Chromium (VI) induces insulin resistance in 3T3-L1 adipocytes through elevated reactive oxygen species generation

Reactive oxygen species (ROS) have been proposed to be involved in the development of insulin resistance, although the exact molecular link between ROS and insulin resistance remains to be determined. Chromium (Cr(VI)) is known as an inducer of ROS. Therefore, this study examined whether Cr(VI) could induce insulin resistance. It demonstrated that Cr(VI) treatment significantly inhibited insulin-stimulated glucose uptake and attenuated insulin signalling. Moreover, Cr(VI) treatment markedly increased the intracellular levels of superoxide anion, hydrogen peroxide and hydroxyl radical. N-acetylcysteine, superoxide dismutase and catalase can block the ROS generation and alleviate the insulin resistance induced by Cr(VI) treatment. In addition, Cr(VI) treatment induced endoplasmic reticulum (ER) stress and JNK activation and these effects were diminished by N-acetylcysteine. These results suggested that ROS generation through Cr(VI) treatment cause ER stress, JNK activation and insulin resistance in adipocytes. Therefore, the oxidative stress could be a potential interventional target for insulin-resistance related diseases.     

Chronic insulin treatment causes insulin resistance in 3T3-L1 adipocytes through oxidative stress

Insulin resistance and hyperinsulinemia are commonly present in obesity and pre-diabetes, and hyperinsulinemia is both a marker and a cause for insulin resistance. However, the molecular link between hyperinsulinemia and insulin resistance remains elusive. The present study examined the effect of chronic insulin treatment on the reactive oxygen species (ROS) production, insulin signalling and insulin-stimulated glucose uptake in 3T3-L1 adipocytes. The results showed that chronic insulin treatment significantly increased the intracellular generation of superoxide anion, hydrogen peroxide and hydroxyl radical. ROS induced by chronic insulin treatment inhibited insulin signalling and glucose uptake, induced endoplasmic reticulum (ER) stress and JNK activation. Furthermore, these effects were reversed by antioxidants N-acetylcysteine, superoxide dismutase or catalase. These results suggested that ROS, ER stress and JNK pathway are involved in insulin resistance induced by chronic insulin treatment. Therefore, oxidative stress could be a potential interventional target for hyperinsulinemia-induced insulin resistance and related diseases.     

Chronic insulin treatment causes insulin resistance in 3T3-L1 adipocytes through oxidative stress

Insulin resistance and hyperinsulinemia are commonly present in obesity and pre-diabetes, and hyperinsulinemia is both a marker and a cause for insulin resistance. However, the molecular link between hyperinsulinemia and insulin resistance remains elusive. The present study examined the effect of chronic insulin treatment on the reactive oxygen species (ROS) production, insulin signalling and insulin-stimulated glucose uptake in 3T3-L1 adipocytes. The results showed that chronic insulin treatment significantly increased the intracellular generation of superoxide anion, hydrogen peroxide and hydroxyl radical. ROS induced by chronic insulin treatment inhibited insulin signalling and glucose uptake, induced endoplasmic reticulum (ER) stress and JNK activation. Furthermore, these effects were reversed by antioxidants N-acetylcysteine, superoxide dismutase or catalase. These results suggested that ROS, ER stress and JNK pathway are involved in insulin resistance induced by chronic insulin treatment. Therefore, oxidative stress could be a potential interventional target for hyperinsulinemia-induced insulin resistance and related diseases.     

Endoplasmic reticulum stress-mediated apoptosis of EBV-transformed B cells by cross-linking of CD70 is dependent upon generation of reactive oxygen species and activation of p38 MAPK and JNK pathway

CD70 is expressed in normal activated immune cells as well as in several types of tumors. It has been established that anti-CD70 mAb induces complement-dependent death of CD70(+) tumor cells, but how anti-CD70 mAb affects the intrinsic signaling is poorly defined. In this report, we show that ligation of CD70 expressed on EBV-transformed B cells using anti-CD70 mAb induced production of reactive oxygen species (ROS) and subsequent apoptosis. We observed an early expression of endoplasmic reticulum (ER) stress response genes that preceded the release of apoptotic molecules from the mitochondria and the cleavage of caspases. CD70-induced apoptosis was inhibited by pretreatment with the ER stress inhibitor salubrinal, ROS quencher N-acetylcysteine, and Ca(2+) chelator BAPTA. We supposed that ROS generation might be the first event of CD70-induced apoptosis because N-acetylcysteine blocked increases of ROS and Ca(2+), but BAPTA did not block ROS generation. We also found that CD70 stimulation activated JNK and p38 MAPK. JNK inhibitor SP600125 and p38 inhibitor SB203580 effectively blocked upregulation of ER stress-related genes and cleavage of caspases. Inhibition of ROS generation completely blocked phosphorylation of JNK and p38 MAPK and induction of ER stress-related genes. Taken together, we concluded that cross-linking of CD70 on EBV-transformed B cells triggered ER stress-mediated apoptosis via ROS generation and JNK and p38 MAPK pathway activation. Our report reveals alternate mechanisms of direct apoptosis through CD70 signaling and provides data supporting CD70 as a viable target for an Ab-based therapy against EBV-related tumors.     

