AIM To ascertain whether cholestasis affects the expression of two CYP3 A isoforms(CYP3 A1 and CYP3 A2) and of pregnane X receptor(PXR) and constitutive androstane receptor(CAR).METHODS Cholestasis was induced by bile...AIM To ascertain whether cholestasis affects the expression of two CYP3 A isoforms(CYP3 A1 and CYP3 A2) and of pregnane X receptor(PXR) and constitutive androstane receptor(CAR).METHODS Cholestasis was induced by bile duct ligation in 16 male Wistar rats; whereas 8 sham-operated rats were used as controls. Severity of cholestasis was assessed on histological examination of liver sections, and serum concentrations of albumin, AST, ALT, GGT, ALPK and bilirubin. Gene and protein expressions of PXR, CAR, CYP3 A1 and CYP3 A2 were assessed by means of q RT-PCR and Western blot, respectively. Alterations in CYP3 A activity were measured by calculating the kinetic parameters of 4-OH and 1'-OH-midazolam hydroxylation, marker reactions for CYP3 A enzymes.RESULTS The m RNA and protein expression of CYP3 A1 increased significantly in mild cholestasis(P < 0.01). At variance, m RNA and protein expression of CYP3 A2 didn't change in mild cholestasis, whereas the expression and activity of both CYP3 A1 and CYP3 A2 decreased dramatically when cholestasis became severe. Consistently with these observations, the nuclear expression of both PXR and CAR, which was measured because they both translocate into the cell nucleus after their activation, virtually disappeared in the late stage of cholestatic injury, after an initial increase. These results indicate that early-and late-stage cholestasis affects CYP3 Amediated drug metabolism differently, probably as consequence of the different activation of PXR and CAR.CONCLUSION Early-and late-stage cholestasis affects CYP3 Amediated drug metabolism differently. PXR and CAR might be targeted therapeutically to promote CYP3 Amediated liver detoxification.展开更多
This study aimed to determine the regulatory mechanism of dietary zinc lactate(ZL)supplementation on intestinal oxidative stress damage in a paraquat(PQ)-induced piglet model.Twenty-eight piglets(mean body weight 9.51...This study aimed to determine the regulatory mechanism of dietary zinc lactate(ZL)supplementation on intestinal oxidative stress damage in a paraquat(PQ)-induced piglet model.Twenty-eight piglets(mean body weight 9.51±0.23 kg)weaned at 28 d of age were randomly divided into control,ZL,PQ,and ZL+PQ groups(n=7 in each group).The ZL-supplemented diet had little effect on growth performance under normal physiological conditions.However,under PQ challenge,ZL supplementation significantly improved average daily gain(P<0.05)and reduced the frequency of diarrhea.ZL improved intestinal morphology and ultrastructure by significantly increasing the expression level of the jejunal tight junction protein,zonula occludens-1(ZO-1)(P<0.05),and intestinal zinc transport and absorption in PQ-induced piglets,which reduced intestinal permeability.ZL supplementation also enhanced the expression of antioxidant and antiinflammatory factor-related genes and decreased inflammatory cytokine expression and secretion in PQinduced piglets.Furthermore,ZL treatment significantly inhibited the activation of constitutive androstane receptor(CAR)signaling(P<0.01)in PQ-induced piglets and altered the structure of the gut microbiota,especially by significantly increasing the abundance of beneficial gut microbes,including UCG_002,Ruminococcus,Rikenellaceae_RC9_gut_group,Christensenellaceae_R_7_group,Treponema,unclassified_Christensenellaceae,and unclassified_Erysipelotrichaceae(P<0.05).These data reveal that pre-administration of ZL to piglets can suppress intestinal oxidative stress by improving antioxidant and anti-inflammatory capacity and regulating the crosstalk between CAR signaling and gut microbiota.展开更多
