成人外周血平滑肌祖细胞与成熟血管平滑肌细胞生物学特性的比较

Comparison of biological properties between adult peripheral blood derived smooth muscle progenitor cells and mature vessel smooth muscle cells

背景:血管平滑肌细胞是构成血管壁组织结构及维持血管张力的主要细胞成分。研究表明从外周血获得的平滑肌祖细胞可分化为血管平滑肌细胞。目的:课题特点在于选择成人外周血来源的平滑肌祖细胞,并与成熟血管平滑肌细胞进行细胞形态、增殖潜能等方面异同点的比较。设计、时间及地点:以细胞为培养对象,观察性实验,于2008-10/2009-04在江西省分子医学重点实验室完成。材料:采取6名健康志愿者外周血20mL,6名下肢大隐静脉曲张患者大隐静脉。方法:分离成人外周血单个核细胞,用纤粘连蛋白、血小板衍生生长因子BB和EGM培养基诱导培养;同期用胶原酶消化大隐静脉获得平滑肌细胞,用高糖DMEM培养基培养。主要观察指标:观察细胞形态,用细胞免疫荧光法和反转录-聚合酶链反应法检测α-SMA和calponin1的表达,四甲基偶氮唑盐法检测细胞增殖能力。结果:成人外周血中平滑肌祖细胞具有血管平滑肌细胞的生物学特性,表达α-SMA和calponin1,平滑肌祖细胞增殖能力明显强于血管平滑肌细胞的增殖能力。结论:成人外周血中单核细胞在血小板衍生生长因子BB的刺激下可以分化为平滑肌祖细胞,且平滑肌祖细胞较平滑肌细胞增殖旺盛,长时间培养不易老化,具有发展为一种理想的血管组织工程种子细胞来源的可能性。

BACKGROUND： Vascular smooth muscle is the main cell component of constructing vascular wall tissue structure and maintaining vascular tension. Studies have shown that smooth muscle progenitor cells obtained from peripheral blood can differentiate into vascular smooth muscle cells. OBJECTIVE： To compare smooth muscle progenitor cells derived from human peripheral blood with mature vessel wall smooth muscle cells in cell morphology and proliferation potential. DESIGN, TIME AND SETTING： Cell culture and observation trials were performed at the Key Laboratory of Molecular Medicine in Jiangxi province from October 2008 to April 2009. MATERIALS： Peripheral blood 20 mL were adopted from 6 healthy volunteers, and great saphenous vein were sourced from 6 patients with varicose vein. METHODS： Mononuclear cells were isolated from human peripheral blood and cultured in fibronectin, platelet derived growth factor-BB and EGM-2 medium. At the same time, the smooth muscle cells were obtained from great saphenous vein with the collagenase digestion method. High-glucose DMEM medium was used for the culture. MAIN OUTCOME MEASURES： Cellular morphology; Cellular immumofluorescence method and RT-PCR method were used to detect the expression ofα-SMA and calponinl. MTT method was used to analyze the proliferation activity. RESULTS： The smooth muscle progenitor cells derived from human peripheral blood exhibited biological characteristics of vascular smooth muscle cells, they were found to express α-SMA and calponinl. The proliferation activity of smooth muscle progenitor cells was also greater than vascular smooth muscle cells. CONCLUSION： Peripheral blood mononuclear cells in the stimulus of platelet-derived growth factor can differentiate into smooth muscle progenitor cells. Compared to smooth muscle cells, smooth muscle progenitor cells proliferate better, and are not easy to age in a long time of cultivation. Smooth muscle progenitor cells are a ideal cell source of tissue engineered vessel.