Constitutive Activation of Epidermal Growth Factor Receptor Promotes Tumorigenesis of Cr(VI)-transformed Cells through Decreased Reactive Oxygen Species and Apoptosis Resistance Development

Hexavalent chromium (Cr(VI)) compounds are well-established lung carcinogens. Epidermal growth factor receptor (EGFR) is a tyrosine kinase transmembrane receptor that regulates cell survival, tumor invasion, and angiogenesis. Our results show that chronic exposure of human bronchial epithelial (BEAS-2B) cells to Cr(VI) is able to cause malignant cell transformation. These transformed cells exhibit apoptosis resistance with reduced poly ADP-ribose polymerase cleavage (C-PARP) and Bax expression and enhanced expressions of Bcl-2 and Bcl-xL. These transformed cells also exhibit reduced capacity of reactive oxygen species (ROS) generation along with elevated expression of antioxidant manganese superoxide dismutase 2 (SOD2). The expression of this antioxidant was also elevated in lung tumor tissue from a worker exposed to Cr(VI) for 19 years. EGFR was activated in Cr(VI)-transformed BEAS-2B cells, lung tissue from animals exposed to Cr(VI) particles, and human lung tumor tissue. Further study indicates that constitutive activation of EGFR in Cr(VI)-transformed cells was due to increased binding to its ligand amphiregulin (AREG). Inhibition of EGFR or AREG increased Bax expression and reduced Bcl-2 expression, resulting in reduced apoptosis resistance. Furthermore, inhibition of AREG or EGFR restored capacity of ROS generation and decreased SOD2 expression. PI3K/AKT was activated, which depended on EGFR in Cr(VI)-transformed BEAS-2B cells. Inhibition of PI3K/AKT increased ROS generation and reduced SOD2 expression, resulting in reduced apoptosis resistance with commitment increase in Bax expression and reduction of Bcl-2 expression. Xenograft mouse tumor study further demonstrates the essential role of EGFR in tumorigenesis of Cr(VI)-transformed cells. In summary, the present study suggests that ligand-dependent constitutive activation of EGFR causes reduced ROS generation and increased antioxidant expression, leading to development of apoptosis resistance, contributing to Cr(VI)-induced tumorigenesis.     

Propranolol improves impaired hepatic phosphatidylinositol 3-kinase/akt signaling after burn injury

Severe burn injury is associated with induction of the hepatic endoplasmic reticulum (ER) stress response. ER stress leads to activation of c-Jun N-terminal kinase (JNK), suppression of insulin receptor signaling via phosphorylation of insulin receptor substrate 1 and subsequent insulin resistance. Marked and sustained increases in catecholamines are prominent after a burn. Here, we show that administration of propranolol, a nonselective β1/2 adrenergic receptor antagonist, attenuates ER stress and JNK activation. Attenuation of ER stress by propranolol results in increased insulin sensitivity, as determined by activation of hepatic phosphatidylinositol 3-kinase and Akt. We conclude that catecholamine release is responsible for the ER stress response and impaired insulin receptor signaling after burn injury.     

β-lapachone significantly increases the effect of ionizing radiation to cause mitochondrial apoptosis via JNK activation in cancer cells

Background

β-lapachone (β-lap), has been known to cause NQO1-dependnet death in cancer cells and sensitize cancer cells to ionizing radiation (IR). We investigated the mechanisms underlying the radiosensitization caused by β-lap.

