Following acute and chronic liver injury,hepatic stellate cells (HSCs) become activated to undergo a phenotypic transformation into myofibroblast-like cells and lose their retinol content,but the mechanisms of retinoi...Following acute and chronic liver injury,hepatic stellate cells (HSCs) become activated to undergo a phenotypic transformation into myofibroblast-like cells and lose their retinol content,but the mechanisms of retinoid loss and its potential roles in HSCs activation and liver fibrosis are not understood.The influence of retinoids on HSCs and hepatic fibrosis remains controversial.The purpose of this study was to evaluate the effects of all-trans retinoid acid (ATRA) on cell proliferation,mRNA expression of collagen genes [procollagen α1 (Ⅰ),procollagen α1 (Ⅲ)],profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),fibrolytic genes (MMP-3,MMP-13) and the upstream element (JNK and AP-1) in the rat hepatic stellate cell line (CFSC-2G).Cell proliferation was evaluated by measuring BrdU incorporation.The mRNA expression levels of collagen genes [procollagen α1 (Ⅰ),procollagen α1 (Ⅲ)],profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),and fibrolytic genes (MMP-3,MMP-13) were quantitatively detected by using real-time PCR.The mRNA expression of JNK and AP-1 was quantified by RT-PCR.The results showed that ATRA inhibited HSCs proliferation and diminished the mRNA expression of collagen genes [procollagen α1 (Ⅰ),procollagen α1 (Ⅲ)] and profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),and significantly stimulated the mRNA expression of MMP-3 and MMP-13 in HSCs by suppressing the mRNA expression of JNK and AP-1.These findings suggested that ATRA could inhibit proliferation and collagen production of HSCs via the suppression of active protein-1 and c-Jun N-terminal kinase signal,then decrease the mRNAs expression of profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),and significantly induce the mRNA expression of MMP-3 and MMP-13.展开更多
Background and Aims:Nonalcoholic fatty liver disease(NAFLD)is a common chronic liver disease caused by overnutrition.Impaired autophagy is closely related to NAFLD progression.Recently,ubiquitin-specific peptidase-10(...Background and Aims:Nonalcoholic fatty liver disease(NAFLD)is a common chronic liver disease caused by overnutrition.Impaired autophagy is closely related to NAFLD progression.Recently,ubiquitin-specific peptidase-10(USP10)was reported to ameliorate hepatic steatosis,but the underlying mechanism is still unclear.In view of the potential effects of USP10 on autophagy,we investigated whether USP10 alleviated steatosis through autophagy.Methods:HepG2 cells were treated with palmitic acid(PA)to model NAFLD in vitro.Lentivirus was used to regulate USP10 level in cells.Autophagic regulators were used to autophagic progression in cells.Western blotting,real-time fluorescence quantitative polymerase chain reaction,lipid drop staining and immunofluorescent staining were performed to determine the effect of USP10 on lipid autophagy.Student’s t-test and Tukey’s post hoc test were used to compare the means among groups.Results:PA induced cellular steatosis with dependance on autophagy.USP10 overexpression alleviated PA-induced steatosis,restored autophagic activity,promoted autophagic flux,including synthesis and degradation of autophagosomes,and lipid-targeted autophagy.In the presence of autophagy inhibitors,the protective effectiveness of USP10 on steatosis decreased.Furthermore,the specific inhibitor to C-jun N-terminal protein kinase-1(JNK1),DB07268,abolished USP10-induced autophagy.However,during early stage inhibition of JNK1,compensatory expression of tuberous sclerosis complex-2(TSC2)maintained autophagy.The degree of TSC2-to-JNK1 compensation was positively associated with USP10 level.Functionally,JNK1 and TSC2 were involved in the lipid-lowering effect of USP10.Conclusions:USP10 alleviated hepatocellular steatosis in autophagy-dependent manner.JNK1/TSC2 signaling pathways were required for USP10-induced autophagy.展开更多
AIM To investigated the relationships between HER2, c-Jun N-terminal kinase(JNK) and protein kinase B(AKT) with respect to metastatic potential of HER2-positive gastric cancer(GC) cells.METHODS Immunohistochemistry wa...AIM To investigated the relationships between HER2, c-Jun N-terminal kinase(JNK) and protein kinase B(AKT) with respect to metastatic potential of HER2-positive gastric cancer(GC) cells.METHODS Immunohistochemistry was performed on tissue array slides containing 423 human GC specimens. Using HER2-positve GC cell lines SNU-216 and NCI-N87, HER2 expression was silenced by RNA interference, and the activations of JNK and AKT were suppressed by SP600125 and LY294002, respectively. Transwell assay, Western blot, semi-quantitative reverse transcriptionpolymerase chain reaction and immunofluorescence staining were used in cell culture experiments. RESULTS In GC specimens, HER2, JNK, and AKT activations were positively correlated with each other. In vitro analysis revealed a positive regulatory feedback loop between HER2 and JNK in GC cell lines and the role of JNK as a downstream effector of AKT in the HER2/AKT signaling pathway. JNK inhibition suppressed migratory capacity through reversing EMT and dual inhibition of JNK and AKT induced a more profound effect on cancer cell motility.CONCLUSION HER2, JNK and AKT in human GC specimens are positively associated with each other. JNK and AKT, downstream effectors of HER2, co-operatively contribute to the metastatic potential of HER2-positive GC cells. Thus, targeting of these two molecules in combination with HER2 downregulation may be a good approach to combat HER2-positive GC.展开更多
