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目的探讨碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)对紫外线(ultraviolet,UVA)诱导的巩膜细胞凋亡的影响及其对凋亡抑制的发生机制。方法用组织块培养法培养豚鼠后极部巩膜细胞。利用紫外线诱导细胞凋亡模型,将培养的巩膜细胞分为对照组(无UVA光照)、单纯UVA组(仅暴露于UAA照射),并于UVA照射前加不同剂量的bFGF(UVA+10μg/L bFGF组,UVA+20μg/L bFGF组,UVA+40μg/L bFGF组)。以流式细胞仪检测细胞凋亡,以RT-PCR法和Western-blot技术测定B细胞淋巴瘤-白血病-2基因(B-cell lymphoma/leukemia 2,bcl-2)的表达。结果组织培养后1~2周开始有细胞长出,3~4周融合,免疫荧光鉴定细胞为巩膜成纤维细胞。流式细胞仪检测显示,bFGF能够抑制UVA诱导的巩膜细胞凋亡,对巩膜细胞有保护作用,且bFGF浓度高于20μg/L时抑制作用更加显著;RT-PCR法、Western-blot检测显示,UVA能够引起凋亡抑制基因bcl-2表达下调,但bFGF能够抑制UVA的下调作用,且随着bFGF浓度的升高其抑制效果逐渐增强,巩膜细胞bcl-2基因的表达也逐渐增加。结论外源性bFGF能够通过上调凋亡抑制基因bcl-2的表达而抑制巩膜细胞的凋亡。
Objective To investigate the effect of basic fibroblast growth factor (bFGF) on apoptosis of scleral cells induced by ultraviolet (UVA) and its mechanism of inhibiting apoptosis. Methods The posterior pole scleral cells were cultured with tissue culture method. The cultured scleral cells were divided into control group (no UVA light), UVA group (exposed to UAA only), and different doses of bFGF (UVA + 10μg / L bFGF group, UVA + 20μg / L bFGF group, UVA + 40μg / L bFGF group). Cell apoptosis was detected by flow cytometry, and the expression of B-cell lymphoma / leukemia-2 gene (Bcl-2) was determined by RT-PCR and Western-blot. Results After 1 ~ 2 weeks after the culture, the cells grew out, fused for 3 to 4 weeks, and the cells identified by immunofluorescence were scleral fibroblasts. Flow cytometry showed that bFGF could inhibit the apoptosis of scleral cells induced by UVA and had a protective effect on scleral cells, and the inhibition was more significant when the concentration of bFGF was higher than 20μg / L. The results of RT-PCR and Western- UVA could induce the down-regulation of bcl-2, but bFGF could inhibit the down-regulation of UVA. With the increase of bFGF concentration, the inhibitory effect was enhanced and the expression of bcl-2 gene in scleral cells increased gradually. Conclusion Exogenous bFGF can inhibit the apoptosis of scleral cells by up-regulating the expression of bcl-2.