养血生发胶囊对小鼠肝毒性"量-时-毒"关系的病理学研究

目的研究养血生发胶囊单次给药对小鼠造成的肝毒性在病理学上的损伤程度,为养血生发胶囊肝毒性损伤提供病理组织学依据。方法采用常规HE染色方法,通过光镜下阅片观察和病理图像分析,观察养血生发胶囊单次灌胃对小鼠肝毒性量-时-毒关系研究中的病理学变化。结果 时-毒关系研究:给药后2h检测发现,在给药后6～24h表现较明显。量-毒关系研究:在2.50~12.00g.kg-1之间出现不同程度的病理组织学损伤,本品导致的主要病理改变为:肝细胞嗜酸性变、玻璃样变、脂肪变、空泡变等。结论单次给小鼠灌胃不同剂量的养血生发胶囊可造成急性肝损伤,并呈现一定的量-时-毒关系,并可导致肝细胞出现嗜酸性变、玻璃样变、脂肪变、空泡变等病理学改变。     

益肾乌发口服液对小鼠急性肝毒性"量-时-毒"关系研究

目的 研究益肾乌发口服液单次给药致小鼠急性肝毒性的"量-时-毒"关系.方法 "时-毒"关系研究:小鼠灌胃一定剂量的益肾乌发口服液,分别于给药后不同时间检测血清丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、总胆红素(TBI)水平及肝、脾、胸腺等脏器指数的变化."量-毒"关系研究:给小鼠灌胃不同剂量的益肾乌发口服液,于给药后2h按"时-毒"研究方法对小鼠进行相应处理.结果 小鼠血清ALT、AST、TBI水平均在灌胃较高剂量的益肾乌发口服液后2h达到高峰,给药后12h肝脏指数升至最高,给药后72h可恢复至正常值;剂量在(4.46～25)mL·kg-1之间肝脏可产生明显病理损伤,且随着剂量增大,ALT、AST、TBI水平升高显著.结论 小鼠单次灌胃一定剂量的益肾乌发口服液可造成急性肝毒性损伤,并呈现明显的"量-时-毒"关系.     

半夏水提组分对小鼠肝毒性"量-时-毒"关系研究

目的 考察半夏水提组分单次给药小鼠肝毒性"量-时-毒"关系.方法 单次给药"时-毒"关系研究:制备62.5 g·kg-1半夏水提组分,按25 mL·kg-1给小鼠灌胃给药1次,给药后按处理时间点依次为0(即空白对照组),0.5,1,2,4,8,12,24,48,72h,于相应时间点取血,测定血清丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST),取肝组织,常规染色,光学显微镜下观察其组织形态学变化;单次给药"量-毒"关系研究:设定半夏水提组分82.5 g·kg-1、70.1 g·kg-1、59.6g·kg-1、50.7 g·kg-1、43.1 g·kg-1、36.6g·kg-16个剂量组,小鼠灌胃给药1次,空白组给蒸馏水,于给药后4h按上述方法测定血清ALT、AST及肝组织镜检.结果 半夏肝毒性"时-毒"关系的研究显示小鼠单次灌胃62.5 g·kg-1的半夏水提组分血清ALT、AST值随时间的不同造成的肝损害的程度也不同,毒性高峰出现在给药后4h,持续约48h.肝组织病理形态学检查显示,半夏水提组分在给药后4h对肝组织产生明显损伤.半夏肝毒性"量-毒"关系的研究显示小鼠单次灌胃较高剂量半夏水提组分可使血清ALT、AST值显著升高.肝组织病理形态学检查显示给予较高剂量的半夏水提组分后,肝细胞可出现不同程度的水肿、脂肪变性以及部分点状坏死,而低剂量给小鼠灌胃,肝组织病理形态学未见异常改变.结论 单次给予半夏水提组分可造成小鼠肝毒性损伤,并呈现一定的"量-时-毒"关系.     

