AIM： To elucidate the interaction between non- parenchymal cells, extracellular matrix and oval cells during the restituting process of liver injury induced by partial hepatectomy （PH）. METHODS： We examined the lo...AIM： To elucidate the interaction between non- parenchymal cells, extracellular matrix and oval cells during the restituting process of liver injury induced by partial hepatectomy （PH）. METHODS： We examined the localization of oval cells, non-parenchymal cells, and the extracellular matrix components using immunohistochemical and double immunofluorescent analysis during the proliferation and differentiation of oval cells in N-2- acetylaminofluorene （2-AAF）/PH rat model. RESULTS： By day 2 after PH, small oval cells began to proliferate around the portal area. Most of stellate cells and laminin were present along the hepatic sinusoids in the periportal area. Kupffer cells and fibronectin markedly increased in the whole hepatic Iobule. From day 4 to 9, oval cells spread further into hepatic parenchyma, closely associated with stellate cells, fibronectin and laminin. Kupffer cells admixed with oval cells by day 6 and then decreased in the periportal zone. From day 12 to 15, most of hepatic stellate cells （HSCs）, laminin and fibronectin located around the small hepatocyte nodus, and minority of them appeared in the nodus. Kupffer cells were mainly limited in the pericentral sinusoids. After day 18, the normal liver Iobule structures began to recover.CONCLUSION： Local hepatic microenvironment may participate in the oval cell-mediated liver regeneration through the cell-cell and cell-matrix interactions.展开更多
AIM： To evaluate the effect of intrahepatic transplantation of hepatic oval cells （HOC） on fulminant hepatic failure （FHF） in rats.METHODS： HOC obtained from rats were labeled with green fluocescent protein （GF...AIM： To evaluate the effect of intrahepatic transplantation of hepatic oval cells （HOC） on fulminant hepatic failure （FHF） in rats.METHODS： HOC obtained from rats were labeled with green fluocescent protein （GFP） or 5, 6- carboxyfluorescein diacetate succinmidyl ester （CFDASE）. Cell fluorescence was observed under fluorescent microscope at 6, 24, 48 and 72 h after labeling. CFDA- SE labeled HOC （5 × 10^6 cells each rat） were injected into livers of rats with FHF induced by D-galactosamine. Serum albumin （ALB）, alanine aminotransferase （ALT）, aspartate aminotransferase （AST） and total bilirubin （TBil） levels were measured at different time points. Liver function of rats was examined on days 3, 7, 14 and 21 after HOC transplantation.RESULTS： The positive rate of GFP and CFDA-SE labeled HOC was 10% and 90%, respectively, with no significant change in cell viabilities. The survival rate was higher in HOC transplantation group than in control group, especially 48 （9/15 vs 6/15） and 72 h （9/15 vs 4/15） after HOC transplantation. The serum ALT, AST and TBil levels were decreased while the serum AIb level was increased after HOC transplantation. Fluorescence became faded and diffused in liver tissues, suggesting that proliferation and differentiation occur in transplanted HOC.CONCLUSION： CFDA-SE is superior to GFP in labeling HOC, although fluorescence intensity is decreased progressively with cell division. HOC transplantation can improve the liver function and increase the survival rate of recipients.展开更多
AIM: To detect immunohistochemically the presence of oval cells in chronic viral hepatitis with antibody against c-kit. METHODS: We detected oval cells in paraffin embedded liver sections of 3 normal controls and 26 l...AIM: To detect immunohistochemically the presence of oval cells in chronic viral hepatitis with antibody against c-kit. METHODS: We detected oval cells in paraffin embedded liver sections of 3 normal controls and 26 liver samples from patients with chronic viral hepatitis, using immunohistochemistry with antibodies against c-kit, piclass glutathione S-transferase (pi-GST) and cytokeratins 19 (CK19). RESULTS: Oval cells were not