Objective To explore the molecular mechanism of plasmalemmal vesicle-associated protein(PLVAP)regulating high glucose(HG)induced endocytosis dysfunction in human liver sinusoidal endothelial cells(HLSECs)through Kelch-like epichlorohydrin ECH-associated protein 1(Keap1)/nuclear factor erythroid 2-related factor 2(Nrf2)/Maf protein.Methods HLSECs were cultured in vitro and divided into normal control(NC)group,high glucose(HG)group,PLVAP overexpression recombinant vector(LV-PLVAP)group,lentivirus empty vector(LV-CON)group,HG+LV-PLVAP+Nrf2 inhibitor(ML385)group,HG+LV-PLVAP+Maf inhibitor(Mafenide)group,HG+LV-CON+ML385 group and HG+LV-CON+Mafenide group.Cell activity was detected by CCK-8 assay.The transfection efficiency of LV-PLVAP was observed by fluorescence microscopy.The fluorescence expression of PLVAP and Nrf2 was detected by immunofluorescence.RT-PCR and western blot were used to detect the mRNA and protein expression of PLVAP,Keap1,Nrf2,and Maf,Caveolin-1(CAV-1).Results A lot of green fluorescence appeared in LV-PLVAP and LV-CON groups,however,no green fluorescence was shown in NC group.Immunofluorescence results showed that PLVAP was expressed in cell membrane;Nrf2 was expressed in cytoplasm,and in HG group,Nrf2 was expressed in nucleus.Compared with NC group,the expression of PLVAP,Keap1 mRNA and protein decreased in HG group(P<0.05 or P<0.01),while the expression of Nrf2,Maf and CAV-1 mRNA and protein increased in HG group(P<0.01).Compared with HG group,the expression of PLVAP,Keap1,Nrf2 mRNA and protein increased in HG+LV-PLVAP group(P<0.01),while the expression of Maf,CAV-1 mRNA and protein decreased in HG+LV-PLVAP group(P<0.01).Compared with HG+LV-PLVAP group,the expression of CAV-1 mRNA and protein increased in HG+LV-PLVAP+ML385 and HG+LV-PLVAP+Mafenide groups(P<0.05 or P<0.01).Conclusions PLVAP regulates the expression of CAV-1 through the Keap1/Nrf2/Maf pathway and then regulates HG induced endocytosis dys-function in HLSECs.Over expression of PLVAP alleviates this pathological reaction.

Objective To explore the molecular mechanism of plasmalemmal vesicle-associated protein(PLVAP)regulating high glucose(HG)induced endocytosis dysfunction in human liver sinusoidal endothelial cells(HLSECs)through Kelch-like epichlorohydrin ECH-associated protein 1(Keap1)/nuclear factor erythroid 2-related factor 2(Nrf2)/Maf protein.Methods HLSECs were cultured in vitro and divided into normal control(NC)group,high glucose(HG)group,PLVAP overexpression recombinant vector(LV-PLVAP)group,lentivirus empty vector(LV-CON)group,HG+LV-PLVAP+Nrf2 inhibitor(ML385)group,HG+LV-PLVAP+Maf inhibitor(Mafenide)group,HG+LV-CON+ML385 group and HG+LV-CON+Mafenide group.Cell activity was detected by CCK-8 assay.The transfection efficiency of LV-PLVAP was observed by fluorescence microscopy.The fluorescence expression of PLVAP and Nrf2 was detected by immunofluorescence.RT-PCR and western blot were used to detect the mRNA and protein expression of PLVAP,Keap1,Nrf2,and Maf,Caveolin-1(CAV-1).Results A lot of green fluorescence appeared in LV-PLVAP and LV-CON groups,however,no green fluorescence was shown in NC group.Immunofluorescence results showed that PLVAP was expressed in cell membrane;Nrf2 was expressed in cytoplasm,and in HG group,Nrf2 was expressed in nucleus.Compared with NC group,the expression of PLVAP,Keap1 mRNA and protein decreased in HG group(P<0.05 or P<0.01),while the expression of Nrf2,Maf and CAV-1 mRNA and protein increased in HG group(P<0.01).Compared with HG group,the expression of PLVAP,Keap1,Nrf2 mRNA and protein increased in HG+LV-PLVAP group(P<0.01),while the expression of Maf,CAV-1 mRNA and protein decreased in HG+LV-PLVAP group(P<0.01).Compared with HG+LV-PLVAP group,the expression of CAV-1 mRNA and protein increased in HG+LV-PLVAP+ML385 and HG+LV-PLVAP+Mafenide groups(P<0.05 or P<0.01).Conclusions PLVAP regulates the expression of CAV-1 through the Keap1/Nrf2/Maf pathway and then regulates HG induced endocytosis dys-function in HLSECs.Over expression of PLVAP alleviates this pathological reaction.

Objective To investigate the mechanism of action of the long non-coding RNA(lncRNA)taurine up-regulating factor 1(TUG1)with high glucose(HG)-induced cellular pyroptosisin microglial cell.Methods Mouse BV2 cells were cultured and divided into normal control(NC),HG,lentiviral empty vector(sh-Con),TUG1 lentiviral gene silencing vector(sh-TUG1),HG+sh-Con and HG+sh-TUG1 group.RT-PCR was used to detect the expression and transfection efficiency of TUG1 mRNA.Nucleotide-binding oligomerized structural domain-like receptor protein 3(NLRP3),cysteoaspartate protease-1(Caspase-1),and ghrelin D(GSDMD),IL-18,IL-1β mRNA and protein expression were detected by RT-PCR and Western blot.Results TUG1 mRNA expression was higher in HG group than in NC group(P＜0.05).After transfection,a lot of green fluorescence appeared in sh-TUG1 and sh-Con group,while no green fluorescence was observed in NC group.The expression of TUG1 mRNA was lower in sh-TUG1 group than in NC group(P＜0.05).Accordingly,the recombinant lentivirus successfully infected BV2 cell.The expressions of the mRNA and protein of NLRP3,Caspase-1,GSDMD,IL-18 and L-1β were higher in HG,HG+sh-Con groups than in NC group(P＜0.05).The expressions of the mRNA and protein of NLRP3,Caspase-1,GSDMD and IL-18 and L-1β were lower in HG+shTUG1 group than in HG,HG+sh-Con groups(P＜0.05).Conclusions TUG1 is involved in high glucose induced pyroptosis in microglia and leads to inflammatory response.Silencing TUG1 can inhibit the pathological reaction.