Gradual alterations of cell’s physiology and functions due to age or exposure to various stresses lead to the conversion of normal cells to senescent cells.Once becoming senescent,the cell stops dividing permanently ...Gradual alterations of cell’s physiology and functions due to age or exposure to various stresses lead to the conversion of normal cells to senescent cells.Once becoming senescent,the cell stops dividing permanently but remains metabolically active.Cellular senescence does not have a single marker but is characterized mainly by a combination of multiple markers,such as,morphological changes,expression of cell cycle inhibitors,senescence associatedβ-galactosidase activity,and changes in nuclear membrane.When cells in an organ become senescent,the entire organism can be affected.This may occur through the senescence-associated secretory phenotype(SASP).SASP may exert beneficial or harmful effects on the microenvironment of tissues.Research on senescence has become a very exciting field in cell biology since the link between age-related diseases,including cancer,and senescence has been established.The loss of regenerative and homeostatic capacity of the liver over the age is somehow connected to cellular senescence.The major contributors of senescence properties in the liver are hepatocytes and cholangiocytes.Senescent cells in the liver have been implicated in the etiology of chronic liver diseases including cirrhosis and hepatocellular carcinoma and in the interference of liver regeneration.This review summarizes recently reported findings in the understanding of the molecular mechanisms of senescence and its relationship with liver diseases.展开更多
Cholangiocytes, the epithelial cells lining the biliary ducts, are the target cells in several liver diseases. Cholangiopathies and cholangiocarcinoma generate interest in many scientists since the genesis. The develo...Cholangiocytes, the epithelial cells lining the biliary ducts, are the target cells in several liver diseases. Cholangiopathies and cholangiocarcinoma generate interest in many scientists since the genesis. The developing mechanisms, and the therapeutic tools of these diseases are still undefined. Several studies demonstrate that many hormones, neuropeptides and neurotransmitters regulate malignant and non-malignant cholangiocyte pathophysiology in the course of chronic biliary diseases. The aim of this review is to present the findings of several studies published in the recent years that contributed to clarifying the role of nervous and neuroendocrine regulation of the pathophysiologic events associated with cholestasis and cholangiocarcinoma development. This manuscript is organized into two parts. The first part offers an overview of the innervation of the liver and the origin of neuroendocrine hormones, neurotransmitters and neuropeptides affecting cholangiocyte function and metabolism. The first section also reviews the effects played by several neuroendocrine hormones and nervous system on cholangiocyte growth, survival and functional activity in the course of cholestasis. In the second section, we summarize the results of some studies describing the role of nervous system and neuroendocrine hormones in the regulation of malignant cholangiocyte growth.展开更多
Objective:Tripterygium glycoside(TG)is widely used in clinical practice for its multiple bioactivities including anti-inflammatory and immunosuppressive effects.However,emerging studies have frequently reported TG-ind...Objective:Tripterygium glycoside(TG)is widely used in clinical practice for its multiple bioactivities including anti-inflammatory and immunosuppressive effects.However,emerging studies have frequently reported TG-induced adverse reactions to multiple organs,especially liver.Here,this study aimed to investigate the mechanism of liver damage induced by TG and explore representative components to reflect TG hepatotoxicity.Methods:Network pharmacology was used to determine the potential targets of bile duct injury caused by TG.Next,the hepatotoxic effects of TG,triptolide(TP)and celastrol(CEL)were investigated and compared in vivo and in vitro.Liver function was determined by measuring serum transaminase and histopathology staining.The cell proliferation and apoptosis were determined by cell viability assay,scratch assay and flow cytometry.The expression of gene of interest was determined by qPCR and Western blot.Results:Based on the network pharmacological analysis of 12 bioactive ingredients found in TG,a total of35 targets and 15 pathways related to bile duct injury were obtained.Both TG and TP resulted in cholangiocyte damage and liver injury,as illustrated by increased levels of serum transaminase and oxidative stress,stimulated portal edema and lymphocytic infiltration and decreased expression of cholangiocyte marker,cytoskeletal 19.In addition,TG and TP inhibited cell proliferation and migration,arrested cell cycle and promoted Caspase-dependent apoptosis of cholangiocytes via suppressing the phosphorylation of extracellular regulated protein kinases 1/2(ERK1/2)and protein kinase B(AKT).While,CEL at equivalent dosage had no obvious hepatotoxicity.Conclusion:We revealed that TG-stimulated liver injury was specifically characterized by cholangiocyte damage and TP might be the decisive ingredient to reflect TG hepatotoxicity.Our results not only provide novel insights into the mechanism underlying the hepatotoxicity effects of TG but also offer reference for clinical rational use of TG.展开更多
