cytochrome-c-t and Chemical-and-Drug-Induced-Liver-Injury

Cyclosporine-A (CsA) is a powerful immunosuppressive agent and hepatotoxicity results from CsA treatment. This study aimed to elucidate the effectiveness of tyrosine kinase inhibitor nilotinib against CsA-induced hepatotoxicity and the underlying molecular mechanisms. Male Sprague-Dawley rats were allocated into four groups and received drugs for 28 days as follows: Control group: received vehicle, Nilotinib group: received nilotinib (20 mg/kg orally), CsA group: received CsA by subcutaneous injection (20 mg/kg daily), CsA-nilotinib: received nilotinib and CsA. Serum lactate dehydrogenase (LDH), liver function biomarkers, hepatic levels of oxidative stress biomarkers, nuclear factor erythroid-2 like-2 (Nrf2), total antioxidant capacity (TAC), interleukin-2 (IL-2), IL-1β, IL-6, and cytochrome-C were assessed. Additionally, the protein levels and mRNA expression of Bcl2 associated X protein (Bax), caspase-3, nuclear factor-κB (NF-κB), hemoxygenase-1 (HO-1) were measured. Moreover, liver tissues were assessed histopathologically using hematoxylin-eosin and Masson trichrome stain. Nilotinib treatment decreased serum LDH, alanine aminotransferase, aspartate aminotransferase, and γ-glutamyltransferase (γ-GT), hepatic malondialdehyde, and cytochrome-C. It also increased superoxide dismutase, reduced glutathione, glutathione reductase, glutathione peroxidase, glutathione-S-transferase (GST), TAC, and Nrf2 compared to CsA-injected rats. In addition, nilotinib decreased NF-κB, IL-1β, IL-6, Bax, and caspase-3, while elevated IL-2 and immunoexpression of HO-1. Additionally, mRNA expression of Bax and caspase-3 was elevated and that of HO-1 and inhibitory protein κB-α was reduced in the nilotinib-treated group. Moreover, nilotinib significantly attenuated CsA-induced histopathological alterations. Nilotinib may have a promising role as a hepato-protective through its antiapoptotic, antioxidant, and anti-inflammatory effects.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Caspase 3; Chemical and Drug Induced Liver Injury; Cyclosporine; Cytochromes c; Male; Protein Kinase Inhibitors; Pyrimidines; Rats; Rats, Sprague-Dawley; Signal Transduction

Cisplatin (Cis) is an effective cancer therapy commonly employed in many therapeutic regimens. However, treatment regimens that contain either a high dose or cumulative doses of Cis could trigger liver damage. A unique study demonstrated that captopril (Cap) protects against Cis-induced liver toxicity, but only some liver function enzymes and some antioxidant enzymes were investigated in that study. Our study aims to elucidate the protective mechanism of Cap against Cis liver toxicity. Acute liver toxicity was induced in rats by injecting a single Cis dose (7.5 mg/kg) in three groups (n = 6). Two groups were pre-treated with low (50 mg/kg) and high (100 mg/kg) Cap doses for one week before Cis injection, and the third group was injected with Cis only. The high Cap dose significantly improved liver function markers (ALT, AST, and ALP) and hepatic tissue pathology. The low Cap dose significantly improved ALP and, to a lesser extent, hepatic tissue pathology. Both Cap doses significantly decreased liver contents of MDA, IL-1β, and cleaved caspase-3; and liver protein expression of TNF-α, Bax, and caspase-3. The high Cap dose significantly increased liver contents of GSH, GPx, CAT, and SOD, and the liver protein expression of Bcl2. Moreover, only the high Cap dose significantly decreased liver IL-6 content and cytochrome C protein expression. Cap did not inhibit the antitumor impact of Cis against HCT116 cancer cells. Therefore, Cap restricts Cis-induced liver toxicity by reducing inflammation and apoptosis and augmenting the antioxidant system.

Topics: Angiotensin-Converting Enzyme Inhibitors; Animals; Anti-Inflammatory Agents; Antineoplastic Agents; Antioxidants; Apoptosis; bcl-2-Associated X Protein; Captopril; Caspase 3; Cell Line, Tumor; Chemical and Drug Induced Liver Injury; Cisplatin; Cytochromes c; Down-Regulation; Humans; Liver Function Tests; Male; Oxidative Stress; Rats; Rats, Wistar; Signal Transduction

Acrylamide (ACR) is a heat-induced carcinogen substance that is found in some foods due to cooking or other thermal processes. The aim of present study was to assess the probable protective effects of morin against ACR-induced hepatorenal toxicity in rats. The rats were treated with ACR (38.27 mg/kg b.w., p.o.) alone or with morin (50 and 100 mg/kg b.w., p.o.) for 10 consecutive days. Morin treatment attenuated the ACR-induced liver and kidney tissue injury by diminishing the serum AST, ALP, ALT, urea and creatinine levels. Morin increased activities of SOD, CAT and GPx and levels of GSH, and suppressed lipid peroxidation in ACR induced tissues. Histopathological changes and immunohistochemical expressions of p53, EGFR, nephrin and AQP2 in the ACR-induced liver and kidney tissues were decreased after administration of morin. In addition, morin reversed the changes in levels of apoptotic, autophagic and inflammatory parameters such as caspase-3, bax, bcl-2, cytochrome c, beclin-1, LC3A, LC3B, p38α MAPK, NF-κB, IL-1β, IL-6, TNF-α and COX-2 in the ACR-induced toxicity. Morin also affected the protein levels by regulating the PI3K/Akt/mTOR signaling pathway and thus alleviated ACR-induced apoptosis and autophagy. Overall, these findings may shed some lights on new approaches for the treatment of ACR-induced hepatotoxicity and nephrotoxicity.

