissues Treated with Aspirin.

Our recent studies have demonstrated that aspirin treatment prevents inflammatory and oxidative stress-induced alterations in mitochondrial function, improves glucose tolerance and pancreatic endocrine function and preserves tissue-specific glutathione (GSH)-dependent redox homeostasis in Goto-Kakizaki (GK) diabetic rats. In the current study, we have investigated the mechanism of action of aspirin in maintaining mitochondrial bioenergetics and redox metabolism in the liver and kidneys of GK rats. Aspirin reduced the production of reactive oxygen species (ROS) and oxidative stress-induced changes in GSH metabolism. Aspirin treatment also improved mitochondrial respiratory function and energy metabolism, in addition to regulating the expression of cell signaling proteins that were altered in diabetic animals. Ultrastructural electron microscopy studies revealed decreased accumulation of glycogen in the liver of aspirin-treated diabetic rats. Hypertrophic podocytes with irregular fusion of foot processes in the renal glomerulus and detached microvilli, condensed nuclei and degenerated mitochondria observed in the proximal convoluted tubules of GK rats were partially restored by aspirin. These results provide additional evidence to support our previous observation of moderation of diabetic complications by aspirin treatment in GK rats and may have implications for cautious use of aspirin in the therapeutic management of diabetes.

Insights into British Columbian Hospital Pharmacists Perspectives on the Discharge Process.

Background: Transitions of care represent a vulnerable time for patients where unintended therapeutic changes are common and inadequate communication of information frequently results in medication errors. Pharmacists have a large impact on the success of patients during these care transitions; however, their role and experiences are largely absent from the literature. Objectives: The purpose of this study was to gain a greater understanding of British Columbian hospital pharmacists' perceptions of the hospital discharge process and their role in it. Methods: A qualitative study utilizing focus groups and key informant interviews of British Columbian hospital pharmacists was conducted from April to May 2021. Questions asked during interviews were developed following a detailed literature search and included questions regarding the use of frequently studied interventions. Interview sessions were transcribed and then thematically analyzed using both NVivo software and manual coding. Results: Three focus groups with a total of 20 participants and one key informant interview were conducted. Six themes were identified through data analysis: (1) overall perspectives; (2) important roles of pharmacists in discharges; (3) patient education; (4) barriers to optimal discharges; (5) solutions to current barriers; and (6) prioritization. Conclusions and Relevance: Pharmacists play a vital role in patient discharges but due to limited resources and inadequate staffing models, they are often unable to be optimally involved. Understanding the thoughts and perceptions of pharmacists on the discharge process can help us better allocate limited resources to ensure patients receive optimal care.

Azadirachtin Attenuates Lipopolysaccharide-Induced ROS Production, DNA Damage, and Apoptosis by Regulating JNK/Akt and AMPK/mTOR-Dependent Pathways in Rin-5F Pancreatic Beta Cells.

Pancreatic inflammation and the resulting cellular responses have been implicated in pancreatitis, diabetes, and pancreatic cancer. Inflammatory responses due to the bacterial endotoxin, lipopolysaccharide (LPS), have been demonstrated to alter cellular metabolism, autophagy, apoptosis, and cell proliferation in different cell populations, and hence increases the risks for organ toxicity including cancer. The exact molecular mechanism is however not clear. In the present study, we investigated the role and mechanism of an antioxidant, azadirachtin (AZD), a limonoid extracted from the neem tree (Azadirachta indica), against LPS-induced oxidative stress in the pancreatic ß-cell line, Rin-5F. We demonstrated that cells treated with LPS (1 µg/mL for 24 h) showed increased reactive oxygen species (ROS) production, DNA damage, cell cycle arrest, and apoptosis. Our results also showed that LPS induced alterations in the AMP-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathways, suppressing autophagy and augmenting apoptosis. Treatment with Azadirachtin (25 µM for 24 h), on the other hand, rendered some degree of protection to the pancreatic cells from apoptosis by inducing the autophagy signals required for cell survival. These results may have significance in elucidating the mechanisms of pancreatic ß-cell survival and death by balancing the molecular communication between autophagy and apoptosis under inflammatory and pathological conditions.

