下调脂肪酸结合蛋白5对人皮肤角质形成细胞放射损伤的影响  

Effect of down-regulation of FABP5 on radiation damage of human keratinocytes

作　　者：管鸿丹 郑榕 官秉洁 林玉萍 王弼思 徐本华[1] 宋建元[1] Guan Hongdan;Zheng Rong;Guan Bingjie;Lin Yuping;Wang Bisi;Xu Benhua;Song Jianyuan(Department of Radiation Oncology,Fujian Medical University Union Hospital,Fujian Key Laboratory of Intelligent Imaging and Precision Radiotherapy for Tumors(Fujian Medical University),Clinical Research Center for Radiology and Radiotherapy of Fujian Province(Digestive,Hematologicaland Breast Malignancies),Fuzhou 350001,China)

机构地区：[1]福建医科大学附属协和医院放疗科福建省肿瘤智能影像与精准放疗重点实验室福建省消化、血液系统与乳腺恶性肿瘤放射与治疗临床医学研究中心,福州350001

出　　处：《中华放射医学与防护杂志》2023年第1期8-14,共7页Chinese Journal of Radiological Medicine and Protection

基　　金：福建省卫生健康中青年骨干人才培养项目(2019-ZQN-44)。

摘　　要：目的探究下调脂肪酸结合蛋白5(FABP5)对皮肤细胞放射损伤的影响及其相关机制。方法构建下调FABP5的慢病毒载体,将慢病毒感染人皮肤角质形成细胞(HaCaT)并检测转染效率。将细胞分为空白对照组、下调FABP5组、单纯照射组、下调FABP5+照射组。予6 MV X射线照射后,CCK-8法测定细胞增殖活力,划痕实验检测细胞迁移,流式细胞术分析细胞凋亡,克隆形成实验检测放射敏感性,免疫印迹检测细胞中PARP1、γ-H2AX、AKT、p-AKT的蛋白表达水平。结果成功构建下调FABP5的HaCaT细胞,从RNA水平(t=25.14,P<0.05)和蛋白水平(t=20.06,P<0.05)验证均达到下调FABP5的目的。下调FABP5组细胞增殖(t=3.55、5.88、3.18,P<0.05)、迁移能力(t=15.44,P<0.05)显著减弱,其放射增敏比为0.782。下调FABP5+照射组细胞凋亡率较单纯照射组显著减少[(9.82±1.45)%vs.(22.05±6.71)%,t=3.08,P<0.05]。下调FABP5+照射组细胞的PARP1和γ-H2AX蛋白水平分别为0.04±0.04、0.11±0.06和0.26±0.11、0.22±0.07,均低于单纯照射组的0.21±0.10、0.52±0.22和0.57±0.06、0.43±0.02(t=2.83、3.07、4.50、5.33,P<0.05),而p-AKT蛋白水平高于单纯照射组(t=-16.24~3.02,P<0.05)。结论下调FABP5抑制皮肤细胞增殖、迁移,增强放射抵抗性,减少辐射诱导皮肤细胞凋亡和DNA损伤。这可能是通过磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(AKT)信号通路来抑制皮肤细胞放射敏感性,为放射性皮肤损伤防护提供一种新思路。Objective To investigate the effects of down-regulation of FABP5(fatty acid binding protein 5)on radiation damage of skin cells,and explore underlying mechanism.Methods A lentiviral vector with down-regulated FABP5 was constructed to infect human immortalized keratinocytes(HaCaT)cells,and the transfection efficiency was examined.The HaCaT cells were divided into blank control group,FABP5 down-regulation group(FABP5),radiation group(IR),and FABP5 down-regulation combined with radiation group(FABP5+IR).After 6 MV X-ray radiation,cell proliferation viability was measured by CCK-8 assay,cell migration was detected by scratch assay,apoptosis was analyzed by flow cytometry,radiosensitivity was evaluated by cloning formation assay,and the cellular protein expressions of PARP1,γ-H2AX,AKT and p-AKT were detected by Western blot.Results FABP5 was successfully knocked-down in both RNA level(t=25.14,P<0.05)and protein level(t=20.06,P<0.05).The down-regulation of FABP5 decreased the abilities of cells proliferation(t=3.55,5.88,3.18,P<0.05)and migration(t=15.44,P<0.05),but increased cell resistance to irradiation with a radiosensitization ratio of 0.782.The apoptosis rate of FABP5+IR group was significantly lower than IR group(22.05±6.71)%vs.(9.82±1.45)%,t=3.08,P<0.05.The protein levels of PARP1 andγ-H2AX in FABP5+IR group were also lower than those in the IR group 0.04±0.04,0.11±0.06,0.26±0.11,0.22±0.07,0.21±0.10,0.52±0.22,0.57±0.06,0.43±0.02(t=2.83,3.07,4.50,5.33,P<0.05),while the protein level of p-Akt in FABP5+IR group was higher than that in IR group(t=-16.24—3.02,P<0.05).Conclusions Down-regulation of FABP5 inhibited cell proliferation and migration,increased radioresistance,and reduced radiation-induced apoptosis and DNA damage of skin cells probably through PI3K/AKT signaling pathway.

关 键 词：皮肤 脂肪酸结合蛋白5 放射敏感性 辐射防护 PI3K/AKT信号通路 

分 类 号：R818[医药卫生—放射医学]