Biliary cryptosporidiosis caused by Cryptosporidium is life-threatening in immunocompromised individuals,particularly those with acquired immune deficiency syndrome(AIDS),resulting in AIDS cholangiopathy.However,studi...Biliary cryptosporidiosis caused by Cryptosporidium is life-threatening in immunocompromised individuals,particularly those with acquired immune deficiency syndrome(AIDS),resulting in AIDS cholangiopathy.However,studies of biliary cryptosporidiosis have lagged due to the lack of in vitro models that allow complete pathogen development.Here we describe procedures for the generation of mouse cholangiocyte organoids(Chol-orgs).The Chol-orgs,which expressed stem and biliary cell markers,could be clonally expanded for three months and stored in liquid nitrogen for more than one year.Combined with cell differentiation using the air-liquid interface(ALI)approach,we established a culture system for C.parvum.ALI cultures using Chol-orgs have supported50-fold amplification of the pathogen and generated viable oocysts in vitro.In addition,we analyzed the transcriptome of Chol-ALI cultures infected with the IId subtype of C.parvum to characterize host cell responses.RNA-seq analysis revealed that C.parvum upregulated immune and inflammatory responses and downregulated metabolic and cell proliferation signaling pathways in Cholorgs.A similar system using bovine cholangiocytes also supported the complete development of C.parvum in vitro.These in vitro models provide convenient methods to study biliary cryptosporidiosis and other hepatic infections and to develop effective therapies for AIDS cholangiopathy.展开更多
Objective:This study explores the mechanism of action of Danhongqing formula(DHQ),a compoundbased Chinese medicine formula,in the treatment of cholestatic liver fibrosis.Methods:In vivo experiments were conducted usin...Objective:This study explores the mechanism of action of Danhongqing formula(DHQ),a compoundbased Chinese medicine formula,in the treatment of cholestatic liver fibrosis.Methods:In vivo experiments were conducted using 8-week-old multidrug resistance protein 2 knockout(Mdr2-/-)mice as an animal model of cholestatic liver fibrosis.DHQ was administered orally for 8 weeks,and its impact on cholestatic liver fibrosis was evaluated by assessing liver function,liver histopathology,and the expression of liver fibrosis-related proteins.Real-time polymerase chain reaction,Western blot,immunohistochemistry and other methods were used to observe the effects of DHQ on long non-coding RNA H19(H19)and signal transducer and activator of transcription 3(STAT3)phosphorylation in the liver tissue of Mdr2-/-mice.In addition,cholangiocytes and hepatic stellate cells(HSCs)were cultured in vitro to measure the effects of bile acids on cholangiocyte injury and H19 expression.Cholangiocytes overexpressing H19 were constructed,and a conditioned medium containing H19 was collected to measure its effects on STAT3 protein expression and cell activation.The intervention effect of DHQ on these processes was also investigated.HSCs overexpressing H19 were constructed to measure the impact of H19 on cell activation and assess the intervention effect of DHQ.Results:DHQ alleviated liver injury,ductular reaction,and fibrosis in Mdr2-/-mice,and inhibited H19expression,STAT3 expression and STAT3 phosphorylation.This formula also reduced hydrophobic bile acid-induced cholangiocyte injury and the upregulation of H19,inhibited the activation of HSCs induced by cholangiocyte-derived conditioned medium,and decreased the expression of activation markers in HSCs.The overexpression of H19 in a human HSC line confirmed that H19 promoted STAT3 phosphorylation and HSC activation,and DHQ was able to successfully inhibit these effects.Conclusion:DHQ effectively alleviated spontaneous cholestatic liver fibrosis in Mdr2-/-mice by inhibiting H19 upregulation in chol展开更多
Cytosolic Ca^2＋ is an important second messenger in virtually every type of cell. Moreover, Ca^2＋ generally regulates multiple activities within individual cells. This article reviews the cellular machinery that is ...Cytosolic Ca^2＋ is an important second messenger in virtually every type of cell. Moreover, Ca^2＋ generally regulates multiple activities within individual cells. This article reviews the cellular machinery that is responsible for Ca^2＋ signaling in cholangiocytes. In addition, two Ca^2＋-mediated events in cholangiocytes are discussed： bicarbonate secretion and apoptosis. Finally, emerging evidence is reviewed that Ca^2＋ signaling is involved in the pathogenesis of diseases affecting the biliary tree and that Ca^2＋ signaling pathways can be manipulated to therapeutic advantage in the treatment of cholestatic disorders.展开更多