Topics: Acrylamide; Acute Kidney Injury; Animals; Autophagy; bcl-2-Associated X Protein; Beclin-1; Biomarkers; Caspase 3; Chemical and Drug Induced Liver Injury; Cyclooxygenase 2; Cytochromes c; Cytokines; Disease Models, Animal; Flavonoids; Kidney; Lipid Peroxidation; Liver; Male; Microtubule-Associated Proteins; Mitogen-Activated Protein Kinase 14; NF-kappa B; Oxidative Stress; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; TOR Serine-Threonine Kinases

Herbal drug-induced liver injury has been reported worldwide and gained global attention. Thousands of hepatic sinusoidal obstruction syndrome (HSOS) cases have been reported after consumption of herbal medicines and preparations containing pyrrolizidine alkaloids (PAs), which are natural phytotoxins globally distributed. And herbal medicines, such as Gynura japonica, are the current leading cause of PA-induced HSOS. The present study aimed to reveal the mechanism underlying the hepatotoxicity of seneciphylline (Seph), a main PA in G. japonica. Results showed that Seph induced severe liver injury through apoptosis in mice (70 mg/kg Seph, orally) and primary mouse and human hepatocytes (5-50 μM Seph). Further research uncovered that Seph induced apoptosis by disrupting mitochondrial homeostasis, inducing mitochondrial depolarization, mitochondrial membrane potential (MMP) loss, and cytochrome c (Cyt c) release and activating c-Jun N-terminal kinase (JNK). The Seph-induced apoptosis in hepatocytes could be alleviated by Mdivi-1 (50 μM, a dynamin-related protein 1 inhibitor), as well as SP600125 (25 μM, a specific JNK inhibitor) and ZVAD-fmk (50 μM, a general caspase inhibitor). Moreover, the Seph-induced MMP loss in hepatocytes was also rescued by Mdivi-1. In conclusion, Seph induced liver toxicity via activating mitochondrial-mediated apoptosis in mice and primary hepatocytes. Our results provide further information on Seph detoxification and herbal medicines containing Seph such as G. japonica.

Topics: Animals; Apoptosis; Cells, Cultured; Chemical and Drug Induced Liver Injury; Cytochromes c; Drugs, Chinese Herbal; Dynamins; Hepatocytes; Humans; JNK Mitogen-Activated Protein Kinases; Liver; Male; Membrane Potential, Mitochondrial; Mice, Inbred C57BL; Mitochondria, Liver; Primary Cell Culture; Pyrrolizidine Alkaloids; Signal Transduction

The hepatotoxicity of Tripterygium wilfordii Hook. f. (TW), due to the presence of triptolide (TP), limits its therapeutic potential. Based on the traditional Chinese medicine theory, the theory of "Yi lei xiang zhi" was proposed that Chinese herbs with different efficacy can restrict each other to achieve the least adverse reactions.. To observe the effects of Catapol (CAT) and Panax notoginseng saponins (PNS), active ingredients in Rehmannia glutinosa (RG) and Panax notoginseng (PN) respectively, on reducing TP-induced hepatotoxicity, and further to explore the mechanisms.. The human hepatic cell line L-02 was cultured and treated with CAT, PNS or Combinations, and then treated with TP. The cytotoxic assay, the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH), apoptosis, mitochondrial membrane potential and the expressions of NF-E2-related factor 1 (Nrf1) and its downstream targets were detected. Rats were treated with TP, TP + CAT, TP + PNS, or the combinations for 4 weeks. The levels of ALT, AST and LDH in serum, apoptosis of liver cells, mitochondria injury and the protein expressions of Caspase 3 and Nrf1 were investigated.. CAT, PNS or CAT+PNS pre-treatment inhibited TP-induced toxicity in L-02 cells, distinctly decreased the apoptosis, alleviated the reduction of mitochondrial membrane potential, and modulated the expressions of Nrf1 and its downstream target, the mitochondrial transcription factor A (TFAM) and cytochrome C (Cyt-C). CAT, PNS or CAT+PNS inhibited the TP-induced hepatotoxicity in SD rats by reducing the mitochondria injury, decreasing the cells apoptosis and increasing the Nrf1 protein expression. Noticeably, TP + PNS + CAT combinations exhibited more effective than any single ingredient alone.. PNS and CAT were able to effectively attenuate TP-induced hepatotoxicity. The efficiency benefits from their modulating Nrf1 and its downstream genes TFAM and Cyt-C, and further influencing mitochondrial functions and cells apoptosis. The combination is more effective than single ingredient alone.

Topics: Animals; Apoptosis; Biomarkers; Caspase 3; Cell Line; Chemical and Drug Induced Liver Injury; Cytochromes c; Cytoprotection; Disease Models, Animal; Diterpenes; DNA-Binding Proteins; Dose-Response Relationship, Drug; Drug Therapy, Combination; Drugs, Chinese Herbal; Epoxy Compounds; Female; Humans; Liver; Membrane Potential, Mitochondrial; Mitochondria, Liver; Mitochondrial Proteins; NF-E2-Related Factor 1; Panax; Phenanthrenes; Phytotherapy; Plants, Medicinal; Quaternary Ammonium Compounds; Rats, Sprague-Dawley; Saponins; Transcription Factors

Industrial and agricultural developments in recent years have resulted in the excessive discharge of arsenic into the environment, making arsenic toxicity a major worldwide concern. Oxidative stress is considered the primary mechanism for arsenic toxicity. The main objective of this study was to evaluate acetyl-l-carnitine's (ALC) protective ability against the arsenic-induced hepatotoxicity. For this purpose, male Wistar rats were distributed randomly into 5 groups of 8 rats each: control, arsenic (5 mg/kg) and arsenic plus ALC (5 mg/kg; 100, 200, 300 mg/kg). The animals were gavaged for 21 consecutive days. Liver tissue samples were extracted 24 h after the last treatment and were later analyzed for biochemical and histological alterations. The arsenic-induced oxidative damage was confirmed by elevation of malondialdehyde (MDA), a lipid peroxidation byproduct, as well as depletion in physiological antioxidant content such as superoxide dismutase (SOD) and catalase (CAT). Furthermore, alterations in mitochondrial functions including a significant decrease of mitochondrial outer membrane potential and reactive oxygen species (ROS) generation increase, mitochondrial swelling, release of cytochrome c and consequent activation of caspase-3 and caspase-9 and initiation of apoptosis, was observed following arsenic administration. Moreover, the inflammation was confirmed by the overexpression of inflammatory mediators such as NF-ĸB and IL-1 and IL-6. The present study demonstrated that ALC ameliorates arsenic-induced oxidative damage, mitochondrial dysfunction, apoptosis, inflammation and histological damage. ALC's protective features against arsenic hepatotoxicity may be due to this agent's antioxidant and anti-inflammatory properties as well as its stabilizing effects on mitochondrial function.

