潘志国 邵玉 万军 耿焱 陈镜合 苏磊

DOI：10.3760/cma.j.issn.1671-0282.2014.08.008

基金項目：全军医学科学技术研究“十二五”发展计划重点项目（BWS12J018）；国家自然科学基金（81071529，81101467，81101406）

作者单位：510010 广州，广州军区广州总医院重症医学科 全军热区创伤救治与组织修复重点实验室

通信作者：苏磊，Email： slei_icu@163.com

【摘要】目的 研究热打击对培养人骨骼肌细胞（HSKMC）通透性、细胞骨架及细胞周期影响。

访法 流式细胞仪钙离子内流检测热打击对HSKMC细胞膜通透性的影响，考马斯亮蓝R-250染色法检测热打击对HSKMC细胞骨架影响，流式细胞仪检测热打击对HSKMC细胞周期改变。结果 给予培养人HSKMC细胞不同温度梯度热打击1 h后， 43 ℃热打击组钙离子流式检测的中位数为91.63，37 ℃对照组中位数为22.98。同对照组相比，随着热打击程度加强，显微镜高倍镜下可见骨骼肌细胞骨架逐渐出现变粗、变短、出现明显的应力纤维。骨骼肌细胞在热打击情况下，各组G0/G1期DNA含量在44.13～62.98之间，与正常对照组对比，当细胞离开热打击环境后培养18 h前G0/G1期DNA含量明显高于对照组，培养第18 小时恢复才同对照组基本相同。

结论 热打击的情况下造成细胞钙离子内流效应，导致细胞内钙超载。可改变HSKMC骨架结构，使骨架失去正常的网状有序排列结构，间隙增大。细胞周期出现阻滞，细胞被阻滞在G0/G1期。

【关键词】热打击；骨骼肌细胞；通透性；钙超载；细胞骨架；细胞周期；考马斯亮蓝染色；流式检测

The effect of heat stress on the permeability， cytoskeleton and cell cycle of human skeletal muscle cellPan Zhiguo， Shao Yu， Wan Jun， Geng Yan， Chen Jinghe， Su Lei.ICU Department of Guangzhou General Hospital of Guangzhou Military Command， Guangzhou 510010， China

Corresponding author：Su Lei， Email： slei_icu@163.com

【Abstract】Objective To study the effect of heat stress on the permeability， cytoskeleton and cell cycle of human skeletal muscle cell （HSKMC）. Methods The HSKMC membrane permeability was detected by calcium ion inflow with flow cytometer， the cytoskeleton was stained by CBB 250， and the cell cycle was determined by flow cytometer.Results After 1 h of heat stress on human HSKMC cells under different temperature gradient， the median level of calcium ion was 91.63 in 43 ℃ heat stress group compared with 22.98 in 37 ℃control group. As temperature increased， thicker and shorter cytoskeleton and stress fiber were shown under the high power lens of microscope. The DNA expression of skeleton cells at G0/G1 stage was 44.13-62.98 in groups under heat stress. Compared with normal control group， DNA expression was much higher in heat stress group， when HSKMC was cultured under 37℃ temperature for another 18 h， it kept decreasing DNA expression to a similar level as control group.Conclusions Heat stress can cause calcium iron inflow resulting in intracellular calcium overload， and affect the cytoskeleton leading to loss of normal web ordered arrangement and increased gap in HSKMC cells， which give rise to blocking cell cycle into G0/G1 stage.

【Key words】Heat stress；Skeletal muscle cells；Permeability；Calcium overload；Cytoskeleton；Cell cycles；Coomassie brilliant blue； Coomassie brilliant blue；Flow cytometry

中暑的特征为中心体温过高伴随着系统炎症反应致多脏器功能障碍，患者中心体温可超过40 ℃，导致机体主要脏器如心、肝、肺、肾、肠、血管和骨骼肌等发生一系列损伤^[1-7]。……