内皮唾液酸蛋白在人增生性瘢痕中的表达及其对成纤维细胞表型的调控  

Expression of endosialin in human hypertrophic scars and its regulation on fibroblast phenotype

作　　者：张清怡 张丽霞 韩东晖 焦晓春 郑朝 郭凯 杨云舒 Zhang Qingyi;Zhang Lixia;Han Donghui;Jiao Xiaochun;Zheng Zhao;Guo Kai;Yang Yunshu(Department of Burns and Cutaneous Surgery,Burn Center of PLA,the First Affiliated Hospital of Air Force Medical University,Xi'an 710032,China;The Third Student Battalion,School of Basic Medical Sciences of Air Force Medical University,Xi'an 710032,China;Department of Urology,the First Affiliated Hospital of Air Force Medical University,Xi'an 710032,China)

机构地区：[1]空军军医大学第一附属医院全军烧伤中心,烧伤与皮肤外科,西安710032 [2]空军军医大学基础医学院学员三大队,西安710032 [3]空军军医大学第一附属医院泌尿外科,西安710032

出　　处：《中华烧伤与创面修复杂志》2023年第12期1168-1174,共7页Chinese Journal of Burns And Wounds

基　　金：国家自然科学基金青年科学基金项目(81501666,82302809);陕西省重点研发计划(2022SF-399)。

摘　　要：目的探讨内皮唾液酸蛋白即CD248在人增生性瘢痕(HS)中的表达及其对人HS成纤维细胞(HSF)表型的调控作用。方法采用实验研究方法。2023年3—5月,空军军医大学第一附属医院烧伤与皮肤外科收治3例HS患儿,其中女2例、男1例,年龄1岁10个月~2岁。取前述患儿术中切除的HS组织及行全厚皮移植术后剩余的全厚皮片即正常皮肤组织,进行后续实验。取前述2种组织,行苏木精-伊红染色后观察组织结构,行Masson染色后观察组织中胶原分布情况,行免疫组织化学染色后观测组织中CD248表达情况。取HS组织,采用组织块培养法提取原代HSF,取第3~5代HSF进行后续实验。将HSF按随机数字表法分为免疫球蛋白G78(IgG78)处理组与IgG对照组,分别用终物质的量浓度为200 nmol/L的人源性CD248单克隆抗体IgG78与人源性IgG对照抗体处理24 h,采用实时荧光定量反转录PCR法检测细胞中Ⅰ型胶原蛋白和α平滑肌肌动蛋白(α-SMA)的mRNA表达,采用蛋白质印迹法检测细胞中Ⅰ型胶原蛋白和α-SMA的蛋白表达,采用免疫荧光法检测细胞中Ⅰ型胶原蛋白和α-SMA的定位和蛋白表达情况。各实验样本数均为3。对数据行配对样本t检验和独立样本t检验。结果相较于正常皮肤组织,HS组织中表皮与真皮层均明显增厚,真皮层中胶原大量蓄积且排布紊乱。HS组织中CD248表达量明显高于正常皮肤组织(t=5.29,P<0.05)。处理24 h,IgG78处理组HSF中Ⅰ型胶原蛋白、α-SMA的mRNA表达量分别为0.39±0.05、0.56±0.09,分别明显低于IgG对照组的1.00±0.07、1.00±0.08(t值分别为11.87、6.49,P值均<0.05);IgG78处理组HSF中Ⅰ型胶原蛋白、α-SMA的蛋白表达量分别为0.617±0.011、0.67±0.14,分别明显低于IgG对照组的1.259±0.052、1.23±0.16(t值分别为20.92、4.52,P值均<0.05)。处理24 h,免疫荧光染色显示,Ⅰ型胶原蛋白、α-SMA主要定位在2组HSF的细胞质中,IgG78处理组HSF中Ⅰ型胶原蛋白、α-SMAObjective To explore the expression of endosialin,i.e.,CD248 in human hypertrophic scars(HSs)and its regulatory effect on the phenotype of hypertrophic scar fibroblasts(HSFs).Methods The method of experimental research was used.From March to May,2023,3 pediatric patients with HS were admitted to the Department of Burns and Cutaneous Surgery of the First Affiliated Hospital of Air Force Medical University,including 2 females and 1 male,aged one year ten months to two years.The HS tissue resected during the surgery and the remaining full-thickness skin graft,i.e.,normal skin tissue after full-thickness skin grafting were collected from the aforementioned pediatric patients for subsequent experiments.Using the aforementioned two types of tissue,the histological structures were observed by hematoxylin-eosin staining,collagen distribution was observed by Masson staining,and the expression of CD248 was observed and measured by immunohistochemical staining.The primary HSFs were isolated from HS tissue using explant culture technique,and the 3rd to 5th passages of HSFs were used in subsequent experiments.According to the random number table,HSFs were divided into immunoglobulin G78(IgG78)-treated group and IgG control group,which were treated with 200 nmol/L human CD248 monoclonal antibody IgG78 and human IgG control antibody for 24 h,respectively.The mRNA expressions of collagen type Ⅰ(ColⅠ)and α-smooth muscle actin(α-SMA)in HSFs were measured by real-time fluorescence quantitative reverse transcription polymerase chain reaction,the protein expressions of ColⅠand α-SMA in HSFs were detected by Western blotting,and the intracellular location and protein expressions of ColⅠandα-SMA were detected by immunofluorescence method.The number of samples in each experiment was 3.Data were statistically analyzed with paired sample t test and independent sample t test.Results Compared with those in normal skin tissue,the epidermis and dermis in HS tissue were significantly thicker,with massive accumulation and disordered

关 键 词：瘢痕 成纤维细胞 细胞外基质 细胞转分化 内皮唾液酸蛋白 胶原合成 

分 类 号：R726.2[医药卫生—儿科]