低强度次声对体外培养的骨样细胞骨架蛋白F-actin表达的动态影响

关键词：次声 小鼠成骨样细胞 聚合态肌动蛋白 激光扫描共聚焦显微镜

Keywords:infrasound  osteoblast MC3T3  F-actin  laser scanning confocal microscope

摘要：

      目的：探讨4Hz／100dB、12Hz／100dB、20Hz／100dB的次声作用后，小鼠成骨样细胞MC3T3的细胞骨架F—actin表达的改变。方法：将MC3T3接种于细胞玻片上，并分为对照组和4Hz／100dB、12Hz／100dB、20Hz／100dB的次声暴露组。对照组无次声输出，其他各组接受次声作用30min／d。第3d于暴露后2h、4h、8h不同时间，对细胞进行F—actin的免疫荧光染色．应用激光扫描共聚焦显微镜．观察细胞F—actin的表达改变，测定单个细胞F—actin的平均荧光强度。结果：对照组细胞大部分荧光物质呈弥漫状态，胞膜荧光较强，胞浆内少量肌动蛋白纤维丝，方向不规则，长短不一；不同频率次声作用后2h．各次声作用组均可看到胞浆中微丝F—actin明显粗大纤长，荧光物质大多为较长的粗大应力丝，沿细胞纵轴排列较多，其中20Hz，100dB组的荧光强度增强较对照组具有显著意义（P〈0．05）；在次声作用后4h和8h．次声作用组的细胞F—actin仍处于较高表达状态．其中12Hz，100dB组和20Hz，100dB组的荧光强度增高较对照组具有显著意义（P〈0．05）。不同频率次声作用组细胞的F—actin变化趋势较一致，各组在各时间点未见明显差异（P〉0．05）。结论：4HZ、12Hz和20Hz的100dB的次声作用30min／d后，可诱导F—actin表达的增强，这种改变在8h后仍未见减弱。

Abstract:

      Objective:To study the alterations in expression of F-actin of osteoblast MC3T3 exposed by low intensity infrasound. Method: After the primary MC3T3 were cultured on coverglass,the cells were divided into four groups. Cells in experimental groups were exposed by infrasound with output intensity of 100dB and frequency of 4Hz,12Hz and 20Hz individually for 30min/d. The cells in control group were managed in the environment as experimental groups but without infrasound output. Laser scanning confocal microscope was used to examine the changes of F-actin after immunofluorescent staining and the photos were taken for further analysis of the cells` average fluorescence by spectrofluorometric quantification. Result:In control group,most fluorescein-labelled substance was in diffusion states. Less actin filaments were tenuous and short without direction. Correspondingly F-actin in the cells of experimental groups was thick and long and showed in longitudinal arrangement at the 2nd hour after exposure. The intensity of fluorescence was significantly increased in 20Hz/100dB group(P<0.05). More F-actin expression in experimental groups were being kept obviously increasing in 12Hz/100dB group and 20Hz/100dB group than the control at the 4th hour and 8th hours after infrasound exposure(P<0.05). However, there was no significant difference in F-actin expression between the experimental groups(P>0.05). Anyway cells in experimental groups showed synchronous changes. Conclusion:F-actin expression can be increased in osteoblast MC3T3 exposed 30min/d by low intensity infrasound with output intensity of 100dB and frequency of 4Hz,12Hz,20Hz. The expression increasing of F-actin still can be founded at the 8th hour after exposed under the low intensity infrasound.