尖锐湿疣患者角质形成细胞的培养及其生物学特性鉴定

Culture and biological characters of keratinocytes of patients with condyloma acuminatum

目的对尖锐湿疣(condyloma acuminatum,CA)患者皮损处角质形成细胞进行培养及鉴定,测定β-catenin m RNA表达情况,分析细胞周期的变化。方法采用两步消化法对皮损细胞进行消化,获得原代角质形成细胞后,进行体外无血清培养基培养。镜下观察细胞形态,并进行免疫组化染色鉴定;绘制细胞生长曲线;MTT法检测细胞增殖能力;流式细胞仪测定细胞周期。结果镜下可见细胞为多角形或不规则形,呈铺路石状,细胞生长旺盛,轮廓清晰,折光性好,胞浆均被染成红色,为角蛋白阳性,于第3天开始进入对数生长期,第7、8天进入平台期。获得的角质形成细胞可稳定增殖,在CA的发病过程中,β-catenin m RNA的表达异常,细胞周期发生改变。结论 CA的发病过程中β-catenin m RNA的表达异常及细胞周期的改变可为CA的临床治疗提供了理论依据。

Objective To culture and identify the keratinocytes in skin lesion sites of patients with condyloma acuminatum（CA）, determine the expression of β-catenin m RNA and analyze the cell cycle. Methods The cells in lesion sites were digested by two-step digestion method, and the obtained primary keratinocytes were cultured in vitro in serum-free medium, observed for morphology under microscope and identified by immunohistochemical staining, based on which the cell growth curve was plotted, the cell proliferation ability was determined by MTT assay, and cell cycle by flow cytometry. Results The cells under microscope were mainly in polygonal or irregular shape and in flagstone-like array,which grew well with clear outline and good refractivity, and the cytoplasm was stained red, indicating the characters of keratinocytes. The cells entered to the logarithmic growth phase on the 3rd day and to the platform phase on days 7 and 8after culture. The obtained keratinocytes were proliferated stably, in which the β-catenin was expressed abnormally during pathogenesis of CA, while the cell cycle changed. Conclusion Abnormal expression of β-catenin m RNA and change of cell cycle during pathogenesis of CA provided a theoretical basis for the clinical treatment of CA.