养血生发胶囊对小鼠肝毒性"量-时-毒"关系研究

目的 研究养血生发胶囊致小鼠肝毒性的"量-时-毒"关系.方法 "时-毒"关系研究:取小鼠按不同时间点分组,单次灌胃给予一定剂量养血生发胶囊,观察给药后小鼠毒性反应,分别于药后不同时间检测血清丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、总胆红素(TBI),计算肝、肾、脾、胸腺脏器指数."量-毒"关系研究:取小鼠按不同剂量分组,单次灌胃给予不同剂量的养血生发胶囊分别于给药后2h按"时-毒"研究方法对小鼠进行相应处理.结果 小鼠灌胃较高剂量养血生发胶囊后,血清ALT,AST活力在2h达到高峰,持续时间均约达8h;给药后12h肝脏指数升至最高,药后48h可恢复至正常值.养血生发胶囊剂量在(2.50～12.00)g·kg-1之间、对肝组织产生明显损伤,且随着剂量增大,ALT、AST、TBI升高显著.结论 小鼠单次灌胃给予一定剂量的养血生发胶囊可造成急性肝损伤,并呈现一定的"量-时-毒"关系.关于其肝脏损伤的机制有待进一步研究.     

山豆根不同组分多次给药对小鼠肝毒性"量-时-毒"关系研究

目的 研究山豆根不同组分多次给药致小鼠肝毒性的"量-时-毒"关系.方法 连续7天给小鼠灌胃不同剂量的山豆根水提、醇提组分,观察小鼠一般状况,分别于给药后第1、3、7天检测小鼠血清丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、碱性磷酸酶(AKP)的活性和总胆红素(TBI)、白蛋白(ALB)的含量,计算肝脏指数,并观察肝组织形态变化.结果 在给药后第1天山豆根水提、醇提组分低剂量组未对小鼠肝脏造成明显损伤,高、中剂量组会使小鼠血清中ALT、AST、AKP活性升高,TBI含量升高,ALB含量下降;给药后第3天山豆根水提、醇提各剂量组小鼠出现烦躁,体重增长缓慢等症状,血清中上述肝功能指标变化明显,肝体比值增大,并见部分肝细胞脂肪变性、气球样变,点状坏死等病理变化;给药后第7天观察上述症状依次加重.水提组分对上述指标的影响比醇提组分明显.结论 多次给小鼠高、中剂量的山豆根水提组分和醇提组分均可造成明显的肝损伤,且水提组分的肝毒性大于醇提组分,并呈现明显的肝毒性"量-时-毒"关系.     

半夏酸水渗漉液单次给药对小鼠肝毒性"量-时-毒"关系研究

目的 考察半夏酸水渗漉提取样品单次给药对小鼠肝毒性"量-时-毒"关系.方法 单次给药"时-毒"关系研究:制备1.77 g·kg-1半夏酸水渗漉液,取小鼠按不同时间点分组,25 mL·kg-1给小鼠灌胃给药1次,观察给药后小鼠死亡情况和毒性反应,分别于给药后相应时间点取血,测定血清丙氨酸氨基转移酶(ATL)、天门冬氨酸氨基转移酶(AST)水平,计算肝体比值,另取肝组织,常规染色,光学显微镜下观察其组织形态学变化;单次给药"量-毒"关系研究:取小鼠按不同剂量分组,单次灌胃给予不同剂量的半夏酸水渗漉液2.68g·kg-1、2.14g·kg-1、1.72g·kg-1、1.37g·kg-1、1.10 g·kg-1、0.88 g·kg-1,空白组给同体积蒸馏水,于给药后2h按"时-毒"研究方法对小鼠进行相应处理.结果 半夏肝毒性"时-毒"关系的研究显示小鼠单次灌胃1.77 g·kg-1 的半夏酸水渗漉液,血清ALT、AST.值随时间的不同造成的肝损害的程度也不同,毒性高峰出现在给药后2h,持续时间约达72h.肝组织病理形态学检查显示,半夏酸水渗漉液在给药后2～4h对肝组织产生明显损伤.半夏肝毒性"量-毒"关系的研究显示小鼠单次灌胃较高剂量半夏酸水渗漉液对肝组织产生明显损伤,且随着剂量的增大,可使血清ALT、AST值显著升高.肝组织病理形态学检查显示给予较高剂量的半夏酸水渗漉液后,可致肝细胞部分出现灶状坏死,而低剂量给小鼠灌胃,肝组织病理形态学未见异常改变.结论 单次给予较高剂量的半夏酸水渗漉液可造成小鼠急性肝损伤,毒性出现早、持续时间长,具体表现为血清ALT、AST值升高甚或肝组织病理形态学的改变,且呈明显的时毒、量毒关系.     

