OBJECTIVE:To study the effects of domoic acid (DA) on membrane function of primary cultured rat glial cell.METHODS:After the glial cells were treated with 6.4 x 10(-2), 6.4 x 10(-3) and 6.4 x 10(-4) micromol/L DA for 24 h, the activities of Na(+)-K(+)-ATPase and Ca(2+)-Mg(2+)-ATPase, the membrane fluidity and the permeability were measured to reflect the membrane function.RESULTS:After treatment of DA for 24 h, the activities of Na(+)-K(+)-ATPase and Ca(2+)-Mg(2+)-ATPase were inhibited significantly, the membrane fluidity decreased and the membrane permeability increased. The fluorescence polarization and microviscosity in the low, middle and high dosage treatment groups were 0.0626 +/- 0.0051, 0.0685 +/- 0.0097, 0.0648 +/- 0.0086 and 0.3154 +/- 0.0298, 0.3510 +/- 0.0571, 0.3286 +/- 0.0504 respectively, compared with the control group (0.0481 +/- 0.0069 and 0.2338 +/- 0.0372) (P < 0.01).CONCLUSION:DA has obvious effects on membrane function of rat glial cells and may cause further injury on the cells.

Toosendanin (Tsn), a triterpenoid extracted from Melia toosendan Sieb et Zucc, possesses different pharmacological effects in human and important values in agriculture. However, liver injury has been reported when toosendanin or Melia-family plants, which contain toosendanin are applied. The mechanism by which toosendanin induces liver injury remains largely unknown. Here we reported that toosendanin induced primary rat hepatocytes death by mitochondrial dysfunction and caspase activation. Toosendanin led to decrease of mitochondrial membrane potential, fall in intracellular ATP level, release of cytochrome c to cytoplasm, activation of caspase-8, 9, and 3 and ultimately cell death. Level of reactive oxygen species (ROS) was also increased in hepatocytes after incubation with toosendanin. Catalase, the H2O2-decomposing enzyme, can prevent the reduction in ATP level and protect hepatocytes from toosendanin-induced death. The ERK1/2 (p44/42 MAP kinases) and JNK (c-Jun N-terminal kinase) were activated, but p38 MAPK was not activated by toosendanin. Inhibition of ERK1/2 activation sensitized hepatocytes to death and increased activity of caspase-9 and 3 in response to toosendanin. Inhibition of JNK attenuated toosendanin-induced cell death. These results suggested that toosendanin causes death of primary rat hepatocytes by mitochondrial dysfunction and caspase activation. Generation of ROS and MAP kinases activation might be involved in this process.

Aristolochic acid (AA), a natural nephrotoxin and carcinogen, can induce a progressive tubulointerstitial nephropathy. However, the mechanism by which AA causes renal injury remains largely unknown. Here we reported that the mitochondrial permeability transition (MPT) plays an important role in the renal injury induced by aristolochic acid I (AAI). We found that in the presence of Ca(2+), AAI caused mitochondrial swelling, leakage of Ca(2+), membrane depolarization, and release of cytochrome c in isolated kidney mitochondria. These alterations were suppressed by cyclosporin A (CsA), an agent known to inhibit MPT. Culture of HK-2 cell, a human renal tubular epithelial cell line for 24 h with AAI caused a decrease in cellular ATP, mitochondrial membrane depolarization, cytochrome c release, and increase of caspase 3 activity. These toxic effects of AAI were attenuated by CsA and bongkrekic acid (BA), another specific MPT inhibitor. Furthermore, AAI greatly inhibited the activity of mitochondrial adenine nucleotide translocator (ANT) in isolated mitochondria. We suggested that ANT may mediate, at least in part, the AAI-induced MPT. Taken together, these results suggested that MPT plays a critical role in the pathogenesis of HK-2 cell injury induced by AAI and implied that MPT might contribute to human nephrotoxicity of aristolochic acid.