Effect of Aspirin on Mitochondrial Dysfunction and Stress in the Pancreas and Heart of Goto-Kakizaki Diabetic Rats.

Our previous study in Goto-Kakizaki (GK) type 2 diabetic rats provided significant evidence that aspirin treatment improves pancreatic ß-cell function by reducing inflammatory responses and improving glucose tolerance. In the present study, we aimed to elucidate the mechanism of action of aspirin on the pathophysiology and progression of type 2 diabetic complications in the heart and pancreas of insulin-resistant GK rats. Aspirin treatment demonstrated a reduction in mitochondrial reactive oxygen species (ROS) production and lipid peroxidation, accompanied by improved redox homeostasis. Furthermore, the recovery of metabolic and mitochondrial functions, as well as cytochrome P450 enzyme activities, which were altered in the pancreas and heart of GK rats, were observed. Aspirin treatment brought the activity of CYP 2E1 to the control level in both tissues, whereas the CYP 3A4 level decreased only in the pancreas. This suggests the tissue-specific differential metabolism of substrates in these rats. The recovery of redox homeostasis could be the key target in the improvement of oxidative-stress-dependent alterations in mitochondrial functions which, in turn, facilitated improved energy metabolism in these tissues in the aspirin-treated GK rats. These results may have implications in determining the therapeutic use of aspirin, either alone or in combination with other clinically approved therapies, in insulin-resistant type 2 diabetes.

A magnetoencephalography dataset for motor and cognitive imagery-based brain-computer interface.

Recent advancements in magnetoencephalography (MEG)-based brain-computer interfaces (BCIs) have shown great potential. However, the performance of current MEG-BCI systems is still inadequate and one of the main reasons for this is the unavailability of open-source MEG-BCI datasets. MEG systems are expensive and hence MEG datasets are not readily available for researchers to develop effective and efficient BCI-related signal processing algorithms. In this work, we release a 306-channel MEG-BCI data recorded at 1KHz sampling frequency during four mental imagery tasks (i.e. hand imagery, feet imagery, subtraction imagery, and word generation imagery). The dataset contains two sessions of MEG recordings performed on separate days from 17 healthy participants using a typical BCI imagery paradigm. The current dataset will be the only publicly available MEG imagery BCI dataset as per our knowledge. The dataset can be used by the scientific community towards the development of novel pattern recognition machine learning methods to detect brain activities related to motor imagery and cognitive imagery tasks using MEG signals.

Mitigation of Glucolipotoxicity-Induced Apoptosis, Mitochondrial Dysfunction, and Metabolic Stress by N-Acetyl Cysteine in Pancreatic ß-Cells.

Glucolipotoxicity caused by hyperglycemia and hyperlipidemia are the common features of diabetes-induced complications. Metabolic adaptation, particularly in energy metabolism; mitochondrial dysfunction; and increased inflammatory and oxidative stress responses are considered to be the main characteristics of diabetes and metabolic syndrome. However, due to various fluctuating endogenous and exogenous stimuli, the precise role of these factors under in vivo conditions is not clearly understood. In the present study, we used pancreatic ß-cells, Rin-5F, to elucidate the molecular and metabolic changes in glucolipotoxicity. Cells treated with high glucose (25 mM) and high palmitic acid (up to 0.3 mM) for 24 h exhibited increased caspase/poly-ADP ribose polymerase (PARP)-dependent apoptosis followed by DNA fragmentation, alterations in mitochondrial membrane permeability, and bioenergetics, accompanied by alterations in glycolytic and mitochondrial energy metabolism. Our results also demonstrated alterations in the expression of mammalian target of rapamycin (mTOR)/5' adenosine monophosphate-activated protein kinase (AMPK)-dependent apoptotic and autophagy markers. Furthermore, pre-treatment of cells with 10 mM N-acetyl cysteine attenuated the deleterious effects of high glucose and high palmitic acid with improved cellular functions and survival. These results suggest that the presence of high energy metabolites enhance mitochondrial dysfunction and apoptosis by suppressing autophagy and adapting energy metabolism, mediated, at least in part, via enhanced oxidative DNA damage and mTOR/AMPK-dependent cell signaling.

