Silencing GRP78 Expression Enhances The Sensitivity Of HOS Cells To MPPα-PDT Therapy

Objective: To observe the role of glucose regulated protein 78(GRP78)in the treatment of pyropheophorbide-α methyl ester-mediated photodynamic therapy(MPPα-PDT)on human osteosarcoma HOS cells.Methods:We observed the HOS apoptotic morphology and apoptosis after MPPα-PDT by Hoechst 33258 apoptosis staining and flow cytometry(FCM),respectively.HOS osteosarcoma cells were treated by MPPα-PDT for 3,6,12 and 24 h,respectively.The expression of GRP78 protein was detected by western blotting and immunofluorescence.HOS osteosarcoma cells were transfected with si RNA-GRP78 and divided into the Control group,si RNA-GRP78 group,si RNA-GRP78+MPPα-PDT group and MPPα-PDT group.Cell viability was detected by cell counting kit-8(CCK-8)assay.Cell proliferation-related proteins(Ki67 and PCNA),apoptosis-related proteins(Cleaved caspase 3 and Cleaved PARP)and Wnt/β-catenin signaling pathway related proteins were detected by western blotting.The apoptosis rate was analyzed by flow cytometry.Reactive oxygen species(ROS)levels were detected by flow cytometry and confocal fluorescence microscope.Results:Hoechst 33258 apoptosis staining showed the morphological changes of apoptosis in MPPα-PDT,such as bright blue and nuclear mitosis and nuclear pyknosis.Compared with the Control group,MPPαgroup and LED group,flow cytometry demonstrated that the apoptosis rate in the MPPα-PDT 24 h group was increased(P < 0.05).After successful transfection with si RNA-GRP78,HOS cell proliferation was decreased(P < 0.05),and apoptosis-related proteins were increased(P <0.05),Wnt/β-catenin-related proteins were decreased(P < 0.05),and ROS levels were increased(P < 0.05).Conlusion:MPPα-PDT can induce HOS cell apoptosis and increase GRP78 expression.After effective silencing of GRP78 expression by si RNA-GRP78,HOS cells proliferation activity decrease,apoptosis increase,and increase the sensitivity of HOS cells to MPPα-PDT.The mechanism may be related to inhibiting the Wnt/β-catenin signaling pathway activation and increasing ROS levels.