observed in normal livers. In chronic viral hepatitis, hepatic oval cells were located predominantly in the periportal region and fibrosis septa,characterized by an ovoid nucleus, small size,and scant cytoplasm. Antibody against stem cell factor receptor, c-kit, had higher sensitivity and specificity than pi-GST and CK19. About 50%-70% of c-kit positive oval cells were stained positively for either pi-GST or CK19. CONCLUSION: Oval cells are frequently detected in human livers with chronic viral hepatitis, suggesting that oval cell proliferation is associated with the liver regeneration in this condition.展开更多
Objective To acquire oval cells （progenitor stem cells ） from adult rat liver of different models including diabetic rats. Methods Thirty Sprague-Dawley （ SD ） rats were divided into 5 groups randomly： control, 2...Objective To acquire oval cells （progenitor stem cells ） from adult rat liver of different models including diabetic rats. Methods Thirty Sprague-Dawley （ SD ） rats were divided into 5 groups randomly： control, 2-acetylaminofluorene （ 2-AAF ）, 2-AAF ＋ partial hepatectomy （ PH ）, 2-AAF ＋ carbon tetrachloride （ CCl4 ）, and diabetic groups. As two-step collagenase perfusion protocol of Seglen, oval cells were isolated by Percoll density gradient centrifugation. Thy1. 1 positive cells were sorted by flow cytometry, and then cultured in Dulbecco＇s minimum Eagle＇s medium （DMEM）. Immunofluorescence staining was applied to labelling Thyl. 1. Results Different rates of Thy1.1 positive oval cells were found in different rat model groups ： 0. 5 % in 2-AAF, 0. 3% in 2-AAF ＋ PH, 0. 2% in 2-AAF ＋ CCl4, 0. 1% in diabetic, and 0. 0% in control. Isolated cells adhered to plate with fusiform or polygon as epithelial cells. Conclusion Progenitor stem cells exist in injured liver tissue including those from diabetic rats.展开更多
Summary： Oval cells have a potential to differentiate into a variety of cell lineages including hepatocytes and biliary epithelia. Several models have been established to activate the oval cells by incorporating a va...Summary： Oval cells have a potential to differentiate into a variety of cell lineages including hepatocytes and biliary epithelia. Several models have been established to activate the oval cells by incorporating a variety of toxins and carcinogens, alone or combined with surgical treatment. Those models are obviously not suitable for the study on human hepatic oval cells. It is necessary to establish a new and efficient model to study the human hepatic oval cells. In this study, the hepatocyte growth factor（HGF） and epidermal growth factor（EGF） were used to induce differentiation of mouse embryonic stem（ES） cells into hepatic oval cells. We first confirmed that hepatic oval cells derived from ES cells, which are bipotential, do exist during the course of mouse ES cells＇ differentiation into hepatic parenchymal cells. RT-PCR and transmission electron microscopy were applied in this study. The ratio of Sca-1＋/CD34＋ cells sorted by FACS in the induction group was increased from day 4 and reached the maximum on the day 8, whereas that in the control group remained at a low level. The differentiation ratio of Sca-1＋/CD34＋ cells in the induction group was significantly higher than that in the control group. About 92.48% of the sorted Sca-1＋/CD34＋ cells on the day 8 were A6 positive. Highly purified A6＋/Sca-1＋/CD34＋ hepatic oval cells derived from ES cells could be obtained by FACS. The differentiation ratio of hepatic oval cells in the induction group（up to 4.46%） was significantly higher than that in the control group. The number of hepatic oval cells could be increased significantly by HGF and EGF. The study also examined the ultrastructures of ES-derived hepatic oval cells＇ membrane surface by atomic force microscopy. The ES-derived hepatic oval cells cultured and sorted by our protocols may be available for the future clinical application.展开更多
