过表达生长停滞特异性蛋白6的骨髓间充质干细胞对糖尿病小鼠全层皮肤缺损创面的影响及其机制  

Influence and mechanism of bone marrow mesenchymal stem cells overexpressing growth arrest specific 6 on full-thickness skin defect wounds in diabetic mice

作　　者：刘培[1] 王超[1] 魏绮键 李玉腾 崔丽君 王昌钏 张帆[1] 马玲[1] 田轩 Liu Pei;Wang Chao;Wei Qijian;Li Yuteng;Cui Lijun;Wang Changchuan;Zhang Fan;Ma Ling;Tian Xuan(Department of Plastic Surgery,Qilu Hospital of Shandong University,Jinan 250012,China;Department of Surgery,Mudan District Hospital of Traditional Chinese Medicine of Heze City,Heze 274000,China)

机构地区：[1]山东大学齐鲁医院整形外科,济南250012 [2]菏泽市牡丹区中医医院外科,菏泽274000

出　　处：《中华烧伤与创面修复杂志》2025年第2期145-154,共10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(82400767);山东省自然科学基金青年基金项目(ZR2023QH243)。

摘　　要：目的 探讨过表达生长停滞特异性蛋白6(GAS6)的骨髓间充质干细胞(BMSC),即GAS6/BMSC对糖尿病小鼠全层皮肤缺损创面的影响及其机制.方法 该研究为实验研究.将12只8周龄雄性C57BU6J小鼠按照随机数字表法分为仅造成全层皮肤缺损的对照创面组与造成糖尿病全层皮肤缺损的糖尿病创面组,每组6只小鼠.伤后3、7、14、21d,计算创面愈合率.伤后21d,收集创面组织标本,行苏木精-伊红染色观察组织病理学情况;行Masson染色检测胶原沉积情况;行免疫组织化学染色检测增殖细胞核抗原(PCNA)阳性细胞数和CD31阳性细胞数,分别表示细胞增殖情况和毛细血管密度;行免疫荧光染色检测F4/80和髓过氧化物酶(MPO)双阳性细胞数,表示胞葬情况.取2只4周龄雄性C57BU6J小鼠,提取BMSC,通过腺病毒转染构建GAS6/BMSC并成功鉴定.取18只8周龄雄性C57BU6J小鼠制成糖尿病全层皮肤缺损创面模型后,按照随机数字表法将其分为磷酸盐缓冲液(PBS)组、BMSC组和GAS6/BMSC组(每组6只小鼠),伤后即刻于3组小鼠创面局部分别注射PBS、BMSC单细胞悬液、GAS6/BMSC单细胞悬液,于前述实验相同时间点计算创面愈合率、检测细胞增殖情况和毛细血管密度及胞葬情况.结果 伤后3、7、14、21d,糖尿病创面组小鼠创面愈合率均显著低于对照创面组(t值分别为7.99、8.62、9.80、5.85,P<0.05).与对照创面组相比,糖尿病创面组小鼠伤后21 d的创面组织中大量炎症细胞浸润,胶原沉积减少.伤后21 d,糖尿病创面组小鼠创面组织中PCNA阳性细胞数和CD31阳性细胞数均显著少于对照创面组(t值分别为6.61、5.38,P<0.05).伤后21d,糖尿病创面组小鼠创面组织中F4/80和MPO双阳性细胞数为(3.3±0.8)个,较对照创面组的(12.7±1.8)个显著减少(t=11.00,P<0.05).伤后14、21 d,BMSC组小鼠创面愈合率均显著高于PBS组(P<0.05);伤后3、7、14、21 d,GAS6/BMSC组小鼠创面愈合率均显著高于BMSC组(P<0.05).ObjectiveTo investigate the influence and mechanism of bone marrow mesenchymal stem cells(BMSCs)overexpressing growth arrest specific 6,i.e.GAS6/BMSCs on full-thickness skin defect wounds in diabetic mice.MethodsThis study was an experimental study.Twelve 8-week-old male C57BL/6J mice were divided into a control wound group with only full-thickness skin defects and a diabetic wound group with diabetic full-thickness skin defects according to the random number table method,with 6 mice in each group.The wound healing rates were calculated at 3,7,14,and 21 days after injury.At 21 days after injury,wound tissue specimens were collected for hematoxylin-eosin staining to observe the histopathological conditions;Masson staining was performed to detect collagen deposition;immunohistochemical staining was performed to detect the number of proliferating cell nuclear antigen(PCNA)-positive cells and CD31-positive cells,representing cell proliferation and capillary density,respectively;immunofluorescence staining was performed to detect the number of F4/80 and myeloperoxidase(MPO)double-positive cells,indicating efferocytosis.Two 4-week-old male C57BL/6J mice were used to extract BMSCs,and GAS6/BMSCs were constructed through adenovirus transfection and successfully identified.Eighteen 8-week-old male C57BL/6J mice were used to create diabetic full-thickness skin defect wound models and divided into phosphate buffered solution(PBS)group,BMSC group,and GAS6/BMSC group(with 6 mice in each group)according to the random number table method.Immediately after injury,PBS,BMSC single-cell suspension,and GAS6/BMSC single-cell suspension were injected locally into the wounds of the three groups of mice,respectively.The wound healing rates were calculated,and the cell proliferation,capillary density,and efferocytosis were detected at the same time points as the previous experiments.ResultsAt 3,7,14,and 21 days after injury,the wound healing rates of mice in diabetic wound group were significantly lower than those in control wound group(

关 键 词：糖尿病 2型 溃疡 间充质干细胞 巨噬细胞 胞葬作用 生长停滞特异性蛋白6 创面修复 

分 类 号：R73[医药卫生—肿瘤]