中性粒细胞来源外泌体对大鼠全层皮肤缺损愈合的影响  

Effects of neutrophil-derived exosomes on full-thickness wound healing in rats

作　　者：李若尘 王长虹 崔毅鹏 刘蔚[1] 张晨[1] LI Ruochen;WANG Changhong;CUI Yipeng;LIU Wei;ZHANG Chen(Department of Osteoarticular Surgery,The Second Affiliated Hospital of Xi’an Jiaotong University,Xi’an 710004,China)

机构地区：[1]西安交通大学第二附属医院骨关节外科,陕西西安710004

出　　处：《西安交通大学学报（医学版）》2022年第6期833-839,共7页Journal of Xi’an Jiaotong University（Medical Sciences）

基　　金：国家自然科学基金面上项目(No.81772346)。

摘　　要：目的探讨中性粒细胞来源外泌体对皮肤缺损愈合的影响。方法①抽取10名健康志愿者的静脉血,分离纯化中性粒细胞,用差速离心法从中性粒细胞上清液中制备外泌体。②制备全层皮肤缺损的大鼠创伤模型,将其随机分为3组:对照组、50μg/mL外泌体组和100μg/mL外泌体组,用CCK8实验测定血管内皮细胞增殖情况,用划痕实验测定血管内皮细胞的迁移情况,探讨中性粒细胞来源外泌体对内皮细胞体外管状结构形成的影响;伤后7、14 d,观察皮肤创面愈合情况,比较不同处理组的创面愈合率;伤后14 d,对各组创面组织进行HE染色和Masson染色,观察胶原纤维的排列情况。结果①中性粒细胞来源外泌体为直径约100 nm的圆形或椭圆形囊泡。②伤后12 h,通过划痕实验测得各组创面中血管内皮细胞愈合率相近(P>0.05)。随着时间延长,24 h时,100μg/mL中性粒细胞来源外泌体处理组愈合率高于对照组(P<0.01)和50μg/mL中性粒细胞来源外泌体处理组(P<0.05);伤后12 h,通过CCK8实验测得,50μg/mL和100μg/mL中性粒细胞来源外泌体处理组血管内皮细胞增殖数量高于对照组(P<0.01),24 h时这两组的结果仍高于对照组(P<0.01)。管腔形成实验结果表明,中性粒细胞来源外泌体能促进血管内皮细胞体外管状结构形成(P<0.01)。③伤后7、14 d,与对照组相比,50、100μg/mL外泌体组的创面愈合率均增高(P<0.01),且100μg/mL外泌体组的创面愈合率高于50μg/mL外泌体组(P<0.01)。伤后14 d,与对照组相比,100μg/mL外泌体组创面新形成的表皮和真皮更长,胶原纤维数量增多且排列方向高度一致。结论中性粒细胞来源外泌体通过促进新生血管内皮细胞的增殖、迁移和胶原纤维形成,从而加速大鼠全层皮肤缺损愈合。Objective To investigate the effects of neutrophil-derived exosomes on skin wound healing.Method①The venous blood was taken from 10 healthy volunteers to separate and purify neutrophils,and exosomes were prepared from neutrophil supernatants using differential centrifugation on Sepharose columns.②The rat trauma model with full-thickness skin defect was prepared and randomly divided into three groups:control group,50μg/mL exosomes group,and 100μg/mL exosomes group.The CCK8 test and the scratch test were used to determine the proliferation and migration of vascular endothelial cells,respectively,which was used to investigate the effects of neutrophil-derived exosomes on the formation of endothelial cell tubular structures in vitro.On day 7 and day 14 after surgery,the skin wound healing was observed and the wound healing rate in different treatment groups was compared.On day 14 after surgery,the wound tissues of each group were stained with HE and Masson to observe the arrangement of collagen fibers.Results①Neutrophil-derived exosomes were round or oval vesicles with diameter about 100 nm.②On post injury hour(PIH)12,the scratch test suggested that wound healing rates in the three groups were close(P>0.05),while wound healing rate of 100μg/mL neutrophil-derived exosomes in the treatment group was higher than that in the control group(P<0.01)and 50μg/mL exosomes group(P<0.05)on PIH 24.On PIH 12,the result of CCK8 assay showed that 50 and 100μg/mL exosomes groups had enhanced proliferative activity of vascular endothelial cells(P<0.01)and the results were still higher in both groups than in the control group on PID 24.The results of tube formation assay showed that neutrophil-derived exosomes could promote the formation of tubular structures of endothelial cells in vitro(P<0.01).③On day 7 and 14 after surgery,compared with the control group,the wound healing rates of the 50 and 100μg/mL exosomes groups were higher(P<0.01).The wound healing rate of 100μg/mL exosomes group was higher than that of 50μg/

关 键 词：中性粒细胞 创伤和损伤 外泌体 伤口愈合 

分 类 号：R622[医药卫生—整形外科]