Methodology/Principal Findings

β-lap enhanced the effect of IR to cause clonogenic cells in NQO1⁺-MDA-MB-231 cells but not in NQO1⁻-MDA-MB-231 cells. β-lap caused apoptosis only in NQO1⁺ cells and not in NQO1⁻ cells and it markedly increased IR-induced apoptosis only in NQO1⁺ cells. Combined treatment of NQO1⁺ cells induced ROS generation, triggered ER stress and stimulated activation of ERK and JNK. Inhibition of ROS generation by NAC effectively attenuated the activation of ERK and JNK, induction of ER stress, and subsequent apoptosis. Importantly, inhibition of ERK abolished ROS generation and ER stress, whereas inhibition of JNK did not, indicating that positive feedback regulation between ERK activation and ROS generation triggers ER stress in response to combined treatment. Furthermore, prevention of ER stress completely blocked combination treatment-induced JNK activation and subsequent apoptotic cell death. In addition, combined treatment efficiently induced the mitochondrial translocation of cleaved Bax, disrupted mitochondrial membrane potential, and the nuclear translocation of AIF, all of which were efficiently blocked by a JNK inhibitor. Caspases 3, 8 and 9 were activated by combined treatment but inhibition of these caspases did not abolish apoptosis indicating caspase activation played a minor role in the induction of apoptosis.

Conclusions/Significance

β-lap causes NQO1-dependent radiosensitization of cancer cells. When NQO1⁺ cells are treated with combination of IR and β-lap, positive feedback regulation between ERK and ROS leads to ER stress causing JNK activation and mitochondrial translocation of cleaved Bax. The resultant decrease in mitochondrial membrane leads to translocation of AIF and apoptosis.     

Hyperosmotic stress-dependent NFkappaB activation is regulated by reactive oxygen species and IGF-1 in cultured cardiomyocytes

We have recently shown that hyperosmotic stress activates p65/RelB NFkappaB in cultured cardiomyocytes with dichotomic actions on caspase activation and cell death. It remains unexplored how NFkappaB is regulated in cultured rat cardiomyocytes exposed to hyperosmotic stress. We study here: (a) if hyperosmotic stress triggers reactive oxygen species (ROS) generation and in turn whether they regulate NFkappaB and (b) if insulin-like growth factor-1 (IGF-1) modulates ROS production and NFkappaB activation in hyperosmotically-stressed cardiomyocytes. The results showed that hyperosmotic stress generated ROS in cultured cardiac myocytes, in particular the hydroxyl and superoxide species, which were inhibited by N-acetylcysteine (NAC). Hyperosmotic stress-induced NFkappaB activation as determined by IkappaBalpha degradation and NFkappaB DNA binding. NFkappaB activation and procaspase-3 and -9 fragmentation were prevented by NAC and IGF-1. However, this growth factor did not decrease ROS generation induced by hyperosmotic stress, suggesting that its actions over NFkappaB and caspase activation may be due to modulation of events downstream of ROS generation. We conclude that hyperosmotic stress induces ROS, which in turn activates NFkappaB and caspases. IGF-1 prevents NFkappaB activation by a ROS-independent mechanism.     

Estrogen-dependent inhibition of dextrose-induced endoplasmic reticulum stress and superoxide generation in endothelial cells

Increased oxidative stress and endoplasmic reticulum stress (ER stress) have been implicated in atherosclerosis. Estrogens have potent antioxidant activity but their effects on ER stress have not been well studied. Therefore, we studied the effects of estradiol and related sex steroids on dextrose-induced ER stress and superoxide (SO) generation in human umbilical vein endothelial cells (HUVECs). Oxidative stress was measured using hydroethidine fluorescence and MCLA chemiluminescence. ER stress was measured with an ER stress-sensitive secreted alkaline phosphatase (ES-TRAP) assay and by Western blot analysis of the expression of GRP78, JNK1, and phosphorylated JNK1, markers for ER stress. A supraphysiological dextrose concentration (27.5mM) increased ER stress and SO generation compared to treatment with a physiological concentration (5.5mM) of dextrose. In the presence of estradiol or testosterone (T), ER stress and SO generation were significantly reduced. In contrast to T-treated cells, dihydrotestosterone and 5-methyltestosterone were ineffective at alleviating ER stress or SO generation. When HUVECs were treated with T and the aromatase inhibitor 4-hydroxy-4-androstene-3,17-dione, T was no longer effective at suppressing ER stress or inhibiting SO generation. Changes in GRP78 expression and JNK activity in HUVECs support the results obtained in the ES-TRAP assay. These results indicate that dextrose-induced endoplasmic reticulum stress and superoxide generation are reversed by estradiol and testosterone; however, the latter requires aromatase-dependent conversion to estradiol.     