Polycystic liver diseases(PCLDs) are a heterogeneous group of genetic disorders characterized by the development of multiple fluid-filled cysts in the liver,which derive from cholangiocytes,the epithelial cells lining...Polycystic liver diseases(PCLDs) are a heterogeneous group of genetic disorders characterized by the development of multiple fluid-filled cysts in the liver,which derive from cholangiocytes,the epithelial cells lining the bile ducts.When these cysts grow,symptoms such as abdominal distension,nausea,and abdominal pain may occur.PCLDs may exist isolated(i.e.,autosomal dominant polycystic liver disease,ADPLD) or in combination with renal cystogenesis(i.e.,autosomal dominant polycystic kidney disease and autosomal recessive polycystic liver disease).The exact prevalence of PCLDs is unknown,but is estimated to occur in approximately 1:1000 persons.Although the pathogenesis of each form of PCLD appears to be different,increasing evidences indicate that hepatic cystogenesis is a phenomenon that may involve somatic loss of heterozygosity(LOH) in those pathological conditions inherited in a dominant form.A recent report,using highly sophisticated methodology,demonstrated that ADPLD patients with a germline mutation in the protein kinase C substrate 80K-H(PRKCSH) gene mostly develop hepatic cystogenesis through a second somatic mutation.While hepatocystin,the PRKCSH-encoding protein,was absent in the hepatic cysts with LOH,it was still expressed in the heterozygous cysts.On the other hand,no additional trans-heterozygous mutations on the SEC63 homolog(S.cerevisiae /SEC63) gene(also involved in the development of PCLDs) were observed.These data indicate that PCLD is recessive at the cellular level,and point out the important role of hepatocystin loss in cystogenesis.In this commentary,we discuss the knowledge regarding the role of somatic second-hit mutations in the development of PCLDs,and the most relevant findings have been highlighted.展开更多
肝内胆管缺失是指肝内胆管数量的减少或消失,是胆管树基本病理改变之一。免疫紊乱、肿瘤、感染、药物、缺血、遗传等因素都有可能引起胆管缺失。临床上,通过肝活组织检查,在10个及以上门管区的标本里,发现50%以上的小叶间胆管缺失,即可...肝内胆管缺失是指肝内胆管数量的减少或消失,是胆管树基本病理改变之一。免疫紊乱、肿瘤、感染、药物、缺血、遗传等因素都有可能引起胆管缺失。临床上,通过肝活组织检查,在10个及以上门管区的标本里,发现50%以上的小叶间胆管缺失,即可确诊[1]。胆管缺失的预后取决于病因和损伤程度。晚期出现不可逆的广泛胆管缺失甚至胆管消失时,该病理综合征称为胆管消失综合征(vanishing bile duct syndrome,VBDS),仅发生在0.5%小胆管病[2]。随后,可进展至广泛的胆管纤维化或肝硬化。有趣的是,也有部分患者表现为胆管上皮细胞再生,在数月或数年后得到恢复。本文总结了胆管缺失发病的分子机制,并重点强调近年来免疫介导的胆管病和胆管缺失的研究进展。展开更多
Primary biliary cholangitis(PBC)is a chronic cholestatic liver disease that is observed more frequently in middle-aged women.This disorder is considered an autoimmune disease,since liver injury is sustained by the pre...Primary biliary cholangitis(PBC)is a chronic cholestatic liver disease that is observed more frequently in middle-aged women.This disorder is considered an autoimmune disease,since liver injury is sustained by the presence of selfdirected antimitochondrial antibodies targeting the bile duct cells.The prognosis may vary depending on an early diagnosis and response to therapy.However,nearly a third of patients can progress to liver cirrhosis,thus requiring a liver transplant.Traditional immunosuppressive therapies,commonly employed for other autoimmune diseases,have limited effects on PBC.In fact,dramatic functional changes that occur in the biliary epithelium in the course of inflammation play a major role in perpetuating the injury.In this minireview,after a background on the disease and possible predisposing factors,the sequential cooperation of cellular/molecular events leading to end-stage PBC is discussed in detail.The rise and maintenance of the autoimmune process,as well as the response of the biliary epithelia during inflammatory injury,are key factors in the progression of the disease.The so-called“ductular reaction(DR)”,intended as a reactive expansion of cells with biliary phenotype,is a process frequently observed in PBC and partially understood.However,recent findings suggest a strict relationship between this pathological picture and the progression to liver fibrosis,cell senescence,and loss of biliary ducts.All these issues(onset of chronic inflammation,changes in secretive and proliferative biliary functions,DR,and its relationship with other pathological events)are discussed in this manuscript in an attempt to provide a snapshot,for clinicians and researchers,of the most relevant and sequential contributors to the progression of this human cholestatic disease.We believe that interpreting this disorder as a multistep process may help identify possible therapeutic targets to prevent evolution to severe disease.展开更多
Developing functional ductal organoids(FDOs)is essential for liver regenerative medicine.We aimed to construct FDOs with biliary tree networks in rat decellularized liver scaffolds(DLSs)with primary cholangiocytes iso...Developing functional ductal organoids(FDOs)is essential for liver regenerative medicine.We aimed to construct FDOs with biliary tree networks in rat decellularized liver scaffolds(DLSs)with primary cholangiocytes isolated from mouse bile ducts.The developed FDOs were dynamically characterized by functional assays and metabolomics for bioprocess clarification.FDOs were reconstructed in DLSs retaining native structure and bioactive factors with mouse primary cholangiocytes expressing enriched biomarkers.Morphological assessment showed that biliary tree-like structures gradually formed from day 3 to day 14.The cholangiocytes in FDOs maintained high viability and expressed 11 specific biomarkers.Basal-apical polarity was observed at day 14 with immunostaining for E-cadherin and acetylatedα-tubulin.The rhodamine 123 transport assay and active collection of cholyl-lysyl-fluorescein exhibited the specific functions of bile secretion and transportation at day 14 compared to those in monolayer and hydrogel culture systems.The metabolomics analysis with 1075 peak pairs showed that serotonin,as a key molecule of the tryptophan metabolism pathway linked to biliary tree reconstruction,was specifically expressed in FDOs during the whole period of culture.Such FDOs with biliary tree networks and serotonin expression may be applied for disease modeling and drug screening,which paves the way for future clinical therapeutic applications.展开更多