Topics: Acetylcarnitine; Animals; Anti-Inflammatory Agents; Antioxidants; Apoptosis; Arsenic; Caspase 3; Caspase 9; Catalase; Chemical and Drug Induced Liver Injury; Cytochromes c; Glutathione; Interleukin-1beta; Interleukin-6; Liver; Male; Malondialdehyde; Mitochondria; Oxidative Stress; Rats, Wistar; Reactive Oxygen Species; Superoxide Dismutase; Transcription Factor RelA

The detoxifying effect of pyridoxine against acetaminophen (APAP)-induced hepatotoxicity was investigated. HepG2 cells were co-treated with APAP and pyridoxine to compare with betaine or methionine for 24 h. LDH, ALT and AST activities were measured to determine direct cells damage in vitro and in vivo. Lipid peroxidation, antioxidant enzymes activity, and glutathione level were measured. Cytochrome c releaseand procaspase-3, cleaved caspase-3, Bcl-2, or Bax protein levels were measured to determine APAP-induced apoptotic cell death. Pyridoxine treatment significantly increased cell viability and decreased leakage of LDH activity against APAP-induced hepatotoxicity in HepG2 cells. ALT and AST activities were dose-dependently reduced by pyridoxine treatment compared to APAP-treated group. Significant increases in activities of GST and GPx were observed after co-treatment with APAP and pyridoxine. Although APAP-induced Nrf2 and HO-1 expression levels were gradually reduced in HepG2 cells by pyridoxine treatment, induction of antioxidant enzymes activities were dose-dependently increased. These protected effects of pyridoxine against APAP-induced hepatoxicity were closely associated with suppression of APAP-induced oxidative stress and apoptotic cell death in HepG2 cells. These data indicated that the protective action of pyridoxine against hepatic cell injuries was involved in the direct antioxidant activity which provides a pivotal mechanism for its potential hepatoprotective action.

Topics: Acetaminophen; Alanine Transaminase; Animals; Aspartate Aminotransferases; Chemical and Drug Induced Liver Injury; Cytochromes c; Glutathione; Glutathione Peroxidase; Hep G2 Cells; Humans; Lipid Peroxidation; Liver; Male; Malondialdehyde; Mice, Inbred ICR; Oxidative Stress; Pyridoxine

The FDA has approved 31 small-molecule kinase inhibitors (KIs) for human use as of November 2016, with six having black box warnings for hepatotoxicity (BBW-H) in product labeling. The precise mechanisms and risk factors for KI-induced hepatotoxicity are poorly understood. Here, the 31 KIs were tested in isolated rat liver mitochondria, an in vitro system recently proposed to be a useful tool to predict drug-induced hepatotoxicity in humans. The KIs were incubated with mitochondria or submitochondrial particles at concentrations ranging from therapeutic maximal blood concentrations (Cmax) levels to 100-fold Cmax levels. Ten endpoints were measured, including oxygen consumption rate, inner membrane potential, cytochrome c release, swelling, reactive oxygen species, and individual respiratory chain complex (I-V) activities. Of the 31 KIs examined only three including sorafenib, regorafenib and pazopanib, all of which are hepatotoxic, caused significant mitochondrial toxicity at concentrations equal to the Cmax, indicating that mitochondrial toxicity likely contributes to the pathogenesis of hepatotoxicity associated with these KIs. At concentrations equal to 100-fold Cmax, 18 KIs were found to be toxic to mitochondria, and among six KIs with BBW-H, mitochondrial injury was induced by regorafenib, lapatinib, idelalisib, and pazopanib, but not ponatinib, or sunitinib. Mitochondrial liability at 100-fold Cmax had a positive predictive power (PPV) of 72% and negative predictive power (NPV) of 33% in predicting human KI hepatotoxicity as defined by product labeling, with the sensitivity and specificity being 62% and 44%, respectively. Similar predictive power was obtained using the criterion of Cmax ≥1.1 µM or daily dose ≥100 mg. Mitochondrial liability at 1-2.5-fold Cmax showed a 100% PPV and specificity, though the NPV and sensitivity were 32% and 14%, respectively. These data provide novel mechanistic insights into KI hepatotoxicity and indicate that mitochondrial toxicity at therapeutic levels can help identify hepatotoxic KIs.

Topics: Animals; Chemical and Drug Induced Liver Injury; Cytochromes c; Dose-Response Relationship, Drug; Drug Labeling; Female; Male; Membrane Potential, Mitochondrial; Mitochondria, Liver; Oxygen; Predictive Value of Tests; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Risk Factors; Sensitivity and Specificity; Species Specificity

Ethanol-induced liver injury is associated with oxidative stress and hepatocyte apoptosis. We previously demonstrated that SIRT1/p66Shc pathway activation attenuates hepatocyte apoptosis in liver ischemia/reperfusion. The current study aimed to investigate whether carnosic acid (CA), a natural antioxidant, can inhibit acute ethanol-induced apoptosis of hepatocytes and to determine the effect of SIRT1/p66Shc on this process. Our results showed that CA pretreatment significantly reduced ethanol-induced histologic damage, serum aminotransferase activity, and oxidative stress in rats. Importantly, CA pretreatment increased SIRT1 expression following ethanol exposure. Furthermore, p66Shc expression was negatively correlated with SIRT1 expression. Consistent with the results demonstrating p66Shc inhibition, CA pretreatment inhibited the release of cytochrome C and apoptosis-inducing factor (AIF) from mitochondria. After exposing L02 cells to ethanol, the increased SIRT1 expression induced by CA was abrogated by pharmacologic SIRT1 inhibition or the use of siRNA against SIRT1. Additionally, SIRT1 inhibition significantly abrogated the suppression of p66Shc expression and mitochondrial translocation induced by CA. Accordingly, CA-induced decreases in the release of cytochrome C and AIF and in mitochondrial apoptosis were nearly abolished by SIRT1 knockdown. These data indicated that CA-activated SIRT1 is protective against ethanol treatment. In summary, CA attenuates acute ethanol-induced liver injury via a SIRT1/p66Shc-mediated mitochondrial pathway.

Topics: Abietanes; Animals; Antioxidants; Apoptosis; Apoptosis Inducing Factor; Chemical and Drug Induced Liver Injury; Cytochromes c; Ethanol; Hepatocytes; Liver; Male; Mitochondria; Oxidative Stress; Rats; Rats, Wistar; RNA, Small Interfering; Sirtuin 1; Src Homology 2 Domain-Containing, Transforming Protein 1