Background: Baicalin (BA) exhibits an anti-inflammatory effect in vivo and in vitro and is used to treat chronic hepatitis. However, the mechanism by which BA exerts the liver-protective effect remains largely unknown. Aims: The present study reports that BA inhibits cytokine production and hepatocyte apoptosis to protect mice from liver injury induced by concanavalin A (Con A), a T-cell-dependent liver injury model. Results: Con A injection of mice induced severe immune responses and extensive hepatocellular apoptosis within 24 h. Pretreatment of 200 or 100 mg/kg BA markedly reduced serum aminotransferase activities, protected hepatoycte apoptosis and reduced the increase of plasma cytokine levels, including tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma) and interleukin-6 (IL-6). Furthermore, BA pretreatment decreased tissue myeloperoxidase activity and lipid peroxidation, but increased the superoxide dismutase level. In vitro studies indicated that the beneficial effect of BA was associated with reduced cytokine production from lymphocytes and reduced TNF-alpha-induced hepatocyte apoptosis. Conclusion: These results suggest that BA has therapeutic potential for T-cell-mediated liver injury.

Deltamethrin is a type II synthetic pyrethroid insecticide and is used extensively. Numerous studies have demonstrated that deltamethrin can cause severe central nervous system symptoms. The mechanism of neuro‐toxicity caused by deltamethrin is still poorly understood. This study investigated the effect of deltamethrin on the mitochondria‐mediated apoptosis pathway in rat brain. The cortex and hippocampus mitochondrial membrane potential, permeability transition, expression of cytochrome c and activity of cytochrome c oxidase was assayed at 5, 24, 48 and 72 h following deltamethrin treatment. The study observed that the membrane potential, change of absorbance at 540 nm (A 540 ), intensities of cytochrome c in mitochondria and activity of cytochrome c oxidase in the cortex and hippocampus of treated groups were significantly lower than those of the control group. The results of the study demonstrated that deltamethrin induced a decreased mitochondrial membrane potential and increased permeability, and reduced the expression of cytochrome c, thus depressing the activity of cytochrome c oxidase significantly. It indicates that deltamethrin may have an effect on mitochondria‐mediated apoptosis of nerve cells in the rat brain. Copyright © 2007 John Wiley & Sons, Ltd.

Aim: Angiotensin II is believed to play an important role in tissue repair and remodeling in lungs by the angiotensin type I (AT1) receptor via a number of potential mechanisms. However, the role of the AT1 receptor in early lung injury has not been characterized. Methods: Bleomycin-induced pulmonary fibrosis (PF) in rats was utilized to value the treatment with valsartan, an AT1 receptor antagonist, by measurement of body weight, wet weight of the left lung, hydroxyproline content, mRNA expression of collagen I/III, and the degree of fibrosis in lung tissues on d 21. Tissue injury in the early phase was assessed on d 1, 3 and 7 by apoptosis, malondialdehyde content, myeloperoxidase activity, inflammatory cell count and protein content. Angiotensin converting enzyme (ACE) activity and the AT1 receptor in lung tissues were analyzed by biochemistry method and Western blotting, respectively. Results: Valsartan ameliorated PF induced by bleomycin in the rats on d 21. After bleomycin was injected intratracheally, increases in the lung AT1 receptor and ACE activity were observed by d 1, 3 and 7. Lung injury deteriorated in the early phase. Valsartan reduced the increase of the AT1 receptor, ACE activity and lung injury induced by bleomycin in the early phase. Conclusion: These observations suggest that angiotensin II may play a potent role in early lung injury via the AT1 receptor. AT1 receptor antagonists should be assessed as potential new therapies for fibrotic lung disease.