Fas-stimulated generation of reactive oxygen species or exogenous oxidative stress sensitize cells to Fas-mediated apoptosis

Inhibition of Fas-mediated apoptosis in B cell lymphomas by thiol antioxidants (glutathione and N-acetylcysteine) supported previous studies, suggesting that Fas-stimulated ROS generation may play a role in Fas-mediated apoptosis. Thus, the goal of the current study was to determine if Fas stimulation could induce ROS generation and what role, if any, it played in apoptosis. Fas crosslinking induced rapid generation of ROS (within 15 min) well before the appearance of characteristic apoptotic changes. Overexpression of catalase or superoxide dismutase suggested that Fas induced production of both superoxide anion and hydrogen peroxide. ROS generation was only observed, however, in cells that were sensitive to apoptosis and not in B cells inherently resistant to anti-Fas or in those in which resistance was induced by B cell receptor crosslinking. The exogenous addition of 250 microM hydrogen peroxide could reverse the resistant phenotype and sensitize cells to Fas-induced apoptosis. In Fas-sensitive cells, depletion of endogenous antioxidant defenses with buthionine sulfoximine increased the sensitivity to Fas-induced apoptosis, while overexpression of antioxidant enzymes and antiapoptotic proteins suggested a role for Fas-induced production of hydrogen peroxide in apoptosis. Further analysis suggested a redox-sensitive step early in Fas signaling at the level of initiator caspase (caspase-8) activation. Thus, the data suggest that the level of oxidative stress, either from exogenous sources or generated endogenously upon receptor stimulation, regulates the sensitivity to Fas-mediated apoptosis.     

Induction of apoptosis by casticin in cervical cancer cells: reactive oxygen species-dependent sustained activation of Jun N-terminal kinase

Casticin, a polymethoxyflavone from Fructus viticis used as an anti-inflammatory agent in Chinese traditional medicine, has been reported to have anti-cancer activity. The purpose of this study was to examine the apoptotic activity of casticin on human cervical cancer cells and its molecular mechanism. We revealed a novel mechanism by which casticin-induced apoptosis occurs and showed for the first time that the apoptosis induced by casticin is mediated through generation of reactive oxygen species (ROS) and sustained activation of c-Jun N-terminal kinase (JNK) in HeLa cells. Casticin markedly increased the levels of intracellular ROS and induced the expression of phosphorylated JNK and c-Jun protein. Pre-treatment with N-acetylcysteine and SP600125 effectively attenuated induction of apoptosis by casticin in HeLa cells. Moreover, casticin induced ROS production and apoptotic cell death in other cervical cancer cell lines, such as CasKi and SiHa. Importantly, casticin did not cause generation of ROS or induction of apoptosis in normal human peripheral blood mononuclear cells and embryonic kidney epithelium 293 cells. These results suggest that ROS generation and sustained JNK activation by casticin play a role in casticin-induced apoptosis and raise the possibility that treatment with casticin might be promising as a new therapy against human cervical cancer.     

Dissociation of inositol-requiring enzyme (IRE1α)-mediated c-Jun N-terminal kinase activation from hepatic insulin resistance in conditional X-box-binding protein-1 (XBP1) knock-out mice

Hepatic insulin resistance has been attributed to both increased endoplasmic reticulum (ER) stress and accumulation of intracellular lipids, specifically diacylglycerol (DAG). The ER stress response protein, X-box-binding protein-1 (XBP1), was recently shown to regulate hepatic lipogenesis, suggesting that hepatic insulin resistance in models of ER stress may result from defective lipid storage, as opposed to ER-specific stress signals. Studies were designed to dissociate liver lipid accumulation and activation of ER stress signaling pathways, which would allow us to delineate the individual contributions of ER stress and hepatic lipid content to the pathogenesis of hepatic insulin resistance. Conditional XBP1 knock-out (XBP1Δ) and control mice were fed fructose chow for 1 week. Determinants of whole-body energy balance, weight, and composition were determined. Hepatic lipids including triglyceride, DAGs, and ceramide were measured, alongside markers of ER stress. Whole-body and tissue-specific insulin sensitivity were determined by hyperinsulinemic-euglycemic clamp studies. Hepatic ER stress signaling was increased in fructose chow-fed XBP1Δ mice as reflected by increased phosphorylated eIF2α, HSPA5 mRNA, and a 2-fold increase in hepatic JNK activity. Despite JNK activation, XBP1Δ displayed increased hepatic insulin sensitivity during hyperinsulinemic-euglycemic clamp studies, which was associated with increased insulin-stimulated IRS2 tyrosine phosphorylation, reduced hepatic DAG content, and reduced PKCε activity. These studies demonstrate that ER stress and IRE1α-mediated JNK activation can be disassociated from hepatic insulin resistance and support the hypothesis that hepatic insulin resistance in models of ER stress may be secondary to ER stress modulation of hepatic lipogenesis.     

