海藻-昆布药对对大鼠肝微粒体代谢酶的影响及肝毒性研究

目的 发现不同剂量海藻-昆布药对提取物对大鼠肝微粒体代谢酶的诱导或抑制作用, 预测服用海藻-昆布药对时可能出现的药物-药物相互作用及肝脏毒性。方法 雌雄各半SD大鼠18只, 被随机分为海藻-昆布药对低、高剂量组和对照组, 低、高剂量组大鼠分别ig给予海藻-昆布药对提取物10.8、86.4 g/(kg·d), 连续经口给药15 d后麻醉处死, 取肝组织制备肝微粒体及HE染色石蜡切片。通过肝微粒体体外孵育方法测定3种肝脏CYP450同工酶特异性底物非那西丁(CYP1A2)、氯唑沙宗(CYP2E1)及咪达唑仑(CYP3A4)的降解和代谢产物生成量来评价肝药酶的诱导或抑制作用, 并以光镜下的组织病理切片检查来考察其肝毒性。结果 低剂量组大鼠无显著诱导或抑制3种CYP450代谢酶亚型1A2、2E1和3A4现象, 肝组织出现了肝窦扩张、轻度水肿等适应性改变, 高剂量组能显著诱导CYP3A4亚型, 但也不能显著的诱导或抑制肝微粒体代谢酶CYP1A2、CYP2E1亚型, 肝组织出现了脂肪变、点状坏死等可逆性损伤。结论 海藻-昆布药对具有诱导肝微粒体代谢酶CYP3A4的作用和轻微的肝细胞毒性, 高剂量经口给药能引起有临床意义的CYP450酶的诱导现象和肝脏损伤并可能导致不期望的药物-药物相互作用。     

阿尔茨海默病转基因小鼠肝内核受体和细胞色素氧化酶基因转录谱变化

目的探讨阿尔茨海默病(AD)病变与肝Ⅰ相药酶基因表达的相关性,为早期和长期药物干预AD病程提供实验参考。方法制备B6.129-PS1M146V/APPSwe/Tau P301L三重转基因AD模型小鼠(3×Tg AD小鼠)及同系非转基因正常小鼠(NTg小鼠)肝组织总RNA和总蛋白质样品,逆转录PCR/实时荧光定量PCR技术检测肝核因子4(HNF4)等7种核受体和细胞色素氧化酶P4501a2(Cyp1a2)等7种CYP基因mRNA相对表达量,比较基因转录谱差异;Western蛋白印迹法检测技术检测差异显著的目的基因蛋白质水平。结果与NTg小鼠相比,3×Tg AD小鼠HNF4、雌激素受体(ERα)及组成型雄甾烷受体均下调近66%(P<0.01),孕烷X受体和芳香烃受体分别上调1.84和1.64倍(P<0.05),糖皮质激素受体及受氧化应激激活的核因子E2相关因子未见显著变化;主要药物代谢酶Cyp2b10及Cyp2a5 mRNA分别降低至正常小鼠水平的3%(即下降97%,P<0.01)和30%(P<0.05),Cyp2e1、Cyp2j9、Cyp3a11基因的转录水平则分别上调了2.26倍、2.42倍和3.66倍(P<0.05);Cyp1a2和Cyp2d22基因转录在两种小鼠中无明显差异。Western蛋白印迹分析证实,3×Tg AD小鼠ERα和CYP2a5明显下降(P<0.05)而Cyp3a11表达则明显上升(P<0.05),蛋白质表达变化的趋势与其相应mRNA水平改变一致。结论 3×Tg AD小鼠肝内核受体基因及CYP酶基因转录谱发生显著改变,提示AD病变可能干扰机体肝代谢功能,对机体内源性及外源性化合物生物转运产生影响。     

