摘要
目的探讨槲皮素抑制瘢痕疙瘩成纤维细胞增殖和阻滞细胞周期机制。方法人瘢痕疙瘩成纤维细胞购自上海生命科学研究院细胞资源中心。分别以10μmol/L及40μmol/L浓度槲皮素处理细胞(记为10μmol/L槲皮素组、40μmol/L槲皮素组);分别将miR-NC、微小核糖核酸200c(miR-200c)转染至瘢痕疙瘩成纤维细胞(记为miR-NC组、miR-200c组);将抗miR-NC、抗miR-200c分别转染至瘢痕疙瘩成纤维细胞,再用40μmol/L浓度槲皮素处理(记为40μmol/L槲皮素+抗miR-NC组、40μmol/L槲皮素+抗miR-200c组)。细胞计数试剂盒-8(CCK-8)检测细胞增殖;流式细胞仪检测细胞周期;实时荧光定量聚合酶链式反应(qRT-PCR)检测miR-200c表达水平;蛋白质印迹法检测磷酸化雷帕霉素的哺乳动物靶标(p-mTOR)蛋白表达。两组间比较采用t检验;多组间比较用单因素方差分析(两组间比较用LSD-t检验)。结果10μmol/L及40μmol/L浓度槲皮素组瘢痕疙瘩成纤维细胞增殖率、S期显著低于Control组[增殖率分别(66.85±7.04)%、(43.86±5.73)%比(98.65±9.64)%,S期分别(19.47±2.43)%、(13.87±2.22)%比(28.95±3.86)%],G0/G1期及miR-200c表达表达显著高于Control组[G0/G1期分别(50.44±4.43)%、(59.45±6.22)%比(40.68±6.86)%,miR-200c表达分别1.74±0.33、2.64±0.48比1.00±0.12],差异有有统计学意义(F=386.976、243.858、376.886、185.643,P<0.05)。miR-200c组瘢痕疙瘩成纤维细胞miR-200c表达和G0/G1期显著高于miR-NC组细胞[miR-200c表达为2.50±0.37比1.02±0.10,G0/G1期为(58.46±3.11)%比(42.57±5.97)%],细胞增殖率、S期和p-mTOR蛋白表达显著低于miR-NC组细胞[细胞增殖率为(48.49±4.98)%比(98.56±9.56)%,S期为(15.05±3.11)%比(27.84±2.58)%,p-mTOR蛋白表达为0.33±0.03比0.89±0.08],差异有统计学意义(t=19.064、39.764、34.864、28.979、14.176,P<0.05)。40μmol/L槲皮素+抗miR-200c组瘢痕疙瘩成纤维细胞miR-200c表达、G0/G1期显著低于40μmol/L槲皮素+抗miR-NC组细胞[miR-200c表达为0.37�
Objective To study the mechanism of Quercetin inhibiting keloid fibroblast’s proliferation and arresting cell cycle.Methods Human keloid fibroblasts were purchased from the cell resource center of shanghai institutes for biological sciences.Cells were treated with quercetin at concentrations of 10μmol/L and 40μmol/L respectively(10μmol/L quercetin group and 40μmol/L quercetin group);miR-NC and Micro-ribonucleic acid-200c(miR-200c)were transfected into keloid fibroblasts(miR-NC group and miR-200c group);anti-miR-NC and anti-miR-200c were transfected into keloid fibroblasts respectively and treated with 40μmol/L quercetin(40μmol/L quercetin+anti-miR-NC group,40μmol/L quercetin+anti-miR-200c group).Cell counting kit-8(CCK-8)was used to detect cell proliferation.Flow cytometry was used to detect cell cycle;Quantitative real-time polymerase chain reaction(qRT-PCR)was used to detect the expression of miR-200c.The protein expression of phospho-mammalian target of rapamycin(p-mTOR)was detected by Western blotting.The independent samples t test was used for comparison between two groups;the one-way analysis of variance was used for comparison between multiple groups(LSD-t test was used for comparison between two groups).Results The proliferation rate and S phase of keloid fibroblasts in the 10μmol/L and 40μmol/L quercetin groups were significantly lower than those in the Control group[proliferation rates were(66.85±7.04)%,(43.86±5.73)%vs.(98.65±9.64)%,S phase were(19.47±2.43)%,(13.87±2.22)%vs.(28.95±3.86)%],while the expression of G0/G1 and miR-200c were significantly higher[G0/G1 phase were(50.44±4.43)%,(59.45±6.22)%vs.(40.68±6.86)%,miR-200c expression were 1.74±0.33,2.64±0.48 vs.1.00±0.12,respectively],the differences were statistically significant(F=386.976,243.858,376.886,185.643,P<0.05).The miR-200c expression and G0/G1 phase in the miR-200c group were significantly higher than those in the miR-NC group[miR-200c expression was 2.50±0.37 vs.1.02±0.10,G0/G1 phase was(58.46±3.11)%vs.(42.57±5.9
作者
任拥媛
李钢
李迎辉
赵羽佳
白杨
张士歆
Ren Yongyuan;Li Gang;Li Yinghui;Zhao Yujia;Bai Yang;Zhang Shixin(Depatment of Plastic Surgery and Medical Cosmetic,the Second Hospital of Tianjin Medical University,Tianjin 300211,China;State Key Laboratory of Experimental Hematology,Institute of Hematology&Blood Diseases Hospital,China Academy of Medical Sciences&Peking Union Medical College,Tianjin 300202,China)
出处
《中华实验外科杂志》
CAS
北大核心
2022年第11期2109-2112,共4页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金面上项目(81970105)。
