茵栀黄颗粒对雌激素诱导的胆汁瘀积大鼠肝细胞膜多药耐药相关转运体Mrp1~4的调节作用

目的考察茵栀黄颗粒对胆汁瘀积大鼠肝脏转运体多药耐药相关蛋白1⁴(Mrp14(Mrp1⁴)表达的影响。方法 Wistar雄性大鼠20只,随机分为4组,即正常组、模型组、对照组和茵栀黄组,每组5只。大鼠颈部皮下连续注射苯甲酸雌二醇[EB,5 mg/(kg·d)]造模。Western blot实验考察茵栀黄颗粒对肝脏转运体Mrp14)表达的影响。方法 Wistar雄性大鼠20只,随机分为4组,即正常组、模型组、对照组和茵栀黄组,每组5只。大鼠颈部皮下连续注射苯甲酸雌二醇[EB,5 mg/(kg·d)]造模。Western blot实验考察茵栀黄颗粒对肝脏转运体Mrp1⁴的调节作用。结果与对照组相比,茵栀黄组肝细胞膜转运体Mrp14的调节作用。结果与对照组相比,茵栀黄组肝细胞膜转运体Mrp1³的表达均显著增加(P<0.05),而Mrp4的表达差异无统计学意义(P>0.05)。结论茵栀黄颗粒能明显上调胆汁瘀积模型大鼠肝细胞膜转运体Mrp13的表达均显著增加(P<0.05),而Mrp4的表达差异无统计学意义(P>0.05)。结论茵栀黄颗粒能明显上调胆汁瘀积模型大鼠肝细胞膜转运体Mrp1³的表达,但对Mrp4的表达无影响。     

吲达帕胺对大鼠替米沙坦药代动力学的影响及性别差异(英文)

目的研究吲达帕胺对雌性和雄性大鼠替米沙坦药代动力学的影响。方法Wistar大鼠随机分为替米沙坦单用和替米沙坦与吲哒帕胺联用组,雌雄各半,分别单次ig给予替米沙坦3.6mg·kg-1或替米沙坦3.6mg·kg-1+吲达帕胺0.135mg·kg-1。96h内定时取血后,采用反相高效液相色谱-荧光检测法测定血浆中替米沙坦浓度。结果无论雄性雌性大鼠,与替米沙坦单用组相比,联用吲达帕胺后替米沙坦的主要药代动力学参数无显著性改变;而不同性别大鼠替米沙坦的药代动力学有显著差异,无论替米沙坦单用或联用吲达帕胺组,雌性大鼠替米沙坦的主要药代动力学参数AUC和cmax值均显著高于雄性,而血浆清除率显著低于雄性。结论替米沙坦与吲达帕胺联用对大鼠替米沙坦的药代动力学无明显影响;而无论替米沙坦单用或联用吲达帕胺,雌性和雄性大鼠替米沙坦的药代动力学存在显著的性别差异。     

2-巯基苯并咪唑SD大鼠灌胃给药的毒性研究

目的:研究2-巯基苯并咪唑(MBI)对 SD 大鼠所产生的毒性反应的性质和程度、剂量和毒性效应的关系、无毒反应剂量以及毒性的主要靶器官。方法:采用 SD 大鼠,灌胃给予 MBI,分为溶媒对照组和3个给药组(MBI 2 mg·kg~(-1),MBI 10 mg·kg~(-1),MBI 50 mg·kg~(-1)),连续给药28 d,期间对体重、摄食量进行监测。给药结束次日解剖取材,测定血液学及血清生化学指标,并对大鼠主要脏器进行大体观察和组织病理学检查。结果:50 mg·kg~(-1)MBI 使大鼠的体重增长及摄食量的增加受到明显抑制;10 mg·kg~(-1)和*或50 mg·kg~(-1)MBI 导致雌雄大鼠甲状腺、脑、肝脏重量明显升高,胸腺、脾脏重量显著下降;使雄性大鼠垂体、肺、睾丸重量明显升高,雌性肾上腺、颚下腺重量显著下降,肾脏重量显著升高;50 mg·kg~(-1)剂量引起大鼠 AST、LDH、CRE、BUN、CHO 等血清生化水平的显著变化;组织病理学检查发现,10 mg·kg~(-1)以上剂量的 MBI 可引起雄性和雌性动物甲状腺增生性病变、垂体前叶β细胞肿胀以及雄性动物胸腺萎缩;50 mg·kg~(-1)MBI 可引起雄性和雌性动物肝损伤,肾上腺皮质细胞的脂肪变性,以及雌性动物肾盂扩张。结论:MBI 毒性作用的主要靶器官为甲状腺;MBI 在 SD 大鼠的最大无毒反应剂量为2 mg·kg~(-1)。     