柴胡总皂苷对小鼠肠道首过效应和肝脏Cyp3a、Cyp2e1的影响

目的:研究柴胡总皂苷对小鼠肠道首过效应(Cyp3a,P-糖蛋白)和肝脏细胞色素氧化酶(Cyp3a,Cyp2e1)的影响。方法:供试物灌胃给小鼠2次/d,连续3 d。实验当日,对乙酰氨基酚(Acetaminophen,APAP;P-gp底物)以50 mg/kg经口投予后60 min断头采血,并摘取肝脏和全段小肠。以分光光度法测定血中APAP浓度;用梯度离心法分离小鼠肝/肠微粒体;以分光光度法检测微粒体中Cyp3a/Cyp2e1活性;以实时荧光定量法测定Cyp3a11/Cyp2e1 mRNA在小鼠肝脏中的表达。结果:血中APAP浓度测定结果和P-糖蛋白偶联的ATP酶活性测定结果显示,柴胡总皂苷各剂量组与对照组之间均无统计学差异(P>0.05);在肝脏和肠道微粒体实验中不论以氨基吡啉还是以红霉素为底物测定Cyp3a,仅有柴胡总皂苷高剂量组(150 mg/kg)的Cyp3a活性显著高于对照组(P<0.05);仅有柴胡总皂苷中剂量组(75 mg/kg)的Cyp2e1活性显著低于对照组(P<0.05);RT-PCR结果显示,仅有柴胡总皂苷高剂量(150 mg/kg)时能够诱导Cyp3a11在肝脏中的表达。结论:柴胡总皂苷对小鼠肝脏和肠道中的Cyp3a具有一定的诱导作用,对肝脏中的Cyp2e1具有一定的抑制作用,对小鼠肠道P-糖蛋白的转运活性无影响。     

丁香酸和柠檬苦素对小鼠肝脏CYP450酶的影响

目的 研究丁香酸和柠檬苦素对小鼠肝脏细胞色素P450主要亚型mRNA及蛋白表达水平的影响.方法 C57BL/6小鼠随机分为空白对照组、丁香酸组、柠檬苦素组和苯巴比妥组,连续灌胃给药2周,末次给药后处死,提取小鼠肝脏总RNA及肝微粒体,荧光定量聚合酶链式反应(PCR)技术和蛋白质免疫印迹(Western blot)测定CYP450酶主要亚型mRNA和蛋白表达水平.结果 在mRNA水平上,丁香酸对Cyp1a2、Cyp2c37、Cyp2d9 mRNA表达没有明显作用,柠檬苦素对Cyp1a2 mRNA的表达有显著诱导作用;在蛋白水平上,丁香酸对CYP1A1、CYP1A2、CYP3A、CYP2D和CYP2E1蛋白的表达有明显的诱导作用,柠檬苦素对CYP1A1、CYP1A2、CYP2A、CYP2D和CYP2E1有显著的诱导,对CYP2B和CYP2C蛋白表达产生抑制作用.结论 丁香酸和柠檬苦素对细胞色素P450主要亚型均具有不同程度的诱导和抑制作用.     

川芎嗪对肝药物代谢酶CYP3A4的影响及机制研究

目的探讨川芎嗪对HepG2细胞中肝药物代谢酶CYP3A4的影响及其作用机制。方法采用不同浓度的川芎嗪作用于HepG2细胞48 h,MTT法检测细胞活力,并选取对细胞存活率影响较小的浓度进行实验。设置空白组、酮唑康组、不同浓度川芎嗪组以及酮唑康和不同浓度川芎嗪共同给药组,CYP3A4和PXR转染各组细胞,采用荧光素酶报告基因技术检测各给药组对CYP3A4和PXR酶活性的影响。并提取各组细胞蛋白,采用Western blot法检测HepG2细胞的CYP3A4和PXR蛋白表达水平。结果川芎嗪可以使CYP3A4酶活性增加,并呈剂量依赖性,还可使CYP3A4蛋白表达水平上调;川芎嗪可剂量依赖性地增加PXR转录活性,并使PXR蛋白水平升高。结论川芎嗪能通过激活PXR受体而对药物代谢酶CYP3A4起诱导作用。     

