糖尿病足患者创缘皮肤组织中成纤维细胞与角质形成细胞的相互作用及其机制  

Interaction between fibroblasts and keratinocytes in the wound edge skin tissue of a diabetic foot patient and the mechanism

作　　者：阮琼芳[1] 章思语 席毛毛 阮晶晶[2] 刘淑华 李炳辉[3] 谢卫国[2] Ruan Qiongfang;Zhang Siyu;Xi Maomao;Ruan Jingjing;Liu Shuhua;Li Binghui;Xie Weiguo(Institute of Burns,Tongren Hospital of Wuhan University&Wuhan Third Hospital,Wuhan 430060,China;Department of Burns,Tongren Hospital of Wuhan University&Wuhan Third Hospital,Wuhan 430060,China;Department of Wound Repair,Liyuan Hospital Affiliated to Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430077,China)

机构地区：[1]武汉大学附属同仁医院暨武汉市第三医院烧伤研究所,武汉430060 [2]武汉大学附属同仁医院暨武汉市第三医院烧伤科,武汉430060 [3]华中科技大学同济医学院附属梨园医院创面修复科,武汉430077

出　　处：《中华烧伤与创面修复杂志》2024年第8期762-771,共10页Chinese Journal of Burns And Wounds

基　　金：武汉市科技局知识创新项目(2023020201010192);湖北省自然科学基金面上项目(2021CFB532);湖北省卫健委科研项目(WJ2021M260);上海王正国创伤医学发展基金会生长因子复兴计划(SZYZ-TR-10)。

摘　　要：目的探讨糖尿病足患者创缘皮肤组织中成纤维细胞(Fb)与角质形成细胞(KC)的相互作用及其机制。方法该研究为实验研究。取华中科技大学同济医学院附属梨园医院创面修复科2021年8月收治的1例糖尿病足患者(男,33岁)和该院手外科2021年9月收治的1例急性足外伤患者(男,50岁)创缘皮肤组织,行单细胞转录组测序,分析Fb亚群中趋化因子配体和KC亚群中趋化因子受体的相互作用。收集常规培养和用高浓度葡萄糖培养7 d的人包皮Fb(HFF)的上清液,分别作为正常条件培养基(CM)和高糖CM。取HaCaT细胞,分为用正常CM培养的正常CM组和用高糖CM培养的高糖CM组,进行划痕试验,并计算划痕后24、48 h细胞迁移率(样本数为3)。利用液相悬浮芯片检测2种CM中细胞因子含量(样本数为5)。取HFF,分为常规培养的正常组和用高浓度葡萄糖培养的高糖组,培养7 d,采用实时荧光定量反转录PCR法检测CXC趋化因子配体1(CXCL1)、CXCL2、CXCL8和CXCL12的mRNA表达(样本数为6)。取正常CM组和高糖CM组HaCaT细胞,采用蛋白质印迹法检测培养48 h细胞中CXC趋化因子受体4(CXCR4)的蛋白表达(样本数为3)。取HaCaT细胞,分为正常CM组、高糖CM组、正常CM+CXCL12组、高糖CM+CXCL12组,前2组细胞处理同前,后2组细胞分别采用含重组人CXCL12的正常CM和高糖CM培养,行划痕试验并计算划痕后24、48 h细胞迁移率,采用蛋白质印迹法检测培养48 h细胞中CXCR4蛋白表达(样本数均为3)。结果相较于急性足外伤创缘皮肤组织,糖尿病足创缘皮肤组织Fb亚群中的趋化因子配体(CXCL1、CXCL2、CXCL3、CXCL8、CXCL12)和KC亚群中的趋化因子受体(CXCR2和CXCR4)间的相互作用明显减弱。划痕后24、48 h,高糖CM组HaCaT细胞迁移率均明显低于正常CM组(t值分别为23.50、15.65,P<0.05)。相较于正常CM,高糖CM中的CXCL1含量明显增多(P<0.05),CXCL12含量明显减少(P<0.05)。培养7 d,相较于正常组,高糖�Objective To investigate the interaction between fibroblasts(Fb)and keratinocytes(KC)in the wound edge skin tissue of a diabetic foot patient and the mechanism.Methods This was an experimental research.The wound edge skin tissue from a diabetic foot patient(male and 33 years old)admitted to the Department of Wound Repair of Liyuan Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology in August 2021 and from an acute foot injury patient(male and 50 years old)admitted to the Department of Hand Surgery of the hospital in September 2021 was collected.The single-cell transcriptome sequencing was performed to analyze the interaction between chemokine ligands of Fb subgroup and chemokine receptors of KC subgroup.The supernatant was collected after human foreskin fibroblast(HFF)was cultured routinely and with high concentration of glucose for 7 days as normal conditioned medium(CM)and high glucose CM,respectively.HaCaT cells were collected and divided into normal CM group cultured with normal CM and high glucose CM group cultured with high glucose CM,the scratch test was performed to calculate the cell migration rates at 24 and 48 h after scratch(n=3).The content of cytokines in the two kinds of CM was detected by liquid suspension chip(n=5).HFF was collected and divided into normal group cultured routinely and high glucose group cultured with high concentration of glucose for 7 days,and the mRNA expressions of C-X-C motif chemokine ligand 1(CXCL1),CXCL2,CXCL8,and CXCL12 were detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction(n=6).HaCaT cells in normal CM group and high glucose CM group were collected to detect the protein expressions of C-X-C motif chemokine receptor 4(CXCR4)in cells cultured for 48 h by Western blotting(n=3).HaCaT cells were collected and divided into normal CM group,high glucose CM group,normal CM+CXCL12 group,and high glucose CM+CXCL12 group.The first two groups of cells were treated as before,and the latter two gro

关 键 词：细胞间通讯 趋化因子 CXC 受体 趋化因子 成纤维细胞 角质形成细胞 细胞迁移 创面修复 

分 类 号：R587.2[医药卫生—内分泌]