左氧氟沙星静脉给药对正常与肺炎大鼠肺组织中的药动学差异比较

目的:比较左氧氟沙星静脉给药在正常及肺炎大鼠肺组织中的药动学差异。方法:取SD雄性大鼠24只分为肺炎组及正常组;肺炎组大鼠予建立肺炎链球菌模型,模拟左氧氟沙星人体400 mg/d静脉给药;采用微透析技术对两组大鼠的血液及肺组织同步取样,分别测定其游离左氧氟沙星组织中浓度。结果:(1)静脉给药后,正常组大鼠肺组织游离药物浓度始终低于血药浓度,左氧氟沙星在肺组织中的平均穿透率(PR)为0.47±0.11,而肺炎组大鼠自20min后,肺组织药物浓度与血药浓度相近,PR为1.011 1±0.21(P<0.001);(2)左氧氟沙星在正常组大鼠血液中的消除半衰期(t_(1/2)为(1.04±0.81)h,平均驻留时间(MRT)为(1.76±0.72)h,而在肺炎组大鼠血液中分别为(3.83±0.93)h和(4.49±1.56)h,两组数据经比较其差异有统计学意义(P<0.05),但峰浓度(c_(max))及曲线下面积(AUC)经比较其差异有统计学意义(P<0.05);(3)与正常组大鼠相比,左氧氟沙星在肺炎组大鼠肺组织中c_(max)和AUC_(0~∞)显著增高,即(15.90±5.45)mg/L vs(28.08±9.88)mg/L和(13.20±2.82)mg/(h·L)vs(56.21±18.11)mg/(h·L)(均P<0.05);t_(1/2)和MRT明显延长,即(0.86±0.58)h vs(3.88±1.24)h和(2.16±1.07)h vs(4.80±1.79)h(均P<0.05)。结论:左氧氟沙星静脉给药后在肺炎组大鼠肺组织中的游离药物浓度及AUC值显著升高,穿透率明显高于正常组大鼠,可达到清除肺炎链球菌的有效浓度;感染者(肺炎组)局部组织中所获得的药动学参数比正常者(正常组)更有意义。     

甘草酸单铵保护利福平致大鼠肝损伤的机制初探

目的: 本文基于肝细胞膜转运体多药耐药相关蛋白2(multidrug resistance protein 2, Mrp2)和Na⁺-牛磺酸钠共转运体(sodium taurocholate cotransporting polypeptide, Ntcp),初步探究甘草酸单铵(monoammonium glycyrrhizinate,MAG)对利福平(rifampicin,RIF)致大鼠肝损伤的保护作用及机制。方法: Wistar雄性大鼠随机分为3组:对照组(control组):灌胃等容量的生理盐水;RIF肝损伤组(RIF组):灌胃RIF 60 mg·kg^-1·d^-1;MAG治疗组(MAG+RIF组):灌胃MAG 45 mg·kg^-1·d^-1 3 h后灌胃RIF 60 mg·kg^-1·d^-1。给药第7,14,21天时,各组分别随机取5只大鼠分离血清测定生化指标;取肝组织做病理切片,观察肝脏组织病理学变化并进行肝组织学活动指数(HAI)评分;采用 Western blotting法检测肝组织Mrp2和Ntcp蛋白表达量。结果: 与对照组相比,RIF组给药7,14,21 d时,其部分血清生化指标呈明显上升趋势,肝脏病理学HAI分值显著增加(P<0.01);MAG+RIF组血清生化指标与对照组之间无显著差异,与RIF组相比其HAI评分显著降低(P<0.05)。给药7,14,21 d时,RIF组较对照组Mrp2的表达均显著升高(P<0.05),而MAG+RIF组Mrp2的表达均显著低于RIF组(P<0.05)。给药期间,各组Ntcp的表达均无显著差异(P>0.05)。结论: 利福平肝损伤机制可能与其上调Mrp2有关,MAG可保护利福平诱导的肝损伤,且其保肝作用可能与其下调Mrp2有关。     

