基于线粒体氧化应激通路探讨牡蛎肽减轻大鼠神经性勃起功能障碍的作用

Study on the effect of oyster peptide on neurogenic erectile dysfunction in rats based on mitochondrial oxidative stress pathway

目的基于线粒体氧化应激通路探讨牡蛎肽减轻大鼠神经性勃起功能障碍(NED)的作用。方法50只SD雄性大鼠建立NED模型并按照随机数字表分为模型组,西地那非组,牡蛎肽低、中、高剂量组,另取10只SD雄性大鼠记为假手术组。西地那非组予以5 mg/kg西地那非溶于1 mL/100 g生理盐水中灌胃,牡蛎肽低、中、高剂量组予以75、150、300 mg/kg牡蛎肽溶于等量生理盐水中灌胃,模型组和假手术组均仅予以等量生理盐水灌胃,每天1次,共4周。生物机能实验系统测定各组平均颈动脉压(MAP)、阴茎海绵体内压(ICP),并计算ICP/MAP;酶联免疫吸附测定(ELISA)检测各组阴茎海绵体组织氧化应激指标及血管内皮功能指标活力;苏木素-伊红(H-E)染色观察大鼠阴茎海绵体组织病理改变;透射电镜观察大鼠阴茎海绵体细胞超微结构;实时-反转录定量聚合酶链反应(RT-qPCR)检测大鼠阴茎海绵体组织核因子类红细胞-相关因子2(Nrf2)、血红素氧合酶-1(HO-1)信使核糖核酸(mRNA)表达;蛋白质免疫印迹法(WB)检测大鼠阴茎海绵体组织Nrf2、HO-1蛋白表达。结果模型组大鼠阴茎海绵体组织平滑肌细胞数量减少,内皮细胞严重损伤,微小血管管壁增厚,成纤维样,电镜下可见线粒体高度肿胀,嵴消失或呈空泡化,西地那非组和牡蛎肽3剂量组阴茎海绵体组织损伤均有不同程度改善;与假手术组比,模型组、西地那非组和牡蛎肽3剂量组ICP和ICP/MAP,阴茎海绵体组织NO、SOD含量,GSH-PX、eNOS活性及Nrf2、HO-1 mRNA及蛋白表达降低,MDA含量升高(P<0.05);与模型组比,西地那非组和牡蛎肽3剂量组ICP和ICP/MAP,阴茎海绵体组织NO、SOD含量、GSH-PX、eNOS活性及Nrf2、HO-1 mRNA及蛋白表达均升高,MDA含量降低(P<0.05),其中牡蛎肽作用呈剂量依赖性。结论牡蛎肽对NED大鼠勃起功能有显著改善作用,推测是基于线粒体氧化应激通路抑制Nrf2、HO-1的表达实现此作用的。

Objective To investigate the effect of oyster peptide on neurogenic erectile dysfunction(NED)in rats based on mitochondrial oxidative stress pathway.Methods 50 SD male rats were established NED model and randomly divided into model group,sildenafil group,low,medium and high dose oyster peptide groups.Another 10 SD male rats were recorded as sham operation group.Sildenafil group was given 5 mg/kg sildenafil dissolved in 1 mL/100 g normal saline,oyster peptide low,medium and high dose groups were given 75,150 and 300 mg/kg oyster peptide dissolved in the same amount of normal saline,model group and sham operation group were only given the same amount of normal saline once a day for 4weeks.Mean carotid pressure(map)and intracavernosal pressure(ICP)were measured by biological function experimental system,and ICP/MAP was calculated.Enzyme linked immunosorbent assay(ELISA)was used to detect the activities of oxidative stress indexes and vascular endothelial function indexes.Hematoxylin eosin(H-E)staining was used to observe the histopathological changes of rat corpus cavernosum.The ultrastructure of rat penile cavernous cells was observed by transmission electron microscope.Real time reverse transcription quantitative polymerase chain reaction(RT-qPCR)was used to detect the expression of nuclear factor erythroid related factor 2(Nrf2)and heme oxygenase-1(HO-1)messenger RNA(mRNA)in rat penile cavernous tissues.The protein expressions of Nrf2 and HO-1 in rat penile cavernous tissues were detected by Western blot(WB).Results In the model group,the number of smooth muscle cells in penile cavernous tissues decreased,endothelial cells were seriously damaged,microvascular wall thickened and fibroblast like.Under electron microscope,mitochondria were highly swollen,cristae disappeared or vacuolized.The damage of penile cavernous tissues in sildenafil group and oyster peptide 3 dose group were improved in varying degrees.Compared with the sham operation group,ICP and ICP/MAP,NO,SOD content,GSH-PX,eNOS activity,Nrf2,HO-1 mRNA and protein expression in penile cavernous tissu decreased and MDA content increased in the model group,sildenafil group and oyster peptide 3-dose group(P<0.05).Compared with the model group,ICP and ICP/MAP,NO,SOD content,GSH-PX,eNOS activity,Nrf2,HO-1 mRNA and protein expression in penile cavernous tissue were increased and MDA content was decreased in sildenafil group and oyster peptide 3dose group(P<0.05).The effect of oyster peptide was dose-dependent.Conclusion Oyster peptide can significantly improve the erectile function of NED rats.It is speculated that this effect is based on the inhibition of Nrf2 and HO-1 expression by mitochondrial oxidative stress pathway.