AIM:To investigate the ultrastructure of oval cells in children with chronic hepatitis B,with special emphasis on their location in areas of collagen fibroplasia. METHODS:Morphological investigations were conducted on...AIM:To investigate the ultrastructure of oval cells in children with chronic hepatitis B,with special emphasis on their location in areas of collagen fibroplasia. METHODS:Morphological investigations were conducted on biopsy material obtained from 40 children,aged 3-16 years with chronic hepatitis B. The stage of fibrosis was assessed histologically using the arbitrary semiquantitative numerical scoring system proposed by Ishak et al. The material for ultrastructural investigation was fixed in glutaraldehyde and paraformaldehyde and processed for transmission-electron microscopic analysis. RESULTS:Ultrastructural examination of biopsy specimens obtained from children with chronic hepatitis B showed the presence of two types of oval cells,the hepatic progenitor cells and intermediate hepatic-like cells. These cells were present in the parenchyma and were seen most commonly in areas of intense periportal fibrosis (at least stage 2 according to Ishak et al) and in the vicinity of the limiting plate of the lobule. The activated nonparenchymal hepatic cells,i.e. transformed hepatic stellate cells and Kupffer cells were seen in close proximity to the intermediate hepatic-like cells. CONCLUSION:We found a distinct relationship between the prevalence of oval cells (hepatic progenitor cells and intermediate hepatocyte-like cells) and fibrosis stage in pediatric patients with chronic hepatitis B.展开更多
AIM:To investigate the effect of the neuropeptides bombesin(BBS)and neurotensin(NT)on oval cell proliferation in partially hepatectomized rats not pretreated with a known hepatocyte inhibitor.METHODS:Seventy male Wist...AIM:To investigate the effect of the neuropeptides bombesin(BBS)and neurotensin(NT)on oval cell proliferation in partially hepatectomized rats not pretreated with a known hepatocyte inhibitor.METHODS:Seventy male Wistar rats were randomly divided into five groups:Ⅰ=controls,Ⅱ=sham operated,Ⅲ=partial hepatectomy 70%(PHx),Ⅳ=PHx+ BBS(30μg/kg per day),Ⅴ=PHx+NT(300μg/kg per day).Forty eight hours after liver resection,portal en-dotoxin levels and hepatic glutathione redox state were determined.α-fetoprotein(AFP)mRNA(in situ hybridisation),cytokeratin-19 and Ki67 antigen expression (immunohistochemistry)and apoptosis(TUNEL)were evaluated on liver tissue samples.Cells with morphological features of oval cells that were cytokeratin-19 (+)and AFP mRNA(+)were scored in morphometric analysis and their proliferation was recorded.In addition,the proliferation and apoptotic rates of hepatocytes were determined.RESULTS:In the control and sham operated groups,oval cells were significantly less compared to groups Ⅲ,ⅣandⅤ(P<0.001).The neuropeptides BBS and NT significantly increased the proliferation of oval cells compared to groupⅢ(P<0.001).In addition,BBS and NT induced a significant increase of hepatocyte proliferation(P<0.001),whereas it decreased their apoptotic activity(P<0.001)compared to groupⅢ.BBS and NT significantly decreased portal endotoxemia (P<0.001)and increased the hepatic GSH:GSSG ratio (P<0.05 and P<0.001,respectively)compared to groupⅢ.CONCLUSION:BBS and NT stimulated oval cell proliferation in a model of liver regeneration,without use of concomitant suppression of hepatocyte proliferation as oval cell activation stimuli,and improved the hepatocyte regenerative response.This peptides-induced combined stimulation of oval cell and hepatocyte proliferation might serve as a possible treatment modality for several liver diseases.展开更多