The Effects of Palmitate on Hepatic Insulin Resistance Are Mediated by NADPH Oxidase 3-derived Reactive Oxygen Species through JNK and p38MAPK Pathways

Elevated plasma free fatty acid (FFA) levels in obesity may play a pathogenic role in the development of insulin resistance. However, molecular mechanisms linking FFA to insulin resistance remain poorly understood. Oxidative stress acts as a link between FFA and hepatic insulin resistance. NADPH oxidase 3 (NOX3)-derived reactive oxygen species (ROS) may mediate the effect of TNF-α on hepatocytes, in particular the drop in cellular glycogen content. In the present study, we define the critical role of NOX3-derived ROS in insulin resistance in db/db mice and HepG2 cells treated with palmitate. The db/db mice displayed increased serum FFA levels, excess generation of ROS, and up-regulation of NOX3 expression, accompanied by increased lipid accumulation and impaired glycogen content in the liver. Similar results were obtained from palmitate-treated HepG2 cells. The exposure of palmitate elevated ROS production and NOX3 expression and, in turn, increased gluconeogenesis and reduced glycogen content in HepG2 cells. We found that palmitate induced hepatic insulin resistance through JNK and p38^MAPK pathways, which are rescued by siRNA-mediated NOX3 reduction. In conclusion, our data demonstrate a critical role of NOX3-derived ROS in palmitate-induced insulin resistance in hepatocytes, indicating that NOX3 is the predominant source of palmitate-induced ROS generation and that NOX3-derived ROS may drive palmitate-induced hepatic insulin resistance through JNK and p38^MAPK pathways.     

NOX3-derived reactive oxygen species promote TNF-α-induced reductions in hepatocyte glycogen levels via a JNK pathway

TNF-α-induced insulin resistance is associated with generation of reactive oxygen species (ROS). This study aims at defining the link between ROS production and hepatic insulin resistance. Treatment with TNF-α increased ROS generation through activating NADPH oxidase 3 (NOX3) in HepG2 hepatocytes. Down-regulation of NOX3 using siRNA prevented TNF-α-induced decrease of cellular glycogen. In the cells treated with TNF-α, there were NOX3-dependent activation of JNK, inhibition of IRS1 and phosphorylation of AKT/PKB and GSK. In conclusion, the effects of TNF-α on hepatic insulin resistance appear to be, at least in part, mediated by NOX3-derived ROS through a JNK pathway.     

Angiotensin II-induced skeletal muscle insulin resistance mediated by NF-kappaB activation via NADPH oxidase

Reduced insulin sensitivity is a key factor in the pathogenesis of type 2 diabetes and hypertension. Skeletal muscle insulin resistance is particularly important for its major role in insulin-mediated glucose disposal. Angiotensin II (ANG II) is integral in regulating blood pressure and plays a role in the pathogenesis of hypertension. In addition, we have documented that ANG II-induced skeletal muscle insulin resistance is associated with generation of reactive oxygen species (ROS). However, the linkage between ROS and insulin resistance in skeletal muscle remains unclear. To explore potential mechanisms, we employed the transgenic TG(mRen2)27 (Ren-2) hypertensive rat, which harbors the mouse renin transgene and exhibits elevated tissue ANG II levels, and skeletal muscle cell culture. Compared with Sprague-Dawley normotensive control rats, Ren-2 skeletal muscle exhibited significantly increased oxidative stress, NF-kappaB activation, and TNF-alpha expression, which were attenuated by in vivo treatment with an angiotensin type 1 receptor blocker (valsartan) or SOD/catalase mimetic (tempol). Moreover, ANG II treatment of L6 myotubes induced NF-kappaB activation and TNF-alpha production and decreased insulin-stimulated Akt activation and GLUT-4 glucose transporter translocation to plasma membranes. These effects were markedly diminished by treatment of myotubes with valsartan, the antioxidant N-acetylcysteine, NADPH oxidase-inhibiting peptide (gp91 ds-tat), or NF-kappaB inhibitor (MG-132). Similarly, NF-kappaB p65 small interfering RNA reduced NF-kappaB p65 subunit expression and nuclear translocation and TNF-alpha production but improved insulin-stimulated phosphorylation (Ser(473)) of Akt and translocation of GLUT-4. These findings suggest that NF-kappaB plays an important role in ANG II/ROS-induced skeletal muscle insulin resistance.     