山豆根不同组分对小鼠急性肝毒性“量-时-毒”关系研究

目的研究山豆根水提组分及醇提组分单次给药致小鼠急性肝毒性的＂量-时-毒＂关系。方法 ＂时-毒＂关系研究：小鼠灌胃一定剂量的山豆根水提、醇提组分,分别于给药后不同时间检测血清丙氨酸氨基转移酶（ALT）、门冬氨酸氨基转移酶（AST）水平及肝、脾、胸腺等脏器指数的变化,光学显微镜下进行肝脏病理学检查。＂量-毒＂关系研究：给小鼠灌胃不同剂量的山豆根水提、醇提组分,分别于药后6h、4h按＂时-毒＂研究方法对小鼠进行相应处理。结果小鼠血清ALT、AST水平分别在灌胃较高剂量的山豆根水提、醇提组分后6h、4h达到高峰,给药后12h肝脏指数均升至最高,给药后48h均可恢复至正常值;山豆根水提组分剂量在（1.789～3.494）g·kg^-1之间、醇提组分在（2.779～5.427）g·kg^-1之间,肝脏可产生明显病理损伤,且随着剂量增大ALT、AST水平升高显著。光学显微镜下观察小鼠肝细胞出现不同程度的核固缩、变性等病理改变。结论小鼠单次灌胃一定剂量的山豆根水提组分或醇提组分可造成急性肝毒性损伤,并呈现明显的＂量-时-毒＂关系。     

北豆根不同组分单次给药对小鼠肝毒性“量-时-毒”关系研究

目的观察北豆根不同组分单次给药对小鼠肝毒性"量-时-毒"关系的影响。方法取小鼠按不同时间点或不同剂量分组,观察给药后小鼠死亡情况和毒性反应,分别于药后不同时间检测血清谷丙转氨酶(ALT)、谷草转氨酶(AST),计算肝脏指数,光镜下观察肝组织的病理变化。结果小鼠灌胃较高剂量的北豆根水提组分和醇提组分后,血清ALT、AST活力在4 h达到高峰,持续时间均约达48 h;水提组分给药6 h后即可出现肝脏明显肿大,肝脏指数升高,其中24 h肝脏指数升高最为明显;光镜下观察北豆根水提组分6h后,肝小叶结构尚还正常,个别细胞变性坏死;12h后大部分细胞核固缩,出现广泛的气球样变、玻璃样变、嗜酸性变性等,有假小叶形成;72h后肝细胞形态基本恢复正常,仍有个别细胞核固缩;醇提组分给药4 h后肝脏明显肿大,肝脏指数升高,其中24 h肝脏指数升高最为明显,光镜下观察北豆根醇组分6 h后,肝小叶结构尚还正常,部分肝细胞核固缩;12 h后,大部分细胞核固缩,间质细胞变性坏死;72h后部分肝细胞变性坏死。结论北豆根水提组分剂量在46.05~61.4g.kg-1之间、醇提组分在8.45~15.02g.kg-1之间对肝组织产生明显损伤,光镜下观察出现不同程度的肝细胞损伤;且随着剂量增大,ALT、AST升高显著,病理损伤更严重。小鼠单次灌胃给予一定剂量的北豆根水提组分或醇提组分可造成急性肝损伤,并呈现一定的"量-时-毒"关系。关于其肝脏损伤的机制有待进一步研究。     

益肾乌发口服液对小鼠急性毒性实验研究

目的 探讨益肾乌发口服液对小鼠急性毒性的影响.方法 采用经典的急性毒性试验方法,进行益肾乌发口服液对小鼠的急性毒性研究.结果 按生药量计算益肾乌发口服液对小鼠的最大给药量(MLD)为47.5g·kg-1·d-1.相当于人日用剂量的280.0倍;给药后各小鼠急毒主要表现为安静、怠动、腹泻、毛色不华等毒性症状,给药后6天体重增长缓慢,与空白对照组有明显差异,给药后8天逐渐恢复,余未见明显异常,末次给药后2h血ALT、AST水平明显增高,肝体比值增加.结论 益肾乌发口服液小鼠急毒虽未做出LD50和MTD,且无一死亡,但药后有肝功能损伤、肝体比值增加,提示在超大剂量灌胃后具有一定的肝毒性,与临床报道相一致.     