Endoplasmic reticulum (ER) stress is involved in the development of several liver diseases and tumors. This study investigated the underlying mechanisms of α-naphthyl isothiocyanate (ANIT)-induced liver injury with cholestasis in mice and found ER stress contributes to the injury. All animals were randomly divided into three groups. In the ANIT-intoxicated group, mice were intragastrically given 100mg/kg ANIT (dissolved in corn oil), while the other groups received an equal volume of vehicle as control. After 24 and 48h of ANIT administration, blood samples and liver tissues of all animals were collected for serum biochemistry and hepatic histopathological examinations to evaluate liver injuries with cholestasis. Hepatocellular apoptosis was assessed by the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. The expression of hepatic ER stress-related markers was determined by real-time PCR, immunohistochemical assay and Western blot. ANIT was found to significantly induce liver injury with cholestasis compared with control mice as evidenced by the increase of serum transaminases and total bilirubin (TBil), and histopathological changes in mice. ANIT remarkably induced hepatocellular apoptosis, upregulated the expression of caspase-9 and cytochrome c, and inhibited the gene and protein expression of proliferating cell nuclear antigen (PCNA). The gene expression of ER stress-related markers, including glucose-regulated protein 78 (GRP78), protein kinase R-like ER kinase (PERK), eukaryotic initiation factor 2α (eIF2α), inositol requiring enzyme-1α (IRE-1α) and activating transcription factor 6 (ATF6) was upregulated by ANIT in mice. ANIT also upregulated the protein expression of GRP78 and activated the phosphorylation of IRE1. These results suggested that ANIT induced liver injury with cholestasis partly due to its ability to activate the ER stress pathway.

Topics: Animals; Apoptosis; Caspase 9; Chemical and Drug Induced Liver Injury; Cholestasis; Cytochromes c; Endoplasmic Reticulum Chaperone BiP; Endoplasmic Reticulum Stress; Isocyanates; Liver; Male; Mice; Mice, Inbred ICR; Naphthalenes

To assess the effects of dihydromyricetin (DHM) as a hepatoprotective candidate in reducing hepatic injury and accelerating hepatocyte proliferation after carbon tetrachloride (CCl4) treatment.. C57 BL/6 mice were used in this study. Mice were orally administered with DHM (150 mg/kg) for 4 d after CCl4 treatment. Serum and liver tissue samples were collected on days 1, 2, 3, 5 and 7 after CCl4 treatment. The anti-inflammatory effect of DHM was assessed directly by hepatic histology detection and indirectly by serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), albumin, and superoxide dismutase (SOD). Inflammatory cytokines, such as interleukin (IL)-1β, IL-6 and tumor necrosis factor-α (TNF-α), were detected using ELISA kits. Proliferating cell nuclear antigen (PCNA) staining was used to evaluate the role of DHM in promoting hepatocyte proliferation. Hepatocyte apoptosis was measured by TUNEL assay. Furthermore, apoptosis proteins Caspases-3, 6, 8, and 9 were detected by Western blot. SP600125 were used to confirm whether DHM regulated liver regeneration through JNK/TNF-α pathways.. DHM showed a strong anti-inflammatory effect on CCl4-induced liver injury in mice. DHM could significantly decrease serum ALT, AST, IL-1β, IL-6 and TNF-α and increase serum albumin, SOD and liver SOD compared to the control group after CCl4 treatment (P < 0.05). PCNA results indicated that DHM could significantly increase the number of PCNA positive cells compared to the control (348.9 ± 56.0 vs 107.1 ± 31.4, P < 0.01). TUNEL assay showed that DHM dramatically reduced the number of apoptotic cells after CCl4 treatment compared to the control (365.4 ± 99.4 vs 90.5 ± 13.8, P < 0.01). Caspase activity detection showed that DHM could reduce the activities of Caspases- 8, 3, 6 and 9 compared to the control (P < 0.05). The results of Western blot showed that DHM increased the expression of JNK and decreased TNF-α expression. However, DHM could not affect TNF-α expression after SP600125 treatment. Furthermore, DHM could significantly improve the survival rate of acute liver failure (ALF) mice (73.3% vs 20.0%, P < 0.0001), and SP600125 could inhibit the effect of DHM.. These findings demonstrate that DHM alleviates CCl4-induced liver injury, suggesting that DHM is a promising candidate for reversing liver injury and ALF.

Topics: Animals; Anti-Inflammatory Agents; Biomarkers; Carbon Tetrachloride; Caspase Inhibitors; Cell Proliferation; Chemical and Drug Induced Liver Injury; Cytochromes c; Disease Models, Animal; Flavonols; Inflammation Mediators; JNK Mitogen-Activated Protein Kinases; Liver; Liver Failure, Acute; Liver Regeneration; Male; Mice, Inbred C57BL; Mitochondria, Liver; Protein Kinase Inhibitors; Signal Transduction; Time Factors; Tumor Necrosis Factor-alpha

Cryptotanshinone from Salvia miltiorrhiza Bunge was investigated for hepatoprotective effects in d-galactosamine (GalN)/lipopolysaccharide (LPS)-induced fulminant hepatic failure. Cryptotanshinone (20 or 40 mg/kg) was orally administered 12 and 1h prior to GalN (700 mg/kg)/LPS (10 μg/kg) injection. The increased mortality and TNF-α levels by GalN/LPS were declined by cryptotanshinone pretreatment. In addition, cryptotanshinone attenuated GalN/LPS-induced apoptosis, characterized by the blockade of caspase-3, -8, and -9 activation, as well as the release of cytochrome c from the mitochondria. In addition, cryptotanshinone significantly suppressed JNK, ERK and p38 phosphorylation induced by GalN/LPS, and phosphorylation of TAK1 as well. Furthermore, cryptotanshinone significantly inhibited the activation of NF-κB and suppressed the production of proinflammatory cytokines. These findings suggested that hepatoprotective effect of cryptotanshinone is likely associated with its anti-apoptotic activity and the down-regulation of MAPKs and NF-κB associated at least in part with suppressing TAK1 phosphorylation.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Aspartate Aminotransferases; Caspases; Chemical and Drug Induced Liver Injury; Cytochromes c; Galactosamine; Lipopolysaccharides; Liver; Liver Failure, Acute; Male; MAP Kinase Kinase Kinases; Mice; Mice, Inbred C57BL; Mitochondria; Mitogen-Activated Protein Kinases; NF-kappa B; Phenanthrenes; Phytotherapy; Plant Extracts; Salvia miltiorrhiza; Tumor Necrosis Factor-alpha

Doxorubicin (Dox) is an effective chemotherapeutic agent, but known to cause cardiac and hepatic toxicity. Mechanisms of toxicity have not been clearly identified, but shown to involve oxidative stress and mitochondrial dysfunction. However, antioxidant supplementation has only shown modest protection from Dox-induced toxicity in clinical trials. Therefore, further research is required to discern alternative mechanisms that may also play an important role in Dox-induced toxicity. Thus, we aimed to investigate the role of mitochondrial fusion and fission in Dox-induced hepatic toxicity, which has not yet been investigated. Six-week-old male F344 rats were injected IP with 20 mg/kg of Dox or saline. Once administered, both groups of animals were fasted with no food or water until sacrifice 24 h later. Dox decreased content of primary regulators of mitochondrial fusion (OPA1, MFN1, and MFN2) with no effect on regulators of fission (DRP1 and FIS1), thus shifting the balance favoring mitochondrial fission. Moreover, it was determined that mitochondrial fission was likely not coupled to cell proliferation or cytochrome c release leading to the activation of mitochondrial-mediated apoptotic signaling. Rather, mitochondrial fission may be coupled to mitophagy and may be an adaptive response to protect against Dox-induced hepatic toxicity. This is the first study to report the role of altered mitochondrial dynamics and mitophagy machinery in Dox-induced hepatic injury.