Objective:To explore the mechanism of action of Dahuang Lingxian Formula in alleviating the inflammatory response of bile duct cells in LPS-induced intrahepatic bile duct inflammation model rats based on IL-6/JAK/STAT...Objective:To explore the mechanism of action of Dahuang Lingxian Formula in alleviating the inflammatory response of bile duct cells in LPS-induced intrahepatic bile duct inflammation model rats based on IL-6/JAK/STAT3 signaling pathway.Methods:Fifty SD rats were randomly divided into five groups,blank group,model group,choling tablets(0.5 g/kg),and low and high concentration groups(2.4 g/kg and 4.8 g/kg)of Dahuang Lingxian Formula,ten rats in each group.Except for the blank group,the rats in each group were injected with 1.25 mg/kg LPS at the common bile duct at one time to construct an animal model of intrahepatic bile duct infection.After gavage on day 8,liver tissues were taken from rats at the hepatic hilum,and the histopathological changes of the hepatic hilum and biliary tree were observed by HE staining.The expression levels of serum glutamic alanine transaminase(ALT),glutamic oxalacetic transaminase(AST),malondialdehyde(MDA)and superoxide dismutase(SOD)were measured by biochemical method.The expression levels of interleukin 6(IL-6),Janus protein tyrosine kinase 2(JAK2),signal transducer and activator of transcription 3(STAT3)in rat serum were measured by enzyme-linked immunosorbent assay(ELISA).Protein immunoblotting(WB)and real-time fluorescence quantitative PCR(RT-qPCR)were used to detect the expression levels of IL-6,JAK2,STAT3 protein and mRNA in biliary tree tissues.Results:①Compared with the blank group,the structures such as interlobular bile ducts in the hepatic sinusoids and portal duct area of the model rats were destroyed,and inflammatory cells infiltrated around them.The expression of ALT,AST,MDA,IL-6,JAK2 and STAT3 in the serum increased significantly,the expression level of SOD decreased,and the expression levels of IL-6,JAK2 and STAT3 proteins and mRNA increased.②Compared with the model group,the degree of liver pathological damage in rats in the Chiling Ning tablet group and the low and high concentration groups of Dahuang Lingxian Formula were improved,which could significantly reduc展开更多
目的:研究原发性胆汁性胆管炎(primary biliary cholangitis,PBC)中发生CD8+细胞毒性T细胞(cytotoxic T lymphocyte,CTL)穿入现象及其意义。方法:收集53例PBC患者肝穿刺标本,采用H&E染色和免疫荧光染色方法,在光学显微镜和激光共聚...目的:研究原发性胆汁性胆管炎(primary biliary cholangitis,PBC)中发生CD8+细胞毒性T细胞(cytotoxic T lymphocyte,CTL)穿入现象及其意义。方法:收集53例PBC患者肝穿刺标本,采用H&E染色和免疫荧光染色方法,在光学显微镜和激光共聚焦显微镜下观察PBC肝组织中CTL的穿入及穿入宿主细胞的类型,并分析与病程的关系。结果:53例PBC患者中10例(18.9%)发生淋巴细胞穿入肝细胞,15例(28.3%)发生淋巴细胞穿入胆管上皮,仅1例患者同时存在淋巴细胞穿入肝细胞和胆管上皮细胞。免疫荧光染色证实穿入的淋巴细胞是CD8阳性细胞毒性T细胞(CTL)。早期PBC患者中4例(22.2%)、晚期PBC患者中6例(17.1%)发生CTL穿入肝细胞,两组差异无统计学意义(P>0.05)。早期PBC患者中9例(50.0%)、晚期PBC患者中6例(17.1%)发生CTL穿入胆管上皮,两组差异有统计学意义(Z=2.52,P<0.05)。结论:在PBC肝组织中,CTL可穿入肝细胞和胆管上皮细胞,CTL穿入胆管上皮细胞并导致其发生凋亡可能是PBC发病过程中胆管损伤的机制之一。展开更多
背景与目的:采用体外获得性表达外源发状分裂相关增强子-1(hairy and enhancer of split1,Hes1)基因的方法,探讨Hes1在肝干细胞分化以及胆管上皮细胞发育中的作用。材料与方法:通过PCR方法从小鼠基因组中克隆Hes1基因片段,构建表达载体p...背景与目的:采用体外获得性表达外源发状分裂相关增强子-1(hairy and enhancer of split1,Hes1)基因的方法,探讨Hes1在肝干细胞分化以及胆管上皮细胞发育中的作用。材料与方法:通过PCR方法从小鼠基因组中克隆Hes1基因片段,构建表达载体pEGFP-C1-Hes1和pcDNA3.1-Hes1,将2种表达载体分别转染肝原始细胞系(LEPCs),应用RT_PCR和Real-timePCR技术检测胆管细胞分子标志物CK19、GGT,胆管上皮细胞相关转录因子HNF6、HNF1β,肝细胞分子标志物GS、BGP和肝卵圆细胞的分子标志物Thy-1的表达,并在荧光显微镜下观察EGFP标记的LEPCs细胞系荧光强度的变化。结果:成功构建表达载体pEGFP-C1-Hes1和pcDNA3.1-Hes1。RT-PCR和Real_timePCR检测均表明胆管细胞分子标志CK19、GGT表达量上调;胆管上皮细胞相关转录因子HNF6、HNF1β表达上调;肝细胞分子标志GS、BGP表达量下调;卵圆细胞的分子标志Thy-1表达量下调;LEPCs细胞系绿色荧光增强。结论:初步证明小鼠肝原始细胞经Hes1的表达诱导后可向胆管上皮细胞方向分化,推测Hes1为胆管上皮细胞分化的转录调控因子。展开更多
文摘Gradual alterations of cell’s physiology and functions due to age or exposure to various stresses lead to the conversion of normal cells to senescent cells.Once becoming senescent,the cell stops dividing permanently but remains metabolically active.Cellular senescence does not have a single marker but is characterized mainly by a combination of multiple markers,such as,morphological changes,expression of cell cycle inhibitors,senescence associatedβ-galactosidase activity,and changes in nuclear membrane.When cells in an organ become senescent,the entire organism can be affected.This may occur through the senescence-associated secretory phenotype(SASP).SASP may exert beneficial or harmful effects on the microenvironment of tissues.Research on senescence has become a very exciting field in cell biology since the link between age-related diseases,including cancer,and senescence has been established.The loss of regenerative and homeostatic capacity of the liver over the age is somehow connected to cellular senescence.The major contributors of senescence properties in the liver are hepatocytes and cholangiocytes.Senescent cells in the liver have been implicated in the etiology of chronic liver diseases including cirrhosis and hepatocellular carcinoma and in the interference of liver regeneration.This review summarizes recently reported findings in the understanding of the molecular mechanisms of senescence and its relationship with liver diseases.