N-acetyl cysteine attenuates oxidative stress and glutathione-dependent redox imbalance caused by high glucose/high palmitic acid treatment in pancreatic Rin-5F cells.

Elevated levels of glucose and fatty acids are the main characteristics of diabetes, obesity and other metabolic disorders, associated with increased oxidative stress, mitochondrial dysfunction and inflammation. Once the primary pathogenesis of diabetes is established, which is potentially linked to both genetic and environmental factors, hyperglycemia and hyperlipidemia exert further destructive and/or toxic effects on ß-cells. The concept of glucolipotoxicity has arisen from the combination of deleterious effects of chronic elevation of glucose and fatty acid levels on pancreatic ß- cell function and/or survival. Though numerous studies have been conducted in this field, the exact molecular mechanisms and causative factors still need to be established. The aim of the present work was to elucidate the molecular mechanisms of oxidative stress, and inflammatory/antioxidant responses in the presence of high concentrations of glucose/fatty acids in a cell-culture system using an insulin-secreting pancreatic ß-cell line (Rin-5F) and to study the effects of the antioxidant, N-acetyl cysteine (NAC) on ß-cell toxicity. In our study, we investigated the molecular mechanism of cytotoxicity in the presence of high glucose (up to 25 mM) and high palmitic acid (up to 0.3 mM) on Rin-5F cells. Our results suggest that the cellular and molecular mechanisms underlying ß-cell toxicity are mediated by increased oxidative stress, imbalance of redox homeostasis, glutathione (GSH) metabolism and alterations in inflammatory responses. Pre-treatment with NAC attenuated oxidative stress and alterations in GSH metabolism associated with ß-cells cytotoxicity.

Augmentation of Glucotoxicity, Oxidative Stress, Apoptosis and Mitochondrial Dysfunction in HepG2 Cells by Palmitic Acid.

Hyperglycemia and hyperlipidemia are the hallmarks of diabetes and obesity. Experimental and epidemiological studies have suggested that dietary management and caloric restriction are beneficial in reducing the complications of diabesity. Studies have suggested that increased availability of energy metabolites like glucose and saturated fatty acids induces metabolic, oxidative, and mitochondrial stress, accompanied by inflammation that may lead to chronic complications in diabetes. In the present study, we used human hepatoma HepG2 cells to investigate the effects of high glucose (25 mM) and high palmitic acid (up to 0.3 mM) on metabolic-, inflammatory-, and redox-stress-associated alterations in these cells. Our results showed increased lipid, protein, and DNA damage, leading to caspase-dependent apoptosis and mitochondrial dysfunction. Glucolipotoxicity increased ROS production and redox stress appeared to alter mitochondrial membrane potential and bioenergetics. Our results also demonstrate the enhanced ability of cytochrome P450s-dependent drug metabolism and antioxidant adaptation in HepG2 cells treated with palmitic acid, which was further augmented with high glucose. Altered NF-kB/AMPK/mTOR-dependent cell signaling and inflammatory (IL6/TNF-α) responses were also observed. Our results suggest that the presence of high-energy metabolites enhances apoptosis while suppressing autophagy by inducing inflammatory and oxidative stress responses that may be responsible for alterations in cell signaling and metabolism.

Predictors of objectively measured physical activity in 12-month-old infants: A study of linked birth cohort data with electronic health records.

