中华皮肤科杂志 ›› 2016, Vol. 49 ›› Issue (6): 396-401.

• 论著 • 上一篇    下一篇

系统性红斑狼疮患者骨髓间充质干细胞的生物学特征及其p27kip1/PTEN蛋白表达

王智琴1,彭学标2   

  1. 1. 南方医大学南方医院
    2. 南方医科大学南方医院风湿科
  • 收稿日期:2015-08-26 修回日期:2015-11-12 发布日期:2016-05-31
  • 通讯作者: 彭学标 E-mail:pengxuebiao@126.com
  • 基金资助:

    广东省自然科学基金项目

Biological characteristics of and protein s of p27kip1/PTEN in bone marrow mesenchymal stem cells from patients with systemic lupus erythematosus

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  • Received:2015-08-26 Revised:2015-11-12 Published:2016-05-31

摘要:

目的 探讨系统性红斑狼疮(SLE)患者骨髓间充质干细胞(BMSC)的生物学行为,证实SLE患者BMSC是衰老的干细胞,并探讨其衰老的可能机制。 方法 用密度梯度离心和贴壁分离法对6例SLE患者和8例健康人BMSC进行分离培养。光学显微镜观察BMSC形态学改变及生长状况并绘制生长曲线,诱导成骨、成脂分化检测BMSC分化能力,流式细胞仪鉴定BMSC表面标志物及分析细胞凋亡和细胞周期,划痕试验检测BMSC迁移能力,免疫荧光法及蛋白印迹法分析BMSC内p27kip1/PTEN的分布及表达。 结果 SLE患者组BMSC形态宽大、扁平、呈多角形,与健康对照组相比,其生长速率、迁移及成骨成脂分化能力均明显降低。SLE组处于早期(Q2)、中晚期(Q4)凋亡的BMSC细胞比例(17.98% ± 3.26%,16.80% ± 9.63%)显著高于健康对照组(8.23% ± 3.25%,3.33% ± 2.21%),差异有统计学意义(t Q2 = 3.91,P Q2 = 0.011;t Q4 = 2.99,P Q4 = 0.048)。SLE组BMSC阻滞于G0/G1期的细胞百分比显著高于健康对照组(92.34% ± 5.80%比78.65% ± 3.22%,t = 3.635,P = 0.015),同时其S期所占细胞百分比明显低于健康对照组(0.86% ± 1.72%比5.06% ± 1.874%;t = 3.084,P = 0.027)。SLE患者BMSC经内参均一化后的p27、PTEN蛋白表达水平(即p27/β肌动蛋白,PTEN/β肌动蛋白)均高于健康对照者,差异有统计学意义(t p27= 2.784,P P27 = 0.039;t PTEN= 4.812,P PTEN = 0.041)。 结论 SLE患者BMSCs表现出衰老特征,可能与细胞内p27kip1/PTEN表达水平升高有关。

Abstract:

Wang Zhiqin, Peng Xuebiao Department of Dermatology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China Corresponding author: Peng Xuebiao, Email: pengxuebiao@126.com 【Abstract】 Objective To evaluate biological behaviors of bone marrow mesenchymal stem cells (BMSCs) from patients with systemic lupus erythematosus (SLE), to confirm that the BMSCs are aging stems cells, and to explore mechanisms underlying their aging. Methods BMSCs were isolated from bone marrow of 6 patients with SLE (patient group) and 8 healthy human controls (control group) by density-gradient centrifugation and plastic adherence, and cultured in vitro. Optical microscopy was conducted to observe morphological changes and growth of BMSCs, and growth curves were drawn. The differentiation ability of BMSCs was evaluated through culture of them with adipogenic and osteogenic induction medium. Flow cytometry was performed to identify cellular surface markers and to analyze cell cycle and apoptosis, and scratch assay to assess the migration ability of BMSCs. Immunofluorescence assay and Western blot analysis were carried out to analyze the distribution and of p27kip1/PTEN in BMSCs respectively. Results Patient-derived BMSCs, which had a broad, flat and polygonal shape, showed decreased growth rate, migration activity as well as adipogenic and osteogenic ability compared with those from the controls. There was a significant increase in the proportion of BMSCs in early stage (17.98% ± 3.26% vs. 8.23% ± 3.25%, t = 3.91, P = 0.011) as well as in middle to late stages (16.80% ± 9.63% vs. 3.33% ± 2.21%, t = 2.99, P = 0.048) of apoptosis in the patient group compared with the control group. Moreover, compared with the control group, the patient group showed a significantly higher proportion of BMSCs arrested in the G0/G1 phase (92.34% ± 5.80% vs. 78.65% ± 3.22%, t = 3.635, P = 0.015), but a lower proportion in the S phase (0.86% ± 1.72% vs. 5.06% ± 1.874%, t = 3.084, P = 0.027). The protein s of p27 and PTEN were significantly higher in the patient group than in the control group (p27/β-actin: t = 2.784, P = 0.039; PTEN/β-actin: t = 4.812, P = 0.041). Conclusion The BMSCs from SLE patients exhibit senescence-related features, which may be associated with elevated levels of p27kip1/PTEN.

中图分类号: 

  • R751

引用本文

王智琴 彭学标. 系统性红斑狼疮患者骨髓间充质干细胞的生物学特征及其p27kip1/PTEN蛋白表达[J]. 中华皮肤科杂志, 2016,49(6):396-401. doi: