Jingxiang Bai

Mount Sinai School of Medicine, Manhattan, NY, United States

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Publications (10)51.32 Total impact

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    ABSTRACT: Regulation and downstream effects of mitochondrial protein S-glutathionylation in response to oxidative stress are poorly understood. The study aim was to determine whether anti-oxidants such as catalase and estradiol alter mitochondrial protein S-glutathionylation and in turn affect apoptosis following ultraviolet B (UV-B) light irradiation. HeLa cells were transduced with increasing amounts of adenovirus encoding catalase (Ad-Cat) and β-galactosidase (Ad-Lac Z) or pre-incubated with estradiol before induction of apoptosis by UV-B light exposure. Inhibition of mitochondrial protein S-glutathionylation was assessed using autoantibodies specific for the non-S-glutathionylated form of PDC-E2. The percentage of apoptotic cells following UV-B irradiation were not significantly different between mock cells (cells with no virus infection) and Ad-Cat and Ad-Lac Z infected cells at all viral doses (all p > 0.050). Autoantibody staining of non-S-glutathionylated PDC-E2 in apoptotic cells was three times greater in only Ad-Cat infected cells compared to only Ad-Lac Z infected cells (81.3 ± 16.7 vs 26 ± 7.2 %, respectively, p = 0.030). Similarly estradiol treatment (33 and 100 nM) also significantly increased PDC-E2 staining in apoptotic cells compared to non-treated cells (both p < 0.010). The percentage of apoptotic cells was not significantly different with any of the estradiol concentrations (all p > 0.100). The observed procaspase 12 cleavage following UV-B irradiation suggests that a mitochondrial-independent apoptotic pathway was activated. In conclusion, following an apoptotic stimulus, estradiol may inhibit mitochondrial protein S-glutathionylation without inhibiting apoptosis. This effect may play a role in ninefold greater prevalence of autoantibodies against PDC-E2 in women with primary biliary cirrhosis.
    Molecular and Cellular Biochemistry 06/2012; 367(1-2):51-8. · 2.33 Impact Factor
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    ABSTRACT: Cytochrome P450 2E1 (CYP2E1) is suggested to play a role in alcoholic liver disease, which includes alcoholic fatty liver, alcoholic hepatitis, and alcoholic cirrhosis. In this study, we investigated whether CYP2E1 plays a role in experimental alcoholic fatty liver in an oral ethanol-feeding model. After 4 weeks of ethanol feeding, macrovesicular fat accumulation and accumulation of triglyceride in liver were observed in wild-type mice but not in CYP2E1-knockout mice. In contrast, free fatty acids (FFAs) were increased in CYP2E1-knockout mice but not in wild-type mice. CYP2E1 was induced by ethanol in wild-type mice, and oxidative stress induced by ethanol was higher in wild-type mice than in CYP2E1-knockout mice. Peroxisome proliferator-activated receptor alpha (PPARalpha), a regulator of fatty acid oxidation, was up-regulated in CYP2E1-knockout mice fed ethanol but not in wild-type mice. A PPARalpha target gene, acyl CoA oxidase, was decreased by ethanol in wild-type but not in CYP2E1-knockout mice. Chlormethiazole, an inhibitor of CYP2E1, lowered macrovesicular fat accumulation, inhibited oxidative stress, and up-regulated PPARalpha protein level in wild-type mice fed ethanol. The introduction of CYP2E1 to CYP2E1-knockout mice via an adenovirus restored macrovesicular fat accumulation. These results indicate that CYP2E1 contributes to experimental alcoholic fatty liver in this model and suggest that CYP2E1-derived oxidative stress may inhibit oxidation of fatty acids by preventing up-regulation of PPARalpha by ethanol, resulting in fatty liver.
    Hepatology 06/2008; 47(5):1483-94. · 12.00 Impact Factor
  • Jingxiang Bai, Arthur I Cederbaum
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    ABSTRACT: Cycloheximide (CHX), an inhibitor of protein synthesis, has been reported to prevent cell death in a wide variety of cell types and produced by different apoptotic stimuli. However, the mechanisms by which CHX protects cells from apoptosis are still unclear. In this study, we investigated whether p53 plays a role in the protection by CHX against serum withdrawal-induced apoptosis. Deprivation of serum from the culture medium causes apoptosis in HepG2 cells, and CHX dramatically protects cells from death. p53, p21, and Bax protein levels were elevated, and cell cycle arrest was produced after serum withdrawal. CHX abolished this elevation of p53, p21, and Bax as well as the cell cycle arrest induced by serum deprivation. The p53 inhibitor pifithrin-alpha protects HepG2 cells against apoptosis induced by serum withdrawal. HepG2 cells expressing a dominant negative form of mutant p53 and Hep3B cells lacking p53 were resistant to serum withdrawal-induced apoptosis. Lowering of p53 by small interfering RNA protects HepG2 cells from serum withdrawal-induced apoptosis. p53 phosphorylation was induced by serum withdrawal and other chemotherapeutic reagents such as actinomycin D, doxorubicin, and etoposide. CHX decreases the levels of phosphorylated p53 (pp53) even in the presence of a proteasome inhibitor, which maintains the total p53 levels, whereas it does not affect the dephosphorylation of pp53. These results suggest the possibility that kinases that phosphorylate p53 might be affected by CHX administration. In summary, CHX protects HepG2 cells from serum withdrawal-induced apoptosis through inhibiting the synthesis of p53 and the phosphorylation of p53.
    Journal of Pharmacology and Experimental Therapeutics 01/2007; 319(3):1435-43. · 3.89 Impact Factor
  • Jingxiang Bai, Arthur I Cederbaum
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    ABSTRACT: Induction of cytochrome P450 2E1 by ethanol is believed to be one of the central pathways by which ethanol generates a state of oxidative stress and causes hepatotoxicity. In order to evaluate the biochemical and toxicological actions of CYP2E1 and its sensitization of hepatotoxin-induced injury, an adenovirus which can mediate overexpression of CYP2E1 was constructed. Injecting this virus into mice through the tail vein elevated CYP2E1 protein and activity twofold in the liver of the mice compared with the mice injected with Ad-LacZ or saline. Transaminase levels were dramatically increased in mice injected with the CYP2E1 adenovirus. Histological evaluation of liver specimens of mice injected with Ad-2E1 showed liver cell injury. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) assay demonstrated that more cells were stained positively in the liver of the mice infected with Ad-2E1 than in the liver of the mice infected with Ad-LacZ. 3-Nitrotyrosine protein adducts and protein carbonyl adducts were increased in the liver of the mice infected with Ad-2E1 compared with Ad-LacZ. This potentiated toxicity most likely reflects interactions between CYP2E1- and adenovirus-mediated toxicity pathways. These results show that adenovirus-mediated overexpression of CYP2E1 could induce liver toxicity in mice and suggests a mechanism involving oxidative/nitrosative stress.
    Toxicological Sciences 07/2006; 91(2):365-71. · 4.33 Impact Factor
  • Jingxiang Bai, Arthur I Cederbaum
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    ABSTRACT: Induction of CYP2E1 by ethanol is one mechanism by which ethanol causes oxidative stress and alcohol liver disease. Although CYP2E1 is predominantly found in the endoplasmic reticulum, it is also located in rat hepatic mitochondria. In the current study, chronic alcohol consumption induced rat hepatic mitochondrial CYP2E1. To study the role of mitochondrial targeted CYP2E1 in generating oxidative stress and causing damage to mitochondria, HepG2 lines overexpressing CYP2E1 in mitochondria (mE10 and mE27 cells) were established by transfecting a plasmid containing human CYP2E1 cDNA lacking the hydrophobic endoplasmic reticulum targeting signal sequence into HepG2 cells followed by G418 selection. A 40-kDa catalytically active NH2-terminally truncated form of CYP2E1 (mtCYP2E1) was detected in the mitochondrial compartment in these cells by Western blot analysis. Cell death caused by depletion of GSH by buthionine sulfoximine (BSO) was increased in mE10 and mE27 cells as compared with cells transfected with empty vector (pCI-neo). Antioxidants were able to abolish the loss of cell viability. Increased levels of reactive oxygen species and mitochondrial 3-nitrotyrosine and 4-hydroxynonenal protein adducts and decreased mitochondrial aconitase activity and mitochondrial membrane potential were observed in mE10 and mE27 cells treated with BSO. The mitochondrial membrane stabilizer, cyclosporine A, was also able to protect these cells from BSO toxicity. These results revealed that CYP2E1 in the mitochondrial compartment could induce oxidative stress in the mitochondria, damage mitochondria membrane potential, and cause a loss of cell viability. The accumulation of CYP2E1 in hepatic mitochondria induced by ethanol consumption might play an important role in alcohol liver disease.
    Journal of Biological Chemistry 03/2006; 281(8):5128-36. · 4.65 Impact Factor
  • Jingxiang Bai, Arthur I Cederbaum
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    ABSTRACT: To study the biochemical and toxicological properties of cytochrome P450 2E1 (CYP2E1), an adenovirus containing human CYP2E1 cDNA (Ad-CYP2E1) was constructed and was shown to successfully mediate the overexpression of CYP2E1 in HepG2 cells. Acetaminophen (APAP) toxicity to HepG2 cells infected with Ad-CYP2E1 was characterized as a preliminary proof of principle experiment to validate the functionality of the CYP2E1 adenovirus. Compared with cells infected with Ad-LacZ, HepG2 cells infected with Ad-CYP2E1 were more sensitive to APAP induced necrosis and apoptosis when the cells were depleted of intracellular reduced glutathione (GSH). The APAP cytotoxicity was dependent on both the concentration of APAP and the multiplicity of infection of the Ad-CYP2E1 virus. Apoptosis induced by APAP in HepG2 cells overexpressing CYP2E1 was caspase dependent and could be inhibited by the pan-caspase inhibitor Z-VAD-fmk. After treatment with APAP, mitochondrial membrane potential was dramatically decreased in the CYP2E1-expressing cells. APAP protein adducts were elevated in HepG2 cells infected with Ad-CYP2E1 compared with that in cells infected with Ad-LacZ; two bands around 90 KD were found only in the CYP2E1-expressing cells. These results demonstrate that adenovirus-mediated overexpression of human CYP2E1 activates APAP to reactive metabolites which damage mitochondria, form protein adducts, and result in toxicity to HepG2 cells. The Ad-CYP2E1 may be useful for studies designed to investigate the role of CYP2E1 in APAP and alcoholic liver injury and to further characterize the actions and effects of CYP2E1.
    Molecular and Cellular Biochemistry 08/2004; 262(1-2):165-76. · 2.33 Impact Factor
  • Jingxiang Bai, Arthur I Cederbaum
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    ABSTRACT: Oxidants such as H(2)O(2) play a role in the toxicity of certain DNA-damaging agents, a process that often involves the tumor suppressor p53. H(2)O(2) is rapidly degraded by catalase, which protects cells against oxidant injury. To study the effect of catalase on apoptosis induced by DNA-damaging agents, HepG2 cells were infected with adenovirus containing the cDNA of catalase (Ad-Cat). Forty-eight hours after infection, catalase protein and activity was increased 7-10-fold compared with control cells infected with Ad-LacZ. After treatment with Vp16 or mitomycin C, control cells underwent apoptosis in a p53-dependent manner; however, overexpression of catalase inhibited this apoptosis. Basal levels as well as Vp16- or mitomycin C-stimulated levels of p53 and p21 protein were decreased in the catalase-overexpressing cells as compared with control cells; however, p53 mRNA levels were not decreased by catalase. There was no difference in p53 protein synthesis between catalase-overexpressing cells and control cells. However, pulse-chase experiments indicated that p53 protein degradation was enhanced in the catalase-overexpressing cells. Proteasome inhibitors but not calpeptin prevented the catalase-mediated decrease of p53 content. Whereas Vp16 increased, catalase overexpression decreased the phosphorylation of p53. The protein phosphatase inhibitor okadaic acid did not prevent the catalase-mediated down-regulation of p53 or phosphorylated p53. These results demonstrate that catalase protects HepG2 cells from apoptosis induced by DNA-damaging agents in association with decreasing p53 phosphorylation; the latter may lead to an acceleration in the degradation of p53 protein by the proteasome complex. This suggests that the level of catalase may play a critical role in cell-induced resistance to the effects of anti-cancer drugs which up-regulate p53.
    Journal of Biological Chemistry 03/2003; 278(7):4660-7. · 4.65 Impact Factor
  • Jingxiang Bai, Joseph A Odin
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    ABSTRACT: Apoptosis is a normal physiologic form of cell death that follows activation of either an intrinsic or extrinsic pathway. In the intrinsic, various stimuli, such as oxidative stress, lead to mitochondrial dysfunction and the release of pro-apoptotic factors. Ligand binding to cell surface death receptors, such as Fas, activates the extrinsic pathway. Due to the rapid clearance of apoptotic cells, detection and quantification of apoptotic cells is prone to underestimation. In the liver, the importance of apoptosis is evident both during development and homeostasis of the biliary tree. Apoptosis also plays a prominent role in liver pathogenesis. Induction of the extrinsic apoptotic pathway by cytotoxic lymphocytes predominates in autoimmune liver diseases, viral hepatitis, and liver allograft rejection. Biliary cell apoptosis is highly regulated by bcl-2 family members. Both the extrinsic and intrinsic pathways are active in alcohol-related liver disease. Overexpression of anti-apoptotic proteins and FasL allow liver tumor cells to evade tumor specific cytotoxic lymphocytes. Agents that modulate apoptosis may be of future therapeutic benefit in a number of liver diseases.
    Autoimmunity Reviews 02/2003; 2(1):36-42. · 7.98 Impact Factor
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    ABSTRACT: Induction of cytochrome P450 CYP2E1 by ethanol appears to be one of the mechanisms by which ethanol creates a state of oxidative stress. Glutathione (GSH) is a key cellular antioxidant that detoxifies reactive oxygen species. Depletion of GSH, especially mitochondrial GSH, is believed to play a role in the ethanol-induced liver injury. Previous results reported that depletion of GSH by buthionine-(S,R)-sulfoximine (BSO) treatment caused apoptosis and necrosis in HepG2 cells, which overexpress CYP2E1. In the current work, adenoviral infection with vectors that resulted in expression of catalase either in the cytosol or mitochondrial compartments was able to abolish the loss of mitochondrial membrane potential or damage to mitochondria observed in HepG2 cells overexpressing CYP2E1 that were treated with BSO. Loss of cell viability, either necrotic or apoptotic, was also prevented by the catalase overexpression after infection with the adenoviral vectors. The protective effects of catalase were associated with the suppression of the increase in the production of reactive oxygen species and of mitochondrial lipid peroxidation observed after GSH depletion. These results reveal a prominent role for H(2)O(2) as a mediator in the cytotoxicity observed after depletion of GSH in HepG2 cells overexpressing CYP2E1. Damage to mitochondria may be a critical step for cellular toxicity by CYP2E1-derived reactive oxygen species.
    Journal of Pharmacology and Experimental Therapeutics 05/2002; 301(1):111-8. · 3.89 Impact Factor
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    ABSTRACT: Several studies have shown that pyruvate can scavenge H(2)O(2) and protect from H(2)O(2)-mediated cell injury. Mitochondria are critical participants in the control of apoptotic and necrotic cell death. Mitochondrial GSH plays an important role in the maintenance of cell functions and viability by metabolism of oxygen free radicals generated by the respiratory chain. Since loss of GSH, especially mitochondrial GSH, is associated with increased production of reactive oxygen species and cell toxicity, the ability of pyruvate to protect against these actions was evaluated. Adding pyruvate to HepG2 cells depleted of GSH by treatment with l-buthionine sulfoximine (BSO) surprisingly caused loss of viability after 24 and 48 h of incubation. Anoxia, treatment with antioxidants, and infection with cytosolic catalase, and interestingly, catalase expressed in the mitochondrial compartment were able to rescue the HepG2 cells from this pyruvate plus BSO injury, suggesting a key role for H(2)O(2), and lipid peroxides as mediators in the cytotoxicity. This toxicity and cell death observed was linked to damage to the mitochondria as evidenced by the increased lipid peroxidation in total homogenate and mitochondrial fraction, loss of mitochondrial membrane potential, and a decrease in protein-sulfhydryl groups. The type of cell death observed under these conditions was a mixture of apoptosis and necrosis. These results suggest that the protective ability of pyruvate against oxidant damage requires a functional GSH pool, especially in the mitochondrial compartment, and that in the absence of GSH, pyruvate increases cell injury by damaging the mitochondria, presumably as a consequence of enhanced electron flow and reactive oxygen production by the respiratory chain.
    Free Radical Biology and Medicine 02/2002; 32(1):73-83. · 5.27 Impact Factor