文摘Following acute and chronic liver injury,hepatic stellate cells (HSCs) become activated to undergo a phenotypic transformation into myofibroblast-like cells and lose their retinol content,but the mechanisms of retinoid loss and its potential roles in HSCs activation and liver fibrosis are not understood.The influence of retinoids on HSCs and hepatic fibrosis remains controversial.The purpose of this study was to evaluate the effects of all-trans retinoid acid (ATRA) on cell proliferation,mRNA expression of collagen genes [procollagen α1 (Ⅰ),procollagen α1 (Ⅲ)],profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),fibrolytic genes (MMP-3,MMP-13) and the upstream element (JNK and AP-1) in the rat hepatic stellate cell line (CFSC-2G).Cell proliferation was evaluated by measuring BrdU incorporation.The mRNA expression levels of collagen genes [procollagen α1 (Ⅰ),procollagen α1 (Ⅲ)],profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),and fibrolytic genes (MMP-3,MMP-13) were quantitatively detected by using real-time PCR.The mRNA expression of JNK and AP-1 was quantified by RT-PCR.The results showed that ATRA inhibited HSCs proliferation and diminished the mRNA expression of collagen genes [procollagen α1 (Ⅰ),procollagen α1 (Ⅲ)] and profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),and significantly stimulated the mRNA expression of MMP-3 and MMP-13 in HSCs by suppressing the mRNA expression of JNK and AP-1.These findings suggested that ATRA could inhibit proliferation and collagen production of HSCs via the suppression of active protein-1 and c-Jun N-terminal kinase signal,then decrease the mRNAs expression of profibrogenic genes (TGF-β 1,CTGF,MMP-2,TIMP-1,TIMP-2,PAI-1),and significantly induce the mRNA expression of MMP-3 and MMP-13.
文摘Background and Aims:Nonalcoholic fatty liver disease(NAFLD)is a common chronic liver disease caused by overnutrition.Impaired autophagy is closely related to NAFLD progression.Recently,ubiquitin-specific peptidase-10(USP10)was reported to ameliorate hepatic steatosis,but the underlying mechanism is still unclear.In view of the potential effects of USP10 on autophagy,we investigated whether USP10 alleviated steatosis through autophagy.Methods:HepG2 cells were treated with palmitic acid(PA)to model NAFLD in vitro.Lentivirus was used to regulate USP10 level in cells.Autophagic regulators were used to autophagic progression in cells.Western blotting,real-time fluorescence quantitative polymerase chain reaction,lipid drop staining and immunofluorescent staining were performed to determine the effect of USP10 on lipid autophagy.Student’s t-test and Tukey’s post hoc test were used to compare the means among groups.Results:PA induced cellular steatosis with dependance on autophagy.USP10 overexpression alleviated PA-induced steatosis,restored autophagic activity,promoted autophagic flux,including synthesis and degradation of autophagosomes,and lipid-targeted autophagy.In the presence of autophagy inhibitors,the protective effectiveness of USP10 on steatosis decreased.Furthermore,the specific inhibitor to C-jun N-terminal protein kinase-1(JNK1),DB07268,abolished USP10-induced autophagy.However,during early stage inhibition of JNK1,compensatory expression of tuberous sclerosis complex-2(TSC2)maintained autophagy.The degree of TSC2-to-JNK1 compensation was positively associated with USP10 level.Functionally,JNK1 and TSC2 were involved in the lipid-lowering effect of USP10.Conclusions:USP10 alleviated hepatocellular steatosis in autophagy-dependent manner.JNK1/TSC2 signaling pathways were required for USP10-induced autophagy.
基金Supported by SNUH Research Fund,Grant NO 04-2016-0220the Education and Research Encouragement Fund of Seoul National University Hospital(2015)
文摘AIM To investigated the relationships between HER2, c-Jun N-terminal kinase(JNK) and protein kinase B(AKT) with respect to metastatic potential of HER2-positive gastric cancer(GC) cells.METHODS Immunohistochemistry was performed on tissue array slides containing 423 human GC specimens. Using HER2-positve GC cell lines SNU-216 and NCI-N87, HER2 expression was silenced by RNA interference, and the activations of JNK and AKT were suppressed by SP600125 and LY294002, respectively. Transwell assay, Western blot, semi-quantitative reverse transcriptionpolymerase chain reaction and immunofluorescence staining were used in cell culture experiments. RESULTS In GC specimens, HER2, JNK, and AKT activations were positively correlated with each other. In vitro analysis revealed a positive regulatory feedback loop between HER2 and JNK in GC cell lines and the role of JNK as a downstream effector of AKT in the HER2/AKT signaling pathway. JNK inhibition suppressed migratory capacity through reversing EMT and dual inhibition of JNK and AKT induced a more profound effect on cancer cell motility.CONCLUSION HER2, JNK and AKT in human GC specimens are positively associated with each other. JNK and AKT, downstream effectors of HER2, co-operatively contribute to the metastatic potential of HER2-positive GC cells. Thus, targeting of these two molecules in combination with HER2 downregulation may be a good approach to combat HER2-positive GC.
