玻璃体腔注射HGF-MSCs对糖尿病大鼠视网膜HGF表达的影响

doi:10.6040/j.issn.1673-3770.0.2021.121

摘要/Abstract

摘要： 目的观察玻璃体腔注射肝细胞生长因子基因修饰的人脐带间充质干细胞(HGF-MSCs)对糖尿病大鼠视网膜组织肝细胞生长因子(HGF)表达的影响,探讨HGF-MSCs对大鼠Ⅱ型糖尿病性视网膜病变的作用。方法体外分离、培养人脐带间充质干细胞(hUCMSCs)并鉴定。采用感染复数值(MOI)50的慢病毒载体转染hUCMSCs。4周龄SD雄性大鼠,按照随机数字表法分为正常组(8只),糖尿病组(60只),糖尿病组又分为糖尿病对照组、PBS 组、MSCs组和 HGF-MSCs组(每组15只)。糖尿病组大鼠采用腹腔注射链脲佐菌素(STZ)建立糖尿病大鼠模型。建模3个月后对糖尿病组进行后续实验。糖尿病对照组不行特殊治疗,PBS 组玻璃体腔注射磷酸盐缓冲液,MSCs组玻璃体腔注射hUCMSCs,HGF-MSCs组玻璃体腔注射HGF-MSCs。干预2、4、6周后行荧光定量PCR检测大鼠视网膜组织HGF mRNA的表达;Western blot检测大鼠视网膜组织HGF的表达。结果培养的脐带间充质干细胞可以诱导分化为成骨细胞、成脂肪细胞。MOI 50 的慢病毒成功转染hUCMSCs。大鼠视网膜荧光定量PCR和Western blot结果显示:在同一时间点,与其他组相比,正常组大鼠视网膜的HGF及其 mRNA的表达明显降低,差异具有统计学意义(P<0.05)。在同一时间点,与其他组相比,HGF-MSCs组大鼠视网膜的HGF 及其mRNA的表达明显增加,差异具有统计学意义(P<0.05)。不同时间点各组大鼠视网膜HGF 及其mRNA的表达差异没有统计学意义(P>0.05)。结论糖尿病大鼠视网膜HGF 及其mRNA的表达增加。单纯玻璃体腔注射hUCMSCs,不影响糖尿病大鼠视网膜HGF 及其mRNA的表达。玻璃体腔注射HGF修饰的hUCMSCs,可以促进糖尿病大鼠视网膜HGF 及其mRNA的表达,可能与HGF相互作用减轻糖尿病视网膜病变的早期病变发展。

关键词: 糖尿病视网膜病变, 玻璃体腔注射, HGF, 慢病毒转染, hUCMSCs

Abstract: Objective To observe the effect of intravitreal injection of human umbilical cord mesenchymal stem cells(hUCMSCs)modified by the hepatocyte growth factor gene(HGF-MSCs)on the hepatocyte growth factor(HGF)expression in retina tissue of diabetic rats, and to explore the effect of HGF-MSCs on diabetic retinopathy in rats. Methods hUCMSCs were isolated, cultured, and identified. hUCMSCs were transfected with lentiviral vectors with a multiplicity of infection(MOI)of 50. Four-week-old Sprague-Dawley rats were randomly divided into the normal(15 rats)and diabetes(60 rats)groups. The diabetic rat models were established by intraperitoneal injection of streptozotocin(STZ). Three months after the diabetic rat models were confirmed successfully, the diabetes group was further divided into the diabetes control(no injection), PBS(intravitreal injection of PBS), MSCs(intravitreal injection of MSCs), and HGF-MSCs(intravitreal injection of HGF-MSCs)groups. After 2, 4, and 6 weeks of intravitreal injection, the HGF mRNA expression in rat retina was detected through real-time PCR. The expression of HGF in the rat retina was assessed through Western blot. Results The cultured hUCMSCs can be differentiated into osteoblasts and adipocytes. MOI 50 lentivirus successfully transfected hUCMSCs. Two weeks after intravitreal injection, quantitative real-time PCR and Western blot showed that, at the same time point, the expression of HGF and HGF mRNA in the normal group decreased compared to that of the other groups, and the difference was statistically significant(P<0.05). Compared with other groups at the same time point, the expression of HGF and HGF mRNA in HGF-MSCs group increased, and the difference was statistically significant(P<0.05). No significant difference in the expression of HGF and HGF mRNA was observed in the same group at different time points(P>0.05). Conclusion The HGF and HGF mRNA expression increased in diabetic rat retina. Intravitreal injection of hUCMSCs did not affect the HGF and HGF mRNA expression in diabetic rat retina. Intravitreal injection of hUCMSCs modified by HGF increased the HGF and HGF mRNA expression in diabetic rats. HGF-MSCs may interact with HGF to alleviate early diabetic retinopathy.

Key words: Diabetic retinopathy, Intravitreal injection, HGF, Transfection, hUCMSCs

中图分类号:

R774.1

王惠, 王俊,孙乙,于腾飞,朱玉广, 朱艳. 玻璃体腔注射HGF-MSCs对糖尿病大鼠视网膜HGF表达的影响[J]. 山东大学耳鼻喉眼学报, 2022, 36(2): 72-77.

WANG Hui, WANG Jun, SUN Yi, YU Tengfei, ZHU Yuguang, ZHU Yan. Effect of intravitreal injection of HGF-MSCs on the expression of HGF in retina tissue of diabetic rats[J]. Journal of Otolaryngology and Ophthalmology of Shandong University, 2022, 36(2): 72-77.

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