Protective Effects Of Heme Oxygenase-1 Transduced Bone Marrow Mesenchymal Stem Cells On Reduced-size Liver Transplantation: Role Of Autophagy Regulated By ERK/mTOR Signaling

Objective:Heme Oxygenase-1(HO-1)transduced bone marrow mesenchymal stem cells(BM MSCs)could regulate immune and repair tissues of liver transplantation.However,the mechanisms remain mostly unknown.Autophagy is a critical lysosomal pathway that degrades cytoplasmic components to maintain cell homeostasis and provide substrates for energy metabolism.The aim of this study was to explore the effects and related mechanism of autophagy on the protection conferred by HO-1 transduced BM MSCs on 50 % reduced-size liver transplantation in a rat model.Methods:Firstly,BM MSCs were isolated,then were transfected with adenovirus carrying HO-1 gene and an acute rejection model following 50 % reduced-size liver transplantation in rats was established,with recipients divided into three groups receiving treatment with BM MSCs,HO-1 transduced BM MSCS(HO-1/BM MSCs),or normal saline(NS)injected through the dorsal penile vein.Transplanted liver tissues at 0,1,3,5,7,10 and 14 d after transplantation were acquired for further analysis.The ultrastructure of the transplanted livers was observed by projection electron microscopy.Pathological changes and the extent of rejection were evaluated under a light microscope,and acute cellular rejection was classified according to the Banff criteria.The levels of microtubule-associated protein 1 light 3(LC3)and autophagy regulator Beclin-1,ERK,p-ERK,mTOR and p-mTOR proteins were detected by Western blotting.Results:1.BM MSCs were identified by morphology,flow cytometry and the capacity of differentiation,these showed that BM MSCs were high purity and had the ability to differentiate into osteoblasts and adipocytes.In addition,BM MSCs were transfected with adenovirus carrying HO-1 gene,the rate was up to 85 %.Rejection activity indices of the HO-1/BM MSCs group indicated significantly alleviated rejection compared with those of the BM MSCs group and NS group.2.Electron microscopy showed slight nuclear pycnosis,reduced tissue damage and increased numbers of autophagosomes in the HO-1/BM MSCs groups.As time passing by,the expression of autophagy-related proteins LC3 and Beclin-1 increased,the levels in HO-1/BM MSCs group was higher than BM MSCs group and the NS group.It showed that autophagy was involved in the protective effects of HO-1/BM MSCs on liver graft.3.As time passing by,the levels of p-ERK increased in the HO-1/BM MSCs compared with BM MSCs group and NS group,these was accordance with the results of autophagy-related proteins.However,and the levels of mTOR and p-mTOR decreased,these was opposite to the results of autophagy-related proteins and p-ERK.with statistically significant differences.Conclusions:We succeeded to culture BM MSCs and build HO-1/BM MSCs.An acute rejection model following 50 % reduced-size liver transplantation in rats was also established successfully.These showed HO-1/BM MSCs could protect liver graft after reduced-size liver transplantation.The results of the ultrastructure and autophagy-related proteins indicated that autophagy is involved in the protective effects of HO-1/BM MSCs on liver graft after 50 % reduced-size liver transplantation,possibly via upregulation of autophagy-related proteins through the ERK/mTOR signaling pathway.