选择性瞬变感受器电位蛋白V4激动剂GSK1016790A对轴突再生的影响  

Effect of transient receptor potential vanilloid 4 agonist GSK1016790A on axonal regeneration

作　　者：秦绪祯 马进进 李梅梅 王星然 谢计乐 赛吉拉夫 Qin Xuzhen;Ma Jinjin;Li Meimei;Wang Xingran;Xie Jile;Saijilafu(First Affiliated Hospital of Soochow University,Suzhou 215000,Jiangsu Province,China;Institute of Orthopedics,Soochow University,Suzhou 215000,Jiangsu Province,China)

机构地区：[1]苏州大学附属第一医院,江苏省苏州市215000 [2]苏州大学骨科研究所,江苏省苏州市215000

出　　处：《中国组织工程研究》2022年第24期3903-3907,共5页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金(81571189,81772353),项目负责人:赛吉拉夫;国家重点研究发展计划(2016YFC1100203),项目负责人:赛吉拉夫;江苏省创新创业计划,项目负责人:赛吉拉夫。

摘　　要：背景:选择性瞬变感受器电位蛋白V4(transient receptor potential vanilloid 4,TRPV4)激动剂GSK1016790A能够在短时间内激活TRPV4蛋白,表现为Ca^(2+)内流,而蛋白表达量不变。但是在培养过程中,GSK1016790A会使HeLa细胞膜表面TRPV4表达降低,这一现象对神经再生是否产生影响仍不清楚。目的:探究TRPV4激动剂GSK1016790A在不同时期内对神经元轴突再生的影响。方法:取6-8周龄ICR小鼠背根神经节,经过胶原酶和胰酶处理使细胞充分解离,进行背根神经节细胞培养。在培养初期加入GSK1016790A刺激2 h后去除激动剂继续培养3 d,或者加入GSK1016790A持续处理3 d;空白对照组不进行任何处理。采用免疫印迹法检测TRPV4蛋白表达以及轴突再生相关蛋白表达,同时进行TUJ1和TRPV4免疫荧光染色,观察轴突分叉数目、轴突再生长度、细胞存活数变化。结果与结论:①与空白对照组相比,GSK1016790A刺激神经细胞2 h后,TRPV4蛋白变化无明显差异(P>0.05);而刺激神经细胞3 d后TRPV4表达明显降低(P<0.05);②与空白对照组相比,GSK1016790A刺激神经细胞2 h后,轴突分叉数目及轴突长度无明显差异(P>0.05);而刺激神经细胞3 d后轴突分叉数目及轴突长度均明显增加(P<0.05);③与空白对照组相比,GSK1016790A刺激神经细胞2 h或3 d后,细胞存活率均无明显改变;④与空白对照组相比,GSK1016790A刺激神经细胞3 d后,PTEN蛋白表达降低(P<0.05),神经轴突再生显著,这一过程可能经抑制PTEN蛋白表达介导的。BACKGROUND:The transient receptor potential vanilloid 4(TRPV4)agonist GSK1016790A can activate TRPV4 protein and induce Ca^(2+) influx in a short time,while its expression remains unchanged.However,during culture,the decreased expression of TRPV4 on HeLa cell membrane surface was induced by GSK1016790A.Whether the phenomenon had an effect on neural regeneration is still not clear.OBJECTIVE:To explore the effect of TRPV4 agonist GSK1016790A on axonal regeneration in different periods.METHODS:The dorsal root ganglion cells of ICR mouse at 6-8 weeks were treated by collagenase and trypsin for cell culture.In the 2-hour group,the agonist GSK1016790A was added at the initial stage of culture and the cells were treated for 2 hours and the culture was continued for 3 days.The 3-day group was treated with the agonist GSK1016790A for 3 days.The blank control group did not undergo any treatment.The TRPV4 protein expression and axon regeneration related protein expression were examined by western blot assay.TUJ1 and TRPV4 immunofluorescence staining was performed simultaneously.Number of axon branches,length of axon regeneration,and number of cell survival were counted.RESULTS AND CONCLUSION:(1)Compared with the blank control group,there was no significant difference in TRPV4 protein changes after GSK101 stimulated nerve cells for 2 hours(P>0.05);while the expression of TRPV4 decreased significantly after nerve cells were stimulated for 3 days(P<0.05).(2)Compared with the blank control group,after GSK1016790A stimulated nerve cells for 2 hours,there was no significant difference in the number of axon branches and axon length(P>0.05);after stimulating nerve cells for 3 days,the number of axon bifurcations and axon length were increased significantly(P<0.05).(3)Compared with the blank control group,there was no significant change in cell survival rate after GSK1016790A stimulated nerve cells for 2 hours or 3 days.(4)Compared with the blank control group,after GSK1016790A stimulated nerve cells for 3 days,the expression of PTE

关 键 词：选择性瞬变感受器电位蛋白V4 GSK1016790A 背根神经节 神经再生 轴突 PTEN蛋白 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学]