Topics: Animals; Antibiotics, Antineoplastic; Apoptosis; Cell Proliferation; Chemical and Drug Induced Liver Injury; Cytochromes c; Doxorubicin; Male; Mitochondria, Liver; Mitochondrial Dynamics; Mitophagy; Oxidative Stress; Rats; Rats, Inbred F344; Signal Transduction

We attempted to determine whether betanin (from natural pigments) that has anti-oxidant properties would be protective against paraquat-induced liver injury in Sprague-Dawley rats. Paraquat was injected intraperitoneally into rats to induce liver toxicity. The rats were randomly divided into four groups: a control group, a paraquat group, and two groups that received betanin at doses of 25 and 100mg/kg/day three days before and two days after they were administered paraquat. We evaluated liver histopathology, serum liver enzymatic activities, oxidative stress, cytochrome P450 (CYP) 3A2 mRNA expression, and mitochondrial damage. The rats that were injected with paraquat incurred liver injury, evidenced by histological changes and elevated serum aspartate aminotransferase and alanine aminotransferase levels; paraquat also led to oxidative stress, an increase of cytochrome P450 3A2 mRNA expression, and mitochondrial damage, indicated by mitochondrial membrane swelling, reduced mitochondrial cytochrome C, and apoptosis-inducing factor protein levels. Pathological damage and all of the above mentioned markers were lesser in the animals treated with betanin than in those who received paraquat alone. Betanin had a protective effect against paraquat-induced liver damage in rats. The mechanism of the protection appears to be the inhibition of CYP 3A2 expression and protection of mitochondria.

Topics: Animals; Antioxidants; Apoptosis Inducing Factor; Betacyanins; Biomarkers; Chemical and Drug Induced Liver Injury; Cytochrome P-450 CYP3A; Cytochromes c; Enzyme Inhibitors; Liver; Male; Mitochondria; Oxidative Stress; Paraquat; Rats; Rats, Sprague-Dawley; RNA, Messenger

Exposure to high levels of nitrites for a prolonged time have adverse health effects on several organs especially the liver due to oxidative properties. Meanwhile, cod liver oil has been reported to ameliorate organ dysfunction in animal models that involve oxidative stress.. Examine the impact of dietary cod liver oil on sodium nitrite-induced liver damage.. Thirty-two adult male Sprague-Dawely rats were daily treated with sodium nitrite (80 mg/kg) in presence or absence of cod liver oil (5 ml/kg). Morphological changes were assessed in liver sections. Oxidative stress and antioxidant markers were measured in serum and liver homogenates. Liver samples were used for measurements of MCP-1, DNA fragmentation and mitochondrial function.. The hepatoprotective effect of cod liver oil was proved by significant reduction of elevated liver enzymes and normal appearance of hepatocytes. Cod liver oil significantly reduced hepatic malondialdehyde, hydrogen peroxide and superoxide anion (224.3 ± 18.9 nmol/g, 59.3 ± 5.1 and 62.5 ± 5.1 µmol/g, respectively) compared with sodium nitrite (332.5 ± 25.5 nmol/g, 83.1 ± 8.1 and 93.9 ± 6.5 µmol/g, respectively). Cod liver oil restored hepatic cytochrome c oxidase activity after 38% reduction by sodium nitrite. Furthermore, cod liver oil significantly reduced hepatic MCP-1 (79.8 pg/mg) and DNA fragmentation (13.8%) compared with sodium nitrite (168.7 pg/mg and 41.3%, respectively).. Cod liver oil ameliorates sodium nitrite induced hepatic impairment through several mechanisms including attenuation of oxidative stress, blocking MCP-1, reactivation of mitochondrial function and reduction of DNA fragmentation.

Topics: Animals; Antioxidants; Chemical and Drug Induced Liver Injury; Chemokine CCL2; Cod Liver Oil; Cytochromes c; Cytoprotection; Disease Models, Animal; DNA Fragmentation; Hydrogen Peroxide; Liver; Male; Malondialdehyde; Mitochondria, Liver; Oxidative Stress; Rats; Rats, Sprague-Dawley; Sodium Nitrite; Superoxides

Melittin is a cationic, hemolytic peptide that is the main toxic component in the venom of the honey bee (Apis mellifera). Melittin has multiple effects, including anti-bacterial, anti-viral and anti-inflammatory, in various cell types. However, the anti-apoptotic mechanisms of melittin have not been fully elucidated in hepatocytes. Apoptosis contributes to liver inflammation and fibrosis. Knowledge of the apoptotic mechanisms is important to develop new and effective therapies for treatment of cirrhosis, portal hypertension, liver cancer, and other liver diseases. In the present study, we investigated the anti-apoptotic effect of melittin on transforming growth factor (TGF)-β1-induced apoptosis in hepatocytes. TGF-β1-treated hepatocytes were exposed to low doses (0.5 and 1 μg/mL) and high dose (2 μg/mL) of melittin. The low doses significantly protected these cells from DNA damage in TGF-β1-induced apoptosis compared to the high dose. Also, melittin suppressed TGF-β1-induced apoptotic activation of the Bcl-2 family and caspase family of proteins, which resulted in the inhibition of poly-ADP-ribose polymerase (PARP) cleavage. These results demonstrate that TGF-β1 induces hepatocyte apoptosis and that an optimal dose of melittin exerts anti-apoptotic effects against TGF-β1-induced injury to hepatocytes via the mitochondrial pathway. These results suggest that an optimal dose of melittin can serve to protect cells against TGF-β1-mediated injury.

