Regression of liver fibrosis after adipose-derived mesenchymal stem cell transplantation via different approaches: an objective standard comparison

doi:10.3969/j.issn.2095-4344.0624

Abstract

Abstract:

BACKGROUND: The choice of transplant approach may directly affect the therapeutic effect of stem cell transplantation on liver injury.
OBJECTIVE: To compare the therapeutic differences in liver fibrosis rats treated with transplantation of adipose-derived mesenchymal stem cells via caudal vein, portal vein and hepatic artery.
METHODS: Seventy-five male Sprague-Dawley rats were selected, 15 of which were used as normal control without any intervention, and the remaining 60 rats were used to establish liver fibrosis models by subcutaneous injection of carbon tetrachloride. After modeling, these model rats were divided into four groups, in three groups of which, allogeneic adipose-derived mesenchymal stem cell suspension (2 mL, cell concentration of 1×10⁹/L) was transplanted through hepatic artery, portal vein and caudal vein. The model group was injected with the same amount of cell culture medium via caudal vein. After 4 weeks of transplantation, liver function, liver fibrosis indexes and liver histopathology were detected and observed.
RESULTS AND CONCLUSION: (1) Compared with the model group, the serum levels of alanine aminotransferase, alanine aminotransferase and total bilirubin were significantly reduced in the three transplantation groups (P < 0.05). Compared with the caudal vein transplantation group, the above indexes in and hepatic artery and portal vein transplantation groups were significantly lowered (P < 0.05). However, there was no difference between hepatic artery transplantation group and portal vein transplantation group. (2) Compared with the model group, the levels of hyaluronic acid, laminin and type III procollagen in liver tissues decreased significantly in the three cell transplantation groups (P < 0.05). Compared with the caudal vein transplantation group, the above indexes were significantly decreased in the hepatic artery transplantation group and portal vein transplantation group (P < 0.05). However, there was no difference in the above indexes between hepatic artery transplantation group and portal vein transplantation group. (3) Findings from hematoxylin-eosin staining and Masson staining showed typical pathological changes of liver fibrosis in the model group. Liver fibrosis was improved to different extents in the three transplantation groups as compared with the model group. The regression of liver fibrosis in hepatic artery transplantation group and portal vein transplantation group was better than that in caudal vein transplantation group. These findings indicate that adipose-derived mesenchymal stem cells transplanted via caudal vein, hepatic artery and portal vein can improve liver function and regress liver fibrosis in hepatic fibrosis rats. Moreover, cell transplantation via hepatic artery and portal vein is better than that via caudal vein.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Stem Cell Transplantation, Liver Cirrhosis, Tissue Engineering

CLC Number:

R394.2

Gu Chun, Yu Xiao-jiong, Dong Ke, Xiong Wei, Peng Sheng-kun. Regression of liver fibrosis after adipose-derived mesenchymal stem cell transplantation via different approaches: an objective standard comparison[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(29): 4663-4668.

Figures/Tables 1

2.1 脂肪间充质干细胞培养与鉴定结果分离培养的脂肪间充质干细胞接种24 h开始贴壁，细胞多为圆形或多角形，随着时间的延长，细胞胞体变大，生长缓慢，换液后细胞增殖加快，5 d可传代；第3代脂肪间充质干细胞生长旺盛，细胞形态伸展为三角形、多角形，最终趋于长梭形，见图1。第3代脂肪间充质干细胞表面标记物CD90、CD29为阳性表达，CD34、CD45为阴性表达(图2)，并可向脂肪细胞与成骨细胞分化(图3)，说明分离培养的为间充质干细胞。 2.2 实验动物数量分析实验过程中每组均保证15只实验动物，最终75只大鼠进入结果分析。 2.3 肝功能指标细胞移植4周后，与正常对照组相比，模型组大鼠血清谷丙转氨酶、谷草转氨酶和总胆红素水平均显著升高(P < 0.05)；与模型组相比，尾静脉移植组、肝动脉移植组和门静脉移植组大鼠谷丙转氨酶、谷草转氨酶及总胆红素水平均显著降低(P < 0.05)；与尾静脉移植组相比，肝动脉移植组和门静脉移植组大鼠谷丙转氨酶、谷草转氨酶及总胆红素水平均显著降低(P < 0.05)；肝动脉移植组大鼠谷丙转氨酶、谷草转氨酶及总胆红素水平与门静脉移植组接近，见表1。 2.4 肝纤维化指标肝组织透明质酸、层粘连蛋白和Ⅲ型前胶原水平能反映细胞外基质的沉积程度[8]。细胞移植4周后，与正常对照组相比，模型组大鼠肝组织透明质酸、层粘连蛋白和Ⅲ型前胶原水平明显升高(P < 0.05)；与模型组相比，尾静脉移植组、肝动脉移植组和门静脉移植组大鼠肝组织透明质酸、层粘连蛋白和Ⅲ型前胶原水平均显著降低(P < 0.05)；与尾静脉移植组相比，肝动脉移植组和门静脉移植组肝组大鼠织透明质酸、层粘连蛋白和Ⅲ型前胶原水平均显著降低(P < 0.05)；肝动脉移植组大鼠肝组织透明质酸、层粘连蛋白和Ⅲ型前胶原水平与门静脉组接近，见表2。 2.5 肝组织病理变化细胞移植4周后，苏木精-伊红染色结果显示，正常对照组肝组织及肝细胞结构正常，无炎细胞浸润及纤维组织增生；模型组可见大范围的肝细胞变性及坏死，纤维组织增生，在汇管区及中央静脉周围有大量炎性细胞浸润，呈典型的肝纤维化病理改变；与模型组相比，尾静脉移植组、肝动脉移植组和门静脉移植组大鼠肝细胞变性及坏死、炎性细胞浸润及纤维组织增生均有不同程度的改善，其中肝动脉移植组和门静脉移植组改善程度优于尾静脉移植组(图4)。 Masson染色结果显示，模型组肝组织呈现典型的肝纤维化表现，尾静脉移植组、门静脉移植组与肝动脉移植组肝纤维化程度均明显轻于模型组，并且门静脉移植组、肝动脉移植组肝纤维化改善程度优于尾静脉移植组(图4B)。 Chevallier肝纤维化组织学半定量计分系统显示，正常对照组为1.23±0.98，模型组为18.86±3.26，尾静脉移植组为13.25±1.69，门静脉移植组为9.28±3.16，肝动脉移植组为9.43±2.97；模型组评分显著高于正常对照组(P < 0.05)；尾静脉移植组、门静脉移植组、肝动脉移植组评分明显低于模型组(P < 0.05)；门静脉移植组、肝动脉移植组评分明显低于尾静脉移植组(P < 0.05)；门静脉移植组与肝动脉移植组评分差异无显著性意义。"

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