The constitutive androstane receptor （CAR, NRll3） plays a crucial role in the regulation of drug metabolism, energy homeostasis, and cancer development through modulating the transcription of its numerous target gen...The constitutive androstane receptor （CAR, NRll3） plays a crucial role in the regulation of drug metabolism, energy homeostasis, and cancer development through modulating the transcription of its numerous target genes. Different from prototypical nuclear receptors, CAR can be activated by either direct ligand binding or ligand-independent （indirect） mechanisms both initiated with nuclear translocation of CAR from the cytoplasm. In comparison to the well-defined ligand-based activation, indirect activation of CAR appears to be exclusively involved in the nuclear translocation through mecha- nisms yet to be fully understood. Accumulating evi- dence reveals that without activation, CAR forms a protein complex in the cytoplasm where it can be func- tionally affected by multiple signaling pathways. In this review, we discuss recent progresses in our under- standing of the signaling regulation of CAR nuclear accumulation and activation. We expect that this review will also provide greater insight into the similarity and difference between the mechanisms of direct vs. indirect human CAR activation.展开更多
The constitutive androstane receptor(CAR, NR3 I1) belongs to nuclear receptor superfamily.It was reported that CAR agonist TCPOBOP induces hepatomegaly but the underlying mechanism remains largely unknown. Yes-associa...The constitutive androstane receptor(CAR, NR3 I1) belongs to nuclear receptor superfamily.It was reported that CAR agonist TCPOBOP induces hepatomegaly but the underlying mechanism remains largely unknown. Yes-associated protein(YAP) is a potent regulator of organ size. The aim of this study is to explore the role of YAP in CAR activation-induced hepatomegaly and liver regeneration.TCPOBOP-induced CAR activation on hepatomegaly and liver regeneration was evaluated in wildtype(WT) mice, liver-specific YAP-deficient mice, and partial hepatectomy(PHx) mice. The results demonstrate that TCPOBOP can increase the liver-to-body weight ratio in wild-type mice and PHx mice.Hepatocytes enlargement around central vein(CV) area was observed, meanwhile hepatocytesproliferation was promoted as evidenced by the increased number of KI67+cells around portal vein(PV)area. The protein levels of YAP and its downstream targets were upregulated in TCPOBOP-treated mice and YAP translocation can be induced by CAR activation. Co-immunoprecipitation results suggested a potential proteineprotein interaction of CAR and YAP. However, CAR activation-induced hepatomegaly can still be observed in liver-specific YAP-deficient(Yape/e) mice. In summary, CAR activation promotes hepatomegaly and liver regeneration partially by inducing YAP translocation and interaction with YAP signaling pathway, which provides new insights to further understand the physiological functions of CAR.展开更多
The canalicular membrane represents the excretory pole of hepatocytes.Bile is an important route of elimination of potentially toxic endo-and xenobiotics(including drugs and toxins),mediated by the major canalicular t...The canalicular membrane represents the excretory pole of hepatocytes.Bile is an important route of elimination of potentially toxic endo-and xenobiotics(including drugs and toxins),mediated by the major canalicular transporters:multidrug resistance protein 1(MDR1, ABCB1),also known as P-glycoprotein,multidrug resistance-associated protein 2(MRP2,ABCC2),and the breast cancer resistance protein(BCRP,ABCG2).Their activities depend on regulation of expression and proper localization at the canalicular membrane,as regulated by transcriptional and post-transcriptional events,respectively.At transcriptional level,specific nuclear receptors(NR)s modulated by ligands,co-activators and co-repressors,mediate the physiological requirements of these transporters.This complex system is also responsible for alterations occurring in specific liver pathologies.We briefly describe the major ClassⅡNRs, pregnane X receptor(PXR)and constitutive androstane receptor(CAR),and their role in regulating expression of multidrug resistance proteins.Several therapeutic agents regulate the expression of relevant drug transporters through activation/inactivation of these NRs.We provide some representative examples of the action of therapeutic agents modulating liver drug transporters, which in addition,involve CAR or PXR as mediators.展开更多
