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目的探索在海绵体神经损伤的大鼠神经性阴茎勃起功能障碍(NED)模型中,注射BDNF过表达的MSCs在神经再生和修复中的作用。方法建立脑源性神经营养因子(BDNF)过表达的骨髓间充质干细胞(MSCs),通过Western Blot检测BDNF和VEGF-A的表达的水平。通过Elisa检测BDNF和VEGF-A在细胞上清液中的含量。取120只3月龄大鼠随机分为4组:第1组为假手术组(Sham组),其余三组均损伤双侧海绵体神经,第2组(Control组)不再进行干预,第3组(MSC组)和第4组(BDNF-MSC)分别在关腹前注射MSCs和BDNF过表达的MSCs。大鼠在手术造模后12周,通过海绵体神经电刺激并测阴茎海绵体内压(ICP)来评价阴茎勃起功能。取大鼠阴茎海绵体组织行免疫组化评价诱导型NO合酶(iNOS)的表达水平。结果通过绿色免疫荧光试验证实BDNF过表达MSCs制备成功。Western Blot验证结果显示BDNF过表达的MSC细胞中的BDNF和VEGF-A表达水平较MSC细胞明显上调,ELISA验证结果显示BDNF过表达的MSC细胞上清液中的BDNF和VEGF-A表达水平较MSC细胞上清液明显上调。手术后12周,通过海绵体神经电刺激并测ICP来评价阴茎勃起功能,其中假手术组平均最大ICP为(102.20±15.61)cmH_2O,实验对照组为(35.47±13.16),MSC干预组为(43.79±16.94)cmH_2O,BDNF-MSC干预组为(66.04±1 5.33)cmH_2O。免疫组化结果显示,BDNF-MSC干预组大鼠阴茎海绵体组织内iNOS阳性表达细胞数明显高于MSC干预组。结论本研究结果显示局部注射MSCs和BDNF过表达的MSCs对于大鼠的NED具有治疗作用。
Objective To explore the role of MSCs injected with BDNF in the regeneration and repair of nerve in rats with neurogenic erectile dysfunction (NED) induced by spontaneous nerve injury. Methods Bone marrow mesenchymal stem cells (MSCs) overexpressing brain derived neurotrophic factor (BDNF) were established. The expression of BDNF and VEGF-A were detected by Western Blot. The content of BDNF and VEGF-A in the cell supernatant was measured by Elisa. One hundred and twenty three-month-old rats were randomly divided into four groups: the sham operation group (Sham group), the other three groups were injured bilateral cavernous nerve, the second group (Control group) no longer intervene, the first Group 3 (MSC group) and group 4 (BDNF-MSC) were injected with MSCs and BDNF-overexpressing MSCs respectively before transabdominal injection. Twelve weeks after operation, rat erectile function was evaluated by electrical stimulation of the cavernous nerve and intracavernosal pressure (ICP). The expression of inducible nitric oxide synthase (iNOS) was detected by immunohistochemistry in the corpus cavernosum of rats. Results The green fluorescent immunofluorescence assay confirmed the successful preparation of BDNF-overexpressing MSCs. The results of Western Blot showed that BDNF and VEGF-A expression levels in BDNF over-expressed MSC cells were significantly upregulated compared with MSC cells. The results of ELISA showed that BDNF and VEGF-A expression in BDNF-overexpressing MSCs supernatant was higher than that in MSC Cell supernatants were significantly up-regulated. The penile erectile function was assessed by spontaneous nerve stimulation and ICP measurement at 12 weeks after operation. The mean maximal ICP was (102.20 ± 15.61) cmH_2O in sham-operated group, (35.47 ± 13.16) in control group and 43.79 ± 16.94) cmH_2O, and the group of BDNF-MSC intervention was (66.04 ± 1 5.33) cmH_2O. The results of immunohistochemistry showed that the number of iNOS positive cells in the corpus cavernosum of BDNF-MSC intervention group was significantly higher than that in the MSC intervention group. Conclusion The results of this study show that MSCs injected with MSCs and BDNF overexpressing MSCs have a therapeutic effect on NED in rats.