何首乌不同组分单次给药对小鼠肝毒性“量-时-毒”关系研究

目的观察何首乌不同组分单次给药对小鼠肝毒性"量-时-毒"关系的影响。方法 "时-毒"关系研究:取小鼠按不同时间点分组,单次灌胃给予一定剂量的何首乌水提组分、醇提组分,观察给药后小鼠死亡情况和毒性反应,分别于药后不同时间检测血清丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST),计算肝、脾、胸腺脏器指数。"量-毒"关系研究:取小鼠按不同剂量分组,单次灌胃给予不同剂量的何首乌水提组分、醇提组分,分别于药后4h、2h按"时-毒"研究方法对小鼠进行相应处理。结果小鼠灌胃较高剂量何首乌水提组分后,血清ALT、AST活力在4h达到高峰,持续时间均约达24h;给药4h后小鼠出现肝脏明显肿大,肝指数升高,其中4～6h肝脏指数升高较为明显。小鼠灌胃较高剂量何首乌醇提组分后,血清ALT、AST活力在2h达到高峰,持续时间均约达48 h;给药2 h后小鼠出现肝脏明显肿大,肝指数升高,其中2～4h肝脏指数升高较为明显。何首乌水提组分剂量在(5.5～30.75)g.kg-1之间、醇提组分在(8.5～24.5)g.kg-1之间对肝组织产生明显损伤,且随着剂量增大,ALT、AST升高显著。结论小鼠单次灌胃给予一定剂量的何首乌水提组分或醇提组分可造成急性肝损伤,并呈现一定的"量-时-毒"关系。关于其肝脏损伤的机制有待进一步研究。     

益肾乌发口服液中二苯乙烯苷、大黄素、大黄素甲醚的含量测定

目的 建立益肾乌发口服液中2,3,5,4'-四羟基二苯乙烯-2-O-β-D-葡萄糖苷、大黄素、大黄素甲醚含量的测定方法,从指标成分角度来探讨益肾乌发口服液毒性与何首乌的关系,并为益肾乌发口服液的质量控制提供方法.方法 采用HPLC法,C18柱(4.6mm×250mm,5μm);测定二苯乙烯苷的流动相为乙腈-水(25:75),检测波长为320hm,流速为1.0 mL·min-1,柱温:25℃;测定大黄素大黄素甲醚的流动相为甲醇-0.1%磷酸溶液(80:20),检测波长为254nm,流速为1.0 mL·min-1,柱温:25℃.结果 二苯乙烯苷在0.1μg～10μg、大黄素在0.0168μg～1.68μg、大黄素甲醚在0.0208～1.04μg范围内呈良好的线性关系,r分别为0.9989(n=8)、0.9984(n=8)、0.9966(n=8);平均回收率:二苯乙烯苷为99.33%,大黄素为99.998%,大黄素甲醚为99.576%;RSD:二苯乙烯苷为0.68%,大黄素为1.11%,大黄素为1.29%.结论 高效液相色谱法测定益肾乌发口服液中二苯乙烯苷、大黄素、大黄素甲醚的含量方法操作简单、灵敏度高、干扰少、重现性好、回收率高,可用于益肾乌发口服液的含量测定.初步证实益肾乌发口服液的毒性可能与君药制何首乌中二苯乙烯苷、大黄素、大黄素甲醚的含量有关,应控制其剂量范围,避免不良反应的发生.     

不同比例五味子宁神口服液镇静催眠作用研究

目的:寻找五味子宁神口服液镇静催眠作用的最佳配伍比例。方法:通过对小鼠自发活动的影响、对戊巴比妥钠的协同作用、对中枢神经递质的影响等3方面药理学研究,探讨五味子宁神口服液不同配伍比例的镇静催眠作用。结果:五味子宁神口服液以原方用药25%五味子比例作用最好。结论:五味子宁神口服液组方配伍是合理的。     

Comparison of hepatotoxicity and metabolism of butyltin compounds in the liver of mice,rats and guinea pigs