Topics: Animals; Apoptosis; Caspases; Cell Line; Cell Survival; Chemical and Drug Induced Liver Injury; Cytochromes c; Flow Cytometry; Hepatocytes; Immunoblotting; Melitten; Membrane Potential, Mitochondrial; Mice; Mitochondria, Liver; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-bcl-2; Transforming Growth Factor beta1

Although magnolol is cytoprotective against warm ischemia/reperfusion injury, its effect on cold preservation has not been fully investigated. This study aimed at examining whether magnolol maintains the liver graft integrity after cold preservation and elucidating the underlying mechanisms in terms of apoptotic signaling under both normothermic and hypothermic conditions. After being preserved in Ringer's lactate (RL) at 4 degrees C for 6h ex vivo, the magnolol-treated grafts demonstrated significantly higher AST, ALT, and LDH levels in perfusates than those from negative controls. TUNEL staining showed no difference in the number of apoptotic nuclei in both groups, whereas a more intense apoptotic signal in magnolol-treated grafts was shown as compared with the controls. In vitro data showed no significant difference in viability of RL-preserved clone-9 hepatocytes between the magnolol-treated and control groups, while magnolol pretreatment at 30min before cold preservation prominently induced hepatocyte cell death. RT-PCR and Western blotting analyses revealed a suppression in Bcl-2, but an up-regulation in Bax expression in clone-9 cells after magnolol treatment. Magnolol suppressed the ratios of NF-kappaB to I-kappaBalpha protein contents and I-kappaBalpha phosphorylation induced by TNF-alpha, and potentiated mitochondrial cytochrome c release and subsequent caspase-3 cleavage. Conversely, caspase-3 inhibitor attenuated magnolol-induced hepatotoxicity. We concluded that magnolol could not protect liver grafts from cold ischemia/reperfusion injury. High concentration of magnolol under serum-reduced conditions attenuates NF-kappaB-mediated signaling and induces intrinsic apoptotic pathway, thereby inducing in vitro hepatotoxicity.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Biphenyl Compounds; Blotting, Western; Caspase 3; Chemical and Drug Induced Liver Injury; Cold Temperature; Cryopreservation; Cytochromes c; I-kappa B Proteins; In Situ Nick-End Labeling; Lignans; Liver; Liver Transplantation; Magnolia; Male; Mitochondria; NF-kappa B; Plant Bark; Plant Extracts; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Serum; Signal Transduction; Tumor Necrosis Factor-alpha

The emergence of silymarin (SMN) as a natural remedy for liver diseases, coupled with its entry into NIH clinical trial, signifies its hepatoprotective potential. SMN is noted for its ability to interfere with apoptotic signaling while acting as an antioxidant. This in vivo study was designed to explore the hepatotoxic potential of Doxorubicin (Dox), the well-known cardiotoxin, and in particular whether pre-exposures to SMN can prevent hepatotoxicity by reducing Dox-induced free radical mediated oxidative stress, by modulating expression of apoptotic signaling proteins like Bcl-xL, and by minimizing liver cell death occurring by apoptosis or necrosis. Groups of male ICR mice included Control, Dox alone, SMN alone, and Dox with SMN pre/co-treatment. Control and Dox groups received saline i.p. for 14 days. SMN was administered p.o. for 14 days at 16 mg/kg/day. An approximate LD(50) dose of Dox, 60 mg/kg, was administered i.p. on day 12 to animals receiving saline or SMN. Animals were euthanized 48 h later. Dox alone induced frank liver injury (>50-fold increase in serum ALT) and oxidative stress (>20-fold increase in malondialdehyde [MDA]), as well as direct damage to DNA (>15-fold increase in DNA fragmentation). Coincident genomic damage and oxidative stress influenced genomic stability, reflected in increased PARP activity and p53 expression. Decreases in Bcl-xL protein coupled with enhanced accumulation of cytochrome c in the cytosol accompanied elevated indexes of apoptotic and necrotic cell death. Significantly, SMN exposure reduced Dox hepatotoxicity and associated apoptotic and necrotic cell death. The effects of SMN on Dox were broad, including the ability to modulate changes in both Bcl-xL and p53 expression. In animals treated with SMN, tissue Bcl-xL expression exceeded control values after Dox treatment. Taken together, these results demonstrated that SMN (i) reduced, delayed onset, or prevented toxic effects of Dox which are typically associated with hydroxyl radical production, (ii) performed as an antioxidant limiting oxidative stress, (iii) protected the integrity of the genome, and (iv) antagonized apoptotic and necrotic cell death while increasing antiapoptotic Bcl-xL protein levels and minimizing the leakage of proapoptotic cytochrome c from liver mitochondria. These observations demonstrate the protective actions of SMN in liver, and raise the possibility that such protection may extend to other organs during Dox treatment including the h

Topics: Animals; Antibiotics, Antineoplastic; Antioxidants; Apoptosis; bcl-X Protein; Chemical and Drug Induced Liver Injury; Cytochromes c; Deoxyribonucleases; DNA Fragmentation; Doxorubicin; Male; Mice; Mice, Inbred ICR; Mitochondria, Liver; Oxidative Stress; Poly(ADP-ribose) Polymerases; Silymarin; Tumor Suppressor Protein p53

Pyrrolizidine alkaloid (PA) clivorine, isolated from traditional Chinese medicinal plant Ligularia hodgsonii Hook, has been shown to induce apoptosis in hepatocytes via mitochondrial-mediated apoptotic pathway in our previous research. The present study was designed to observe the protection of N-acetyl-cysteine (NAC) on clivorine-induced hepatocytes apoptosis. Our results showed that 5 mM NAC significantly reversed clivorine-induced cytotoxicity via MTT and Trypan Blue staining assay. DNA apoptotic fragmentation analysis and Western-blot results showed that NAC decreased clivorine-induced apoptotic DNA ladder and caspase-3 activation. Further results showed that NAC inhibited clivorine-induced Bcl-xL decrease, mitochondrial cytochrome c release and caspase-9 activation. Intracellular glutathione (GSH) is an important ubiquitous redox-active reducing sulfhydryl (--SH) tripeptide, and our results showed that clivorine (50 microM) decreased cellular GSH amounts and the ratio of GSH/GSSG in the time-dependent manner, while 5 mM NAC obviously reversed this depletion. Further results showed that GSH synthesis inhibitor BSO augmented clivorine-induced cytotoxicity, while exogenous GSH reversed its cytotoxicity on hepatocytes. Clivorine (50 microM) significantly induced cellular reactive oxygen species (ROS) generation. Further results showed that 50 microM Clivorine decreased glutathione peroxidase (GPx) activity and increased glutathione S transferase (GST) activity, which are both GSH-related antioxidant enzymes. Thioredoxin-1 (Trx) is also a ubiquitous redox-active reducing (--SH) protein, and clivorine (50 microM) decreased cellular expression of Trx in a time-dependent manner, while 5 mM NAC reversed this decrease. Taken together, our results demonstrate that the protection of NAC is major via maintaining cellular reduced environment and thus prevents clivorine-induced mitochondrial-mediated hepatocytes apoptosis.