The constitutive androstane receptor (CAR) is a transcription factor that belongs to the nuclear receptor superfamily. CAR binds as a heterodimer with the retinoid X receptor α (RXRα) to CAR response elements (CAREs...The constitutive androstane receptor (CAR) is a transcription factor that belongs to the nuclear receptor superfamily. CAR binds as a heterodimer with the retinoid X receptor α (RXRα) to CAR response elements (CAREs) and regulates the expression of various drug metabolizing enzymes and transporters. To identify CAR/RXRα binding sites in the human genome, we performed a modified yeast one-hybrid assay that enables rapid and efficient identification of genomic targets for DNA-binding proteins. DNA fragments were recovered from positive yeast colonies by PCR and sequenced. A motif enrichment analysis revealed that the most frequent motif was a direct repeat (DR) of RGKTCA-like core sequence spaced by 4 bp. Next, we predicted 149 putative CAR/RXRα binding sites from 414 unique clones, by searching for DRs, everted repeats (ERs) and inverted repeats (IRs) of the RGKTCA-like core motif. Based on gel mobility shift assays, the CAR/RXRα heterodimer could directly interact with the 108 predicted sequences, which included not only classical CAREs but also a wide variety of arrangements. Furthermore, we identified 17 regulatory polymorphisms on the CAR/RXRα-binding sites that may influence individual variation in the expression of CAR-regulated genes. These results provide insights into the molecular mechanisms underlying the physiological and pathological actions of CAR/RXRα het-erodimers.展开更多
The nuclear receptors pregnane X receptor(PXR) and constitutive androstane receptor(CAR) were cloned and/or established as xenobiotic receptors in 1998.Due to their activities in the transcriptional regulation of phas...The nuclear receptors pregnane X receptor(PXR) and constitutive androstane receptor(CAR) were cloned and/or established as xenobiotic receptors in 1998.Due to their activities in the transcriptional regulation of phase I and phase II enzymes as well as drug transporters,PXR and CAR have been defined as the master regulators of xenobiotic responses.The discovery of PXR and CAR provides the essential molecular basis by which drugs and other xenobiotic compounds regulate the expression of xenobiotic enzymes and transporters.This article is intended to provide a historical overview on the discovery of PXR and CAR as xenobiotic receptors.展开更多
基金Supported by the University of Padova,No.CPDA138721/13
文摘AIM To ascertain whether cholestasis affects the expression of two CYP3 A isoforms(CYP3 A1 and CYP3 A2) and of pregnane X receptor(PXR) and constitutive androstane receptor(CAR).METHODS Cholestasis was induced by bile duct ligation in 16 male Wistar rats; whereas 8 sham-operated rats were used as controls. Severity of cholestasis was assessed on histological examination of liver sections, and serum concentrations of albumin, AST, ALT, GGT, ALPK and bilirubin. Gene and protein expressions of PXR, CAR, CYP3 A1 and CYP3 A2 were assessed by means of q RT-PCR and Western blot, respectively. Alterations in CYP3 A activity were measured by calculating the kinetic parameters of 4-OH and 1'-OH-midazolam hydroxylation, marker reactions for CYP3 A enzymes.RESULTS The m RNA and protein expression of CYP3 A1 increased significantly in mild cholestasis(P < 0.01). At variance, m RNA and protein expression of CYP3 A2 didn't change in mild cholestasis, whereas the expression and activity of both CYP3 A1 and CYP3 A2 decreased dramatically when cholestasis became severe. Consistently with these observations, the nuclear expression of both PXR and CAR, which was measured because they both translocate into the cell nucleus after their activation, virtually disappeared in the late stage of cholestatic injury, after an initial increase. These results indicate that early-and late-stage cholestasis affects CYP3 Amediated drug metabolism differently, probably as consequence of the different activation of PXR and CAR.CONCLUSION Early-and late-stage cholestasis affects CYP3 Amediated drug metabolism differently. PXR and CAR might be targeted therapeutically to promote CYP3 Amediated liver detoxification.