垂盆草总黄酮对肝纤维化大鼠肝组织TGF-β 1和Smad7表达的影响

摘 要 目的： 观察垂盆草总黄酮(SSTF)对CCL₄诱导肝纤维化大鼠肝组织TGF-β1和Smad7蛋白及mRNA表达的影响。方法: 60只雄性SD大鼠随机分为正常组、模型组、SSTF低剂量组（100 mg·kg^-1）、SSTF中剂量组(200 mg·kg^-1）、SSTF高剂量组（400 mg·kg^-1）和秋水仙碱阳性药对照组,共6组,每组10只。CCL₄皮下注射诱导大鼠肝纤维化模型,同时SSTF灌胃干预,7周后处死大鼠。采用Masson染色法检测大鼠肝脏组织病理学改变,Western blotting法检测肝脏组织TGF-β1和Smad7蛋白表达,实时定量RT-PCR检测肝脏组织TGF-β1和Smad7mRNA表达。结果: 与模型组比较,SSTF各剂量均能显著降低CCL₄诱导的肝纤维化程度（P<0.05）;中高剂量可以显著减少肝组织中TGF-β1蛋白和mRNA表达（P<0.05）,并且明显增加Smad7蛋白和mRNA表达（P<0.05）。结论:SSTF可有效地逆转实验性大鼠肝纤维化,其机制可能与下调肝组织中TGF-β1蛋白和mRNA表达,及增加Smad7蛋白和mRNA表达有关。     

中药莪术激活PXR及对大鼠肝细胞色素P450 3A的影响

目的考察莪术能否通过体外激活PXR调节细胞色素P4503A4(CYP3A4)的转录表达及对大鼠肝脏CYP3A在酶活性及mRNA表达的诱导作用。方法在HepG2细胞中,采用瞬时共转染报告基因实验研究莪术对PXR介导的CYP3A4的转录调节作用;在大鼠体内,采用紫外分光光度法和RT-PCR技术检测莪术对大鼠肝脏CYP450含量及CYP3A同工酶红霉素N-脱甲基酶(ERD)的活性和CYP3A基因mRNA表达的影响。结果在体外报告基因实验研究中,莪术提取物及其有效成分能不同程度的诱导CYP3A4的表达;在大鼠体内实验中,莪术提取物能够增加CYP450蛋白含量和CYP3A酶活性;在mRNA水平上,莪术提取物能够明显诱导CYP3A1及CYP3A2的基因表达。结论莪术提取物及其有效成分能够明显激活PXR并诱导CYP3A4的转录表达;莪术提取物对大鼠体内CYP3A的酶活性及mRNA表达均有明显诱导作用。     

丙戊酸钠及其代谢产物的体外肝毒性研究

目的 研究丙戊酸钠及3个代谢产物（2-丙基-4-五烯酸、3-羟基丙戊酸、5-羟基丙戊酸）对体外人正常肝细胞L02增殖活性及对肝细胞损伤相关指标的影响。方法 实验分为对照组和实验组,对照组细胞常规培养,实验组加入丙戊酸钠及3个代谢产物,采用CCK-8法检测细胞增殖活性,PCR法检测CYP1A1、CYP1A2、PCNA、Bax及Bcl-2的mRNA相对含量,Western Blotting法检测蛋白表达,同时检测细胞上清液中谷草转氨酶（AST）、谷丙转氨酶（ALT）、乳酸脱氢酶（LDH）的含量。结果 与对照组相比,随着丙戊酸钠及3个代谢产物浓度和时间的增加,对 L02细胞增殖活性的抑制逐渐增强,CYP1A1、CYP1A2及Bax的mRNA相对含量和蛋白表达量升高,PCNA及Bcl-2的 mRNA相对含量和蛋白表达量均有下降,AST、ALT、LDH含量升高。结论 丙戊酸钠及3个代谢产物与肝毒性有关。     

