摘要
目的通过体外研究探索人脂肪干细胞(hADSC)促进海水浸泡创面愈合的可能机制。方法利用人表皮细胞系HaCaT细胞和人工模拟海水建立海水浸泡创面导致细胞损伤的体外模型。从人脂肪组织分离、培养hADSC并进行鉴定,建立HaCaT细胞与hADSC共培养体系。利用活细胞计数试剂盒(CCK-8)、5-乙炔基-2’-脱氧尿苷(EdU)细胞增殖检测试剂盒与细胞划痕实验等评估HaCaT细胞增殖、迁移能力,并通过蛋白质印迹与实时定量PCR技术检测表皮生长因子受体(EGFR)/细胞外调节蛋白激酶(ERK)信号通路活化情况。结果在添加了10%海水的培养液中,HaCaT细胞的增殖活力已受到明显抑制,与不添加海水培养的细胞相比差异有统计学意义(P<0.05)。利用成功分离的hADSC与HaCaT细胞共培养模型,发现添加10%海水培养的HaCaT细胞增殖与迁移能力均低于不添加海水培养的HaCaT细胞及与hADSC共培养且添加10%海水培养的HaCaT细胞(P均<0.05),而不添加海水培养的HaCaT细胞和与hADSC共培养且添加10%海水培养的HaCaT细胞增殖与迁移能力差异无统计学意义(P>0.05)。添加10%海水培养的HaCaT细胞EGFR/ERK信号通路表达受到抑制,与不添加海水培养的HaCaT细胞及与hADSC共培养且添加10%海水培养的HaCaT细胞相比差异有统计学意义(P<0.05);不添加海水培养的HaCaT细胞和与hADSC共培养且添加10%海水培养的HaCaT细胞EGFR/ERK信号通路表达差异无统计学意义(P>0.05)。结论海水可阻碍EGFR/ERK信号通路的激活、抑制HaCaT细胞的增殖与迁移;hADSC可促进EGFR/ERK信号通路的激活,减轻海水对HaCaT细胞增殖与迁移能力的抑制作用。
Objective To explore the possible mechanism of human adipose-derived stem cells(hADSCs)promoting seawater immersion wound healing in vitro.Methods Human epidermal cell line HaCaT cells and artificially simulated seawater were used to establish an in vitro model of cell damage induced by seawater immersion.hADSCs were isolated from human adipose tissues,and a co-culture system of HaCaT cells and hADSCs was established.The proliferation and migration abilities of HaCaT cells were detected by cell counting kit-8(CCK-8),5-ethynyl-2’-deoxyuridine(EdU)cell proliferation detection kit and cell scratch test.The activation levels of epidermal growth factor receptor(EGFR)/extracellular-regulated protein kinase(ERK)signaling pathway were detected by Western blotting and real-time quantitative PCR.Results The proliferation of HaCaT cells cultured with the medium containing 10%artificial seawater was significantly inhibited compared with the cells cultured without artificial seawater(P<0.05).The proliferation and migration abilities of seawater-cultured HaCaT cells were significantly lower than those cultured without seawater and those with hADSCs and seawater(all P<0.05),but there were no significant differences in cell proliferation or migration abilities between the HaCaT cells cultured without seawater and those co-cultured with hADSCs and seawater(P>0.05).The expression of EGFR/ERK signaling pathway in seawater-cultured HaCaT cells was significantly inhibited compared with the cells cultured without seawater and those cocultured with hADSCs and seawater(P<0.05),while the expression of EGFR/ERK signaling pathway was not significantly different between the HaCaT cells cultured without seawater and those co-cultured with hADSCs and seawater(P>0.05).Conclusion Seawater can block the activation of EGFR/ERK signaling pathway and inhibit the proliferation and migration of HaCaT cells.hADSCs can promote the activation of EGFR/ERK signaling pathway and reduce the inhibition effect of seawater against proliferation and migration
作者
熊佳超
嵇铂尧
王留军
刘知晓
宋建星
XIONG Jia-chao;JI Bo-yao;WANG Liu-jun;LIU Zhi-xiao;SONG Jian-xing(Department of Plastic Surgery,Changhai Hospital,Naval Medical University(Second Military Medical University),Shanghai 200433,China;Department of Histology and Embryology,College of Basic Medical Sciences,Naval Medical University(Second Military Medical University),Shanghai 200433,China)
出处
《第二军医大学学报》
CAS
CSCD
北大核心
2019年第10期1062-1068,共7页
Academic Journal of Second Military Medical University
基金
海军军医大学(第二军医大学)长海医院“1255”学科建设计划(CH125550900)~~
关键词
海水
表皮细胞
人脂肪干细胞
细胞增殖
细胞迁移
seawater
epidermal cells
human adipose-derived stem cells
cell proliferation
cell migration
