军事医学
軍事醫學
군사의학
BULLETIN OF THE ACADEMY OF MILITARY MEDICAL SCIENCES
2014年
11期
845-849
,共5页
叶枫%王健%但国蓉%上官陶%赵吉清%赵远鹏%邹仲敏
葉楓%王健%但國蓉%上官陶%趙吉清%趙遠鵬%鄒仲敏
협풍%왕건%단국용%상관도%조길청%조원붕%추중민
半硫芥%角质形成细胞%miR-34 a%细胞迁移%ERK1/2
半硫芥%角質形成細胞%miR-34 a%細胞遷移%ERK1/2
반류개%각질형성세포%miR-34 a%세포천이%ERK1/2
CEES%HaCaT%miR-34a%cell migration%ERK1/2
目的：研究半硫芥（2-氯乙基乙基硫醚，2-chloroethyl ethyl sulfide ，CEES）染毒对角质形成细胞迁移的影响以及miR-34a作用机制。方法体外建立CEES染毒HaCaT细胞模型，采用化学合成miR-34a抑制物转染HaCaT细胞沉默miR-34a，荧光显微镜观察细胞转染效率，细胞划痕实验检测细胞迁移能力，Western 印迹检测细胞分化标志物K5和K10、ERK1／2总蛋白及磷酸化水平改变。结果采用0．5 mmol／L CEES染毒后HaCaT细胞迁移显著降低。细胞形态、干性标志物K5和分化标志物K10没有明显变化。 miR-34 a沉默可以增加细胞迁移能力，迁移相关的信号蛋白ERK1／2通路激活，ERK1／2信号通路抑制剂U0126使用后可以显著降低细胞迁移。结论 miR-34 a沉默可以显著增加角质形成细胞迁移，并部分逆转CEES的抑制作用，miR-34a沉默可能部分是通过ERK1／2信号通路激活引起染毒细胞迁移增加。
目的：研究半硫芥（2-氯乙基乙基硫醚，2-chloroethyl ethyl sulfide ，CEES）染毒對角質形成細胞遷移的影響以及miR-34a作用機製。方法體外建立CEES染毒HaCaT細胞模型，採用化學閤成miR-34a抑製物轉染HaCaT細胞沉默miR-34a，熒光顯微鏡觀察細胞轉染效率，細胞劃痕實驗檢測細胞遷移能力，Western 印跡檢測細胞分化標誌物K5和K10、ERK1／2總蛋白及燐痠化水平改變。結果採用0．5 mmol／L CEES染毒後HaCaT細胞遷移顯著降低。細胞形態、榦性標誌物K5和分化標誌物K10沒有明顯變化。 miR-34 a沉默可以增加細胞遷移能力，遷移相關的信號蛋白ERK1／2通路激活，ERK1／2信號通路抑製劑U0126使用後可以顯著降低細胞遷移。結論 miR-34 a沉默可以顯著增加角質形成細胞遷移，併部分逆轉CEES的抑製作用，miR-34a沉默可能部分是通過ERK1／2信號通路激活引起染毒細胞遷移增加。
목적：연구반류개（2-록을기을기류미，2-chloroethyl ethyl sulfide ，CEES）염독대각질형성세포천이적영향이급miR-34a작용궤제。방법체외건립CEES염독HaCaT세포모형，채용화학합성miR-34a억제물전염HaCaT세포침묵miR-34a，형광현미경관찰세포전염효솔，세포화흔실험검측세포천이능력，Western 인적검측세포분화표지물K5화K10、ERK1／2총단백급린산화수평개변。결과채용0．5 mmol／L CEES염독후HaCaT세포천이현저강저。세포형태、간성표지물K5화분화표지물K10몰유명현변화。 miR-34 a침묵가이증가세포천이능력，천이상관적신호단백ERK1／2통로격활，ERK1／2신호통로억제제U0126사용후가이현저강저세포천이。결론 miR-34 a침묵가이현저증가각질형성세포천이，병부분역전CEES적억제작용，miR-34a침묵가능부분시통과ERK1／2신호통로격활인기염독세포천이증가。
Objective To explore the effect of 2-chloroethyl ethyl sulfide(CEES) poisoning on keratinocyte migration and the regulatory role of microRNA(miR)-34a.Methods MTS was used to detect the viability of cells exposed to CEES in order to select an appropriate dose of CEES exposure in this in vitro model.The protein level of keratin 5 and keratin 10 was detected to assess cell differentiation status .Scratch assay was applied to evaluate cell migration ,and miR-34a silencing in keratinocytes was achieved by transfecting chemically synthesized miR-34a specific miRNA inhibitor.t-ERK1/2 and p-ERK1/2 levels closely related to cell migration were detected using Western blotting .Results An in vitro CEES exposure model of keratinocytes was established at the optimal concentration of 0.5 mmol/L CEES in the viability test , and this dose was chosen to evaluate cell migration changes .The migration of cells was significantly inhibited 24 h after CEES exposure , accompanied by no changes in morphology and keratin 5/10 levels.Silencing of miR-34a significantly increased the migration of cells exposed to CEES , which could be blocked by adding 5 μmol/L U0126 , an ERK1/2 phosphorylation selective inhibitor.Conclusion Silencing of miR-34a can significantly increase keratinocyte migration and partially reverse the inhibition of CEES-caused migration , which could be mediated by ERK 1/2 pathway activation .