The hepatotoxicity of tributyltin chloride (TBTC) and dibutyltin dichloride (DBTC) was compared among mice, rats and guinea pigs in vivo. Further, the metabolism of these butyltin compounds in the liver was also investigated in these species. The oral administration of TBTC and DBTC to mice induced obvious liver injury, as demonstrated by both serodiagnosis and histopathological diagnosis. The concentrations of TBTC and DBTC that induced hepatotoxicity in mice at 24 h after oral administration were 180 and 60 micro mol/kg, respectively. In the case of rats, the liver injury induced by TBTC and DBTC was detected at 24 h by the serodiagnosis, but not by histopathological diagnosis. On the other hand, in guinea pigs, TBTC and DBTC administration did not produce any clear liver injury at 24 h, as evaluated by these two diagnostic methods. Thus, the following ranking was obtained with regard to increasing order of sensitivity to liver injury caused by TBTC and DBTC: mice, rats and guinea pigs. The total butyltin contents in the liver of mice were equivalent at 3 h and 24 h after the administration of TBTC or DBTC; however, the contents in the liver of rats and guinea pigs were relatively lower at 3 h and higher at 24 h than those of mice, although there were no differences between rats and guinea pigs in the total liver butyltin content. Concerning the liver metabolism of these butyltin compounds, the main form of butyltin compounds in these animals treated with TBTC was DBTC within 3 h after oral administration, while the main metabolites at 24 h were different in each species, indicating that the liver metabolism of TBTC might vary by animal type. When the animals were treated with DBTC orally, DBTC was hardly metabolized in the livers of these animals even at 24 h, and the liver levels of DBTC were two times greater in mice and guinea pigs than in rats at 3 h and were lower in mice at 24 h than in rats and guinea pigs. The analysis of cellular distributions of DBTC in the liver at 3 h after the administration showed that the levels of DBTC in the nuclear, microsomal and mitochondrial fractions of mice hepatocytes were relatively higher than in those of rats, which were greater than in those of guinea pigs. These results suggest differences in the sensitivity of mice, rats and guinea pigs to hepatotoxicity caused by butyltin compounds and demonstrate that the difference in the sensitivity of these animals to the hepatotoxicity induced by TBTC and DBTC may be partly due to differences in hepatic metabolism of TBTC and in the distribution of DBTC within cell organelles, respectively.     

The role of Ntcp,Oatp2, Bsep and Mrp2 in liver injury induced by Dioscorea bulbifera L. and Diosbulbin B in mice

Dioscorea bulbifera L. (DB) is a traditional Chinese herb used in thyroid disease and cancer. However, the clinical use of DB remains a challenge due to its hepatotoxicity, which is caused, in part, by the presence of Diosbulbin B (DIOB), a toxin commonly found in DB extracts. As abnormal expression of hepatobiliary transporters plays an important role in drug-induced liver injury, we assessed the hepatotoxicity induced by DB and DIOB, and explored their impacts on hepatobiliary transporter expression levels. Following liquid chromatography-tandem mass analysis of the DIOB content of DB extract, male ICR mice were randomly orally administered DB or DIOB for 14 days. Liver injury was assessed by histopathological and biochemical analysis of liver fuction. The levels of transporter protein and mRNA were determined by western blotting and real-time PCR. Liver function and histopathological analysis indicated that both DB and DIOB could induce liver injury in mice, and that DIOB might be the primary toxic compound in DB. Moreover, down-regulation of Mrp2 blocked the excretion of bilirubin, glutathione disulfide, and bile acids, leading to the accumulation of toxic substrates in the liver and a redox imbalance. We identified down-regulated expression of Mrp2 as potential factors linked to increased serum bilirubin levels and decreased levels of glutathione in the liver and increased liver injury severity. In summary, our study indicates that down-regulation of Mrp2 represents the primary mechanism of DB- and DIOB-induced hepatotoxicity, and provides insight into novel therapies that could be used to prevent DB- and DIOB-mediated liver injury.     

Sex difference in susceptibility to acetaminophen hepatotoxicity is reversed by buthionine sulfoximine

Gender is a factor that influences susceptibility of individuals to drug-induced liver injury in experimental animals and humans. In this study, we investigated the mechanisms underlying resistance of female mice to acetaminophen (APAP)-induced hepatotoxicity. Overnight-fasted male and female CD-1 mice were administered APAP intraperitoneally. A minor increase in serum alanine aminotransferase levels was observed in female mice after APAP administration at a dose that causes severe hepatotoxicity in males. Hepatic glutathione (GSH) depleted rapidly in the both genders prior to development of hepatotoxicity, whereas its recovery was more rapid in female than in male mice. This was consistent with higher induction of hepatic glutamate-cysteine ligase (GCL) in females. Pretreatment of mice with L-buthionine sulfoximine (BSO), an inhibitor of GCL, exaggerated APAP hepatotoxicity only in female mice, resulting in much higher hepatotoxicity in female than in male mice. In addition, hepatic GSH was markedly depleted in BSO-pretreated female mice compared with male mice, which supports severe hepatotoxicity in BSO-pretreated females. APAP treatment highly induced multidrug resistance-associated protein 4 (Mrp4) only in female mice. The resulting high Mrp4 expression could thus contribute to decreased hepatic GSH levels via sinusoidal efflux when GCL is inhibited. In conclusion, resistance to APAP hepatotoxicity in female mice and its reversal by pretreatment with BSO could be attributed to sex differences in disposition of hepatic GSH, which may generally determine susceptibility to drug-induced liver injury.