Protective Effect Of Tacrolimus Through ERS On NRK-52E Cells

The endoplasmic reticulum (ER),an important intracellular organelle of eukaryocyte, is the factory for folding and maturation of newly synthesized transmembrane and secretory proteins. Various stimulates can disorder ER function.Proteins transmission from the ER to Golgi is blocked,and ultimately leading to endoplasmic reticulum stress.Reasons arousing ERS include Ca2+disorder in the cytoplasm,Ischemia and reperfusion,proteinuria,ubiquitin carboxy-terminal hydrolasel inhibitor and so on.GRP78,ORP150and CHOP are the molecular chaperones of ERS.Appropriate ERS triggers the UPR mechanism to protect cells from the damage of harmful factors.UPR protects cells by closeing the protein translation to promote the expression of molecular chaperones,transporting and degradating of misfolded proteins and other mechanisms to reduce the ER load.Those prompts ERS pretreatment has the potential therapeutic role.Strong or lasting ERS causes that cell homeostasis can not be rebuilt and ultimately apoptosis.The study shows that the ERS is related to many diseases closely.In our study, through the establishment of the ERS pretreatment conditions on NRK-52E cell injury model,it investigated that the FK506pretreatment of the d-BSA overload on NRK-52E cells may have protective effect. l.d-BSA overload on NRK-52E cell proliferation, apoptosis rate and ERS:(1)d-BSA overload on NRK-52E cell proliferation, apoptosis rate and ERS:NRK-52E cells induced by different concentrations of d-BSA were divided into normal group and5experimental groups which included1,5,10.20,50mg/ml d-BSA groups.(2)NRK-52E cells were induced by20mg/ml d-BSA which were divided into normal group and3experimental groups by different induces times after induced6,12,24h respectively.Cell proliferation,the rate of apoptosis,GRP78,ORP150and CHOP expression were observed.Results:d-BSA (5,10,20,50mg/ml) overload on NRK-52E cells24h to promote cell proliferation,increased apoptosis rate and increased expression of GRP78, ORP150and CHOP by the d-BSA dose-dependent manner;20m/ml d-BSA overload on NRK-52E cells6,12,24h after were different from the control group;1mg/ml d-BSA overload in NRK-52E cells after the above indicators compared with the control group was no significant.2.The protective effect of FK506pretreatment on d-of BSA overload in NRK-52E cells:(1)10ng/ml FK506incubated normal NRK-52E cells observed after incubated4,8,12,24h,cells of GRP78, ORP150and CHOP expression were observed.(2) FK506(0.1,1,10,20ng/ml)pretreatment in NRK-52E cells after4h adding the final concentration of20mg/ml d-BSA were incubated for24hours,cells of GRP78, ORP150and CHOP expression were observed.(3)10ng/ml FK506pretreatment4h, the final mass concentration of20mg/ml d-of BSA were incubated at different times (6,12,24h).20mg/ml d-BSA group was positive control group,and the normal cultured NRK-52E cells was as the blank control group,cell proliferation,apoptosis rate of GRP78,ORP150and CHOP expression were the observed.Results:10ng/ml FK506incubated normal NRK-52E cells for4h,GRP78,ORP150expression began to increase,the difference was statistically significant compared with the control group (P<0.05);8,12,24h after significant difference (P0.01).CHOP expression after incubation for4h significantly increased compared with the control group (P<0.01).Inhibit the d-BSA (20mg/ml) after FK506(10,20ng/ml)pretreatment overload caused to decrease proliferation,cell apoptosis,CHOP expression and to increase the GRP78,ORP150expression. The changes in the indicators and common incubation FK506concentration was positively correlated.After FK506(10ng/ml) and d-BSA (20mg/ml) incubated for6,12,24h,FK506can inhibit the NRK-52cell proliferation of d-BSA overload-induced,increased cell apoptosis,GRP78,ORP150expression and decreased and CHOP expression.The changes in the indicators and common incubation time was positively correlated. Conlusion:This study founded that the induction time and dose conditions as a significant protective effect of FK506-induced ERS pretreatment on the d-BSA overload in NRK-52Ecells,its effect mechanism may involve inhibition of cell proliferation, decreasing cell apoptosis,increasing of ER chaperone and the inhibition ER excessive stress-related apoptotic pathway.