The liver possesses an extraordinary ability to regenerate after injury.Hepatocyte-driven liver regeneration is the default pathway in response to mild-to-moderate acute liver damage.When replication of mature hepatoc...The liver possesses an extraordinary ability to regenerate after injury.Hepatocyte-driven liver regeneration is the default pathway in response to mild-to-moderate acute liver damage.When replication of mature hepatocytes is blocked,facultative hepatic progenitor cells(HPCs),also referred to as oval cells(OCs)in rodents,are activated.HPC/OCs have the ability to proliferate clonogenically and differentiate into several lineages including hepatocytes and bile ductal epithelia.This is a conserved liver injury response that has been studied in many species ranging from mammals(rat,mouse,and human)to fish.In addition,improper HPC/OC activation is closely associated with fibrotic responses,characterized by myofibroblast activation and extracellular matrix production,in many chronic liver diseases.Matrix remodeling and metalloprotease activities play an important role in the regulation of HPC/OC proliferation and fibrosis progression.Thus,understanding molecular mechanisms underlying HPC/OC activation has therapeutic implications for rational design of anti-fibrotic therapies.展开更多
基金Supported by A grant from National Natural Science Foundation of China
文摘AIM： To elucidate the interaction between non- parenchymal cells, extracellular matrix and oval cells during the restituting process of liver injury induced by partial hepatectomy （PH）. METHODS： We examined the localization of oval cells, non-parenchymal cells, and the extracellular matrix components using immunohistochemical and double immunofluorescent analysis during the proliferation and differentiation of oval cells in N-2- acetylaminofluorene （2-AAF）/PH rat model. RESULTS： By day 2 after PH, small oval cells began to proliferate around the portal area. Most of stellate cells and laminin were present along the hepatic sinusoids in the periportal area. Kupffer cells and fibronectin markedly increased in the whole hepatic Iobule. From day 4 to 9, oval cells spread further into hepatic parenchyma, closely associated with stellate cells, fibronectin and laminin. Kupffer cells admixed with oval cells by day 6 and then decreased in the periportal zone. From day 12 to 15, most of hepatic stellate cells （HSCs）, laminin and fibronectin located around the small hepatocyte nodus, and minority of them appeared in the nodus. Kupffer cells were mainly limited in the pericentral sinusoids. After day 18, the normal liver Iobule structures began to recover.CONCLUSION： Local hepatic microenvironment may participate in the oval cell-mediated liver regeneration through the cell-cell and cell-matrix interactions.
基金Supported by Tianjin Science Committee,Grant No.05SYSYJC02600Tianjin Health Bureau,Grant No.05KYR01
文摘AIM： To evaluate the effect of intrahepatic transplantation of hepatic oval cells （HOC） on fulminant hepatic failure （FHF） in rats.METHODS： HOC obtained from rats were labeled with green fluocescent protein （GFP） or 5, 6- carboxyfluorescein diacetate succinmidyl ester （CFDASE）. Cell fluorescence was observed under fluorescent microscope at 6, 24, 48 and 72 h after labeling. CFDA- SE labeled HOC （5 × 10^6 cells each rat） were injected into livers of rats with FHF induced by D-galactosamine. Serum albumin （ALB）, alanine aminotransferase （ALT）, aspartate aminotransferase （AST） and total bilirubin （TBil） levels were measured at different time points. Liver function of rats was examined on days 3, 7, 14 and 21 after HOC transplantation.RESULTS： The positive rate of GFP and CFDA-SE labeled HOC was 10% and 90%, respectively, with no significant change in cell viabilities. The survival rate was higher in HOC transplantation group than in control group, especially 48 （9/15 vs 6/15） and 72 h （9/15 vs 4/15） after HOC transplantation. The serum ALT, AST and TBil levels were decreased while the serum AIb level was increased after HOC transplantation. Fluorescence became faded and diffused in liver tissues, suggesting that proliferation and differentiation occur in transplanted HOC.CONCLUSION： CFDA-SE is superior to GFP in labeling HOC, although fluorescence intensity is decreased progressively with cell division. HOC transplantation can improve the liver function and increase the survival rate of recipients.