基金supported by the National Natural Science Foundation of China(32172755,32130099)Hunan Key Research and Development Plan(2022NK2023)Shandong Province Taishan Industry Leading Talents Project Blue Talents Project
文摘This study aimed to determine the regulatory mechanism of dietary zinc lactate(ZL)supplementation on intestinal oxidative stress damage in a paraquat(PQ)-induced piglet model.Twenty-eight piglets(mean body weight 9.51±0.23 kg)weaned at 28 d of age were randomly divided into control,ZL,PQ,and ZL+PQ groups(n=7 in each group).The ZL-supplemented diet had little effect on growth performance under normal physiological conditions.However,under PQ challenge,ZL supplementation significantly improved average daily gain(P<0.05)and reduced the frequency of diarrhea.ZL improved intestinal morphology and ultrastructure by significantly increasing the expression level of the jejunal tight junction protein,zonula occludens-1(ZO-1)(P<0.05),and intestinal zinc transport and absorption in PQ-induced piglets,which reduced intestinal permeability.ZL supplementation also enhanced the expression of antioxidant and antiinflammatory factor-related genes and decreased inflammatory cytokine expression and secretion in PQinduced piglets.Furthermore,ZL treatment significantly inhibited the activation of constitutive androstane receptor(CAR)signaling(P<0.01)in PQ-induced piglets and altered the structure of the gut microbiota,especially by significantly increasing the abundance of beneficial gut microbes,including UCG_002,Ruminococcus,Rikenellaceae_RC9_gut_group,Christensenellaceae_R_7_group,Treponema,unclassified_Christensenellaceae,and unclassified_Erysipelotrichaceae(P<0.05).These data reveal that pre-administration of ZL to piglets can suppress intestinal oxidative stress by improving antioxidant and anti-inflammatory capacity and regulating the crosstalk between CAR signaling and gut microbiota.
文摘The constitutive androstane receptor （CAR, NRll3） plays a crucial role in the regulation of drug metabolism, energy homeostasis, and cancer development through modulating the transcription of its numerous target genes. Different from prototypical nuclear receptors, CAR can be activated by either direct ligand binding or ligand-independent （indirect） mechanisms both initiated with nuclear translocation of CAR from the cytoplasm. In comparison to the well-defined ligand-based activation, indirect activation of CAR appears to be exclusively involved in the nuclear translocation through mecha- nisms yet to be fully understood. Accumulating evi- dence reveals that without activation, CAR forms a protein complex in the cytoplasm where it can be func- tionally affected by multiple signaling pathways. In this review, we discuss recent progresses in our under- standing of the signaling regulation of CAR nuclear accumulation and activation. We expect that this review will also provide greater insight into the similarity and difference between the mechanisms of direct vs. indirect human CAR activation.
基金supported by the Natural Science Foundation of China (Grant numbers:82025034 and 81973392)the National Key Research and Development Program (Grant number:2017YFE0109900, China)+5 种基金the Shenzhen Science and Technology Program (Grant number:KQTD20190929174023858, China)the Natural Science Foundation of Guangdong (Grant number:2017A030311018, China)the 111 project (Grant number:B16047, China)the Key Laboratory Foundation of Guangdong Province (Grant number:2017B030314030, China)the Local Innovative and Research Teams Project of Guangdong Pearl River Talents Program (Grant number:2017BT01Y093, China)the National Engineering and Technology Research Center for New drug Druggability Evaluation (Seed Program of Guangdong Province, Grant number:2017B090903004, China)。
文摘The constitutive androstane receptor(CAR, NR3 I1) belongs to nuclear receptor superfamily.It was reported that CAR agonist TCPOBOP induces hepatomegaly but the underlying mechanism remains largely unknown. Yes-associated protein(YAP) is a potent regulator of organ size. The aim of this study is to explore the role of YAP in CAR activation-induced hepatomegaly and liver regeneration.TCPOBOP-induced CAR activation on hepatomegaly and liver regeneration was evaluated in wildtype(WT) mice, liver-specific YAP-deficient mice, and partial hepatectomy(PHx) mice. The results demonstrate that TCPOBOP can increase the liver-to-body weight ratio in wild-type mice and PHx mice.Hepatocytes enlargement around central vein(CV) area was observed, meanwhile hepatocytesproliferation was promoted as evidenced by the increased number of KI67+cells around portal vein(PV)area. The protein levels of YAP and its downstream targets were upregulated in TCPOBOP-treated mice and YAP translocation can be induced by CAR activation. Co-immunoprecipitation results suggested a potential proteineprotein interaction of CAR and YAP. However, CAR activation-induced hepatomegaly can still be observed in liver-specific YAP-deficient(Yape/e) mice. In summary, CAR activation promotes hepatomegaly and liver regeneration partially by inducing YAP translocation and interaction with YAP signaling pathway, which provides new insights to further understand the physiological functions of CAR.