注射用益气复脉(冻干)对CYP450药物代谢酶1A2、2B6和3A4诱导作用的体外研究

目的 评价注射用益气复脉（冻干,YQFM）对人原代肝细胞中的药物代谢酶CYP1A2、2B6和3A4是否具有诱导作用。方法 采用3个单供体的人原代肝细胞分别与不同质量浓度（0.026、0.260和2.600 mg/mL）的YQFM或特异性诱导剂（CYP1A2诱导剂：50 μmol/L奥美拉唑;CYP2B6诱导剂：1 000 μmol/L苯巴比妥;CYP3A4诱导剂：25 μmol/L利福平）于37℃、5% CO₂条件下共孵育3 d,液质联用技术（LC-MS/MS）检测酶活性,实时荧光定量PCR技术检测CYP1A2、2B6和3A4 mRNA表达量。结果 YQFM在测试浓度条件下对人原代肝细胞的CYP1A2、CYP2B6和CYP3A4的酶活性和mRNA的表达均无诱导作用。结论 YQFM不具有显著的基于药物代谢酶诱导的中药和化学药的相互作用。     

四逆散改善Mdr2-/-小鼠胆汁淤积性肝纤维化作用

目的 研究四逆散对Mdr2(Abcb4)基因缺陷(Mdr2^-/-)小鼠胆汁淤积性肝纤维化的缓解作用,并探究其作用机制。方法 C57BL/6J小鼠作为对照组;C57BL/6J背景的Mdr2^-/-小鼠作为模型小鼠,设模型组和四逆散低、高剂量(按生药量计为3.12、6.24 g·kg^-1)组。四逆散组连续3周ig给予四逆散水提物,每天1次,对照组给予纯水。试剂盒法检测血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)、总胆汁酸水平;取肝脏、脾脏称质量,计算肝脏、脾脏系数;结合小鼠肝组织HE染色,Masson染色,纤连蛋白(Fibronectin)、细胞角蛋白19(CK19)的免疫组化染色,明确四逆散对Mdr2^-/-小鼠肝纤维化及胆汁淤积的影响。基于小鼠肝脏转录组学测序技术,挖掘四逆散改善Mdr2^-/-小鼠肝损伤的作用靶点,并通过实时荧光定量PCR(qRT-PCR)法检测肝纤维化[fibronectin(Fn1)、胶原蛋白1(Col1a1)、角蛋白19(krt19)]、炎症[白细胞介素-1β(Il1β)、Il6、肿瘤坏死因子-α(Tnfα)、一氧化氮合成酶(Inos)]、细胞焦亡[凋亡相关斑点样蛋白(Pycard)、Il18]、胆汁酸合成[细胞色素P450家族成员7A1(Cyp7a1)]及转运[胆盐输出泵(Abcb11)、ATP结合盒转运蛋白(Abcc3)、钠离子-牛磺胆酸共转运蛋白(Slc10a1)]相关基因的转录水平。结果 与模型组比较,四逆散高剂量显著降低血清总胆汁酸的水平(P＜0.05);明显缓解了Mdr2^-/-小鼠肝脏中央静脉及胆管周围炎性细胞的浸润和胶原纤维的沉积,并显著抑制胆管反应的发生。转录组学及qRT-PCR结果共同表明,四逆散下调Mdr2^-/-小鼠肝脏纤维化、炎症、细胞焦亡相关基因的转录(P＜0.05);同时,四逆散下调胆汁酸合成关键限速酶调控基因Cyp7a1和调控胆汁酸向肝内转运的基因Slc10a1的转录,并上调调控胆汁酸外排的基因Abcb11、Abcc3的转录。结论 四逆散能缓解Mdr2^-/-小鼠胆汁淤积性肝纤维化,机制可能与其调控炎症反应、细胞焦亡以及胆汁酸的合成和转运有关。     

