组织工程与重建外科杂志

组织工程与重建外科杂志 ›› 2024, Vol. 20 ›› Issue (4): 395-.

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人脐带间充质干细胞恢复糖尿病勃起功能障碍大鼠海绵体神经功能的实验研究

  

  • 出版日期:2024-08-01 发布日期:2024-09-11

Experimental study of human umbilical cord mesenchymal stem cells on recovery of cavernosum nerve function in a rat model of diabetic-induced erectile dysfunction

  • Online:2024-08-01 Published:2024-09-11

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摘要：

目的　探索人脐带间充质干细胞通过再生修复海绵体神经对糖尿病大鼠勃起功能的改善作用。方法　收集 健康足月产新生儿脐带，提取分离人脐带间充质干细胞（hUCMSC）并鉴定表面标志物。通过高脂饮食联合腹腔注射链 脲佐菌素（STZ）的方法诱导大鼠糖尿病，10周后选取高血糖大鼠进行阿扑吗啡（APO）试验，筛查、建立糖尿病引起的勃 起功能障碍（DIED）大鼠模型。hUCMSC移植8周后，通过APO实验及最大海绵体内压/平均动脉压（ICP/MAP）评估大鼠 勃起功能；通过免疫组化法分析大鼠海绵体组织中神经元型一氧化氮合酶（nNOS）水平；通过ELISA法检测各组大鼠阴 茎组织中BDNF、NGF、VEGF和IGF-1神经营养生长因子的表达水平。结果　第4代hUCMSC表面高表达CD73、CD90、CD105，低表达 CD11b、CD19、CD34、CD45、HLA-DR。糖尿病引起的勃起功能障碍大鼠模型制备成功。海绵体内移植hUCMSC 8周后，APO实验表明hUCMSC可增加正常勃起的大鼠数量；中剂量和高剂量hUCMSC治疗后，相对于阴性对 照组，ICP/MAP值升高，阴茎内血流灌注明显改善；免疫组化实验显示中剂量和高剂量hUCMSC治疗后，大鼠阴茎海绵 体中 nNOS水平明显增加，启动大鼠阴茎勃起；ELISA检测结果证实中剂量和高剂量 hUCMSC治疗后，均能不同程度恢 复DIED 大鼠阴茎组织中 BDNF、NGF、VEGF和 IGF-1表达水平。结论　hUCMSC是一种多功能的成体干细胞，经海绵 体移植治疗高脂饮食联合STZ诱导的DIED大鼠是有效的，且有剂量依赖性。hUCMSC治疗DIED的机制与上调BDNF、NGF、VEGF和IGF-1表达水平提供的神经保护和再生修复作用有关。

关键词: 人脐带间充质干细胞, 　糖尿病, 　勃起功能障碍, 　神经元型一氧化氮合酶

Abstract: Objective　 To explore the restoration effects of human umbilical cord mesenchymal stem cells on erectile function in rats by repairing and regenerating cavernous nerve. Methods　Umbilical cords of healthy full-term newborns were collected， human umbilical cord mesenchymal stem cells（ hUCMSC） were isolated and cell surface markers were identified. The rat model of diabetes-induced erection dysfunction （DIED） was established by intra-abdominal streptozotocin （STZ） injection in combination with high-fat diet， and an apomorphine（ APO） test was performed to screen diabetic rat model with erectile dysfunction after 10 weeks. Eight weeks after the transplantation of hUCMSC， erectile function was evaluated by APO test and maximum intracavernous pressure/mean arterial pressure （ICP/MAP） ratio. The levels of neuronal nitric oxide synthase （nNOS） in corpus cavernosum tissues of rats were analyzed by immunohistochemistry analysis. In addition， the expression levels of neurotrophic growth factors including BDNF， NGF， VEGF， and IGF-1 in the corpus cavernosum tissues of SD rats were detected by ELISA. Results　 The hUCMSC was successfully isolated from human umbilical cords， and results of flow cytometry showed that CD73， CD90 and CD105 were highly expressed while CD11b， CD19， CD34， CD45 and HLA-DR were low expressed on the surface of P4 hUCMSC. The rat model of diabetic-induced erectile dysfunction was successfully established. After 8 weeks of hUCMSC therapy， the results of APO test showed that hUCMSC enhanced the erectile function of DIED rats， as evidenced by an increase in the number of complete erection. Compared with the negative group， the blood flow in the penis was improved as indicated by obvious increases in ICP/MAP ratio values in medium-dose and high-dose groups of hUCMSC treatment. Immunohistochemistry results suggested that after the treatment of hUCMSC in medium and high doses， hUCMSC treatment significantly elevated the expression level of nNOS in the corpus cavernosum tissue of DIED rats， then induced penile erection in rats. Furthermore， ELISA test results indicated that the expression levels of BDNF， NGF， VEGF and IGF-1 could be restored in different degrees after treatment with medium- and high-dose of hUCMSC. Conclusion　hUCMSC is a kind of adult pluripotent stem cells， which is effective and dose-dependent in the treatment of DIED rats induced by high fat diet combined with STZ following intracavernosal transplantation. The mechanism of hUCMSC treatment is related to neuroprotective and regenerative repair effects provided by upregulation of the expression levels of BDNF， NGF， VEGF and IGF-1.

Key words: Human umbilical cord mesenchymal stem cells, 　 Diabetes mellitus, 　 Erectile dysfunction, 　 Neuronal nitric oxide synthase