pAdxsi-ERβ腺病毒转染对ERβKO小鼠阴茎海绵体血管内皮的影响

Protective effect of pAdxsi-ERβ adenovirus transfection on penile vascular endothelium in ERβKO mice

目的:探讨ERβ基因过表达对ERβKO小鼠阴茎血管内皮的影响及相关分子机制。方法:选取ERβKO雄性小鼠12只,随机分为两组:ERβKO+TNFα+pAdxsi-ERβ组和ERβKO+TNFα+空病毒组。ERβKO+TNFα+pAdxsi-ERβ组给予ERβ基因重组腺病毒转染处理14 d,ERβKO+TNFα+空病毒组转染空病毒作对照,同时两组均给予TNFα6μg/(kg·d)腹腔注射,连续14 d。采用APO法观察小鼠的阴茎勃起功能;免疫组化检测内皮标志物CD34、vWF的变化情况;RT-PCR、Western印迹和免疫组化检测eNOS-NO通路相关分子表达情况。结果:与ERβKO+TNFα+空病毒组相比,ERβKO+TNFα+pAdxsi-ERβ组小鼠的变化情况如下:①勃起次数增多(2.17±0.41 vs 0.50±0.55,P<0.05),勃起潜伏期缩短[(24.0±1.27)min vs(28.83±1.33)min,P<0.05];②内皮标志物CD34、vWF表达更丰富[(1.50±0.55;1.33±0.52)vs(0.67±0.52;0.50±0.55),P<0.05];③eNOS、Cam表达减少,小窝蛋白-1表达上调(P均<0.05),RT-PCR与Western印迹结果相符合。结论:ERβ基因对阴茎血管内皮具有保护作用,eNOS-NO通路是其发挥作用的机制之一。

Objective: To investigate the effect of the overexpression of the ERβ gene on the penile vascular endothelium of ERβ knockout( ERβKO) mice and its molecular mechanisms. Methods: We randomly divided 12 ERβKO male mice into groups A( ERβKO + TNFα + pAdxsi-ERβ) and B( ERβKO + TNFα + empty virus),the former treated by pAdxsi-ERβ adenovirus transfection,the latter with empty virus,and meanwhile both injected intraperitoneally with TNFα at 6 μg per kg body weight per d for 14 days. Then we observed the erectile function of the mice by APO,determined the changes of the endothelial markers CD34 and vWF by immunohistochemical staining,and detected the expressions of the relevant molecules in the eNOS-NO pathway by RT-PCR,Western blot and immunohistochemistry. Results: Compared with group B,group A showed a significantly increased number of penile erections( 0. 50 ± 0. 55 vs 2. 17 ± 0. 41,P < 0. 05),shortened erectile latency( [28. 83 ± 1. 33]min vs [24. 00 ± 1. 27]min,P < 0. 05),enriched CD34 and vWF markers( 0. 67 ± 0. 52 vs 1. 50 ± 0. 55 and 0. 50 ± 0. 55 vs 1. 33 ± 0. 52,both P < 0. 05), elevated expressions of eNOS and Cam( RT-PCR: 1. 38 ± 0. 03 vs 1. 62 ± 0. 05 and 1. 02 ± 0. 09 vs 1. 42 ± 0. 05,both P < 0. 05; Western blot: 1. 27 ± 0. 04 vs 1. 55 ± 0. 07 and 0. 76 ± 0. 05 vs 0. 95 ± 0. 08,both P < 0. 05),and reduced expression of caveolin-1( RT-PCR: 2. 13 ± 0. 13 vs 1. 72 ± 0. 08,P < 0. 05; Western blot: 3. 99 ± 0. 16 vs 3. 40 ± 0. 14,P < 0. 05). The results of RT-PCR were consistent with those of Western blot. Conclusion: The ERβ gene protects the penile vascular endothelium via the eNOS-NO pathway.