Pregnane X receptor-mediated induction of Cyp3a by black cohosh

Black cohosh (BC) has been widely applied for the treatment of menopausal symptoms. However, increasing concerns about herb–drug interactions demand the need for studies on the influence of BC on cytochrome 450. Cyp3a11 in liver was induced by 7-fold in wild-type mice treated with 500?mg/kg black cohosh for 28 days compared with the control group as assessed by quantitative real-time PCR; no difference was found in small intestine and kidney, suggesting that up-regulation of Cyp3a11 by black cohosh was liver-specific. Western blot, activity assays, and pharmacokinetic analyses established dose- and time-dependent induction of Cyp3a11. To determine the mechanism of Cyp3a11 induction, including the role of pregnane X receptor (PXR) in vivo and in vitro, respectively, in Pxr-null, PXR-humanized, and double transgenic CYP3A4/hPXR mice, cell-based luciferase assays were employed revealing that mouse PXR played a direct role in the induction of Cyp3a11; human PXR was not activated by black cohosh. Overall, these findings demonstrate that induction of Cyp3a11 is liver-specific and involved only mouse PXR, not the human counterpart. Thus, the incidence of herb–drug interaction in patients administered black cohosh may not be mediated by human PXR and CYP3A4.     

Indirect activation of pregnane X receptor in the induction of hepatic CYP3A11 by high-dose rifampicin in mice

Rifampicin (RIF), a typical ligand of human pregnane X receptor (PXR), powerfully induces the expression of cytochrome P450 3A4 (CYP3A4) in humans. Although it is thought that RIF is not a ligand of rodent PXR, treatment with high-dose RIF (e.g. more than 20?mg/kg) increases the expression of CYP3A in the mouse liver. In this study, we investigated whether the induction of CYP3A by high-dose RIF in the mouse liver is mediated via indirect activation of mouse PXR (mPXR). The results showed that high-dose RIF increased the expression of CYP3A11 and other PXR-target genes in the liver of wild-type mice but not PXR-knockout mice. However, the results of reporter gene and ligand-dependent assembly assays showed that RIF does not activate mPXR in a ligand-dependent manner. In addition, high-dose RIF stimulated nuclear accumulation of mPXR in the mouse liver, and geldanamycin and okadaic acid attenuated the induction of Cyp3a11 and other PXR-target genes in primary hepatocytes, suggesting that high-dose RIF triggers nuclear translocation of mPXR. In conclusion, the present study suggests that high-dose RIF stimulates nuclear translocation of mPXR in the liver of mice by indirect activation, resulting in the transactivation of Cyp3a11 and other PXR-target genes.     

Induction of cytochrome P450 3A by paclitaxel in mice: pivotal role of the nuclear xenobiotic receptor, pregnane X receptor.

Paclitaxel, a taxane anti-microtubule agent, is known to induce CYP3A in rat and human hepatocytes. Recent studies suggest that a member of the nuclear receptor family, pregnane X Receptor (PXR), is a key regulator of the expression of CYP3A in different species. We investigated the role of PXR activation, in vitro and in vivo, in mediating Cyp3a induction by paclitaxel. Pregnenolone 16 alpha-carbonitrile (PCN), an antiglucocorticoid, was employed as a positive control for mouse PXR (mPXR) activation in vitro, and Cyp3a induction in vivo. In cell based reporter gene assays paclitaxel and PCN activated mPXR with an EC(50) of 5.6 and 0.27 microM, respectively. Employing PXR wild-type and transgenic mice lacking functional PXR (-/-), we evaluated the expression and activity of CYP3A following treatment with paclitaxel and PCN. Paclitaxel significantly induced CYP3A11 mRNA and immunoreactive CYP3A protein in PXR wild-type mice. Consistent with kinetics of CYP3A induction, the V(max) of testosterone 6 beta-hydroxylation in microsomal fraction increased 15- and 30-fold in paclitaxel- and PCN-treated mice, respectively. The Cyp3a induction response was completely abolished in paclitaxel- and PCN-treated PXR-null mice. This suggests that paclitaxel-mediated CYP3A induction in vivo requires an intact PXR-signaling mechanism. Our study validates the use of PXR activation assays in screening newer taxanes for potential drug interactions that may be related to PXR-target gene induction.     

