摘要
目的通过观察人羊膜间充质干细胞(hAMSCs)对松木屑烟雾溶液致大鼠肺微血管内皮细胞(PMVECs)损伤后增殖、凋亡及炎症反应的影响,探讨hAMSCs对PMVECs的保护作用。方法分离、培养hAMSCs和大鼠PMVECs,分别利用流式细胞术和免疫荧光染色法鉴定hAMSCs和PMVECs。实验分组:Control组(正常培养的PMVECs)、Smoke组(用烟雾致伤液致伤PMVECs)、Smoke+hAMSCs组(用烟雾致伤液致伤PMVECs后,在Transwell培养体系中将hAMSCs与PMVECs共培养)。在共培养后12、24 h,用细胞计数试剂盒-8(CCK-8)检测PMVECs增殖活性、流式细胞术检测PMVECs凋亡、ELISA法检测肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)表达水平。结果成功分离、培养hAMSCs和PMVECs,鉴定hAMSCs表面标志物CD105(95.4%)、CD73(99.8%)和CD90(99.8%)呈强阳性表达,CD34、CD45、CD14、CD19和HLA-DR(共为1.96%)呈微弱表达,PMVECs阳性表达血管内皮细胞标志物CD34,且与西非单叶豆凝集素(BSI)结合也为阳性。在共培养12、24 h观察时间点,与Control组比较,Smoke组中PMVECs增殖活性受抑制(P<0.05),细胞凋亡增加(P<0.05),TNF-α和IL-6表达水平上调(P<0.05);Smoke+hAMSCs组中PMVECs细胞的增殖活性受抑制、细胞凋亡增加及炎症因子表达水平上调现象较Smoke组被逆转(P<0.05)。结论在烟雾溶液致伤PMVECs后,hAMSCs可降低PMVECs炎症因子表达,促进其增殖活性并抑制其凋亡,从而对PMVECs起保护作用。
Objective To investigate the protective effect of human amniotic mesenchymal stem cells(hAMSCs)on pulmonary microvascular endothelial cell(PMVECs)through observing the effects of proliferation,apoptosis and inflammatory response after PMVECs injury induced by pine sawdust smoke solution.Methods HAMSCs and rat PMVECs were isolated and cultured.The flow cytometry and immunofluorescence were used to identify hAMSCs and PMVECs respectively.The experimental grouping:control group(normal cultured PMVECs),smoke group(PMVECs injury induced by pine sawdust smoke solution),smoke+hAMSCs group(after PMVECs was injured by smoke solution,hAMSCs and PMVECs were co-cultured in Transwell culture system).The proliferative activity of PMVECs after co-culture for 12,24 h was mea sured by cell counting kit-8(CCK-8),the apoptosis of PMVECs was measured by flow cy tometry,and the expression levels of TNF-αand IL-6 were detected by enzyme-linked immunosorbent assay(ELISA).Results hAMSCs and PMVECs were successfully isolated and cultured,and the hAMSCs surface markers CD105(95.4%),CD73(99.8%)and CD90(99.8%)were identified as strongly positive expression,while CD34,CD45,CD14,CD19 and HLA-DR were weakly expressed(1.96%in total).The vascular endothelial cell marker CD34 in PMVECs was positively expressed,moreover its combination with aggulutinin BSI was also positive.At the observation time point of 12,24 h co-culture,compared with the Control group,the proliferation activity of PMVECs in the Smoke group was inhibited(P<0.05),and the cellular apoptosis was increased(P<0.05),the TNF-αand IL-6 expression levels were up-regulated(P<0.05);the phenomena of PMVECs proliferation activity inhibition,apoptosis increase and inflammatory factor expression level up-regulation in the Smoke+hAMSCs group were reversed compared with the Smoke group(P<0.05).Conclusion After PMVECs are injured by smoke solution,hAMSCs could decrease the PMVECs inflammatory factors expression,promote its proliferation activity and inhibit its apoptosis,thus play the protective effect on PMVECs.
作者
詹球
崔培
朱秀连
李榕生
蒙凤姬
朱富军
杨福旺
童亚林
辛海明
ZHAN Qiu;CUI Pei;ZHU Xiulian;LI Rongsheng;MENG Fengji;ZHU Fujun;YANG Fuwang;TONG Yalin;XIN Haiming(Animal Laboratory,924 Hospital of Joint Logistics Support Force of People′s Liberation Army,Guilin,Guangxi 541002,China;Burn Plastic Skin Beauty Center,924 Hospital of Joint Logistics Support Force of People′s Liberation Army,Guilin,Guangxi 541002,China)
出处
《重庆医学》
CAS
2024年第23期3527-3534,共8页
Chongqing Medical Journal
基金
广西壮族自治区临床重点专科建设项目(桂卫医发〔2023〕26号)
广西科技基地与人才专项(桂科AD18126016)
广西壮族自治区卫生健康委自筹经费科研项目(Z-C20241561)
广西壮族自治区桂林市自筹经费科技项目(20210102z)
联勤保障部队第九二四医院院内科技计划(GS2020CZ05、GS2020CZ06、SG2020FH08)。
关键词
烟雾吸入性损伤
急性肺损伤
人羊膜间充质干细胞
肺微血管内皮细胞
smoke inhalation injury
acute lung injury
human amniotic mesenchymal stem cells
pulmonary microvascular endothelial cells
作者简介
通信作者:辛海明,E-mail:xinhm123@163.com。