Topics: Acetylcysteine; Apoptosis; bcl-X Protein; Caspase 3; Caspase 9; Cell Line; Cell Survival; Chemical and Drug Induced Liver Injury; Cytochromes c; Cytotoxins; Dose-Response Relationship, Drug; Enzyme Activation; Glutathione; Glutathione Peroxidase; Glutathione Synthase; Glutathione Transferase; Hepatocytes; Humans; Pyrrolizidine Alkaloids; Reactive Oxygen Species; Thioredoxins

Limitations of existing biomarkers to detect liver injury in experimental animals highlight the need for additional tools to predict human toxicity. The utility of cytochrome c (cyt c) as a biomarker in serum and urine was evaluated in two rodent liver injury models. Adult Sprague-Dawley rats treated with acetaminophen or D-galactosamine (GalN) showed dose- and time-dependent histomorphological changes and TUNEL staining in liver consistent with hepatocellular necrosis, apoptosis and inflammation up to 72 h. Matching changes in serum alanine transaminase (ALT), aspartate transaminase (AST) and cyt c peaked at 24 h for either drug at the highest dose, cyt c falling rapidly at 48 hours with ALT and AST remained high. Intracellular transit of cyt c from mitochondria to the cytoplasm in damaged hepatocytes, and then to peripheral circulation, was observed by immunohistochemistry. Correlation coefficients between cyt c and serum diagnostic tests indicate the liver to be the primary source of cyt c. Urinary analysis for cyt c revealed time-dependent increase at 6 h, peaking at 24 h in GalN-treated rats in contrast with irregular patterns of urinary ALT and AST activity. Histological changes detected at 6 h preceded altered ALT, AST and cyt c at 12 and 18 h, respectively, in GalN-treated rats. These studies demonstrate cyt c to be a useful indicator of hepatic injury in rodents and support its utility as a non-invasive predictor of drug-induced hepatotoxicity, when utilized as a potential urinary biomarker.

Topics: Acetaminophen; Acute Disease; Animals; Apoptosis; Biomarkers; Chemical and Drug Induced Liver Injury; Cytochromes c; Disease Models, Animal; Dose-Response Relationship, Drug; Galactosamine; Hepatocytes; Male; Mitochondria; Necrosis; Rats; Rats, Sprague-Dawley

Liver fibrosis and cirrhosis may be reversible, possibly through the selective clearance of activated hepatic stellate cells/myofibroblasts by apoptosis. Hepatic stellate cells transdifferentiate into myofibroblast-phenotype cells in culture, a process that recapitulates hepatic stellate cell activation in vivo. Bakuchiol, a prenylated phenolic terpene isolated from the seed of Psoralea corylifolia L. (Leguminosae), reduced activated hepatic stellate cells when treated to rats during liver injury recovery period as demonstrated by alpha-smooth muscle actin immunostaining in rat liver and induced apoptosis in activated hepatic stellate cells/myofibroblasts as demonstrated by DNA fragmentation, activation of caspase-3, release of cytochrome c into the cytoplasm, translocation of Bax into mitochondria, and the proteolytic cleavage of poly(ADP-ribose) polymerase (PARP) in vitro. Bakuchiol-induced apoptosis was prevented by z-DEVD-fmk, a specific inhibitor of caspase-3, and z-VAD-fmk, a general caspase inhibitor, suggesting that bakuchiol-induced apoptosis occurs through a caspase-3-dependent pathway in vitro. Bakuchiol treatment stimulated the activation of extracellular signal-regulated kinase 1/2 (ERK), c-Jun NH2-terminal protein kinase (JNK), and p38 mitogen-activated protein kinases (MAPK) in vitro. Pretreatment with SP600125 attenuated the bakuchiol-induced translocation of Bax into mitochondria, cytochrome c release into the cytosol, caspase-3 activation, and PARP cleavage. In contrast, preincubation with SB203580, a p38 MAPK inhibitor, and U0126, an ERK inhibitor, had no effect on bakuchiol-induced cell death and caspase-3 activity. Taken together, these findings indicate that bakuchiol induces caspase-3-dependent apoptosis through the activation of JNK, followed by Bax translocation into mitochondria in rat liver myofibroblasts.

Topics: Actins; Animals; Apoptosis; bcl-2-Associated X Protein; Carbon Tetrachloride; Caspase 3; Cells, Cultured; Chemical and Drug Induced Liver Injury; Cytochromes c; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Activation; Fibroblasts; JNK Mitogen-Activated Protein Kinases; Liver; Liver Diseases; Male; MAP Kinase Signaling System; Mitochondria, Liver; Phenols; Protective Agents; Protein Transport; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Time Factors

The purpose of this study was to examine hepatocyte mitochondrion respiratory chain in rats subjected to ethanol and CCl4 administration within 4 weeks to induce an experimental hepatitis. Oxygen consumption was determined as a measure of mitochondrion respiration chain function. The development of liver pathology was accompanied by fat accumulation, fibrosis, triglycerides and lipid peroxidation increase. Respiratory chain characteristics damage was found. Endogenous oxygen consumption by hepatocytes isolated from pathological liver was found 34% higher compared to control. Exogenous malate and pyruvate substrates delivery didn't stimulate cell respiration. Rotenone (the inhibitor of the I complex) decreased 27% oxygen consumption by pathological hepatocytes while dinitrophenol produced 37% cell respiration increase. States 3 (V3) and 4 (V4) mitochondrial respiration with malate + glutamate as substrates were found to be 70 and 56% higher accordingly compared to control level. V3 and Vd (dinitrophenol respiration) for mitochondria from pathological liver didn't differ from control when being tested with malate + glutamate or succinate as substrates. Cytochrome c oxidase activity increased (+ 80%) as compared to control. Administration of hypolipidemic agent simvastatin simultaneously with ethanol and CC14 resulted in decrease liver fat accumulation, fibrosis and peroxidation products. Simvastatin administration caused hepatocyte endogenous respiration decrease while malate + pyruvate, dinitrophenol or rotenone delivery produced oxygen consumption alterations similar to control. However, when isolated mitochondria from liver of simvastatin treated animals being tested the decrease of oxidative phosphorylation coupling for substrates malate + glutamate was found. While simvastatin did not cause changes in cytochrome c oxidase activity. We propose the hypothesis that the NCCR complex in rat mitochondria with experimental toxic hepatitis works extensively on superoxydanion production. Alterations of SCCR, Coenzyme Q-cytochrome c-reductase, cytochrome c oxidase and ATP-synthase activities have an adaptive nature to compensate for impaired NCCR function.