BACKGROUND: Physical activity (PA) levels are associated with long-term health, and levels of PA when young are predictive of adult activity levels. OBJECTIVES: This study examines factors associated with PA levels in 12-month infants. METHOD: One hundred forty-one mother-infant pairs were recruited via a longitudinal birth cohort study (April 2010 to March 2013). The PA level was collected using accelerometers and linked to postnatal notes and electronic medical records via the Secure Anonymised Information Linkage databank. Univariable and multivariable linear regressions were used to examine the factors associated with PA levels. RESULTS: Using univariable analysis, higher PA was associated with the following (P value less than 0.05): being male, larger infant size, healthy maternal blood pressure levels, full-term gestation period, higher consumption of vegetables (infant), lower consumption of juice (infant), low consumption of adult crisps (infant), longer breastfeeding duration, and more movement during sleep (infant) but fewer night wakings. Combined into a multivariable regression model (R2  = 0.654), all factors remained significant, showing lower PA levels were associated with female gender, smaller infant, preterm birth, higher maternal blood pressure, low vegetable consumption, high crisp consumption, and less night movement. CONCLUSION: The PA levels of infants were strongly associated with both gestational and postnatal environmental factors. Healthy behaviours appear to cluster, and a healthy diet was associated with a more active infant. Boys were substantially more active than girls, even at age 12 months. These findings can help inform interventions to promote healthier lives for infants and to understand the determinants of their PA levels.

An EEG-EMG correlation-based brain-computer interface for hand orthosis supported neuro-rehabilitation.

BACKGROUND: Corticomuscular coupling has been investigated for long, to find out the underlying mechanisms behind cortical drives to produce different motor tasks. Although important in rehabilitation perspective, the use of corticomuscular coupling for driving brain-computer interface (BCI)-based neurorehabilitation is much ignored. This is primarily due to the fact that the EEG-EMG coherence popularly used to compute corticomuscular coupling, fails to produce sufficient accuracy in single-trial based prediction of motor tasks in a BCI system. NEW METHOD: In this study, we have introduced a new corticomuscular feature extraction method based on the correlation between band-limited power time-courses (CBPT) associated with EEG and EMG. 16 healthy individuals and 8 hemiplegic patients participated in a BCI-based hand orthosis triggering task, to test the performance of the CBPT method. The healthy population was equally divided into two groups; one experimental group for CBPT-based BCI experiment and another control group for EEG-EMG coherence based BCI experiment. RESULTS: The classification accuracy of the CBPT-based BCI system was found to be 92.81 ±â€¯2.09% for the healthy experimental group and 84.53 ±â€¯4.58% for the patients' group. COMPARISON WITH EXISTING METHOD: The CBPT method significantly (p-value < 0.05) outperformed the conventional EEG-EMG coherence method in terms of classification accuracy. CONCLUSIONS: The experimental results clearly indicate that the EEG-EMG CBPT is a better alternative as a corticomuscular feature to drive a BCI system. Additionally, it is also feasible to use the proposed method to design BCI-based robotic neurorehabilitation paradigms.

Covariate shift estimation based adaptive ensemble learning for handling non-stationarity in motor imagery related EEG-based brain-computer interface.

The non-stationary nature of electroencephalography (EEG) signals makes an EEG-based brain-computer interface (BCI) a dynamic system, thus improving its performance is a challenging task. In addition, it is well-known that due to non-stationarity based covariate shifts, the input data distributions of EEG-based BCI systems change during inter- and intra-session transitions, which poses great difficulty for developments of online adaptive data-driven systems. Ensemble learning approaches have been used previously to tackle this challenge. However, passive scheme based implementation leads to poor efficiency while increasing high computational cost. This paper presents a novel integration of covariate shift estimation and unsupervised adaptive ensemble learning (CSE-UAEL) to tackle non-stationarity in motor-imagery (MI) related EEG classification. The proposed method first employs an exponentially weighted moving average model to detect the covariate shifts in the common spatial pattern features extracted from MI related brain responses. Then, a classifier ensemble was created and updated over time to account for changes in streaming input data distribution wherein new classifiers are added to the ensemble in accordance with estimated shifts. Furthermore, using two publicly available BCI-related EEG datasets, the proposed method was extensively compared with the state-of-the-art single-classifier based passive scheme, single-classifier based active scheme and ensemble based passive schemes. The experimental results show that the proposed active scheme based ensemble learning algorithm significantly enhances the BCI performance in MI classifications.

