兔肌腱细胞的分离、培养及鉴定

背景：肌腱细胞是一种高分化的细胞,其增殖相对缓慢,在体外经多次传代后甚至丧失增殖能力,因此有必要建立肌腱细胞良好的体外分离、培养模式。目的：探讨兔肌腱细胞的分离、培养及鉴定。方法：无菌条件下切取新西兰乳兔趾屈肌腱,显微镜下剥离腱外膜,采用Henderson分步酶消化法分离肌腱细胞,用含体积分数为20%胎牛血清的F-12培养液进行培养、传代。结果与结论：通过不同的酶消化分离可获得较纯肌腱细胞,体外培养细胞表现出良好的细胞增殖能力和传代能力。免疫组织化学染色见分离培养的第2代腱细胞Ⅰ型胶原抗体染色呈阳性,而Ⅲ型胶原抗体染色呈阴性,证明所获细胞为肌腱细胞。提示肌腱细胞能够在体外分离、扩增和传代。

Isolation,culture and identification of rabbit tendon cells

BACKGROUND： Tendon cells are characterized by high differentiation potentials, slow proliferation rate, and even lost proliferation capacity after passage in vitro. Therefore it is necessary to establish the ideal isolation and culture patterns of tendon cells in vitro. OBJECTIVE： To investigate the isolation, culture and identification of tendon cells. METHODS： The flexor tendon of New Zealand fetal rabbits were cut under sterile conditions, the peritenon of flexor tendon was removed by microsurgical technique. Tendon cells were isolated with Henderson-step enzymatic digestion method and cultured in a complete medium consisting of DMEM/F12 and 20% fetal bovine serum for primary culture and passage. RESULTS AND CONCLUSION： The tendon cells with high purity can be successfully isolated by different enzymatic digestion methods, and the cultured cells well proliferated and passaged in vitro. The immunohistochemical staining showed that, passage 2 cells were positive for collagen type I antibody, but negative for collagen type III antibody. These evidences confirmed that the cultured cells are tendon cells. Tendon cells can be isolated, proliferated and passaged in vitro.