基金the MIUR grant, No. 2003060137_004 and by the Fondazione Cariverona 2002 grant "Ambienre e sviluppo sostenibile" to Prof. Benedetti by the "Premio S.I.G.E. 2004" to Dr. Marzioni by the Universita Politecnica delle Marche intramural grants ATBEN00205 to Professor. Benedetti and ATMAROl 105 to Dr. Marzioni, by the "Premio S.I.G.E. 2006" to Dr Fava
文摘Cholangiocytes, the epithelial cells lining the biliary ducts, are the target cells in several liver diseases. Cholangiopathies and cholangiocarcinoma generate interest in many scientists since the genesis. The developing mechanisms, and the therapeutic tools of these diseases are still undefined. Several studies demonstrate that many hormones, neuropeptides and neurotransmitters regulate malignant and non-malignant cholangiocyte pathophysiology in the course of chronic biliary diseases. The aim of this review is to present the findings of several studies published in the recent years that contributed to clarifying the role of nervous and neuroendocrine regulation of the pathophysiologic events associated with cholestasis and cholangiocarcinoma development. This manuscript is organized into two parts. The first part offers an overview of the innervation of the liver and the origin of neuroendocrine hormones, neurotransmitters and neuropeptides affecting cholangiocyte function and metabolism. The first section also reviews the effects played by several neuroendocrine hormones and nervous system on cholangiocyte growth, survival and functional activity in the course of cholestasis. In the second section, we summarize the results of some studies describing the role of nervous system and neuroendocrine hormones in the regulation of malignant cholangiocyte growth.
基金supported by grants from Beijing Nova Program of Science&Technology(Grant No.Z191100001119088 to XL)Beijing Municipal Science&Technology Commission(Grant No.7212174 to XL)+2 种基金National Natural Science Foundation of China(Grant No.82004045 to XL)Young Talents Promotion Project of China Association of Traditional Chinese Medicine(Grant No.2020-QNRC2-01 to XL)Innovation Team and Talents Cultivation Program of National Administration of Traditional Chinese Medicine(Grant No.ZYYCXTD-C-202006 to XL)。
文摘Objective:Tripterygium glycoside(TG)is widely used in clinical practice for its multiple bioactivities including anti-inflammatory and immunosuppressive effects.However,emerging studies have frequently reported TG-induced adverse reactions to multiple organs,especially liver.Here,this study aimed to investigate the mechanism of liver damage induced by TG and explore representative components to reflect TG hepatotoxicity.Methods:Network pharmacology was used to determine the potential targets of bile duct injury caused by TG.Next,the hepatotoxic effects of TG,triptolide(TP)and celastrol(CEL)were investigated and compared in vivo and in vitro.Liver function was determined by measuring serum transaminase and histopathology staining.The cell proliferation and apoptosis were determined by cell viability assay,scratch assay and flow cytometry.The expression of gene of interest was determined by qPCR and Western blot.Results:Based on the network pharmacological analysis of 12 bioactive ingredients found in TG,a total of35 targets and 15 pathways related to bile duct injury were obtained.Both TG and TP resulted in cholangiocyte damage and liver injury,as illustrated by increased levels of serum transaminase and oxidative stress,stimulated portal edema and lymphocytic infiltration and decreased expression of cholangiocyte marker,cytoskeletal 19.In addition,TG and TP inhibited cell proliferation and migration,arrested cell cycle and promoted Caspase-dependent apoptosis of cholangiocytes via suppressing the phosphorylation of extracellular regulated protein kinases 1/2(ERK1/2)and protein kinase B(AKT).While,CEL at equivalent dosage had no obvious hepatotoxicity.Conclusion:We revealed that TG-stimulated liver injury was specifically characterized by cholangiocyte damage and TP might be the decisive ingredient to reflect TG hepatotoxicity.Our results not only provide novel insights into the mechanism underlying the hepatotoxicity effects of TG but also offer reference for clinical rational use of TG.