文摘AIM: To detect immunohistochemically the presence of oval cells in chronic viral hepatitis with antibody against c-kit. METHODS: We detected oval cells in paraffin embedded liver sections of 3 normal controls and 26 liver samples from patients with chronic viral hepatitis, using immunohistochemistry with antibodies against c-kit, piclass glutathione S-transferase (pi-GST) and cytokeratins 19 (CK19). RESULTS: Oval cells were not observed in normal livers. In chronic viral hepatitis, hepatic oval cells were located predominantly in the periportal region and fibrosis septa,characterized by an ovoid nucleus, small size,and scant cytoplasm. Antibody against stem cell factor receptor, c-kit, had higher sensitivity and specificity than pi-GST and CK19. About 50%-70% of c-kit positive oval cells were stained positively for either pi-GST or CK19. CONCLUSION: Oval cells are frequently detected in human livers with chronic viral hepatitis, suggesting that oval cell proliferation is associated with the liver regeneration in this condition.
基金Supported by Shanghai Municipal Education Commission Key Project (07ZZ35)
文摘Objective To acquire oval cells （progenitor stem cells ） from adult rat liver of different models including diabetic rats. Methods Thirty Sprague-Dawley （ SD ） rats were divided into 5 groups randomly： control, 2-acetylaminofluorene （ 2-AAF ）, 2-AAF ＋ partial hepatectomy （ PH ）, 2-AAF ＋ carbon tetrachloride （ CCl4 ）, and diabetic groups. As two-step collagenase perfusion protocol of Seglen, oval cells were isolated by Percoll density gradient centrifugation. Thy1. 1 positive cells were sorted by flow cytometry, and then cultured in Dulbecco＇s minimum Eagle＇s medium （DMEM）. Immunofluorescence staining was applied to labelling Thyl. 1. Results Different rates of Thy1.1 positive oval cells were found in different rat model groups ： 0. 5 % in 2-AAF, 0. 3% in 2-AAF ＋ PH, 0. 2% in 2-AAF ＋ CCl4, 0. 1% in diabetic, and 0. 0% in control. Isolated cells adhered to plate with fusiform or polygon as epithelial cells. Conclusion Progenitor stem cells exist in injured liver tissue including those from diabetic rats.
基金supported by grants from the Special Funds for Major State Basic Research of China(No.2001CB510101)the National Natural Science Foundation of China(No.30230350 and 60278014)the Natural Science Foundation of Guangdong Province(No.36704)
文摘Summary： Oval cells have a potential to differentiate into a variety of cell lineages including hepatocytes and biliary epithelia. Several models have been established to activate the oval cells by incorporating a variety of toxins and carcinogens, alone or combined with surgical treatment. Those models are obviously not suitable for the study on human hepatic oval cells. It is necessary to establish a new and efficient model to study the human hepatic oval cells. In this study, the hepatocyte growth factor（HGF） and epidermal growth factor（EGF） were used to induce differentiation of mouse embryonic stem（ES） cells into hepatic oval cells. We first confirmed that hepatic oval cells derived from ES cells, which are bipotential, do exist during the course of mouse ES cells＇ differentiation into hepatic parenchymal cells. RT-PCR and transmission electron microscopy were applied in this study. The ratio of Sca-1＋/CD34＋ cells sorted by FACS in the induction group was increased from day 4 and reached the maximum on the day 8, whereas that in the control group remained at a low level. The differentiation ratio of Sca-1＋/CD34＋ cells in the induction group was significantly higher than that in the control group. About 92.48% of the sorted Sca-1＋/CD34＋ cells on the day 8 were A6 positive. Highly purified A6＋/Sca-1＋/CD34＋ hepatic oval cells derived from ES cells could be obtained by FACS. The differentiation ratio of hepatic oval cells in the induction group（up to 4.46%） was significantly higher than that in the control group. The number of hepatic oval cells could be increased significantly by HGF and EGF. The study also examined the ultrastructures of ES-derived hepatic oval cells＇ membrane surface by atomic force microscopy. The ES-derived hepatic oval cells cultured and sorted by our protocols may be available for the future clinical application.