NADPH oxidase 2 plays a critical role in dysfunction and apoptosis of pancreatic β-cells induced by very low-density lipoprotein

In type 2 diabetes, pancreatic β-cells cannot secret enough insulin compensate for insulin resistance, which are often accompanied by abnormality in lipid metabolism such as hypertriglyceridemia. It is reported that oxidative stress is involved in pancreatic β-cell dysfunction. However, molecular mechanisms linking between excessive generations of reactive oxygen species (ROS) and β-cell dysfunction and apoptosis induced by high levels of very low-density lipoprotein (VLDL) are poorly understood. In this study, we test the hypothesis that NADPH oxidase 2 (NOX2)-derived ROS may play a key role in dysfunction and apoptosis of pancreatic β-cell induced by VLDL. Our results show that the ApoCIII transgenic mice displayed increased serum TG levels, enhanced generation of ROS and impaired insulin content in pancreatic β-cells. In vitro, the treatment of pancreatic NIT-1 cells with 1?mg/ml VLDL for 12?h stimulated NOX2-derived ROS generation, decreased expression and secretion of insulin. Furthermore, we found that VLDL induced dysfunction and apoptosis of pancreatic β-cells through JNK and p53 pathways, which were rescued by siRNA-mediated NOX2 reduction. In conclusion, our data demonstrate a critical role of NOX2-derived ROS in dysfunction and apoptosis through JNK and p53 pathways in pancreatic β-cells induced by VLDL.     

Suppression of NADPH oxidase 2 substantially restores glucose-induced dysfunction of pancreatic NIT-1 cells

Defects in insulin secretion by pancreatic cells and/or decreased sensitivity of target tissues to insulin action are the key features of type 2 diabetes. It has been shown that excessive generation of reactive oxygen species (ROS) is linked to glucose-induced β-cell dysfunction. However, cellular mechanisms involved in ROS generation in β-cells and the link between ROS and glucose-induced β-cell dysfunction are poorly understood. Here, we demonstrate a key role of NADPH oxidase 2 (NOX2)-derived ROS in the deterioration of β-cell function induced by a high concentration of glucose. Sprague-Dawley rats were fed a high-fat diet for 24 weeks to induce diabetes. Diabetic rats showed increased glucose levels and elevated ROS generation in blood, but decreased insulin content in pancreatic β-cells. In vitro, increased ROS levels in pancreatic NIT-1 cells exposed to high concentrations of glucose (33.3 mmol·L(-1)) were associated with elevated expression of NOX2. Importantly, decreased glucose-induced insulin expression and secretion in NIT-1 cells could be rescued via siRNA-mediated NOX2 reduction. Furthermore, high glucose concentrations led to apoptosis of β-cells by activation of p38MAPK and p53, and dysfunction of β-cells through phosphatase and tensih homolog (PTEN)-dependent Jun N-terminal kinase (JNK) activation and protein kinase B (AKT/PKB) inhibition, which induced the translocation of forkhead box O1 and pancreatic duodenal homeobox-1, followed by reduced insulin expression and secretion. In conclusion, NOX2-derived ROS could play a critical role in high glucose-induced β-cell dysfunction through PTEN-dependent JNK activation and AKT inhibition.     

IRE1α/NOX4 signaling pathway mediates ROS‐dependent activation of hepatic stellate cells in NaAsO2‐induced liver fibrosis

Liver fibrosis is a severe health problem worldwide, and it is characterized by the activation of hepatic stellate cells (HSCs) and excessive deposition of collagen. Prolonged arsenic exposure can induce HSCs activation and liver fibrosis. In the present study, the results showed that chronic NaAsO₂ ingestion could result in liver fibrosis and oxidative stress in Sprague–Dawley rats, along with representative collagen deposition and HSCs activation. In addition, the inositol‐requiring enzyme 1α (IRE1α)–endoplasmic reticulum (ER)‐stress pathway was activated, and the activity of nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) was upregulated in rat livers. Simultaneously, the excessive production of reactive oxygen species (ROS) could induce HSCs activation, and NOX4 played an important role in generating ROS in vitro. Moreover, ER stress occurred with HSCs activation at the same time under NaAsO₂ exposure, and during ER stress, the IRE1α pathway was responsible for NOX4 activation. Therefore, inhibition of IRE1α activation could attenuate the HSCs activation induced by NaAsO₂. In conclusion, the present study manifested that inorganic arsenic exposure could activate HSCs through IRE1α/NOX4‐mediated ROS generation.