基金supported by National Natural Science Foundation of China(32030109)111 Project(D20008)+2 种基金Double First-class Discipline Promotion Project(2023B10564003)supported by National Natural Science Foundation of China(U21A20258)supported by Natural Science Foundation of Guangzhou City(202201010140)。
文摘Biliary cryptosporidiosis caused by Cryptosporidium is life-threatening in immunocompromised individuals,particularly those with acquired immune deficiency syndrome(AIDS),resulting in AIDS cholangiopathy.However,studies of biliary cryptosporidiosis have lagged due to the lack of in vitro models that allow complete pathogen development.Here we describe procedures for the generation of mouse cholangiocyte organoids(Chol-orgs).The Chol-orgs,which expressed stem and biliary cell markers,could be clonally expanded for three months and stored in liquid nitrogen for more than one year.Combined with cell differentiation using the air-liquid interface(ALI)approach,we established a culture system for C.parvum.ALI cultures using Chol-orgs have supported50-fold amplification of the pathogen and generated viable oocysts in vitro.In addition,we analyzed the transcriptome of Chol-ALI cultures infected with the IId subtype of C.parvum to characterize host cell responses.RNA-seq analysis revealed that C.parvum upregulated immune and inflammatory responses and downregulated metabolic and cell proliferation signaling pathways in Cholorgs.A similar system using bovine cholangiocytes also supported the complete development of C.parvum in vitro.These in vitro models provide convenient methods to study biliary cryptosporidiosis and other hepatic infections and to develop effective therapies for AIDS cholangiopathy.
基金supported by grants from the National Natural Science Foundation of China(No.81773980)Project of Science and Technology Commission of Shanghai Municipality(No.15401902600)。
文摘Objective:This study explores the mechanism of action of Danhongqing formula(DHQ),a compoundbased Chinese medicine formula,in the treatment of cholestatic liver fibrosis.Methods:In vivo experiments were conducted using 8-week-old multidrug resistance protein 2 knockout(Mdr2-/-)mice as an animal model of cholestatic liver fibrosis.DHQ was administered orally for 8 weeks,and its impact on cholestatic liver fibrosis was evaluated by assessing liver function,liver histopathology,and the expression of liver fibrosis-related proteins.Real-time polymerase chain reaction,Western blot,immunohistochemistry and other methods were used to observe the effects of DHQ on long non-coding RNA H19(H19)and signal transducer and activator of transcription 3(STAT3)phosphorylation in the liver tissue of Mdr2-/-mice.In addition,cholangiocytes and hepatic stellate cells(HSCs)were cultured in vitro to measure the effects of bile acids on cholangiocyte injury and H19 expression.Cholangiocytes overexpressing H19 were constructed,and a conditioned medium containing H19 was collected to measure its effects on STAT3 protein expression and cell activation.The intervention effect of DHQ on these processes was also investigated.HSCs overexpressing H19 were constructed to measure the impact of H19 on cell activation and assess the intervention effect of DHQ.Results:DHQ alleviated liver injury,ductular reaction,and fibrosis in Mdr2-/-mice,and inhibited H19expression,STAT3 expression and STAT3 phosphorylation.This formula also reduced hydrophobic bile acid-induced cholangiocyte injury and the upregulation of H19,inhibited the activation of HSCs induced by cholangiocyte-derived conditioned medium,and decreased the expression of activation markers in HSCs.The overexpression of H19 in a human HSC line confirmed that H19 promoted STAT3 phosphorylation and HSC activation,and DHQ was able to successfully inhibit these effects.Conclusion:DHQ effectively alleviated spontaneous cholestatic liver fibrosis in Mdr2-/-mice by inhibiting H19 upregulation in chol
基金NIH grants DK61747, DK45710, DK57751, and DK34989, and by a grant-in-aid from the American Heart Association
文摘Cytosolic Ca^2＋ is an important second messenger in virtually every type of cell. Moreover, Ca^2＋ generally regulates multiple activities within individual cells. This article reviews the cellular machinery that is responsible for Ca^2＋ signaling in cholangiocytes. In addition, two Ca^2＋-mediated events in cholangiocytes are discussed： bicarbonate secretion and apoptosis. Finally, emerging evidence is reviewed that Ca^2＋ signaling is involved in the pathogenesis of diseases affecting the biliary tree and that Ca^2＋ signaling pathways can be manipulated to therapeutic advantage in the treatment of cholestatic disorders.
