AIM: We have previously demonstrated that cholangiocytes, the epithelial cells lining intrahepatic bile ducts,encode two functional bile acid transporters via alternative splicing of a single gene to facilitate bile a...AIM: We have previously demonstrated that cholangiocytes, the epithelial cells lining intrahepatic bile ducts,encode two functional bile acid transporters via alternative splicing of a single gene to facilitate bile acid vectorial transport. Cholangiocytes possess ASBT,an apical sodium-dependent bile acid transporter to take up bile acids,and t-ASBT,a basolateral alternatively spliced and truncated form of ASBT to efflux bile acids.Though hepatocyte and ileal bile acid transporters are in part regulated by the flux of bile acids, the effect of alterations in bile acid flux on the expression of t-ASBT in terminal ileocytes remains undear.Thus,we tested the hypothesis that expression of ASBT and t-ASBT in cholangiocytes and ileocytes was regulated by bile acid flux. METHODS: Expression of ASBT and t-ASBT message and protein in cholangiocytes and ileocytes isolated from pair- fed rats given control (C) and 1% taurocholate (TCA) or 5% cholestyramine (CY) enriched diets,were assessed by both quantitative RNase protection assays and quantitative immunoblotting.The data obtained from each of the control groups were pooled to reflect the changes observed following TCA and CY treatments with respect to the control diets. Cholangiocyte taurocholate uptake was determined using a novel microperfusion technique on intrahepatic bile duct units (IBDUs) derived from C,TCA and CY fed rats. RESULTS: In cholangiocytes,both ASBT and t-ASBT message RNA and protein were significantly decreased in response to TCA feeding compared to C diet.In contrast, message and protein of both bile acid transporters significantly increased following CY feeding compared to C diet.In the ileum,TCA feeding significantly up-regulated both ASBT and t-ASBT message and protein compared to C diet,while CY feeding significantly down-regulated message and protein of both bile acid transporters compared to C diet.As anticipated from alterations in cholangiocyte ASBT expression,the uptake of taurocholate in microperfused IBDUs derived from rats on TCA diet decreased 2.7-fold,whereas it increased 1.7-fold in those on CY diet compared to C diet fed groups. CONCLUSION: These data demonstrate that expression of ASBT and t-ASBT in cholangiocytes is regulated by a negative feedback loop while the expression of these transporters in terminal ileum is modified via positive feedback.Thus, while transcriptional regulatory mechanisms in response to alterations in bile acid pool size are operative in both cholangiocytes and ileocytes,each cell type responds differently to bile acid supplementation and depletion.展开更多
Objective:Impaired active fluid transport of al veolar epithelium may involve in the pathogenesis and resolution of alveolar ede ma. The objective of this study was to explore the changes in alveolar epithelia l liqui...Objective:Impaired active fluid transport of al veolar epithelium may involve in the pathogenesis and resolution of alveolar ede ma. The objective of this study was to explore the changes in alveolar epithelia l liquid clearance during lung edema following acute lung injury induced by olei c acid. Methods:Forty-eight Wistar rats were randomly divided into si x groups, i.e., injured, amiloride, ouabain, amiloride plus ouabain and terbutal ine groups. Twenty-four hours after the induction of acute lung injury by intra venous oleic acid ( 0.25 ml/kg), 5% albumin solution with 1.5 μCi 12 5 I-labeled albumin (5 ml/kg) was delivered into both lungs via trachea. Alveo lar liquid clearance (ALC), extravascular lung water (EVLW) content and arterial blood gases were measured one hour thereafter. Results:At 24 h after the infusion of oleic acid, the rats dev eloped pulmonary edema and severe hypoxemia, with EVLW increased by 47.9 % and ALC decreased by 49.2 %. Addition of either 2×10 -3 M amiloride or 5× 10 -4 M ouabain to the instillation further reduced ALC and increased E VLW. ALC increased by approximately 63.7 % and EVLW decreased by 46.9 % wi th improved hypoxemia in the Terbutaline (10 -4 M) group, compared those in injured rats. A significant negative correlation was found between the incremen t of EVLW and the reduction of ALC. Conclusions:Active fluid transport of alveolar epithelium migh t play a role in the pathogenesis of lung edema in acute lung injury.展开更多
The application of nanomedicines in oral drug delivery effectively promotes the drug absorption and transportation through enterocytes.Nevertheless,the absence of mechanism studies on efficacy and safety limits their ...The application of nanomedicines in oral drug delivery effectively promotes the drug absorption and transportation through enterocytes.Nevertheless,the absence of mechanism studies on efficacy and safety limits their final translation in humans.Although the vesicular trafficking has been verified as the general character for transport of nanomedicines,the deeper mechanism in molecular mechanism is still unclear.Moreover,the cellular transport of nanomedicines is a dynamic process involved by different organelles and components.However,most of existing studies just pay attention to the static location of nanomedicines,but neglect the dynamic biological effects on cells caused by them.Here,we prepared gold nanoparticles(Au NPs)as the model and cultured epithelial cell monolayer to explore the nano-bio interactions at the molecular level.The traditional pharmacological inhibition strategy and subcellular imaging technology elucidated the macropinocytosis/endosome/MVB/lysosome pathway during the transportation of Au NPs.Proteomics strategy based on mass spectrometry(MS)was utilized to identify and quantify proteins involved in the cellular transport of nanomedicines.Multiple proteins related to subcellular structure,signal transduction,energy transformation and metabolism regulation were demonstrated to be regulated by nanoparticle transport.These alterations of protein expression clarified the effects of intracellular proteins and verified the conventional findings.More importantly,it revealed a feedback mechanism of cells to the nano-trafficking.We believed that these new regulatory mechanisms provided new insights into the efficient transport of nanomedicines through epithelial barriers.展开更多
