Utilization of adiponectin-modified adipose stem cell transplantation in the rat with nephrotic syndrome

doi:10.3969/j.issn.2095-4344.1588

Abstract

Abstract:

BACKGROUND: The expression of adiponectin gene plays an important role in the pathogenesis of nephrotic syndrome. Transplantation of adipose stem cells can has a certain therapeutic effect in rats with nephrotic syndrome.
OBJECTIVE: To investigate the effect of adiponectin gene-transfected adipose stem cell transplantation on renal function and coagulation function in rats with nephrotic syndrome.
METHODS: (1) Adipose stem cells were cultured in vitro and the cell suspension was prepared. CM-Dil was used as a marker. The third generation of adipose stem cells was used for electrotransfection using adiponectin gene. Western blot was used to detect the relative expression level of adiponectin in the cells at 3 and 28 days of transfection. (2) The random 20 of 80 Sprague-Dawley rats (provided by Beijing Vitone Lihua Animal Experimental Technology Co., Ltd., China) were taken as normal control group without any treatment, and the remaining 60 rats were intravenously injected with doxorubicin to establish nephrotic syndrome models. These rat models were randomized into three groups: model group, adipose stem cell group and adiponectin-transfected adipose stem cell group. Normal control group and model group rats were injected 20 μL of PBS via the tail vein, and rats in the adipose stem cell group and adiponectin-transfected adipose stem cell group were intravenously injected with 20 μL of adipose stem cell suspension and adiponectin-transfected adipose stem cell suspension, respectively, once a day, for 7 continuous days. (3) At 4 weeks after transplantation, a full-automatic biochemical analyzer was used to detect serum biochemical indicators and coagulation indicators, hematoxylin-eosin staining used to observe renal histopathological changes, RT-PCR and Western blue used to detect the expression of heparanase and adiponectin in the kidney tissue.
RESULTS AND CONCLUSION: (1) The high expression of adiponectin in adipose stem cells continued for 28 days. (2) The pathological improvement of adiponectin-transfected adipose stem cell group was superior to that of the adipose stem cell group, and the pathological manifestation in the adipose stem cell group was also slightly improved compared to the model group. (3) Serum levels of total cholesterol, triglyceride, low-density lipoprotein, creatinine, and 24-hour urea nitrogen level were significantly lower in the adipose stem cell group and adiponectin-transfected adipose stem cell group than in the model group (P < 0.05), while the serum levels of total protein, albumin, and high-density lipoprotein were significantly higher than the model group (P < 0.05). The physiological and biochemical indicators of adiponectin-transfected adipose stem cell group were superior to those of adipose stem cell group, and the difference was significant (P < 0.05). The hypercoagulant state of blood in the adipose stem cell group and adiponectin-transfected adipose stem cell group was significantly reduced compared to the model group, and this hypercoagulant state in adiponectin-transfected adipose stem cell group was significantly relieved (P < 0.05). (4) Compared with the model group, the expression levels of heparanase mRNA and protein in the kidney tissue of adipose stem cell group and adiponectin-transfected adipose stem cell group were significantly reduced (P < 0.05), and the expression level of adiponectin increased significantly (P < 0.05). The mRNA and protein expressions of heparanase and adiponectin in the adiponectin-transfected adipose stem cell group were on average superior to those in the adipose stem cell group (P < 0.05). To conclude, adiponectin-transfected adipose stem cell transplantation could significantly improve renal function, blood hypercoagulability and hyperlipidemia in rats with nephrotic syndrome. This effect may be related to the expression levels of heparanase and adiponectin.

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: Nephrosis, Adiponectin, Adipose Tissue, Stem Cells, Tissue Engineering

CLC Number:

Tian Meng, Han Hongling. Utilization of adiponectin-modified adipose stem cell transplantation in the rat with nephrotic syndrome[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(5): 723-728.

Figures/Tables 3

2.3 各组大鼠血清生化指标、凝血指标和24 h尿蛋白含量变化见表2。模型组大鼠的24 h尿蛋白定量及血清总胆固醇、三酰甘油、低密度脂蛋白、肌酐、尿素氮水平与正常组比较均明显升高(P < 0.05)，血清总蛋白、白蛋白、高密度脂蛋白水平则明显降低(P < 0.01)。脂肪干细胞组及脂联素修饰脂肪干细胞组的24 h尿蛋白定量及血清总胆固醇、三酰甘油、低密度脂蛋白、肌酐、尿素氮水平均明显低于模型组(P < 0.05)，而血清总蛋白、白蛋白、高密度脂蛋白水平则明显高于模型组(P < 0.05)。脂联素修饰脂肪干细胞组各项生理生化指标均优于脂肪干细胞组，差异有显著性意义(P < 0.05)。脂肪干细胞组、脂联素修饰脂肪干细胞组大鼠的血液高凝状态较模型组明显缓解，脂联素修饰脂肪干细胞组缓解更明显，各组间差异有显著性意义(P < 0.05)。 2.4 各组大鼠肾组织形态学变化见图3。正常对照组有明显清晰的正常肾小管结构，而模型组肾小管结构破坏较为严重；脂肪干细胞组可见少量的炎性细胞浸润但肾小管结构较为正常；脂联素修饰脂肪干细胞组相对于脂肪干细胞组炎性细胞浸润明显减轻，肾小管结构较为正常。 2.5 荧光显微镜观察CM-Dil标记的脂肪干细胞的存活和分布情况见图4。脂肪干细胞组[(46.58±4.69)个/高倍视野]和脂联素修饰脂肪干细胞组[(98.52±9.42)个/高倍视野]肾脏区域中可见较多的CM-Dil标记阳性细胞，脂联素修饰脂肪干细胞组肾组织中阳性标记细胞数量明显多于脂肪干细胞组，正常对照组和模型组未见CM-Dil标记阳性细胞存在。 2.6 各组大鼠肾组织乙酰肝素酶及脂联素mRNA和蛋白的相对表达水平见表3和图5。模型组乙酰肝素酶mRNA和蛋白表达较正常对照组明显增加(P < 0.05)，脂肪干细胞组和脂联素修饰脂肪干细胞组乙酰肝素酶mRNA和蛋白表达较模型组减少(P < 0.05)；脂联素修饰脂肪干细胞组乙酰肝素酶mRNA和蛋白相对表达量较脂肪干细胞组进一步减少(P < 0.05)。模型组脂联素mRNA和蛋白表达较正常对照组明显减少(P < 0.05)，脂肪干细胞组和脂联素修饰脂肪干细胞组脂联素mRNA和蛋白表达较模型组增加(P < 0.05)；脂联素修饰脂肪干细胞组脂联素mRNA和蛋白相对表达量较脂肪干细胞组进一步增加(P < 0.05)。"

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