甘草酸单铵对异烟肼致大鼠肝损伤的保护作用及其机制

目的:考察甘草酸单铵(MAG)对异烟肼(INH)诱导的大鼠肝损伤的保护作用及其对肝脏转运体Na⁺-牛磺胆酸共转运多肽(Na⁺-taurocholate cotransporting polypeptide,NTCP)及多药耐药相关蛋白2(multidrug resistance protein 2,Mrp2)的调节作用。方法:Wistar雄性大鼠45只,随机分为9组,即对照组,异烟肼肝损伤组(INH组),甘草酸单铵治疗组(MAG组),以上每组又各分为干预7,14,21 d组。INH组大鼠灌胃INH 60 mg·kg^-1·d^-1;MAG组大鼠灌胃MAG 45 mg·kg^-1·d^-1 3 h后灌胃INH。干预结束后,处死大鼠,通过血清生化指标、肝脏病理及Ntcp、Mrp2蛋白表达对各组大鼠进行评价对比。结果:血清生化指标与肝脏病理结果显示,INH诱导的大鼠肝损伤具有时间依赖性,而MAG对INH诱导的大鼠肝损伤具有保护作用,且存在时间依赖性;与INH组相比,MAG干预21 d对肝脏转运体Ntcp及Mrp2表达具有明显下调作用(P<0.05)。结论:MAG对INH诱导的肝损伤的保护作用可能与下调肝细胞膜转运体Ntcp与Mrp2的表达有关。     

小檗碱、巴马汀和药根碱在“三明治”培养大鼠原代肝细胞中的胆汁外排特征

目的研究小檗碱、巴马汀和药根碱在大鼠原代细胞的胆汁外排特征。方法建立"三明治"培养大鼠原代肝细胞(sandwich cultured rat hepatocytes,SCRH)模型,分别加入小檗碱、巴马汀、药根碱,在含Ca²⁺和无Ca²⁺缓冲液中孵育,采用UPLC-MS/MS分别测定3种生物碱的细胞蓄积量,计算胆汁排泄指数和胆汁清除率,评价小檗碱、巴马汀、药根碱在大鼠原代细胞的胆汁外排特征;并考察P-gp、Mrp2抑制剂对3种活性成分在SCRH细胞模型中的外排转运影响。结果随着孵育时间的延长,3种生物碱的细胞蓄积量增加,且含Ca²⁺条件下的细胞蓄积量与无Ca²⁺条件下相比具有明显差异;P-gp抑制剂环孢素A、维拉帕米均能减少小檗碱、巴马汀、药根碱的胆汁排泄,且呈浓度依赖性,Mrp2抑制剂MK571、丙磺舒对3种生物碱的胆汁排泄无明显影响。结论小檗碱、巴马汀、药根碱均经过胆汁外排,P-gp介导了3种生物碱的胆汁外排,Mrp2未参与其胆汁排泄。     

左氧氟沙星羧甲基壳聚糖微球结肠靶向释药的实验研究

目的:检测所制备的左氧氟沙星羧甲基壳聚糖(LVFX/CMC)微球在人工消化液中和大鼠体内结肠靶向释药的性能。方法:以分光光度仪测定微球在人工消化液中的累积释放量,电镜观察微球在人工消化液中形态的改变。SD大鼠60只,随机分为两组,分别以LVFX/CMC微球(含40 mg左氧氟沙星)及等量左氧氟沙星溶剂灌胃,以高效液相色谱法对LVFX/CMC微球和左氧氟沙星(LVFX)灌胃后大鼠盲肠、结肠中药物浓度进行定量检测。结果:左氧氟沙星壳聚糖微球在人工胃液介质中溶解缓慢,2 h仅释药8.62%;在人工小肠液介质中溶解速度稍见加快,6 h释药29.39%,但表现为药物缓释曲线,24 h仅释药42.13%;在人工结肠液中,4 h后释药84.56%,24 h内累积释药量为93%。扫描电镜观察人工胃液中的微球明显溶胀,稍见变形;人工结肠液中的微球溶解,粒径明显减小。灌胃后LVFX/CMC微球组5和9 h时段盲肠、结肠药量明显高于LVFX组。结论:左氧氟沙星羧甲基壳聚糖微球在体外、体内实验中的释放符合结肠靶向释药的特点。     

