Negative pressure promotes epidermal differentiation of bone marrow mesenchymal stem cells

doi:10.3969/j.issn.2095-4344.0418

Abstract

Abstract:

BACKGROUND: As a physical factor, negative pressure can promote the osteogenic differentiation and endothelial differentiation of mesenchymal stem cells. If a negative pressure exerts effects on the epidermal differentiation of mesenchymal stem cells, it will be highly important for the combination use of negative pressure and mesenchymal stem cells in wound healing.
OBJECTIVE: To explore the effect of negative pressure on the epidermal differentiation of bone marrow mesenchymal stem cells.
METHODS: Bone marrow mesenchymal stem cells of New Zealand white rabbits were isolated and cultured. Then, the passage 3 cells were induced for epidermal differentiation under negative pressure (-16.625 kPA, twice a day, once for 4 hours) as experimental group. Another cells induced under no negative pressure were used as control group. After induction, cell growth curve was drawn in each group, and the expression of cytokeratin 5 and cytokeratin 10 mRNA was examined by real-time PCR at 2 weeks after induction.
RESULTS AND CONCLUSION: The cell growth of the experimental group was inhibited, and the mRNA expression of cytokeratin 5 and cytokeratin 10 was significantly increased compared with the control group (P < 0.05). These findings indicate that under the condition of negative pressures, the epidermal differentiation ability of bone marrow mesenchymal stem cells is increased, and in contrary, the cell proliferation is inhibited.

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: Bone Marrow, Mesenchymal Stem Cells, Pressure, Epidermis, Cell Differentiation, Tissue Engineering

CLC Number:

R349.2

Zhao Bo-wen, Xu Qiang, Ge Quan-hu, Li Bo-long, Peng Xin-yu, Zhang Hong-wei, Wu Xiang-wei. Negative pressure promotes epidermal differentiation of bone marrow mesenchymal stem cells[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(1): 101-106.

Figures/Tables 2

2.1 倒置显微镜观察骨髓间充质干细胞的形态刚接种时细胞多为圆形或椭圆形，五六个小时后细胞贴壁呈椭圆形，可见大量圆形淋巴细胞和血细胞未贴壁；48 h后初期贴壁椭圆形细胞开始变形呈棒状或梭形，4-6 d后细胞快速增殖，形成分散的细胞集落；8-10 d后细胞融合至80%-90%，细胞为梭形，呈鱼群样生长(图1A)。第2代骨髓间充质干细胞传代，接种4 h可见细胞贴壁呈现短梭形；24 h后细胞全部贴壁，可见部分细胞变为长梭形，部分细胞变为不规则形，杂细胞较少，细胞壁不光滑可能因传代时胰酶消化以及吹打造成细胞损伤；72 h后可见细胞形态较为均一，均为长梭形，细胞壁光滑，细胞数量较前明显增多(图1B)。 2.2 流式细胞仪鉴定兔骨髓间充质干细胞流式细胞仪检测结果显示，99.4%的细胞表达CD44，98.9%的细胞表达CD29，0%的细胞表达CD34。细胞高表达骨髓间充质干细胞表面抗原CD44、CD29，低表达或不表达骨髓成纤维细胞、造血干细胞表面抗原CD34，见图2-4。 2.3 负压下成表皮诱导的骨髓间充质干细胞生长曲线在负压下诱导第4天时，0 kPa组细胞数为(14.17±0.76)×104个，-16.625 kPa组细胞数为(12.08±0.65) ×104个，两组相比差异有显著性意义(P < 0.05)；在负压下诱导第5-7天，0 kPa组与-16.625 kPa组之间差异有显著性意义(P < 0.05)，负压下细胞增殖受抑制(图5)。 2.4 免疫荧光检测表皮样细胞Pan-Keratin的表达由图6A可见未经过诱导的骨髓间充质干细胞不表达或者稍表达Pan-Keratin；由图6B可见经过成表皮诱导两三周后表皮样细胞表达Pan-Keratin，说明骨髓间充质干细胞可向表皮细胞诱导分化。 2.5 荧光定量PCR检测CK-5、CK-10 mRNA的表达经过负压成表皮诱导14 d后，CK-5、CK-10 mRNA表达量较0 kPa组均有明显增高，差异有显著性意义(P < 0.05)，见图7。"

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