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良性前列腺增生症(BPH)是中老年男性的一种常见病,为探讨性激素对前列腺细胞增殖的作用机理,本课题收集BPH前列腺组织,经硫酸铵分段沉淀、肝素琼脂糖亲和层析,NaCl浓度梯度洗脱,将1.3～1.7mNaCl洗脱液部分(活性最高)合并、冷冻干燥。纯化样品在SDS—PAGE中出现一条与bFGF迁移率一致的主带,MW约为17.000。IEF显示与重组hFGF电泳图象一致。从男性胎儿前列腺分离、培养了人前列腺间质成纤维细胞,经5～7次传代后,移入96孔培养板中以无血清的Ham’sF12为介质进行培养,以~3HTdR掺入法分别观察DHT、E_2和DHT、E_3联合应用及hPGF、bFGF、EGF对细胞增殖反应。实验结果显示①本文所分离纯化的hPGF与bFGF、EGF在加入培养液后三天均呈现强的增殖刺激效应。②DHT也是显示为一种强的增殖刺激剂。但直至第六天才有明显作用,如每天更换培养液则无此效应。③E_2不论单独或与DHT联合应用均无明显增殖刺激效应。作者认为我们纯化的hPFG具有直接刺激效应,而DHT为延缓反应,属于间接作用,可能是通过产生或分泌生长因子而起作用,hPGF可能为其介导物。
Benign prostatic hyperplasia (BPH) is a common disease in middle-aged and elderly men. To explore the mechanism of action of sex hormones on prostatic cell proliferation, BPH prostate tissue was collected, segmented by ammonium sulfate, heparin agarose affinity chromatography, NaCl concentration gradient elution, 1.3 ~ 1.7mNaCl eluate fractions (highest activity) combined, freeze-dried. The purified sample showed a major band consistent with bFGF mobility in SDS-PAGE with a MW of about 17.000. IEF shows consistent with recombinant hFGF electrophoresis images. Human prostate prostatic cells were isolated and cultured human prostate stromal fibroblasts, 5 to 7 times after passage, transferred to 96-well plates in serum-free Ham’s F12 medium for culture, ~ 3HTdR incorporation were observed DHT, E_2 and DHT, E_3 combined with hPGF, bFGF, EGF on cell proliferation response. The experimental results showed that ① hPGF, bFGF and EGF isolated and purified in this study showed a strong proliferative and stimulatory effect for three days after adding the culture solution. DHT is also shown as a strong proliferative stimulant. But until the sixth day had a significant effect, such as daily replacement of culture fluid is no such effect. ③E_2 alone or combined with DHT had no significant proliferative stimulatory effect. The authors believe that our purified hPFG has a direct stimulatory effect, while DHT is a delayed response, which is an indirect effect, probably through the production or secretion of growth factor and hPGF may be its mediator.