氢醌对L-02肝细胞自噬的影响  

The effect of hydroquinone on autophagy of L-02 hepatocytes

作　　者：郑作兵 张伟 熊亮[3] 刘婷婷[3] 吴杨娜 杨嘉辉 胡恭华 Zheng Zuobing;Zhang Wei;Xiong Liang;Liu Tingting;Wu Yangna;Yang Jiahui;Hu Gonghua(School of Public Health,Nanchang University,Nanchang 330006;Jiangxi Provincial Key Laboratory of Preventive Medicine,Nanchang 330006;School of Public Health and Health Management,Gannan Medical University,Ganzhou 341000)

机构地区：[1]南昌大学公共卫生学院,南昌330006 [2]江西省预防医学重点实验室,南昌330006 [3]赣南医学院公共卫生与健康管理学院,赣州341000

出　　处：《安徽医科大学学报》2022年第9期1425-1430,共6页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81960599、81460504、81260434、81060241)。

摘　　要：目的研究氢醌(HQ)对人L-02肝细胞自噬的影响。方法用不同浓度HQ(0、10、20、40和80μmol/L)处理L-02细胞24 h后,采用透射电镜观察自噬体形成;采用mCheery-GFP-LC3B融合蛋白示踪自噬体形成;采用Western blot实验检测自噬标志蛋白LC3、Beclin-1、P62的表达;采用免疫荧光实验检测自噬蛋白LC3、Beclin-1、P62的亚细胞定位及表达。结果透射电镜下,观察到HQ作用后L-02细胞自噬体和溶酶体增多。荧光显微镜下,GFP-LC3B点状黄色斑点数量随HQ剂量增加而增多,而红色片状斑点减少。Western blot结果显示,HQ提高L-02细胞LC3Ⅱ/LC3Ⅰ蛋白的比值水平,各处理组与对照组差异均有统计学意义(P<0.05);HQ对L-02细胞的Beclin-1和P62蛋白表达无明显影响(P>0.05)。免疫荧光实验结果显示,LC3和Beclin-1在L-02细胞的胞核和胞质中均有表达,HQ对L-02细胞内LC3和Beclin-1的亚细胞定位未产生影响,而40、80μmol/L的HQ则可诱导L-02细胞内的P62由胞质向胞核转位。结论HQ诱导L-02细胞的自噬体增多,可能与其导致的自噬体清除障碍有关。Objective To study the effect of hydroquinone on autophagy of human L-02 hepatocytes.Methods L-02 cells were treated with different concentrations of hydroquinone(0,10,20,40 and 80μmol/L)for 24 hours,the formation of autophagosomes was observed by transmission electron microscope;autophagosome formation was traced by mCheery-GFP-LC3B fusion protein.Western blot experiment was used to detect the expression of autophagy marker proteins LC3,Beclin-1,P62;immunofluorescence experiment was used to detect the subcellular localization and expression of autophagy proteins LC3,Beclin-1,P62.Results Under the transmission electron microscope,it was observed that the autophagosomes and lysosomes of L-02 cells increased after the action of hydroquinone.Under the fluorescence microscope,the number of GFP-LC3B punctate yellow spots increased with the increase of hydroquinone dose,while the red flake spots decreased.Western blot results showed that hydroquinone increased the ratio of LC3Ⅱ/LC3Ⅰprotein in L-02 cells,and the difference between treatment groups and the control group was statistically significant(P<0.05).Hydroquinone had no significant effect on Beclin-1 and P62 protein expression in L-02 cells(P>0.05).The results of immunofluorescence experiments showed that LC3 and Beclin-1 were both expressed in the nucleus and cytoplasm of L-02 cells,hydroquinone had no effect on the subcellular localization of LC3 and Beclin-1 in L-02 cells,while 40 and 80μmol/L hydroquinone could induce the translocation of P62 in L-02 cells from the cytoplasm to the nucleus.Conclusion Hydroquinone induces the increase of autophagosomes in L-02 cells,which may be related to the obstacles to autophagosome clearance.

关 键 词：氢醌 肝细胞 自噬 

分 类 号：R114[医药卫生—卫生毒理学]