赛庚啶对大鼠下丘脑-腺垂体-性腺轴和钙调蛋白基因表达的影响(英文)

目的　研究赛庚啶 (Cyp)对SD大鼠生殖系统内分泌功能的影响是否有性别差异。方法　 60只SD大鼠依性别各分为 3组 ,每组 1 0只 ;分别灌胃给予生理盐水 ( 5mL·kg-1·d-1) ,Cyp( 2 .4,4.8mg·kg-1·d-1) ,共 1 4d或 2 1d。放免法测定血清黄体生成素 (LH)、促卵泡激素 (FSH)、雌二醇 (E2 )、孕酮(P)、睾酮 (T)的含量。光电镜观察促性腺激素释放激素细胞、促性腺激素细胞、间质细胞、支持细胞、黄体细胞、颗粒细胞等显微、超微结构的变化。用实时荧光定量PCR技术进行逆转录聚合酶链反应 ,琼脂糖凝胶电泳鉴定扩增产物。结果　Cyp可升高雄性大鼠血清LH、T的含量 ,而对FSH含量无明显影响。Cyp可升高雌性大鼠血清LH的水平 ,降低其FSH ,P ,E2 的含量。电镜发现 ,Cyp促进雄性大鼠分泌功能 ,使雌性大鼠内分泌细胞发生退行性改变。结论Cyp对雌性大鼠下丘脑 腺垂体 卵巢轴内分泌功能有抑制作用 ,而对雄性大鼠下丘脑 腺垂体 睾丸轴内分泌功能有促进作用 ,且使终末靶腺的内分泌细胞超微结构出现相应的变化。Cyp促进雄性大鼠睾丸钙调蛋白mRNA的表达可能与Cyp促进下丘脑 垂体 睾丸轴的内分泌功能有关     

坦索罗辛对大鼠细菌性前列腺炎组织中左氧氟沙星药动学的影响

目的探讨坦索罗辛对急性细菌性前列腺炎模型大鼠前列腺组织中左氧氟沙星药动学的影响。方法将96只急性细菌性前列腺炎模型大鼠随机分为实验组(左氧氟沙星和坦索罗辛联合用药组)和对照组(单用左氧氟沙星组),每组48只,分别在给药后0.125、0.25、0.5、1、2、4、8和12h每个时间点处死6只大鼠,采集两组动物的前列腺制作组织匀浆,用HPLC法测定前列腺组织中左氧氟沙星浓度,3p97软件计算药动学参数。结果实验组和对照组药-时曲线均符合一室模型。主要药动学参数如下:t1/2分别为(4.78±0.75)h和(3.64±0.88)h,tpeak分别为(1.18±0.10)h和(0.81±0.29)h,Cmax分别为(6.16±0.57)μg/g和(3.91±0.62)μg/g,AUC0~12分别为(41.91±1.01)μg h/g和(22.70±3.08)μg h/g。结论坦索罗辛可以明显提高左氧氟沙星在急性细菌性前列腺炎模型大鼠前列腺组织中的药物浓度。     

Up-regulation of Mrp4 expression in kidney of Mrp2-deficient TR- rats

Multidrug resistance-associated proteins (Mrps) are a group of ATP-dependent efflux transporters for organic anions. Mrp2 and Mrp4 are co-localized to the apical (brush-border) membrane domain of renal proximal tubules, where they may function together in the urinary excretion of organic anions. Previous reports showed that urinary excretion of some organic anions is not impaired in transport-deficient (TR-) rats, which lack Mrp2, suggesting that up-regulation of other transporter(s) may compensate for the loss of Mrp2 function. The purpose of this study was to determine whether Mrp4 expression in kidney is altered in TR- rats. Mrp4 mRNA expression was quantified using the high-throughput branched DNA signal amplification assay. Mrp4 protein expression was determined by Western blot and immunohistochemical analysis. Mrp4 mRNA in kidney of TR- rats was 100% higher than normal Wistar rats. Western blot analysis showed a 200% increase in Mrp4 protein expression in kidney of the mutant rats compared to normal rats. Immunohistochemical analysis of Mrp4 protein demonstrated apical localization of Mrp4 on renal proximal tubules, and that the immunoreactivity was more intense in kidney sections from TR- rats than those from normal rats. In summary, the results of the present study demonstrate that renal Mrp4 expression is up-regulated in TR- rats, which may explain why urinary excretion of some organic anions remains normal in the mutant rats.     

