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脂肪组织来源的间充质干细胞移植治疗肝纤维化的效果及可能机制

[Effect of adipose tissue-derived mesenchymal stem cell transplantation in treatment of liver fibrosis and possible mechanism].

作者信息

Tang W P, Hu Q, Dai X M, He G S, Luo J X

机构信息

Division of Hepatobiliary and Pancreatic Surgery, First Affiliated Hospital of University of South China, Hengyang 421001, China.

出版信息

Zhonghua Gan Zang Bing Za Zhi. 2018 Jan 20;26(1):28-33. doi: 10.3760/cma.j.issn.1007-3418.2018.01.008.

Abstract

To investigate the effect of adipose tissue-derived mesenchymal stem cell (ADSC) transplantation in the treatment of liver fibrosis rats and possible mechanism. Subcutaneous adipose tissue in the inguinal region of rats was collected to isolate ADSCs. The rats with liver fibrosis induced by intraperitoneally injected carbon tetrachloride were divided into cell transplantation group and phosphate buffer saline (PBS) injection group, and the rats which were fed normally were enrolled as negative control group. The rats in the cell transplantation group were given tail vein injection of ADSCs, and those in the PBS injection group were given injection of 0.5 ml PBS. At 7 days after transplantation, blood samples were collected from the inferior vena cava to evaluate liver function; liver tissue was collected to measure the protein expression of hepatocyte growth factor (HGF) and alpha-smooth muscle actin (α-SMA); Masson trichrome staining was used to evaluate intrahepatic collagen deposition. Hepatic stellate cells (HSCs) were collected from the rats with liver fibrosis, and indirect co-culture of HSCs and ADSCs was performed in vitro to analyze the influence of ADSCs on the proliferation and apoptosis of HSCs. The independent samples t-test was used for comparison between groups, and an analysis of variance was used for comparison of means between multiple samples. ADSCs were found in liver tissue in the transplantation group, and compared with the PBS injection group, the transplantation group had significant alleviation in hepatocyte necrosis, vacuolization, and area of fibrosis and significant reductions in the serum levels of aminotransferases, while there was no significant difference in the level of albumin between the two groups. Compared with the PBS injection group, the transplantation group had significant upregulation in the protein expression of HGF and significant downregulation in the protein expression of α-SMA (both < 0.05). In vitro co-culture for 72 hours showed that ADSCs inhibited the proliferation of HSCs, and there was a significant difference between the co-culture group and the control group with HSCs cultured alone. Caspase-3 immunostaining showed that after co-culture for 72 hours, there was a significant difference in the apoptosis rate of HSCs between the co-culture group and the control group with HSCs cultured alone (23.42% ± 3.02% vs 14.82% ± 3.93%). ADSC transplantation can upregulate the expression of HGF in the liver, promote the apoptosis of HSCs, and thus alleviate liver fibrosis.

摘要

探讨脂肪组织来源的间充质干细胞(ADSC)移植对肝纤维化大鼠的治疗作用及可能机制。收集大鼠腹股沟区皮下脂肪组织分离ADSCs。将腹腔注射四氯化碳诱导肝纤维化的大鼠分为细胞移植组和磷酸盐缓冲液(PBS)注射组,将正常喂养的大鼠作为阴性对照组。细胞移植组大鼠尾静脉注射ADSCs,PBS注射组大鼠注射0.5 ml PBS。移植后7天,从下腔静脉采集血样评估肝功能;采集肝组织检测肝细胞生长因子(HGF)和α-平滑肌肌动蛋白(α-SMA)的蛋白表达;采用Masson三色染色法评估肝内胶原沉积。从肝纤维化大鼠中分离肝星状细胞(HSCs),体外进行HSCs与ADSCs间接共培养,分析ADSCs对HSCs增殖和凋亡的影响。组间比较采用独立样本t检验,多个样本均数比较采用方差分析。移植组肝组织中发现ADSCs,与PBS注射组相比,移植组肝细胞坏死、空泡化及纤维化面积明显减轻,血清转氨酶水平明显降低,而两组白蛋白水平差异无统计学意义。与PBS注射组相比,移植组HGF蛋白表达明显上调,α-SMA蛋白表达明显下调(均P<0.05)。体外共培养72小时显示,ADSCs抑制HSCs增殖,共培养组与单独培养HSCs的对照组之间差异有统计学意义。Caspase-3免疫染色显示,共培养72小时后,共培养组与单独培养HSCs的对照组之间HSCs凋亡率差异有统计学意义(23.42%±3.02% vs 14.82%±3.93%)。ADSC移植可上调肝脏中HGF的表达,促进HSCs凋亡,从而减轻肝纤维化。

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