After comparing mortality and clinical signs in rats receiving different dexamethasone treatments, we investigated whether 0.5 mg/kg/d dexamethasone could delay pulmonary fibrosis induced by bleomycin and its time course (1, 3, 7, 14, 21 and 28 days). Tissue injury was assessed by apoptosis, lactate dehydrogenase (LDH) release, malondialdehyde content, and protein content; and inflammation was measured in terms of myeloperoxidase (MPO) activity, inflammatory cell count, and the mRNA expression of pro/inflammatory cytokines. Fibrogenic activity was analyzed by measuring the mRNA expression of fibrotic cytokines in tissue, and the promotion of fibroproliferation and synthesis of collagen type I by bronchoalveolar lavage fluids in vitro; and fibrosis was assessed by measuring the hydroxyproline content and collagen-I mRNA expression, and by histology. Bleomycin treatment induced tissue injury, inflammation and fibrogenic activity in lung, and led to fibrosis. Treatment with dexamethasone diminished the extent of fibrosis by strongly reducing inflammation, lung damage, and fibrogenic activity. These results demonstrate that the progression of bleomycin-induced pulmonary fibrosis in rats can be delayed by dexamethasone treatment, which appeared to alleviate not only inflammation but also lung damage and fibrogenic activity, indicating a possible new role for dexamethasone in the treatment of fibrosis.

OBJECTIVE:To study the effects of deltamethrin (DM) on the permeability of mitochondrial membrane and the expression of cytochrome C in brain tissue of rats. METHODS:Wistar rats were randomizedly divided into five groups (including four treated groups and one control group). In the treated groups, DM of 12.5 mg/kg was administered intraperitoneally once in rats and the rats were sacrificed 5, 24, 48 and 72 hours later while in the control group, the salad oil of 5 mg/kg was administered intraperitoneally once. The mitochondria in brain tissue of rats were extracted to measure the membrane permeability and the activity of cytochrome C oxidase as well as the expression of cytochrome C in cortex and hippocampus. RESULTS:After the treatment the permeability of mitochondrial membrane was significantly increased in the treated groups compared with the control group. The expression of cytochrome C was increased in cortex and hippocampus CA1 and CA2 5 h, 24 h and 48 h groups and CA4 24 h group (0.57 +/- 0.04, 0.67 +/- 0.09, 0.58 +/- 0.04) and (0.81 +/- 0.18) (P < 0.05 or P < 0.01) while there was no significant difference in the expression of cytochrome C in cortex and hippocampus CA2 72 h group and CA3 and CA4 5 h, 48 h and 72 h groups between the treated groups and the control group (P > 0.05). The activity of cytochrome C oxidase was inhibited (P < 0.01). CONCLUSION:Deltamethrin can significantly increase the permeability of mitochondrial membrane and the expression of cytochrome C in brain tissue of rats.

Correlation of cytochrome P450 (CYPS) with preneo plastic changes in the early stage of hepatocarcinogenesis is still unclear. To detect the expression of carcinogen-metabolizing related microsomal P450 enzymes, namely the CYP1A1, CYP1A2, CYP2B1/2, CYP2E1, and CYP3A, we performed the medium-term bioas-say of Ito's model in Sprague-Dawley rats. The amount and activity of CYP were assessed by biochemical and immunohistochemical methods in week 8. The correlation between CYP expression and microsomal oxidative stress was investigated by comparing the generation of microsomal lipid peroxidation in the presence or absence of specific CYP inhibitor. In the DEN-2-AAF and 2-AAF alone groups, the expression of CYP1A1 and CYP2E1 were up-regulated and the expression of CYP2B1/2 and CYP1A2 were quite the contrary. Strong staining of CYP2E1 and CYP2B1/2 was found around the centro lobular vein and weak staining in the altered hepatic foci revealed by immunohistochemical procedure. There was no significant change in the activity of CYP3 A among the 4 groups. Altered hepatic tissue bore more microsomal NADPH (nicotinamide adenine dinucleotide phosphate, reduced form)-dependent lipid peroxidation than normal tissue. And the difference among the 4 groups disappeared when CYP2E1 was inhibited. More microsomal lipid peroxidation was generated when incubated with CYP1A inhibitor α-naphthoflavone. CYP altered their expression levels and these alterations can play important roles in the alteration of cell redox status of preneoplastic tissue in the early stage of hepato carcinogenesis.