文摘AIM:To investigate the ultrastructure of oval cells in children with chronic hepatitis B,with special emphasis on their location in areas of collagen fibroplasia. METHODS:Morphological investigations were conducted on biopsy material obtained from 40 children,aged 3-16 years with chronic hepatitis B. The stage of fibrosis was assessed histologically using the arbitrary semiquantitative numerical scoring system proposed by Ishak et al. The material for ultrastructural investigation was fixed in glutaraldehyde and paraformaldehyde and processed for transmission-electron microscopic analysis. RESULTS:Ultrastructural examination of biopsy specimens obtained from children with chronic hepatitis B showed the presence of two types of oval cells,the hepatic progenitor cells and intermediate hepatic-like cells. These cells were present in the parenchyma and were seen most commonly in areas of intense periportal fibrosis (at least stage 2 according to Ishak et al) and in the vicinity of the limiting plate of the lobule. The activated nonparenchymal hepatic cells,i.e. transformed hepatic stellate cells and Kupffer cells were seen in close proximity to the intermediate hepatic-like cells. CONCLUSION:We found a distinct relationship between the prevalence of oval cells (hepatic progenitor cells and intermediate hepatocyte-like cells) and fibrosis stage in pediatric patients with chronic hepatitis B.
文摘AIM:To investigate the effect of the neuropeptides bombesin(BBS)and neurotensin(NT)on oval cell proliferation in partially hepatectomized rats not pretreated with a known hepatocyte inhibitor.METHODS:Seventy male Wistar rats were randomly divided into five groups:Ⅰ=controls,Ⅱ=sham operated,Ⅲ=partial hepatectomy 70%(PHx),Ⅳ=PHx+ BBS(30μg/kg per day),Ⅴ=PHx+NT(300μg/kg per day).Forty eight hours after liver resection,portal en-dotoxin levels and hepatic glutathione redox state were determined.α-fetoprotein(AFP)mRNA(in situ hybridisation),cytokeratin-19 and Ki67 antigen expression (immunohistochemistry)and apoptosis(TUNEL)were evaluated on liver tissue samples.Cells with morphological features of oval cells that were cytokeratin-19 (+)and AFP mRNA(+)were scored in morphometric analysis and their proliferation was recorded.In addition,the proliferation and apoptotic rates of hepatocytes were determined.RESULTS:In the control and sham operated groups,oval cells were significantly less compared to groups Ⅲ,ⅣandⅤ(P<0.001).The neuropeptides BBS and NT significantly increased the proliferation of oval cells compared to groupⅢ(P<0.001).In addition,BBS and NT induced a significant increase of hepatocyte proliferation(P<0.001),whereas it decreased their apoptotic activity(P<0.001)compared to groupⅢ.BBS and NT significantly decreased portal endotoxemia (P<0.001)and increased the hepatic GSH:GSSG ratio (P<0.05 and P<0.001,respectively)compared to groupⅢ.CONCLUSION:BBS and NT stimulated oval cell proliferation in a model of liver regeneration,without use of concomitant suppression of hepatocyte proliferation as oval cell activation stimuli,and improved the hepatocyte regenerative response.This peptides-induced combined stimulation of oval cell and hepatocyte proliferation might serve as a possible treatment modality for several liver diseases.
文摘The liver possesses an extraordinary ability to regenerate after injury.Hepatocyte-driven liver regeneration is the default pathway in response to mild-to-moderate acute liver damage.When replication of mature hepatocytes is blocked,facultative hepatic progenitor cells(HPCs),also referred to as oval cells(OCs)in rodents,are activated.HPC/OCs have the ability to proliferate clonogenically and differentiate into several lineages including hepatocytes and bile ductal epithelia.This is a conserved liver injury response that has been studied in many species ranging from mammals(rat,mouse,and human)to fish.In addition,improper HPC/OC activation is closely associated with fibrotic responses,characterized by myofibroblast activation and extracellular matrix production,in many chronic liver diseases.Matrix remodeling and metalloprotease activities play an important role in the regulation of HPC/OC proliferation and fibrosis progression.Thus,understanding molecular mechanisms underlying HPC/OC activation has therapeutic implications for rational design of anti-fibrotic therapies.
