加热对大鼠表皮干细胞HSP70表达及细胞增殖的影响  

Effects of heat stress on expression of HSP70 and cell propagation of epidermal stem cells in rat

作　　者：王强[1] 邓先见[2] 徐苗苗[1] 程勇[1] 

机构地区：[1]南华大学附属郴州医院,湖南郴州423000 [2]第三军医大学创伤烧伤与复合伤国家重点实验室四室,重庆400042

出　　处：《湘南学院学报（医学版）》2014年第2期5-10,共6页Journal of Xiangnan University（Medical Sciences）

基　　金：第三军医大学创伤;烧伤与复合伤国家重点实验室开放基金资助项目(SKLKF201203)

摘　　要：目的：分离、培养并鉴定SD大鼠表皮干细胞；观察不同温度加热对表皮干细胞热耐受预测因子HSP70表达和细胞增殖的影响。方法体外分离培养大鼠表皮干细胞，用倒置显微镜观察其形态，以角质形成细胞作为对照；采用细胞免疫荧光技术对β1-整合素、干性标志物K19、α6及分化标志物K10进行鉴定；实验对37、41、43、45、47、49℃热处理的表皮干细胞采用MTT法测定细胞吸光度值，计算细胞存活率；Western blot法测定HSP70表达的变化。结果分离的细胞体积小，核质比大，集落生长，呈典型的铺路石样结构；实验组细胞干性标志物K19、β1-整合素及α6呈阳性表达，分化标志物K10表达阴性；随着温度升高，细胞HSP70表达增加，于47℃时表达量达到峰值，同时细胞随温度升高存活率逐渐降低，于45℃～49℃时降低幅度下降成一水平趋势。结论成功建立了SD大鼠表皮干细胞分离、培养方法，升高温度可抑制表皮干细胞增殖能力，细胞存活率下降，提示HSP70表达增加对表皮干细胞热损伤可能具有保护作用。Objecive To isolate and identify the epidermal stem cells from SD rats .Observe the expression of HSP70 and effects of proliferation of epidermal stem cells under different temperatures .Methods The morphology of epidermal stem cells were observed by an inverted microscope .Immunofluorescence technic was used to observe the expression ofβ1-integrin ,keratin19 （CK19 ） ,keratin10 （CK10 ） and α6 .Keratinocytes were served as con-trols .The epidermal stem cells were stimulated at 41℃ ,43℃ ,45℃ ,47℃ and 49℃ for 10 minutes in water bath , respectively .The optical density was tested by MTT ,survival rates were calculated .The fold changes of HSP70 ex-pression were detected by Western Blot test .In the control group ,epidermal stem cells were incubated at 37℃ for 10 minutes .Results Cells showed an epithelial-like appearance under inverted microscopy .Positive expression ofβ1-integrin ,CK19 ,α6 and low expression CK10 of cultured cells were detected by immunofluorescence . In-creased gradually along with a rise of water bath temperature ,the expression of HSP70 in the experimental group was increased ,the level of HSP70 in the experimental group was peaked on 47℃ .Survival rates of EpsC were re-spectively decreased with elevatory temperatures .Conclusions Skin Epidermal stem cells of rats can be success-fully isolated and cultured in vitro ,by heat injury ,the multiplication capacity of epidermal stem cells was inhibit-ed .The high expression of HSP70 may protect epidermal stem cells from heat injury .

关 键 词：表皮干细胞 HSP70 热损伤 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]