Induction of multidrug resistance-associated protein 2 in liver, intestine and kidney of streptozotocin-induced diabetic rats

Many studies have demonstrated that Mrp2 is highly regulated in some physiopathological situations. The aim of this study was to investigate effects of diabetes mellitus on function and expression of multidrug resistance-associated protein 2 (Mrp2) in rat liver, kidney and intestine. Diabetic rats were induced by an intraperitoneal administration of streptozotocin (65?mg/kg) and randomly divided into diabetic (DM) rats and insulin-treated diabetic rats. Sulfobromophthalein (BSP), a substrate of Mrp2, was used to evaluate Mrp2 function in vivo. Data from excretion experiments demonstrated that compared with normal rats, diabetes markedly enhanced BSP excretion via bile, urine and intestinal perfusate, which contributed to the elevated plasma clearance of BSP after intravenous administration of 45 μmol/kg BSP. Western blot results showed higher levels of hepatic, renal and intestinal Mrp2 protein in DM rats, although no difference was observed in renal Mrp2. Insulin treatment partly reversed these alterations. Induction of Mrp2 by diabetes was in parallel with the increase in bile flow, levels of biliary and plasma total bile acid (TBA), and plasma conjugated bilirubin in DM rats. Diabetes may enhance Mrp2 function and expression in liver, kidney and intestine, which might be due to insulin deficiency, increased TBA and conjugated bilirubin.     

Role of constitutive androstane receptor in the in vivo induction of Mrp3 and CYP2B1/2 by phenobarbital.

Phenobarbital (PB) induces the hepatic organic anion transporter, Mrp3. The present study tested the hypothesis that Mrp3 induction by PB is mediated by the constitutive androstane receptor (CAR). PB induction of Mrp3 and CYP2B was examined in lean and obese Zucker rats, male and female Wistar Kyoto (WKY) rats, HepG2 and mouse CAR-expressing HepG2 (g2car-3) cells; HepG2 and g2car-3 cells also were treated with 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP). In obese Zucker rat livers, total and nuclear CAR levels were markedly lower compared with lean rat livers, which correlated with the poor induction of CYP2B1/2 by PB in obese Zucker rats. Mrp3 induction by PB also was impaired in obese Zucker rat livers. Induction of Mrp3 by PB was similar in male and female WKY rat livers, despite the fact that CAR protein levels were significantly lower in female relative to male WKY rat livers. MRP3 levels in both HepG2 and g2car-3 cells were induced to a similar extent in the two cell lines by PB but not by TCPOBOP. In contrast, CYP2B6 levels were measurable and induced by TCPOBOP only in g2car-3 cells. In conclusion, data from WKY rats and HepG2 cells suggest that CAR does not play a key role in PB induction of Mrp3. Impaired induction of Mrp3 by PB in obese Zucker rats is not due solely to CAR deficiency. Interestingly, differences in the constitutive levels of Mrp3 were observed between obese and lean Zucker rats and between male and female WKY rats.     

Association between tubular toxicity of cisplatin and expression of organic cation transporter rOCT2 (Slc22a2) in the rat

Cisplatin is an effective anticancer drug, but has its severe adverse effects, especially nephrotoxicity. The molecular mechanism of cisplatin-induced nephrotoxicity is still not clear. In the present study, we examined the role of rat (r)OCT2, an organic cation transporter predominantly expressed in the kidney, in the tubular toxicity of cisplatin. Using HEK293 cells stably expressing rOCT2 (HEK-rOCT2), we evaluated the cisplatin-induced release of lactate dehydrogenase and the uptake of cisplatin. The release of lactate dehydrogenase and the accumulation of platinum were greater in HEK-rOCT2 cells treated with cisplatin than in mock-transfected cells. Moreover, cimetidine and corticosterone, OCT2 inhibitors, inhibited the cytotoxicity and the transport of cisplatin in HEK-rOCT2 cells. Pharmacokinetics of cisplatin was investigated in male and female rats because the renal expression level of rOCT2 was higher in male than female rats. The renal uptake clearance of cisplatin was greater in male than female rats, while the hepatic uptake clearance was similar between the sexes. In addition, glomerular filtration rate and liver function were unchanged, but N-acetyl-β-d-glucosaminidase activity in the bladder urine and the urine volume were markedly increased 2 days after the administration of 2 mg/kg of cisplatin in male rats. Moreover, cisplatin did not induce the elevation of urinary N-acetyl-β-d-glucosaminidase activity in the castrated male rats whose renal rOCT2 level was lower than that of the sham-operated rats. In conclusion, the present results indicated that renal rOCT2 expression was the major determinant of cisplatin-induced tubular toxicity.     

