Bscl2基因敲除对细胞增殖和DNA损伤修复能力的影响  

Effects of Bscl2 gene knockout on cell proliferation and DNA damage repair capacity

作　　者：张丛俏 黄翔 张伟杨 安智渊 田盼 万能 任来峰[1] 郭晋锋 Zhang Congqiao;Huang Xiang;Zhang Weiyang;An Zhiyuan;Tian Pan;Wan Neng;Reng Laifeng;Guo Jinfeng(Central Laboratory,Cancer Hospital Affiliated of Shanxi Medical University,Taiyuan Shanxi 030001,China;Center of Reproductive Medicine,Shanxi Women and Children Health Hospital,Taiyuan Shanxi 030001,China;Office of the Dean,Cancer Hospital Affiliated of Shanxi Medical University,Taiyuan Shanxi 030001,China)

机构地区：[1]山西医科大学附属肿瘤医院中心实验室,山西太原030001 [2]山西省妇幼保健院生殖医学中心,山西太原030001 [3]山西医科大学附属肿瘤医院办公室,山西太原030001

出　　处：《遵义医科大学学报》2025年第3期234-242,共9页Journal of Zunyi Medical University

基　　金：山西省自然科学基金资助项目(NO:202403021211128);山西省卫生健康委科研基金(NO:2024108)。

摘　　要：目的 探讨Bscl2基因敲除对细胞增殖和DNA损伤修复能力的影响。方法 构建Bscl2基因敲除小鼠模型,胰酶消化法分离胚胎发育14.5 d (E14.5 d)的MEF细胞;免疫荧光染色法对分离培养的MEF细胞进行纯化鉴定;PCR扩增技术结合琼脂糖凝胶电泳对培养细胞及小鼠基因型进行鉴定,并将细胞及小鼠分为野生组(WT wild type)及Bscl2基因敲除组(KO knockout);使用台盼蓝染色法和5-溴脱氧尿嘧啶核苷(BrdU)免疫荧光标记法检测两组细胞增殖能力;用γH2AX免疫荧光染色法评估两组细胞及小鼠组织在双链断裂损伤诱导后的DNA损伤修复能力。结果 分离E14.5 d MEF细胞,成功构建小鼠MEF细胞系。Bscl2基因敲除MEF细胞的增殖能力明显低于野生型,Bscl2基因敲除MEF细胞及小鼠活体组织的DNA损伤修复能力较野生型明显减弱。结论 敲除Bscl2基因抑制小鼠MEF细胞的增殖,降低小鼠细胞及组织的DNA损伤修复能力。Objective This study aims to investigate whether the knockout of the Bscl2 gene affects cell proliferation and DNA damage repair capabilities or not both in vivo and in vitro.Methods Construction of a Bscl2 gene knockout mouse model and MEFs was isolated from fetuses at embryonic day 14.5(E14.5 d)by using trypsin digestion.The isolated and cultured MEFs were purified and identified through immunofluorescence staining.The genotypes of the cultured cells and mice was identified by combining PCR amplification with agarose gel electrophoresis,and cells and mice were categorized into the wild type(WT)and Bscl2-knockout(KO)groups.Trypan blue staining and 5-bromo-2′-deoxyuridine(BrdU)immunofluorescence labeling was performed to evaluate cell proliferation.Assessment of DNA damage repair capacity in two groups of cells and mouse tissues after double-strand break induction was performed byγH2AX immunofluorescence staining.Results MEFs from E14.5 d was successfully isolated to establish a mouse MEF cell line.The proliferation capacity of Bscl2-knockout MEFs was significantly lower than that of the wild type.DNA damage repair capacity in Bscl2-knockout MEFs and mouse tissues was markedly impaired when comparing to the wild type.Conclusion Knockout of the Bscl2 gene inhibits the proliferation of mouse MEFs and reduces the DNA damage repair capacity of mouse cells and tissues.

关 键 词：先天性脂肪营养不良2型基因 胚胎成纤维细胞 细胞增殖 DNA双链断裂 DNA修复 

分 类 号：R589.2[医药卫生—内分泌]