摘要
目的 探讨中波紫外线照射后不同时间点体外培养的HaCaT细胞MAPK信号通路受调控效应.方法 1.5、4.5、7.5、10、20、30、50 mJ/cm2中波紫外线照射HaCaT细胞后8h,对照组细胞除不进行UVB照射外,其他处理同各剂量UVB组,蛋白免疫印迹法测定ERK1/2、JNK和p38蛋白表达及磷酸化水平;50 mJ/cm2中波紫外线照射HaCaT细胞,蛋白免疫印迹法测定照射后2、4、8、12h4个时间点,细胞ERK1/2、JNK和p38蛋白表达及磷酸化水平.使用Quantity One软件计算目的条带吸光度,以相应蛋白条带的吸光度值与GAPDH蛋白内参条带吸光度值的比值表示相对蛋白含量.结果 7.5、10、20、30、50 mJ/cm2 UVB照射HaCaT细胞后8h,ERK1/2、JNK磷酸化水平明显上调（P＜0.05）,20、30、50 mJ/cm2 UVB照射HaCaT细胞后p38磷酸化水平上调显著（P＜0.05）,且都在50 mJ/cm2照射后效应最为显著（P＜0.05）.50 mJ/cm2 UVB照射HaCaT细胞后8h,ERK1/2磷酸化产物水平出现上调,在处理后的12h,与对照组（10.277±0.320）比较,差异有统计学意义（44.844±2.023,P＜ 0.05）,而p38和JNK磷酸化产物水平在照射后2h即开始上调（P＜0.05）,4、8、12h,p38和JNK与对照相比,UVB照射后虽存在着显著地磷酸化水平差异（P＜0.05）,但随时间推移JNK磷酸化产物水平这种上调的趋势逐渐弱化（P＜0.05）.结论 在50 mJ/cm2中波紫外线照射后的HaCaT细胞中,ERK1/2、p38、JNK这3种MAPK信号通路产生了活化效应,具有一定的时间效应差异.
Objective To explore the regulatory effects of ultraviolet B （UVB） radiation on the mitogenactivated protein kinase （MAPK） signaling pathway in cultured HaCaT cells in vitro at different post-radiation time points.Methods Some cultured HaCaT cells were classified into several groups to be exposed to UVB of 1.5,4.5,7.5,10,20,30 and 50 mJ/cm2 for 1,3,5,7,15,20 and 34 seconds respectively,or UVB of 50 mJ/cm2 for 34 seconds,or remain untreated （control group）.Western blot was performed to determine the total and phosphorylation levels of ERK1/2,JNK and p38 in HaCaT cells at 8 hours after exposure to 1.5,4.5,7.5,10,20,30 and 50 mJ/cm2 UVB,as well as at 2,4,8 and 12 hours after exposure to 50 mJ/cm2 UVB.Quantity One software was used to measure the absorbance value of protein bands,and protein levels were expressed as the absorbance ratio of target bands to glyceraldehyde-3-phosphate dehydrogenase （GAPDH）.Statistical analysis was carried out by using one-way analysis of variance,least significant difference （LSD）-t test and multivariate analysis of variance.Results Compared with the control group,there was a significant increase at 8 hours in the phosphorylation levels of ERK 1/2 and JNK in HaCaT cells after exposure to 7.5,10,20,30 and 50 mJ/cm2 UVB （all P 〈 0.05）,and in the phosphorylation level of p38 after exposure to 20,30 and 50 mJ/cm2 UVB （all P 〈 0.05）,with the strongest increase in phosphorylation levels of ERK 1/2,JNK and p38 all observed in HaCaT cells exposed to 50 mJ/cm2 UVB （all P 〈 0.05）.After exposure to 50 mJ/cm2 UVB,the phosphorylation level of ERK1/2 began to increase in HaCaT cells at 8 hours,and significantly increased at 12 hours （44.844 ± 2.023 vs.10.277 ± 0.320,P 〈 0.05） compared with the control group;however,the phosphorylation levels of p38 and JNK increased in HaCaT cells as early as 2 hours after exposure （both P 〈 0.05）,and remained significantly higher compared with the control group at 4,8 and 12 hours （all P 〈 0.05）,while the inc
出处
《中华皮肤科杂志》
CAS
CSCD
北大核心
2015年第6期391-394,共4页
Chinese Journal of Dermatology
基金
国家自然科学基金面上项目（81371755、81171513）
江苏省自然科学基金面上项目（BK20131064）
2013年高等学校博士学科点专项科研基金（20131106120046）
2014年北京协和医学院协和青年科研基金（3332014008）
