Effects of TSG-6 gene expression mesenchymal stem cells on liver transplantation rejection in rats
10.3760/cma.j.issn.1673-4203.2016.07.004
- VernacularTitle:骨髓间充质干细胞表达肿瘤坏死因子刺激蛋白-6对大鼠肝脏移植排斥反应的影响
- Author:
Yong LIU
;
Hao WU
;
Ding CAO
;
Yakun WU
;
Jianping GONG
;
Xianfeng CHEN
- Publication Type:Journal Article
- Keywords:
Mesenchymal stem cells;
Tumor necrosis factors;
Liver transplantation
- From:
International Journal of Surgery
2016;43(7):443-449,封3
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the effects of silencing TSG-6 gene modified bone marrow mesenchymal stem cells (BMSC) transplantation on liver allotransplantation rejection in rats.Methods BMSC and KCs were isolated from rats and cultured in complete medium.We down-regulated TSG-6 expression of BMSC with lentiviral vectors carrying short hairpin RNA (LV3-shTSG-6).TSG-6mRNA and protein level of BMSC were tested respectively by quantitative real-time PCR and western blot analysis.After co-cuhured between BMSC and KCs,the expression of TNF-α and TSG-6 in cell supernatants were tested.Then,we established the orthotopic liver transplantation models in rats,the rats of each group were killed at 1 days,3 days and 7 days after operation.The serum levels of AST,ALT,TBIL,γ-GGT,TNF-α,IL-6 and IL-4 were tested with ELISA in each group,TSG-6mRNA and protein level of liver tissues obtained from each group were tested by quantitative real-time PCR and western blot analysis.The 1iver tissues of each group were stained with HE,then microstructure of liver tissues were observed under light microscope.The postoperative survival rates in other rats of each group were observed.Results The lentivirus transfection efficiency of mesenchymal stem cells was beyond 70 percent;After co-cultured between BMSC and KCs,the expression of TSG-6 in TSG-6-shRNA-BMSC + KCs group supernatant was significantly lower in different time point than that in BMSC + KCs group and TSG-6-NC-BMSC + KCs group (P ＜0.05),and the expression of TNF-α peak in BMSC + KCs group and TSG-6-NC-BMSC + KCs group supernatants were at 6 hours,which were significandy lower than those at 12 hours (P ＜0.05),but the expression of TNF-α in TSG-6-shRNABMSC + KCs group at 12 hours was significantly higher than those in BMSC + KCs group and TSG-6-NC-BMSC +KCs group (P ＜0.05);After transplantation,the serum levels of AST,ALT,TBIL,γ-GGT,TNF-α and IL-6 in TSG-6-shRNA-BMSC group was significantly higher than those in TSG-6-NC-BMSC group and BMSC group in different time point (P ＜ 0.05),but had no significant difference compared with PBS group;The serum levels of IL-4 in TSG-6-shRNA-BMSC group,TSG-6-NC-BMSC group and BMSC group was significantly higher than that in PBS group(P ＜ 0.05),but TSG-6-shRNA-BMSC group compared with TSG-6-NC-BMSC group and BMSC group,the serum levels of IL-4 was significantly lower (P ＜ 0.05);In pathological changes,we found that the degree of liver rejection in TSG-6-shRNA-BMSC group and PBS group were seriously obvious,and were graded Ⅱ-Ⅲ with Banff schedule;Comparation of postoperative survival time in each group,TSG-6-shRNA-BMSC group and PBS group were (16.6 ±4.6) d and (15.4 ± 6.7) d respectively,which were signifcantly lower than those in TSG-6-NCBMSC group (69.6 ± 28.1) d and BMSC group (69.2 ± 28.2) d (P ＜ 0.05).Conclusion Transplantation of BMSC secreted TSG-6 could,to some extent,mitigate acute liver transplantation rejection.
