PDX-1 transfected human adipose-derived mesenchymal stem cells: differentiation into islet-like cells and treatment of type 1 diabetes via cell transplantation

doi:10.3969/j.issn.2095-4344.2017.13.016

Abstract

Abstract:

BACKGROUND: Recent studies have shown that adipose-derived mesenchymal stem cells (ADMSCs) not
only have multilineage differentiation potential, but also exert an important role in blood sugar balance and hormone production.
OBJECTIVE: To observe the differentiation potential of human ADMSCs (hADMSCs) into functional islet-like cells and the therapeutic effect of hADMSCs transplantation in diabetic rats.
METHODS: PDX-1 gene was transfected into hADMSCs by adenovirus. Cell differentiation and insulin secretion were identified and detected by dithizone staining and ELISA, respectively. Twenty male Sprague-Dawley rats were randomly divided into control group (n=4), diabetes group (n=8) and transplantation group (n=8). Rats in the latter two groups were subjected to making diabetic models by 65 mg/kg streptozotocin injection. Afterwards, rats in the transplantation group were given PDX-1 transfected ADMSCs via the tail vein.
RESULTS AND CONCLUSION: At 15 days after transfection, the number of insulin positive cells and insulin secretion were both increased significantly (P < 0.05). Fasting glucose levels in the transplantation group decreased significantly (P < 0.05), while the body weight increased significantly (P < 0.05). In the diabetic group, the fasting glucose level still maintained at a high level, and the body weight of rats was significantly decreased. These results implicated that PDX-1 gene could induce hADMSCs differentiating into functional islet-like cells. PDX-1 transfected ADMSCs transplantation is effective in treating diabetic rats, but the mechanism needs further study.

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: Adipose Tissue, Mesenchymal Stem Cell Transplantation, Genes, Homeobox, Diabetes Mellitus, Tissue Engineering

CLC Number:

R394.2

Shi Guang-jun, Bai Guo-li, Tan Xue-ying, Zhu Ji-jun, Xu Ping . PDX-1 transfected human adipose-derived mesenchymal stem cells: differentiation into islet-like cells and treatment of type 1 diabetes via cell transplantation[J]. Chinese Journal of Tissue Engineering Research, 2017, 21(13): 2062-2067.

Figures/Tables 2

2.1 脂肪间充质干细胞的培养与鉴定结果接种24 h之后换液去除非贴壁细胞；两三天可见细胞形态改变，较均一，成纤维细胞样生长，细胞生长有极性，融合后呈螺旋状(图1)。流式细胞术检测脂肪间充质干细胞高表达分子表型CD29，CD73，CD49d，CD90，低表达分子表型CD14，CD45，CD34，HLA-DR(图2)。脂肪间充质干细胞成脂肪诱导2周后油红O染色阳性，证明其成脂分化能力(图3A)；脂肪间充质干细胞成骨诱导2周后碱性磷酸酶染色呈强阳性反应，证明其成骨分化能力(图3B)。 2.2 转染后细胞PDX-1的表达脂肪间充质干细胞在感染PDX-1 48 h后Western blot法和免疫荧光法均可检测到PDX-1的表达，对照组无PDX-1的表达，且病毒感染后 15 d，仍能检测到PDX-1的表达(图4，5)。 2.3 双硫腙染色结果对转染后细胞进行双硫腙染色，对照组未出现胰岛素阳性细胞，实验组出现双硫腙阳性细胞且被染成棕红色(图6)。 2.4 细胞胰岛素分泌水平转染组和对照组细胞在低糖环境下分泌的胰岛素量差异无显著性意义(P > 0.05)，而在高糖环境下分泌的胰岛素量两组之间差异有显著性意义 (P < 0.05)，对照组细胞在刺激前后分泌的胰岛素量差异无显著性意义(表1)。 2.5 PDX-1转染脂肪间充质干细胞移植对糖尿病大鼠空腹血糖的影响大鼠注射链脲佐菌素48 h后，与对照组相比，模型组大鼠空腹血糖明显增加(P < 0.05)，且明显高于正常值；给药后2周，大鼠空腹血糖仍维持在较高水平，且高于对照组(P < 0.05)，见表2。糖尿病大鼠细胞移植后空腹血糖逐渐下降，14 d时空腹血糖水平明显低于糖尿病组(P < 0.05)。糖尿病组大鼠空腹血糖维持较高水平，未见明显变化，见表3。 2.6 PDX-1转染脂肪间充质干细胞移植对糖尿病大鼠体质量的影响细胞移植后，移植组大鼠体质量逐渐增加，14 d时大鼠体质量明显高于糖尿病组(P < 0.05)，而糖尿病组大鼠体质量持续下降，见表4。"

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