Pregnane X receptor-mediated induction of Cyp3a by black cohosh

Black cohosh (BC) has been widely applied for the treatment of menopausal symptoms. However, increasing concerns about herb-drug interactions demand the need for studies on the influence of BC on cytochrome 450. Cyp3a11 in liver was induced by 7-fold in wild-type mice treated with 500?mg/kg black cohosh for 28 days compared with the control group as assessed by quantitative real-time PCR; no difference was found in small intestine and kidney, suggesting that up-regulation of Cyp3a11 by black cohosh was liver-specific. Western blot, activity assays, and pharmacokinetic analyses established dose- and time-dependent induction of Cyp3a11. To determine the mechanism of Cyp3a11 induction, including the role of pregnane X receptor (PXR) in vivo and in vitro, respectively, in Pxr-null, PXR-humanized, and double transgenic CYP3A4/hPXR mice, cell-based luciferase assays were employed revealing that mouse PXR played a direct role in the induction of Cyp3a11; human PXR was not activated by black cohosh. Overall, these findings demonstrate that induction of Cyp3a11 is liver-specific and involved only mouse PXR, not the human counterpart. Thus, the incidence of herb-drug interaction in patients administered black cohosh may not be mediated by human PXR and CYP3A4.     

Involvement of CAR and PXR in the transcriptional regulation of CYP2B6 gene expression by ingredients from herbal medicines

Abstract

1.?Induction of hepatic drug-metabolizing enzymes can affect drug efficacy and cause toxicity. However, so far, limited information is available regarding the molecular mechanism how herbal medicines induce human CYP2B6, which metabolizes many of the clinically used therapeutics and activates several pro-carcinogens or toxicants. Accumulated evidence suggests that the human constitutive androstane receptor (hCAR) and the human pregnane X receptor (hPXR) play important roles in trans-activation of CYP2B6. In this study, we investigated the effects of 68 Chinese herbal ingredients on the receptor specificity of hPXR/hCAR-mediated CYP2B6 induction by luciferase reporter gene assays in transiently transfected HepG2 cells and on the expression of CYP2B6 in LS174T cells.

2.?The HepG2 cells were transiently transfected with human CYP2B6 luciferase promoter reporter plasmids along with hPXR or hCAR₃. The results indicated that apigenin (Api), curcumol (Cur) and praeruptorin A (Pra A) were identified as potent activators of hPXR, and Pra A was also a ligand of hCAR.

3.?Furthermore, CYP2B6 mRNA expression in LS174T cells treated with the three herbal ingredients was determined by real-time polymerase chain reaction. By combining western blot and LC–MS/MS, CYP2B6 protein expression and catalytic activity induced by the three herbal ingredients were measured.

4.?Our observation showed Api and Cur up-regulated CYP2B6 expression by transactivation of hPXR, and Pra A acted as the ligand of both hPXR and hCAR to induce CYP2B6 expression.     

Induction of CYP3A expression by dehydroepiandrosterone: involvement of the pregnane X receptor.