基金Grants from Agencia Nacional de Promoción Científicay Tecnológica (PICT N° 05-26306)Consejo Nacional de Investigaciones Científicasy Técnicas (PIP N° 6442)Universidad Nacional de Rosario,Argentina
文摘The canalicular membrane represents the excretory pole of hepatocytes.Bile is an important route of elimination of potentially toxic endo-and xenobiotics(including drugs and toxins),mediated by the major canalicular transporters:multidrug resistance protein 1(MDR1, ABCB1),also known as P-glycoprotein,multidrug resistance-associated protein 2(MRP2,ABCC2),and the breast cancer resistance protein(BCRP,ABCG2).Their activities depend on regulation of expression and proper localization at the canalicular membrane,as regulated by transcriptional and post-transcriptional events,respectively.At transcriptional level,specific nuclear receptors(NR)s modulated by ligands,co-activators and co-repressors,mediate the physiological requirements of these transporters.This complex system is also responsible for alterations occurring in specific liver pathologies.We briefly describe the major ClassⅡNRs, pregnane X receptor(PXR)and constitutive androstane receptor(CAR),and their role in regulating expression of multidrug resistance proteins.Several therapeutic agents regulate the expression of relevant drug transporters through activation/inactivation of these NRs.We provide some representative examples of the action of therapeutic agents modulating liver drug transporters, which in addition,involve CAR or PXR as mediators.
文摘The constitutive androstane receptor (CAR) is a transcription factor that belongs to the nuclear receptor superfamily. CAR binds as a heterodimer with the retinoid X receptor α (RXRα) to CAR response elements (CAREs) and regulates the expression of various drug metabolizing enzymes and transporters. To identify CAR/RXRα binding sites in the human genome, we performed a modified yeast one-hybrid assay that enables rapid and efficient identification of genomic targets for DNA-binding proteins. DNA fragments were recovered from positive yeast colonies by PCR and sequenced. A motif enrichment analysis revealed that the most frequent motif was a direct repeat (DR) of RGKTCA-like core sequence spaced by 4 bp. Next, we predicted 149 putative CAR/RXRα binding sites from 414 unique clones, by searching for DRs, everted repeats (ERs) and inverted repeats (IRs) of the RGKTCA-like core motif. Based on gel mobility shift assays, the CAR/RXRα heterodimer could directly interact with the 108 predicted sequences, which included not only classical CAREs but also a wide variety of arrangements. Furthermore, we identified 17 regulatory polymorphisms on the CAR/RXRα-binding sites that may influence individual variation in the expression of CAR-regulated genes. These results provide insights into the molecular mechanisms underlying the physiological and pathological actions of CAR/RXRα het-erodimers.
基金supported in part by the Joseph Koslow Endowed Professorship from the University of Pittsburgh School of Pharmacy
文摘The nuclear receptors pregnane X receptor(PXR) and constitutive androstane receptor(CAR) were cloned and/or established as xenobiotic receptors in 1998.Due to their activities in the transcriptional regulation of phase I and phase II enzymes as well as drug transporters,PXR and CAR have been defined as the master regulators of xenobiotic responses.The discovery of PXR and CAR provides the essential molecular basis by which drugs and other xenobiotic compounds regulate the expression of xenobiotic enzymes and transporters.This article is intended to provide a historical overview on the discovery of PXR and CAR as xenobiotic receptors.
