目的：评价橙皮苷对体外培养的人翼状胬肉成纤维细胞(HPF)增殖的抑制作用及对周期蛋白D(cyclin D)表达的影响。

方法：将人翼状胬肉新鲜组织进行体外贴壁细胞常规培养，并采用免疫荧光染色法进行鉴定。通过MTT法检测不同浓度丝裂霉素C(MMC)(1.5、7.5、30.0μmol/L)和橙皮苷(24、48、64、72、96、120μmol/L)作用不同时间(24、48、72h)对细胞增殖的抑制，筛选适宜的作用浓度和时间。采用Western blot法检测cyclin D在HPF细胞中的相对表达量。

结果：分别采用48、72μmol/L橙皮苷和7.5μmol/L MMC作用于HPF细胞48h，Western blot检测结果显示，空白对照组(正常培养)、MMC组、48μmol/L橙皮苷组、72μmol/L橙皮苷组细胞cyclin D蛋白相对表达量分别为1.20±0.02、0.60±0.03、0.54±0.02、0.45±0.07(F=73.025，P=0.001)，MMC组和橙皮苷组细胞cyclin D蛋白相对表达量均低于空白对照组(P<0.05)，但MMC组和橙皮苷组(48、72μmol/L)细胞cyclin D蛋白相对表达量均无差异(P>0.05)。

结论：橙皮苷能抑制翼状胬肉HPF细胞增殖，该过程与其降低cyclin D的表达有关。

METHODS: The fresh tissue of human pterygium was cultivated by adherent cell culture in vitro and adherent cells were appraised by immune fluorescence staining. HPF cells were treated with hesperidin(24μmol/L, 48μmol/L, 64μmol/L, 72μmol/L, 96μmol/L, 120μmol/L)and MMC(1.5μmol/L, 7.5μmol/L and 30.0μmol/L). The inhibition rate of cell proliferation was detected by MTT assay 24h, 48h and 72h after treatment, and appropriate concentration and time were selected. The relative expression of cyclin D in HPF was detected by Western blot.

RESULTS: When HPF were treated respectively with hesperidin(48μmol/L, 72μmol/L)and MMC(7.5μmol/L)for 48h, Western blot results showed the relative expressions of cyclin D in blank control group(normal culture), MMC group, hesperidin(48μmol/L)group and hesperidin(72μmol/L)group to be 1.20±0.02, 0.60±0.03, 0.54±0.02, 0.45±0.07(F=73.025, P=0.001)respectively. The relative expressions of cyclin D in MMC group and hesperidin group were lower than that of blank control group(P<0.05); while the relative expressions of cyclin D in MMC group and hesperidin(48μmol/L, 72μmol/L)group showed no significant difference(P>0.05).

CONCLUSION:Hesperidin can inhibit the proliferation of HPF by reducing the relative expression of cyclin D.