Cytoprotective Effects of N-Acetylcysteine on Streptozotocin- Induced Oxidative Stress and Apoptosis in RIN-5F Pancreatic ß-Cells.

BACKGROUND/AIMS: Numerous studies have reported overproduction of reactive oxygen species (ROS) and alterations in mitochondrial energy metabolism in the development of diabetes and its complications. The potential protective effects of N-acetylcysteine (NAC) in diabetes have been reported in many therapeutic studies. NAC has been shown to reduce oxidative stress and enhance redox potential in tissues protecting them against oxidative stress associated complications in diabetes. In the current study, we aimed to investigate the molecular mechanisms of the protective action of NAC on STZ-induced toxicity in insulin secreting Rin-5F pancreatic ß-cells. METHODS: Rin-5F cells were grown to 80% confluence and then treated with 10mM STZ for 24h in the presence or absence of 10mM NAC. After sub-cellular fractionation, oxidative stress, GSH-dependent metabolism and mitochondrial respiratory functions were studied using spectrophotometric, flow cytometric and Western blotting techniques. RESULTS: Our results showed that STZ-induced oxidative stress and apoptosis caused inhibition in insulin secretion while NAC treatment restored the redox homeostasis, enhanced insulin secretion in control cells and prevented apoptosis in STZ-treated cells. Moreover, NAC attenuated the inhibition of mitochondrial functions induced by STZ through partial recovery of the mitochondrial enzymes and restoration of membrane potential. STZ-induced DNA damage and expression of apoptotic proteins were significantly inhibited in NAC-treated cells. CONCLUSION: Our results suggest that the cytoprotective action of NAC is mediated via suppression of oxidative stress and apoptosis and restoration of GSH homeostasis and mitochondrial bioenergetics. This study may, thus, help in better understanding the cellular defense mechanisms of pancreatic ß-cells against STZ-induced cytotoxicity.

Cigarette Smoke Toxins-Induced Mitochondrial Dysfunction and Pancreatitis Involves Aryl Hydrocarbon Receptor Mediated Cyp1 Gene Expression: Protective Effects of Resveratrol.

We previously reported that mitochondrial CYP1 enzymes participate in the metabolism of polycyclic aromatic hydrocarbons and other carcinogens leading to mitochondrial dysfunction. In this study, using Cyp1b1-/-, Cyp1a1/1a2-/-, and Cyp1a1/1a2/1b1-/- mice, we observed that cigarette and environmental toxins, namely benzo[a]pyrene (BaP) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), induce pancreatic mitochondrial respiratory dysfunction and pancreatitis. Our results suggest that aryl hydrocarbon receptor (AhR) activation and resultant mitochondrial dysfunction are associated with pancreatic pathology. BaP treatment markedly inhibits pancreatic mitochondrial oxygen consumption rate (OCR), ADP-dependent OCR, and also maximal respiration, in wild-type mice but not in Cyp1a1/1a2-/- and Cyp1a1/1a2/1b1-/- mice. In addition, both BaP and TCDD treatment markedly affected mitochondrial complex IV activity, in addition to causing marked reduction in mitochondrial DNA content. Interestingly, the AhR antagonist resveratrol, attenuated BaP-induced mitochondrial respiratory defects in the pancreas, and reversed pancreatitis, both histologically and biochemically in wild-type mice. These results reveal a novel role for AhR- and AhR-regulated CYP1 enzymes in eliciting mitochondrial dysfunction and cigarette toxin-mediated pancreatic pathology. We propose that increased mitochondrial respiratory dysfunction and oxidative stress are involved in polycyclic aromatic hydrocarbon associated pancreatitis. Resveratrol, a chemo preventive agent and AhR antagonist, and CH-223191, a potent and specific AhR inhibitor, confer protection against BaP-induced mitochondrial dysfunction and pancreatic pathology.