Topics: Animals; Carbon Tetrachloride; Chemical and Drug Induced Liver Injury; Cytochromes c; Dinitrophenols; Electron Transport; Ethanol; Hepatocytes; Malates; Male; Mitochondria; Oxygen Consumption; Pyruvic Acid; Rats; Rats, Wistar; Rotenone; Simvastatin

This study was carried out to evaluate the hepatoprotective activity of glycoprotein isolated from the stems of Ulmus davidiana Nakai (UDN), which has been used as an anti-inflammatory agent in folk medicine. We evaluated lipid peroxidation in glucose/glucose oxidase (G/GO)-induced BNL CL.2 cells and measured thiobarbituric acid reactive substances (TBARS), lactate dehydrogenase (LDH), nitric oxide (NO), antioxidant enzyme (superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx)), activity of cytotoxic-related signals (hepatic cytochrome c, nuclear factor-kappa B (NF-kappaB) and activator protein-1 (AP-1)) and levels of plasma lipids (triglyceride (TG) and total cholesterol (TC)) in carbon tetrachloride (CCl(4,) 1.0 mL kg(-1))-induced A/J mouse. The results in G/GO-induced BNL CL.2 cells showed that UDN glycoprotein had a dose-dependent inhibitory effect on lipid peroxidation. The results in carbon tetrachloride (CCl(4,) 1.0 mL kg(-1))-induced A/J mouse indicated that treatment with UDN glycoprotein (40 mg kg -1) lowered LDH activity and TBARS formation, and increased NO production and antioxidant enzymes activity, compared with control. Also, our finding from CCl(4)-treated mice after pretreatment with UDN glycoprotein demonstrated that the activity of cytotoxic-related signals decreased but the levels of plasma lipids increased, compared with CCl(4) treatment alone. Here, we speculate that UDN glycoprotein has a protective character to CCl(4)-induced mouse liver injury.

Topics: Animals; Antioxidants; Carbon Tetrachloride; Catalase; Cell Line; Chemical and Drug Induced Liver Injury; Cytochromes c; Glutathione Peroxidase; Glycoproteins; Lipid Peroxidation; Lipids; Liver; Liver Diseases; Male; Mice; Mice, Inbred Strains; NF-kappa B; Plant Extracts; Protective Agents; Superoxide Dismutase; Transcription Factor AP-1; Ulmus

Asiatic acid (AA) is one of the triterpenoid components of Terminalia catappa L., which has antioxidative, anti-inflammatory and hepatoprotective activity. This research focused on the mitochondrial protection of AA against acute liver injury induced by lipopolysaccharide (LPS) and D-galactosamine (D-GalN) in mice. It was found that pretreatment with 25, 50 or 100 mg kg(-1) AA significantly blocked the LPS + D-GalN-induced increase in both serum aspartate aminotransferase (sAST) and serum alanine aminotransferase (sALT) levels, which was confirmed by ultrastructural observation under an electron microscope, showing improved nuclear condensation, ameliorated mitochondrion proliferation and less lipid deposition. Meanwhile, different doses of AA could decrease both the transcription and the translation level of voltage-dependent anion channels (VDACs), the most important mitochondrial PTP component protein, and block the translocation of cytochrome c from mitochondria to cytosol. On the other hand, pre-incubation with 25, 50 and 100 microg mL(-1) AA inhibited the Ca(2+)-induced mitochondrial permeability transition (MPT), including mitochondrial swelling, membrane potential dissipation and releasing of matrix Ca(2+) in liver mitochondria separated from normal mice, indicating the direct role of AA on mitochondria. Collectively, the above data suggest that AA could protect liver from damage and the mechanism might be related to up-regulating mitochondrial VDACs and inhibiting the process of MPT.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Calcium; Chemical and Drug Induced Liver Injury; Cytochromes c; Galactosamine; Ion Channels; Lipopolysaccharides; Liver; Male; Membrane Potentials; Mice; Mice, Inbred ICR; Mitochondria, Liver; Mitochondrial Membrane Transport Proteins; Mitochondrial Permeability Transition Pore; Mitochondrial Swelling; Pentacyclic Triterpenes; RNA, Messenger; Triterpenes; Voltage-Dependent Anion Channels

Liver cirrhosis is often preceded by overt signs of hepatitis, including parenchymal cell inflammation and infiltration of polymorphonuclear (PMN) leukocytes. Activated PMNs release both reactive oxygen species and reactive halogen species, including hypochlorous acid (HOCl), which are known to be significantly cytotoxic due to their oxidizing potential. Because the role of mitochondria in the hepatotoxicity attributed to HOCl has not been elucidated, we investigated the effects of HOCl on mitochondrial function in the human hepatoma HepG2 cell line, human fetal liver cells, and isolated rat liver mitochondria. We show here that HOCl induced mitochondrial dysfunction, and apoptosis was dependent on the induction of the mitochondrial permeability transition (MPT), because HOCl induced mitochondrial swelling and collapse of the mitochondrial membrane potential with the concomitant release of cytochrome c. These biochemical events were inhibited by the classical MPT inhibitor cyclosporin A (CSA). Cell death induced by HOCl exhibited several classical hallmarks of apoptosis, including annexin V labeling, caspase activation, chromatin condensation, and cell body shrinkage. The induction of apoptosis by HOCl was further supported by the finding that CSA and caspase inhibitors prevented cell death. For the first time, these results show that HOCl activates the MPT, which leads to the induction of apoptosis and provides a novel insight into the mechanisms of HOCl-mediated cell death at sites of chronic inflammation.

Topics: Animals; Apoptosis; Bongkrekic Acid; Carcinoma, Hepatocellular; Caspase 3; Caspase 7; Caspases; Cell Line; Cell Line, Tumor; Cell Survival; Chemical and Drug Induced Liver Injury; Cyclosporine; Cytochromes c; Humans; Hypochlorous Acid; Liver; Membrane Potentials; Mitochondria, Liver; Mitochondrial Swelling; Permeability; Rats

Topics: Amino Acids; Carbon Tetrachloride; Carbon Tetrachloride Poisoning; Chemical and Drug Induced Liver Injury; Cytochromes; Cytochromes c; Electrons; Hepatitis; Liver; Microscopy; Microscopy, Electron; Mitochondria; Mitochondrial Proteins; Rats; Research; Spectrum Analysis