文摘AIM: We have previously demonstrated that cholangiocytes, the epithelial cells lining intrahepatic bile ducts,encode two functional bile acid transporters via alternative splicing of a single gene to facilitate bile acid vectorial transport. Cholangiocytes possess ASBT,an apical sodium-dependent bile acid transporter to take up bile acids,and t-ASBT,a basolateral alternatively spliced and truncated form of ASBT to efflux bile acids.Though hepatocyte and ileal bile acid transporters are in part regulated by the flux of bile acids, the effect of alterations in bile acid flux on the expression of t-ASBT in terminal ileocytes remains undear.Thus,we tested the hypothesis that expression of ASBT and t-ASBT in cholangiocytes and ileocytes was regulated by bile acid flux. METHODS: Expression of ASBT and t-ASBT message and protein in cholangiocytes and ileocytes isolated from pair- fed rats given control (C) and 1% taurocholate (TCA) or 5% cholestyramine (CY) enriched diets,were assessed by both quantitative RNase protection assays and quantitative immunoblotting.The data obtained from each of the control groups were pooled to reflect the changes observed following TCA and CY treatments with respect to the control diets. Cholangiocyte taurocholate uptake was determined using a novel microperfusion technique on intrahepatic bile duct units (IBDUs) derived from C,TCA and CY fed rats. RESULTS: In cholangiocytes,both ASBT and t-ASBT message RNA and protein were significantly decreased in response to TCA feeding compared to C diet.In contrast, message and protein of both bile acid transporters significantly increased following CY feeding compared to C diet.In the ileum,TCA feeding significantly up-regulated both ASBT and t-ASBT message and protein compared to C diet,while CY feeding significantly down-regulated message and protein of both bile acid transporters compared to C diet.As anticipated from alterations in cholangiocyte ASBT expression,the uptake of taurocholate in microperfused IBDUs derived from rats on TCA diet decreased 2.7-fold,whereas it increased 1.7-fold in those on CY diet compared to C diet fed groups. CONCLUSION: These data demonstrate that expression of ASBT and t-ASBT in cholangiocytes is regulated by a negative feedback loop while the expression of these transporters in terminal ileum is modified via positive feedback.Thus, while transcriptional regulatory mechanisms in response to alterations in bile acid pool size are operative in both cholangiocytes and ileocytes,each cell type responds differently to bile acid supplementation and depletion.
文摘Objective:Impaired active fluid transport of al veolar epithelium may involve in the pathogenesis and resolution of alveolar ede ma. The objective of this study was to explore the changes in alveolar epithelia l liquid clearance during lung edema following acute lung injury induced by olei c acid. Methods:Forty-eight Wistar rats were randomly divided into si x groups, i.e., injured, amiloride, ouabain, amiloride plus ouabain and terbutal ine groups. Twenty-four hours after the induction of acute lung injury by intra venous oleic acid ( 0.25 ml/kg), 5% albumin solution with 1.5 μCi 12 5 I-labeled albumin (5 ml/kg) was delivered into both lungs via trachea. Alveo lar liquid clearance (ALC), extravascular lung water (EVLW) content and arterial blood gases were measured one hour thereafter. Results:At 24 h after the infusion of oleic acid, the rats dev eloped pulmonary edema and severe hypoxemia, with EVLW increased by 47.9 % and ALC decreased by 49.2 %. Addition of either 2×10 -3 M amiloride or 5× 10 -4 M ouabain to the instillation further reduced ALC and increased E VLW. ALC increased by approximately 63.7 % and EVLW decreased by 46.9 % wi th improved hypoxemia in the Terbutaline (10 -4 M) group, compared those in injured rats. A significant negative correlation was found between the incremen t of EVLW and the reduction of ALC. Conclusions:Active fluid transport of alveolar epithelium migh t play a role in the pathogenesis of lung edema in acute lung injury.
基金The National Key R&D Program of China(Grant No.2017YFA0205600)the National Natural Science Foundation of China(Grant No.81690264,81573359 and 81703441)。
文摘The application of nanomedicines in oral drug delivery effectively promotes the drug absorption and transportation through enterocytes.Nevertheless,the absence of mechanism studies on efficacy and safety limits their final translation in humans.Although the vesicular trafficking has been verified as the general character for transport of nanomedicines,the deeper mechanism in molecular mechanism is still unclear.Moreover,the cellular transport of nanomedicines is a dynamic process involved by different organelles and components.However,most of existing studies just pay attention to the static location of nanomedicines,but neglect the dynamic biological effects on cells caused by them.Here,we prepared gold nanoparticles(Au NPs)as the model and cultured epithelial cell monolayer to explore the nano-bio interactions at the molecular level.The traditional pharmacological inhibition strategy and subcellular imaging technology elucidated the macropinocytosis/endosome/MVB/lysosome pathway during the transportation of Au NPs.Proteomics strategy based on mass spectrometry(MS)was utilized to identify and quantify proteins involved in the cellular transport of nanomedicines.Multiple proteins related to subcellular structure,signal transduction,energy transformation and metabolism regulation were demonstrated to be regulated by nanoparticle transport.These alterations of protein expression clarified the effects of intracellular proteins and verified the conventional findings.More importantly,it revealed a feedback mechanism of cells to the nano-trafficking.We believed that these new regulatory mechanisms provided new insights into the efficient transport of nanomedicines through epithelial barriers.