目的应用短期大鼠致肝癌模型,探讨肝脏细胞色素P450酶(CYPs)在早期癌变过程中的作用,为药物快速致癌毒性检测提供致癌机制的理论基础.

目的黄芩甙是一种临床上常用的治疗急慢性肝炎的药物,为了研究其对肝损伤的保护作用及机制,采用刀豆蛋白A(ConA)引起的小鼠急性肝损伤模型及相关的体外实验进行研究.

目的探讨博来霉素(bleomycin,BLM)致大鼠肺纤维化机制. 方法Ⅰ体内试验:雄性SD大鼠,体重220～250 g,60只,随机分成对照组和BLM组(每组50只),用一次性气管内滴注BLM(5 mg/kg,2 mL/kg)的方法复制大鼠肺纤维化模型,对照组滴注生理盐水.

Aim: Correlation of cytochrome P450 (CYPS) with preneo plastic changes in the early stage of hepato carcinogenesis is still unclear. To detect the expression of carcinogen-metabolizing related microsomal P450 enzymes, namely the CYP1A1, CYP1A2, CYP2B1/2, CYP2E1, and CYP3 A, we performed the medium-term bioas-say of Ito's model in Sprague-Dawley rats. Methods: The amount and activity of CYP were assessed by biochemical and immunohistochemical methods in week 8. The correlation between CYP expression and microsomal oxidative stress was investigated by comparing the generation of microsomal lipid peroxidation in the presence or absence of specific CYP inhibitor. Results: In the DEN-2-AAF and 2-AAF alone groups, the expression of CYP1 A1 and CYP2E1 were up-regulated and the expression of CYP2B1/2 and CYP 1A2 were quite the contrary. Strong staining of CYP2E1 and CYP2B1/2 was found around the centro lobular vein and weak staining in the altered hepatic foci revealed by immunohistochemical procedure. There was no significant change in the activity of CYP3 A among the 4 groups. Altered hepatic tissue bore more microsomal NADPH (nicotinamide adenine dinucleotide phosphate, reduced form)-dependent lipid peroxidation than normal tissue. And the difference among the 4 groups disappeared when CYP2E1 was inhibited. More microsomal lipid peroxidation was generated when incubated with CYP1A inhibitor α-naphthoflavone. Conclusion: CYP altered their expression levels and these alterations can play important roles in the alteration of cell redox status of preneoplastic tissue in the early stage of hepato carcinogenesis.

Tetrandrine, a bisbenylisoquinoline alkaloid isolated from the dried root of Stephenia tetrandra (S Moore), possesses a remarkable pharmacological profile. However, the mechanisms of tetrandrine hepatotoxicity remain to be elucidated. In this study, we first proved apoptosis and mitochondrial dysfunction induced by tetrandrine in Sprague–Dawley rat liver in vivo. By further assuming apoptosis as an important mechanism in tetrandrine-induced hepatotoxicity, we focused on mitochondria-initiated apoptosis in primary hepatocytes isolated from Sprague–Dawley male rats. Tetrandrine treatment led to significant release of cytochrome c and downregulation of Bcl-XL accompanied by caspase 3 activation, and ultimately, DNA fragmentation. Caspase 3 activation was markedly inhibited by cyclosporin A (CsA) and Ac-DEVD-CHO. Furthermore, Endo G, a caspase-independent apoptotic protein, was detected for its expression and DNase activity. CsA blocked the release both of Endo G and cytochrome c significantly. Additionally, the generation of reactive oxygen species (ROS) increased in a time-dependent manner corresponding with a fall in intracellular GSH content after 10μM tetrandrine treatment in 4h. Tetrandrine also induced mitochondrial dysfunction indicated by transition of mitochondrial transmembrane potential and decrease of intracellular ATP level. The findings indicated that the caspase-dependent mitochondrial apoptosis pathway was primarily involved in tetrandrine-induced apoptosis in rat primary hepatocytes. In addition, a caspase-independent pathway indicated by Endo G also contributed to apoptosis caused by tetrandrine. Meanwhile, ROS was proved an important inducer in this apoptosis process.

