摘要
目的研究核转运因子CSE1L在化疗后神经母细胞瘤DNA损伤修复过程中的表达变化并初步探索其作用机制,为神经母细胞瘤诊疗提供线索和理论依据。方法通过免疫组化染色方法对19例未化疗和28例经化疗的神经母细胞瘤组织中CSE1L表达水平进行分析。阿霉素处理SH-SY5Y细胞6小时和12小时建立DNA损伤模型,并通过免疫荧光方法检测DNA损伤修复标志物g-H2AX和53BP1在DNA损伤位点的聚集情况;免疫印迹检测阿霉素诱导DNA损伤后CSE1L以及53BP1、FEN-1和EXO-1等DNA损伤相关蛋白的变化水平。慢病毒敲减CSE1L后,免疫印迹检测CSE1L、53BP1、FEN-1和EXO-1蛋白水平变化。结果未化疗组CSE1L阳性率(73.7%)高于化疗组(42.9%)并具有显著差异(卡方检验P=0.037);阿霉素可诱导DNA损伤修复标志物g-H2AX和53BP1在DNA损伤位点的聚集;并诱导53BP1、CSE1L、FEN-1的表达水平在处理6小时先增高、在12小时降低,EXO-1表达持续增高;敲减CSE1L可降低53BP1和FEN-1表达,但促进EXO-1表达。结论 CSE1L在化疗后神经母细胞瘤样本中表达降低,并参与DNA损伤修复过程。
Objective Investigate the role of CSE1L and molecular mechanism in the repair of DNA damage. Methods Immunohistochemical staining was performed to analyze the expression of CSE1L in neuroblastoma tissue. There were 19 cases without chemotherapy and 28 cases with chemotherapy. To establish neuroblastoma DNA damage model, Adriamycin was administrated in SH-SY5 Y cell line for 6 hours and 12 hours. γ-H2AX and 53BP1, the markers for DNA double strand breaks, were observed in DNA damage sites by immunofluorescence. Immunoblotting was used to detect the expression of the related protein of DNA damage, such as CSE1L, 53BP1, FEN-1 and EXO-1 after Adriamycin treatment for 6 hours and 12 hours. Lentivirus introduced CSE1L-knockdown was performed to detect the expression of CSE1L,53BP1 ,FEN-1 and EXO-1 by western blotting. Results The positive rate of CSE1L in non-chemotherapy group （73.7%） is significantly higher than chemotherapy group （42.9 % ） （ chisquare test P = 0. 037 ）. At the sites of DNA damage, Adriamycin induced the aggregation of γ-H2AX and 53BP1. It also induced increased expression of CSE1L, FEN-1 at 6 h, while decreased at 12 h. The expression of EXO-1 increased at 12 h. CSE1L- knockdown inhibited 53BP1 and FEN-1 expression, but promoted the expression of EXO-1. Conclusion The finding indicated that the expression of CSE1L was decreased in the chemotherapy neuroblastoma samples, and it might be involved in the DNA damage repair process.
出处
《标记免疫分析与临床》
CAS
2016年第10期1202-1206,共5页
Labeled Immunoassays and Clinical Medicine
基金
国家自然科学基金(81472369)
北京市科技计划(D131100005313014)
首都医科大学基础-临床科研合作基金(15JL69)
北京市优秀人才培养资助(2015000021469G210)
作者简介
陈峰（1989-），男，博士在读，主要从事儿童头颈部实体肿瘤研究
通讯作者：金雅琼，助理研究员。Tel：010-59616896；E-mail：jyq86824@163.com