Metabolism and disposition of gemfibrozil in Wistar and multidrug resistance-associated protein 2-deficient TR- rats

1. The roles of multidrug resistance-associated protein (Mrp) 2 deficiency and Mrp3 up-regulation were evaluated on the metabolism and disposition of gemfibrozil. 2. Results from in vitro studies in microsomes showed that the hepatic intrinsic clearance (CLint) for the oxidative metabolism of gemfibrozil was slightly higher (1.5-fold) in male TR- rats, which are deficient in Mrp2, than in wild-type Wistar rats, whereas CLint for glucuronidation was similar in both strains. 3. The biliary excretion of intravenously administered [14C]gemfibrozil was significantly impaired in TR-) rats compared with Wistar rats (22 versus 93% of the dose excreted as the acyl glucuronides over 72 h). Additionally, the extent of urinary excretion of radioactivity was much higher in TR- than in Wistar rats (78 versus 2.6% of the dose). 4. There were complex time-dependent changes in the total radioactivity levels and metabolite profiles in plasma, liver and kidney, some of which appeared to be related to the up-regulation of Mrp3. 5. Overall, it was demonstrated that alterations in the expression of the transporters Mrp2 and Mrp3 significantly affected the excretion as well as the secondary metabolism and distribution of [14C]gemfibrozil.     

Hormonal regulation of renal multidrug resistance-associated proteins 3 and 4 (Mrp3 and Mrp4) in mice

Multidrug resistance-associated proteins 3 and 4 (Mrp3 and Mrp4) are expressed at much higher levels in female than male kidney. Sex steroids and sex-specific growth hormone (GH) secretion patterns often mediate gender-predominant gene expression. Thus, three models were used to investigate potential endocrine regulation of Mrp3 and Mrp4: (1) gonadectomized (GNX) mice with 17beta-estradiol (E2) or 5alpha-dihydroxytestosterone (DHT) replacement; (2) hypophysectomized (HPX) mice receiving E2, DHT, or simulated male-pattern (MP) or female-pattern (FP) GH secretion; (3) lit/lit mice, which have a spontaneous mutation in the growth-hormone releasing-hormone (GHRH) receptor, with simulated MP- or FP-GH secretion. GNX and HPX decreased Mrp3 mRNA levels compared with intact females. In both respective models E2 administration increased Mrp3 expression in GNX and HPX mice. DHT markedly repressed Mrp3 from GNX+placebo levels, however, this was not observed in the HPX model. In lit/lit mice, Mrp3 expression was lower than in wild-type controls, and MP-GH and FP-GH simulation slightly increased Mrp3 expression. Whereas GNX increased Mrp4 in males to female levels, HPX actually increased Mrp4 expression in both genders +375% and +66%, respectively. In both models DHT markedly repressed Mrp4. Furthermore, Mrp4 was higher in lit/lit than wild-type male mice, and simulation of MP-GH secretion suppressed female-predominant Mrp4 expression. In conclusion, these data indicate that E2 contributes to higher Mrp3 mRNA expression in females, yet a role for androgens in Mrp3 repression cannot be discounted. In contrast, Mrp4 mRNA is higher in females due to repression by both DHT and MP-GH secretion in males.     

Characterization of transport protein expression in multidrug resistance-associated protein (Mrp) 2-deficient rats.