文摘Polycystic liver diseases(PCLDs) are a heterogeneous group of genetic disorders characterized by the development of multiple fluid-filled cysts in the liver,which derive from cholangiocytes,the epithelial cells lining the bile ducts.When these cysts grow,symptoms such as abdominal distension,nausea,and abdominal pain may occur.PCLDs may exist isolated(i.e.,autosomal dominant polycystic liver disease,ADPLD) or in combination with renal cystogenesis(i.e.,autosomal dominant polycystic kidney disease and autosomal recessive polycystic liver disease).The exact prevalence of PCLDs is unknown,but is estimated to occur in approximately 1:1000 persons.Although the pathogenesis of each form of PCLD appears to be different,increasing evidences indicate that hepatic cystogenesis is a phenomenon that may involve somatic loss of heterozygosity(LOH) in those pathological conditions inherited in a dominant form.A recent report,using highly sophisticated methodology,demonstrated that ADPLD patients with a germline mutation in the protein kinase C substrate 80K-H(PRKCSH) gene mostly develop hepatic cystogenesis through a second somatic mutation.While hepatocystin,the PRKCSH-encoding protein,was absent in the hepatic cysts with LOH,it was still expressed in the heterozygous cysts.On the other hand,no additional trans-heterozygous mutations on the SEC63 homolog(S.cerevisiae /SEC63) gene(also involved in the development of PCLDs) were observed.These data indicate that PCLD is recessive at the cellular level,and point out the important role of hepatocystin loss in cystogenesis.In this commentary,we discuss the knowledge regarding the role of somatic second-hit mutations in the development of PCLDs,and the most relevant findings have been highlighted.
文摘肝内胆管缺失是指肝内胆管数量的减少或消失,是胆管树基本病理改变之一。免疫紊乱、肿瘤、感染、药物、缺血、遗传等因素都有可能引起胆管缺失。临床上,通过肝活组织检查,在10个及以上门管区的标本里,发现50%以上的小叶间胆管缺失,即可确诊[1]。胆管缺失的预后取决于病因和损伤程度。晚期出现不可逆的广泛胆管缺失甚至胆管消失时,该病理综合征称为胆管消失综合征(vanishing bile duct syndrome,VBDS),仅发生在0.5%小胆管病[2]。随后,可进展至广泛的胆管纤维化或肝硬化。有趣的是,也有部分患者表现为胆管上皮细胞再生,在数月或数年后得到恢复。本文总结了胆管缺失发病的分子机制,并重点强调近年来免疫介导的胆管病和胆管缺失的研究进展。
文摘Primary biliary cholangitis(PBC)is a chronic cholestatic liver disease that is observed more frequently in middle-aged women.This disorder is considered an autoimmune disease,since liver injury is sustained by the presence of selfdirected antimitochondrial antibodies targeting the bile duct cells.The prognosis may vary depending on an early diagnosis and response to therapy.However,nearly a third of patients can progress to liver cirrhosis,thus requiring a liver transplant.Traditional immunosuppressive therapies,commonly employed for other autoimmune diseases,have limited effects on PBC.In fact,dramatic functional changes that occur in the biliary epithelium in the course of inflammation play a major role in perpetuating the injury.In this minireview,after a background on the disease and possible predisposing factors,the sequential cooperation of cellular/molecular events leading to end-stage PBC is discussed in detail.The rise and maintenance of the autoimmune process,as well as the response of the biliary epithelia during inflammatory injury,are key factors in the progression of the disease.The so-called“ductular reaction(DR)”,intended as a reactive expansion of cells with biliary phenotype,is a process frequently observed in PBC and partially understood.However,recent findings suggest a strict relationship between this pathological picture and the progression to liver fibrosis,cell senescence,and loss of biliary ducts.All these issues(onset of chronic inflammation,changes in secretive and proliferative biliary functions,DR,and its relationship with other pathological events)are discussed in this manuscript in an attempt to provide a snapshot,for clinicians and researchers,of the most relevant and sequential contributors to the progression of this human cholestatic disease.We believe that interpreting this disorder as a multistep process may help identify possible therapeutic targets to prevent evolution to severe disease.
基金supported by the National Key R&D Program of China(2022YFA1104102,2022YFA1104603,2022YFC2304801)the National Natural Science Foundation of China(81830073,82272426)+1 种基金the National Provincial special support program for high-level personnel recruitment(Ten-thousand Talents Program,2018RA4016)Zhejiang Public Welfare Project(LY21H030007,LGF21H200006).
文摘Developing functional ductal organoids(FDOs)is essential for liver regenerative medicine.We aimed to construct FDOs with biliary tree networks in rat decellularized liver scaffolds(DLSs)with primary cholangiocytes isolated from mouse bile ducts.The developed FDOs were dynamically characterized by functional assays and metabolomics for bioprocess clarification.FDOs were reconstructed in DLSs retaining native structure and bioactive factors with mouse primary cholangiocytes expressing enriched biomarkers.Morphological assessment showed that biliary tree-like structures gradually formed from day 3 to day 14.The cholangiocytes in FDOs maintained high viability and expressed 11 specific biomarkers.Basal-apical polarity was observed at day 14 with immunostaining for E-cadherin and acetylatedα-tubulin.The rhodamine 123 transport assay and active collection of cholyl-lysyl-fluorescein exhibited the specific functions of bile secretion and transportation at day 14 compared to those in monolayer and hydrogel culture systems.The metabolomics analysis with 1075 peak pairs showed that serotonin,as a key molecule of the tryptophan metabolism pathway linked to biliary tree reconstruction,was specifically expressed in FDOs during the whole period of culture.Such FDOs with biliary tree networks and serotonin expression may be applied for disease modeling and drug screening,which paves the way for future clinical therapeutic applications.