化学致癌物质是肝癌病因学中的重要因素,用二乙基亚硝胺(DEN)、苯巴比妥(PB)、二乙酰胺基芴(2-AAF)等已知的致癌物建立的各种致癌模型,目前被广泛应用于评价化合物的潜在致癌性及癌变过程的机制研究.本文介绍了包括中期致癌模型、抵抗性肝细胞(resistant hepatocyte,RH)模型、转基因动物致癌模型、中期致肝癌模型及多器官致癌模型等常用模型有其各自的特点及用途.

The current therapeutic approaches for pulmonary fibrosis, which is characterized by fibroblast proliferation and extracellular matrix remodeling, are unsatisfactory. Feitai, consisting of several herbs, is a folk formula for pulmonary tuberculosis therapy in China. To investigate the effects of Feitai on pulmonary fibrosis, Feitai was administered orally to bleomycin (BLM)-treated rats, and the lung toxicity effects were evaluated according to inflammatory cell count, protein concentration, and lactate dehydrogenase (LDH) activity in the bronchoalveolar lavage fluid (BALF), malondialdehyde level and hydroxyproline content in lung tissue 28 days post-BLM. Serial sections of the lung were stained with hematoxylin and eosin (HE) and Masson trichrome, respectively. The degree of fibrosis was assessed quantitatively using LEICA QWin image analyzer. Results showed that Feitai inhibited BLM-induced lung fibrotic lesions in a dose-dependent manner as reflected by decreased the lung hydroxyproline content and lung fibrosis fraction 28 days after BLM instillation. Treatment with Feitai also significantly ameliorated the BLM-induced lung toxicity effects detected in BALF and lung tissue. The effects in vitro on WI-38 human lung fibroblast cell line showed that Feitai significantly reduced the cell proliferation and transforming growth factor (TGF)–β stimulated type I collagen synthesis. These results strongly demonstrate that Feitai may be useful in the treatment of pulmonary fibrosis.

Pulmonary fibrosis is a common consequence of numerous pulmonary diseases. The current therapeutic approaches for this condition are unsatisfactory. Feitai, a composite formula consisting of several herbs, is used in China as a folk remedy for treating patients with pulmonary tuberculosis. In this study, we extensively investigate the effects and mechanisms of Feitai on bleomycin (BLM)-induced pulmonary fibrosis in rats. One hundred and twenty male Sprague-Dawley rats were randomly divided into four groups, referred to as the saline-water, saline-Feitai, BLM-water, and BLM-Feitai groups. Following a single instillation of BLM (5 mg/kg) or saline, rats were orally administered Feitai at a dose of 3 g/kg body weight or sterilized distilled water once daily. Rats were killed at 7, 14, or 28 d post-BLM. Inflammatory cell count, protein concentration, and lactate dehydrogenase activity in bronchoalveolar lavage fluid were measured, and myeloperoxidase activity and lipid peroxide content in lung homogenates were analyzed. Treatment with Feitai inhibited lung fibrotic progression induced by BLM, as indicated by the decrease in lung hydroproline content and lung fibrosis score at 28 d post-BLM. This was accompanied by significant amelioration of BLM-induced body weight loss, lung edema, and inflammatory response during the development of lung injury in the acute phase. The results strongly indicate the beneficial effects of Feitai in protecting against BLM-induced pulmonary fibrosis. Furthermore, the inflammatory response and lipid peroxidation were inhibited by Feitai, suggesting that the effect of this formula on BLM-induced lung injury and fibrosis is associated with antiinflammatory and antioxidant properties.