Multidrug resistance-associated protein (Mrp) 2-deficient transport-deficient (TR(-)) rats, together with their transport-competent Wistar counterparts (wild type), have been used to examine the contribution of Mrp2 to drug disposition. However, little is known about potential variation in expression of other transport proteins between TR(-) and wild-type rats or whether these differences are tissue-specific. Sections of liver, kidney, brain, duodenum, jejunum, ileum, and colon were obtained from male TR(-) and wild-type Wistar rats. Samples were homogenized in protease inhibitor cocktail and ultracentrifuged at 100,000g for 30 min to obtain membrane fractions. Mrp2, Mrp3, Mrp4, P-glycoprotein, sodium-dependent taurocholate cotransporting polypeptide, organic anion transporting polypeptides 1a1 and 1a4, bile salt export pump, breast cancer resistance protein, ileal bile acid transporter, UDP-glucuronosyl transferase (UGT1a), glyceraldehyde-3-phosphate dehydrogenase, and beta-actin protein expression were determined by Western blot. Mrp3 was significantly up-regulated in the liver ( approximately 6-fold) and kidney ( approximately 3.5-fold) of TR(-) rats compared with wild-type controls. Likewise, the expression of UGT1a enzymes was increased in the liver and kidney of TR(-) rats by approximately 3.5- and approximately 5.5-fold, respectively. Interestingly, Mrp3 expression was down-regulated in the small intestine of TR(-) rats, but expression was similar to wild type in the colon. Mrp4 was expressed to varying extents along the intestine. Expression of some transport proteins and UGT1a enzymes differ significantly between TR(-) and wild-type rats. Therefore, altered drug disposition in TR(-) rats must be interpreted cautiously because up- or down-regulation of other transport proteins may play compensatory roles in the presence of Mrp2 deficiency.     

Metabolism of tris(2-chloroethyl) phosphate in rats and mice.

Tris(2-chloroethyl) phosphate (TRCP), a flame retardant, produces a dose-, sex-, and species-dependent lesion in the hippocampal region of the brain following subchronic oral administration. This lesion is more common and more severe in female F344 rats than in male F344 rats, and is not observed in B6C3F1 mice. The present investigation of the metabolism of TRCP was designed to detect sex and species variations that might account for differences in toxicity. Elimination of TRCP-derived radioactivity was more rapid in mice, which excreted greater than 70% of an oral dose of 175 mg/kg in urine in 8 hr vs. approximately 40% for male or female rats. However, the metabolic profile of TRCP-derived radioactivity in urine was similar for both species. The major metabolite in female rat urine was identified as bis(2-chloroethyl) carboxymethyl phosphate. This metabolite co-chromatographed with the major metabolite found in both male rat and mouse urine. Two additional metabolites identified in female rat urine were bis(2-chloroethyl) hydrogen phosphate and the glucuronide of bis(2-chloroethyl) 2-hydroxyethyl phosphate. These metabolites also cochromatographed with metabolites found in male rat and mouse urine. TRCP metabolism in rats was not induced or inhibited by nine daily 175 mg/kg doses. Toxicity, as evidenced by seizures, was potentiated in male rats pretreated with inhibitors of aldehyde dehydrogenase.     

Efflux transporter expression and acetaminophen metabolite excretion are altered in rodent models of nonalcoholic fatty liver disease.

Efflux transporters are responsible for the excretion of numerous xenobiotics and endobiotics and thus play an essential role in proper liver and kidney function. Nonalcoholic fatty liver diseases (NAFLDs) comprise a spectrum of disorders that range from simple fatty liver (SFL) to nonalcoholic steatohepatitis (NASH). Although the precise events leading to NAFLD are unclear, even less is known about the effects on efflux transporter expression and drug disposition. The purpose of this study was to determine the effect of NAFLD on efflux transporter expression in rat liver as well as on acetaminophen (APAP) metabolite excretion. To simulate SFL and NASH, rats were fed either a high-fat (HF) or a methionine- and choline-deficient (MCD) diet for 8 weeks. In the livers of MCD rats, there were striking increases in both mRNA and protein levels of multidrug resistance-associated protein (Mrp) 3, Mrp4, and breast cancer resistance protein, as well as increased Mrp2 protein. After administration of a nontoxic dose of APAP, biliary concentrations of APAP-sulfate, APAP-glucuronide (APAP-GLUC), and APAP-glutathione were reduced in MCD rats. The effects of the HF diet on both transporter expression and APAP disposition were by comparison far less dramatic than the MCD diet-induced alterations. Whereas APAP-sulfate levels were also decreased in MCD rat plasma, the levels of the Mrp3 substrate APAP-GLUC were elevated. Urinary elimination of APAP metabolites was identical between groups, except for APAP-GLUC, the concentration of which was 80% higher in MCD rats. These studies correlate increased hepatic Mrp3 protein in the MCD model of NASH with increased urinary elimination of APAP-GLUC. Furthermore, the proportional shift in elimination of APAP metabolites from bile to urine indicates that MCD-induced alterations in efflux transporter expression can affect the route of drug elimination.