Dehydroepiandrosterone (DHEA) is a steroid produced by the human adrenal gland. Administration of pharmacological doses of DHEA to rats changes expression of many genes, including the cytochrome P450 family members CYP4A1 and CYP3A23. It is known that induction of CYP4A expression by DHEA requires the peroxisome proliferator-activated receptor alpha (PPAR(alpha)). In the current study, PPAR(alpha)-null mice were used to examine the role of PPAR(alpha) in expression of CYP3A. In wild-type mice, 150 mg/kg DHEA-sulfate induced Cyp4a and Cyp3a11 mRNAs by 5- and 2-fold, respectively. Induction of Cyp4a expression by DHEA-sulfate was not observed in PPAR(alpha)-null mice, whereas induction of Cyp3a11 expression by DHEA-sulfate was similar between genotypes. This suggests that PPAR(alpha) is not involved in induction of Cyp3a11 expression by DHEA. Because expression of CYP3A family members can be induced by activation of another member of the nuclear receptor superfamily, the pregnane X receptor (PXR), we examined the ability of DHEA to activate PXR. In transient transfection assays, DHEA and its metabolites androst-5-ene-3beta,17beta-diol (ADIOL), androst-5-ene-3,17-dione, and androst-4-ene-3,17-dione were activators of PXR. Maximal induction of a PXR-responsive reporter gene of approximately 3-fold was observed at concentrations of 50 to 100 microM, indicating that these steroids are relatively weak activators of PXR. Human and murine PXR exhibited different specificities for DHEA and its metabolites. ADIOL activated reporter gene expression in the presence of murine but not human PXR. Results of these studies suggest that the induction of rodent CYP3A expression upon treatment with high doses of DHEA occurs through activation of PXR.     

阿司匹林联合华法林对HepG2细胞中CYP3A4酶的抑制作用

目的研究阿司匹林联用华法林对肝药物代谢酶CYP3A4活性的影响及其机制。方法不同质量浓度的阿司匹林、华法林及联合用药处理Hep G2细胞48 h,采用MTT法检测细胞存活率;通过荧光素酶报告基因技术检测各组对PXR转录酶活性和酶CYP3A4活性的影响;采用荧光定量PCR法和Western blotting法检测Hep G2细胞的酶CYP3A4的m RNA和蛋白表达水平。结果与对照组比较,阿司匹林组、联合用药组细胞的孕烷X受体(PXR)转录酶活性、酶CYP3A4活性均显著降低(P0.01),CYP3A4 m RNA和蛋白表达水平显著降低(P0.05、0.01),华法林组均无显著差异。结论阿司匹林联用华法林能够抑制药物代谢酶CYP3A4活性,其机制可能是通过抑制PXR受体的m RNA和蛋白表达实现的。     

PXR-mediated induction of human CYP3A4 and mouse Cyp3a11 by the glucocorticoid budesonide

Budesonide, a glucocorticoid with a high first-pass metabolism, is used for the oral treatment of inflammatory bowel disease. Cytochrome P450 3A4 (CYP3A4) is an enzyme involved in the metabolism of numerous drugs, including budesonide. Since inhibition or induction of CYP3A4 is often the cause of drug–drug interactions we analyzed how budesonide affects the activity and expression of this enzyme. CYP3A4 activity was assessed by the metabolism of a luminogenic substrate (luciferin-benzylether) using recombinant human CYP3A4 protein. We observed no inhibition of the metabolism in the presence of budesonide at concentrations up to 25 μM. Induction experiments in human LS180 colon carcinoma cells showed an increased expression of CYP3A4 mRNA after budesonide treatment. Transactivation assays revealed that budesonide activates the CYP3A4 promoter via the pregnane X receptor (PXR). In mice, oral budesonide administration (25 mg/kg) for 4 days induced the murine homolog Cyp3a11 in the intestine 3-fold, whereas liver expression was notably less influenced. In knockout mice devoid of PXR, budesonide-mediated inductions were reduced compared to wild-type mice. In conclusion, we could demonstrate that budesonide is not an efficient inhibitor but rather an inducer of CYP3A via a PXR-mediated mechanism. In vivo, however, oral budesonide administration to mice showed only modest gene induction, which occurred mainly in the intestine. Therefore, the risk for budesonide-mediated drug interactions seems to be low but cannot be ruled out entirely.