白藜芦醇对婴儿血管瘤内皮细胞活性的影响  

Effects of resveratrol on the activity of infantile hemangioma-derived endothelial cells

作　　者：杨开颖 邱桐 龚雪 兰钰茹 周江元 陈思源[2] 吉毅[1] Yang Kaiying;Qiu Tong;Gong Xue;Lan Yuru;Zhou Jiangyuan;Chen Siyuan;Ji Yi(Department of Pediatric Surgery,West China Hospital,Sichuan University,Chengdu 610041,China;Department of Critical Care Medicine,West China Hospital,Sichuan University,Chengdu 610041,China)

机构地区：[1]四川大学华西医院小儿外科,成都610041 [2]四川大学华西医院重症医学科,成都610041

出　　处：《中华皮肤科杂志》2022年第11期990-995,共6页Chinese Journal of Dermatology

基　　金：国家自然科学基金(81401606、81400862);四川省科技厅重点研发项目(2019YFS0322);四川大学优秀青年学者基金(2015SU04A15);四川大学华西医院学科卓越发展1·3·5工程临床研究孵化项目(2019HXFH056、2020HXFH048、ZYJC21060)。

摘　　要：目的:研究葡萄糖转运体1(Glut-1)抑制剂白藜芦醇(RSV)对婴儿血管瘤内皮细胞(HemEC)活性的影响。方法:收集4例增殖期与4例消退期婴儿血管瘤(IH)组织,采用免疫组化观察Glut-1的表达。从4例增殖期IH组织中提取原代HemEC,采用qPCR与Western印迹检测HemEC与对照脐静脉内皮细胞(HUVEC)中Glut-1 mRNA与蛋白的表达水平。体外培养HemEC,分别采用0(对照组)、50、100、200、400、800μmol/L RSV干预24 h,采用CCK-8法检测各组HemEC增殖能力,计算半抑制浓度(IC50)。分别采用Transwell实验、流式细胞仪观察HemEC迁移能力及凋亡水平。组间比较采用t检验或方差分析,组间多重比较采用LSD-t检验。结果:免疫组化显示,增殖期及消退期IH组织中血管内皮细胞Glut-1均阳性,但增殖期表达丰富,消退期表达明显降低。HemEC中Glut-1 mRNA及蛋白表达水平(1.793±0.041、1.959±0.144)均高于HUVEC(0.820±0.073、0.648±0.046,t=16.35、12.28,均P<0.001)。采用Glut-1抑制剂RSV干预HemEC后,0~800μmol/L组HemEC增殖活性差异有统计学意义(F=1043.00,P<0.001),所有RSV干预组HemEC的增殖活性均低于对照组(P<0.05)。GraphPad Prism 8计算得出RSV IC50浓度为150μmol/L,Transwell实验与流式细胞仪检测显示,150μmol/L RSV干预组HemEC组细胞迁移数(61±5)低于对照组(150±6,t=15.11,P<0.001),凋亡率(13.01%±0.45%)高于对照组(3.93%±0.68%,t=19.34,P<0.001)。结论:糖酵解关键酶Glut-1在IH增殖期组织与HemEC中高表达,RSV干预可以抑制HemEC增殖及迁移能力并促进其凋亡。Objective To investigate the effect of the glucose transporter 1(Glut-1)inhibitor resveratrol on the activity of infantile hemangioma(IH)-derived endothelial cells(HemEC).Methods IH tissues were collected from 4 cases of proliferating IH and 4 cases of involuting IH,and immunohistochemical study was performed to determine the Glut-1 expression.Primary HemEC were extracted from 4 proliferating IH tissues,real-time fluorescence-based quantitative PCR(qPCR)and Western blot analysis were performed to determine the mRNA and protein expression of Glut-1 in HemEC and human umbilical vein endothelial cells(HUVEC),respectively.HemEC were cultured in vitro and treated with 0(control group),50,100,200,400 and 800μmol/L resveratrol for 24 hours,respectively.Cell counting kit-8(CCK8)assay was performed to evaluate the proliferative ability of HemEC in the above groups,and the 50%inhibitory concentration(IC50)was calculated.The migratory ability and apoptosis level of HemEC were assessed by Transwell assay and flow cytometry,respectively.Intergroup comparisons were performed using t test or analysis of variance,and multiple comparisons were performed using least significant difference-t test.Results Immunohistochemical study showed that Glut-1 was expressed in vascular endothelial cells derived from both proliferating and involuting IH tissues,and the Glut-1 expression was abundant in the proliferating IH but markedly decreased in the involuting IH tissues.The mRNA and protein expression levels of Glut-1 were significantly higher in HemEC(1.793±0.041,1.959±0.144,respectively)than in HUVEC(0.820±0.073,0.648±0.046,t=16.35,12.28,respectively,both P<0.001).After the treatment with Glut-1 inhibitor resveratrol at different concentrations,the proliferative ability of HemEC significantly differed among the control group,50-,100-,200-,400-and 800-μmol/L resveratrol groups(F=1043.00,P<0.001),and was significantly lower in all the resveratrol groups than in the control group(all P<0.05).The IC50 of resveratrol was calculated to be

关 键 词：血管瘤 人脐静脉内皮细胞 葡萄糖转运体1型 细胞增殖 细胞凋亡 白藜芦醇 

分 类 号：R285.5[医药卫生—中药学]