基金Guangxi Natural Science Foundation(No.2020GXNSFAA238012)Research on Traditional Chinese Medicine Prevention and Treatment of Liver and Bile Related Diseases in the 2021"Qihuang Project"High Level Talent Team Cultivation Project(No.2021006)+1 种基金2020 Guangxi University of Traditional Chinese Medicine First Affiliated Hospital Hospital Hospital Level Doctoral Initiation Fund Project(No.2020BS004)2020 Guangxi University of Traditional Chinese Medicine Introduction Doctoral Research Initiation Fund Project(No.2020BS030)。
文摘Objective:To explore the mechanism of action of Dahuang Lingxian Formula in alleviating the inflammatory response of bile duct cells in LPS-induced intrahepatic bile duct inflammation model rats based on IL-6/JAK/STAT3 signaling pathway.Methods:Fifty SD rats were randomly divided into five groups,blank group,model group,choling tablets(0.5 g/kg),and low and high concentration groups(2.4 g/kg and 4.8 g/kg)of Dahuang Lingxian Formula,ten rats in each group.Except for the blank group,the rats in each group were injected with 1.25 mg/kg LPS at the common bile duct at one time to construct an animal model of intrahepatic bile duct infection.After gavage on day 8,liver tissues were taken from rats at the hepatic hilum,and the histopathological changes of the hepatic hilum and biliary tree were observed by HE staining.The expression levels of serum glutamic alanine transaminase(ALT),glutamic oxalacetic transaminase(AST),malondialdehyde(MDA)and superoxide dismutase(SOD)were measured by biochemical method.The expression levels of interleukin 6(IL-6),Janus protein tyrosine kinase 2(JAK2),signal transducer and activator of transcription 3(STAT3)in rat serum were measured by enzyme-linked immunosorbent assay(ELISA).Protein immunoblotting(WB)and real-time fluorescence quantitative PCR(RT-qPCR)were used to detect the expression levels of IL-6,JAK2,STAT3 protein and mRNA in biliary tree tissues.Results:①Compared with the blank group,the structures such as interlobular bile ducts in the hepatic sinusoids and portal duct area of the model rats were destroyed,and inflammatory cells infiltrated around them.The expression of ALT,AST,MDA,IL-6,JAK2 and STAT3 in the serum increased significantly,the expression level of SOD decreased,and the expression levels of IL-6,JAK2 and STAT3 proteins and mRNA increased.②Compared with the model group,the degree of liver pathological damage in rats in the Chiling Ning tablet group and the low and high concentration groups of Dahuang Lingxian Formula were improved,which could significantly reduc
文摘目的:研究原发性胆汁性胆管炎(primary biliary cholangitis,PBC)中发生CD8+细胞毒性T细胞(cytotoxic T lymphocyte,CTL)穿入现象及其意义。方法:收集53例PBC患者肝穿刺标本,采用H&E染色和免疫荧光染色方法,在光学显微镜和激光共聚焦显微镜下观察PBC肝组织中CTL的穿入及穿入宿主细胞的类型,并分析与病程的关系。结果:53例PBC患者中10例(18.9%)发生淋巴细胞穿入肝细胞,15例(28.3%)发生淋巴细胞穿入胆管上皮,仅1例患者同时存在淋巴细胞穿入肝细胞和胆管上皮细胞。免疫荧光染色证实穿入的淋巴细胞是CD8阳性细胞毒性T细胞(CTL)。早期PBC患者中4例(22.2%)、晚期PBC患者中6例(17.1%)发生CTL穿入肝细胞,两组差异无统计学意义(P>0.05)。早期PBC患者中9例(50.0%)、晚期PBC患者中6例(17.1%)发生CTL穿入胆管上皮,两组差异有统计学意义(Z=2.52,P<0.05)。结论:在PBC肝组织中,CTL可穿入肝细胞和胆管上皮细胞,CTL穿入胆管上皮细胞并导致其发生凋亡可能是PBC发病过程中胆管损伤的机制之一。
文摘背景与目的:采用体外获得性表达外源发状分裂相关增强子-1(hairy and enhancer of split1,Hes1)基因的方法,探讨Hes1在肝干细胞分化以及胆管上皮细胞发育中的作用。材料与方法:通过PCR方法从小鼠基因组中克隆Hes1基因片段,构建表达载体pEGFP-C1-Hes1和pcDNA3.1-Hes1,将2种表达载体分别转染肝原始细胞系(LEPCs),应用RT_PCR和Real-timePCR技术检测胆管细胞分子标志物CK19、GGT,胆管上皮细胞相关转录因子HNF6、HNF1β,肝细胞分子标志物GS、BGP和肝卵圆细胞的分子标志物Thy-1的表达,并在荧光显微镜下观察EGFP标记的LEPCs细胞系荧光强度的变化。结果:成功构建表达载体pEGFP-C1-Hes1和pcDNA3.1-Hes1。RT-PCR和Real_timePCR检测均表明胆管细胞分子标志CK19、GGT表达量上调;胆管上皮细胞相关转录因子HNF6、HNF1β表达上调;肝细胞分子标志GS、BGP表达量下调;卵圆细胞的分子标志Thy-1表达量下调;LEPCs细胞系绿色荧光增强。结论:初步证明小鼠肝原始细胞经Hes1的表达诱导后可向胆管上皮细胞方向分化,推测Hes1为胆管上皮细胞分化的转录调控因子。
