大鼠骨骼肌损伤修复中FZD4时序性变化及其应用

目的以卷曲受体4(FZD4)为例,探索细胞膜上受体蛋白的表达变化能否作为损伤时间推断的指标。方法 78只大鼠随机分为对照组和损伤后4h、8h、12h、16h、20h、24h、28h、32h、36h、40h、44h和48 h组(n=6)。麻醉后,采用重力锤自由落体方式砸伤大鼠右后肢,取股四头肌处挫伤的组织。应用免疫荧光、real-time PCR和western blotting方法检测肌肉中FZD4蛋白及m RNA的表达量。结果免疫荧光结果显示FZD4表达于骨骼肌细胞膜上,胞浆和细胞核内均未见表达;Real-time PCR结果表明FZD4 m RNA在损伤后8h、12h、36h和40h显著上调(P0.05),均超过对照组的两倍;Western blotting结果示FZD4蛋白的表达在8h、36h、40h、44h和48h组明显增加(P0.05),但均未超过对照组的两倍。结论 FZD4 m RNA和蛋白在肌肉挫伤后呈时序性变化规律可作为损伤时间推断的指标,在一定程度上FZD4 m RNA比FZD4更适用于损伤时间推断。

Time-dependent expression of FZD4 in contused skeletal muscle of rat used for wound age estimation

Objective Take frizzled class receptor 4(FZD4) as an example for exploring whether the transmembrane receptor protein is a suitable marker for wound age estimation. Methods A total of 78 Sprague-Dawley rats were divided randomly into the control group and the contusion groups of 4h, 8h, 12h, 16h, 20h, 24h, 28h, 32h, 36h, 40h, 44h, and 48h after injury (n=6). Using a drop-ball technique, a contusion was produced in the right lamb of rats. The samples were dissected from injury zones. Then the expression of FZD4 was detected using immunofluorescence, real-time PCR and western blot, respectively. Results FZD4 was located on the membrane of skeletal muscle cells. Compared to the control group, FZD4 mRNA showed a signiifcant increase (over 2-fold) in 8h, 12h, 36h, and 40h after injury by Real-time PCR (P<0.05), and FZD4 protein showed a statistical up-regulation (less than 2-fold) in 8h, 36h, 40h, 44h, and 48h post contusion by western blotting (P<0.05). Conclusion The expression of FZD4 mRNA and protein are both time-dependent during contused skeletal muscle repair. To some degree, FZD4 mRNA was more suitable than corresponding protein for determining wound age.