| Objective: To observate the repairing effects of vein-transplanting bone marrow mesenchymal stem cells(BMSCs)on testicular injury induced by cadmium and explore its possible mechanisms by mitochondrial pathway.Methods: Adherent culture method was used for purification,amplification to get Wistar rat BMSCs.The cell cycle and surface markers,CD45 and CD90,of the third generation BMSCs were detected by flow cytometry.21 adult male Wistar rats were randomly assigned control group,model group and cell therapy group,7 in each group.Rats were treated with 0,0.4,0.4 mg.kg-1 body weight of Cd Cl2 saline solution respectively,by intraperitoneal injection for 5 times a week,which lasted for 5 weeks.After that,chloromethyl phenyl formamide(CM-Dil)marked BMSCs were transplanted into rats of the cell therapy group,rats of the control and model groups were treated with the same volume of phosphate buffered saline solution.2 weeks later,rat testes were taken out and weighted,organ coefficient was calculated then;HE dyeing was used for pathology histological observation;laser scanning confocal microscope was used to observe BMSCs in rat testicular frozen slice of the cell treatment group;cadmium content in testicular tissue was determinated;TUNEL method was used to detect cell apoptosis in testicular tissue;immunohistochemical method and the Western blot method were used to detect mitochondrial apoptosis-related proteins,including Bim,Bax,Bcl-2,Cytochrome C,Caspase-3,active-Caspase-3 and AIF,mitochondrial autophagy-related proteins,such as Beclin1?PINK1,Parkin,p-Parkin,LC3 B,mitochondrial biogenesis proteins,involved in SIRT1,PGC-1?;real-time quantitative PCR detection of mt DNA copy number was performed.Results: Flow cytometry results showed that most of the third generation of BMSCs were in relatively inactive intermitotic period and cells expressed CD45 lowly,CD90 highly.2 weeks after transplantation,model rats were significantly depressed and sedentary,behavioral and psychological state of cell treating rats were better than that of model rats,and rat body mass of cell treating group was significantly higher than that of the model group(P<0.05).Testicular coefficient of model rats was in no significant difference compared with the control rats.HE staining results showed rat testis seminiferous,tubule atrophy,cells within the tubule arranged disorderly,spermatogenic cell layers and mature spermatogenic cells level decreased,pathological changes in the treatment group improved significantly compared with those in the model group.CM-Dil-labeled(red fluorescent)BMSCs had been observed in the testes of the cell therapy rats by laser scanning confocal microscope.Atomic absorption results showed that the cadmium content in rat testes in the model and the cell therapy groups was significantly higher than that in the control rats,there was no significant difference between the two groups.TUNEL assay revealed that cell apoptotic rates of rat testicular tissue in the cell therapy group was significantly lower than that in the model group(P<0.05).Results of immunohistochemical assay and Western method showed that the expressions of mitochondrial apoptosis-related proteins Bim,Bax,Cytochrome C,Caspase-3,active-Caspase-3 and AIF increased,Bcl-2 reduced significantly in rat testis of model group compared with that in the others(P<0.05);expression levels of mitochondrial autophagy-related proteins Beclin1,PINK1,Parkin,p-Parkin,LC3 B were significantly higher in the model group than that in the other two groups(P<0.05);expression of mitochondrial biogenesis protein SIRT1 reduced,deacetylation level of PGC-1? decreased in model group rats(P<0.05).Mitochondrial DNA copy number was significantly lower in the model group than that in the control and cell treatment group(P<0.05).Conclusion: Cadmium can enter into the rat,accumulate in the rat testis and cause testicular damage;BMSCs can be located on cadmium-induced damaged testis tissue and repair tissue damage;the repair mechanism may be achieved by inhibiting mitochondrial apoptosis,excessive mitochondrial autophagy of testicle tissue and promoting mitochondrial biogenesis. |
PDF Full Text Request