摘要
【目的】研究间充质干细胞(MSC)是否可以通过诱导肺泡巨噬细胞向M2型极化来改善急性肺损伤。【方法】①贴壁法提取脐带MSC,用流式细胞仪分析细胞表型,细胞化学染色法观察细胞体外成骨和成脂肪能力。②小鼠肺泡巨噬细胞系(MH-S细胞)加入脂多糖(LPS)刺激,和MSC隔离共培养,并加入MSC可溶性因子抑制剂,设置LPS组、LPS+MSC组和MSC抑制剂组,共培养48 h后用流式和qPCR技术分析巨噬细胞极化状态。③30只balb/c雄性小鼠随机均分成control组、ALI组、ALI+MSC组(均n=10),LPS滴鼻法建立小鼠急性肺损伤(ALI)模型,用MSC治疗48 h后,肺组织苏木素-伊红(HE)染色进行损伤评估;提取肺泡灌洗液(BALF),其中细胞用流式和qPCR检测M2型巨噬细胞marker CD206、IL-10和Arg1的表达;ELISA检测BALF上清M1型巨噬细胞marker TNF-α。【结果】MSC呈贴壁生长,具有成骨成脂分化能力。MSC可以诱导MH-S细胞向M2型极化,CD206阳性比例明显升高(P <0.05),前列腺素E2(PGE2)抑制剂可以逆转该作用。小鼠ALI模型成功,MSC治疗后,肺组织病理及评分明显减轻。BALF中巨噬细胞CD206阳性细胞的比例,ALI+MSC组较ALI组明显升高。在CD206和IL-10的mRNA水平上,ALI+MSC组高于ALI组(P <0.05)。BALF中炎症因子TNF-α浓度ALI+MSC组较ALI组明显降低(P <0.05)。【结论】脐带MSC通过PGE2诱导巨噬细胞向M2型极化途径有效缓解LPS诱导的小鼠急性肺损伤。
【Objective】To investigate whether mesenchymal stem cells(MSC)can alleviate acute lung injury by inducing alveolar macrophages to polarize to M2 phenotype.【Methods】Umbilical cord MSC was extracted by adherent method and cell phenotypes were analyzed by flow cytometry. The differentiation along osteogenic and adipogenic pathways were assessed by histological staining in vitro. Mouse alveolar macrophage cell line(MH-S cells)which was stimulated by LPS was isolated co-culture with MSC and MSC soluble factor inhibitor was added. We set up three groups(LPS,LPS+MSC,and MSC inhibitor). After being cultured for 48 hours,the macrophage polarization was analyzed by flow cytometry and qPCR. Thirty balb/c male mice were randomly divided into control group(n = 10),ALI group(n = 10),and ALI+MSC group(n = 10). LPS was instilled intranasally to establish acute lung injury model in mice. After treatment with MSC for 48 hours,HE staining of lung tissue was performed for damage assessment. The alveolar lavage fluid(BALF)was obtained and the cells in BALF were analyzed by flow cytometry and qPCR to detect the expression of M2-type macrophage markers including CD206,IL10 and Arg1. The concentration of M1-type macrophage marker TNF-αin the supernatant was measured by ELISA.【Results】MSC showed adherent growth and had the ability of osteogenic and adipogenic differentiation. MSC can induce MH-S cells to polarize to M2 type and with a significant increase of CD206 positive proportion cells(P<0.05). Prostaglandin E2(PGE2) inhibitors can reverse this effect. Mouse ALI model was successful. After treatment with MSC,the pathology and lung injury score was significantly improved. The proportion of CD206 positive macrophages in alveolar lavage fluid in ALI+MSC group was significantly higher than that in ALI group. The expression of CD206 and IL-10 in mRNA level was significantly higher in ALI+MSC group than that in ALI group. The concentration of inflammatory cytokine TNF-α in alveolar lavage fluid was significantly lower in the ALI+MSC group than in the ALI group(P<0.05).【Conclusion】Umbilical cord mesenchymal stem cells can effectively alleviate acute lung injury induced by LPS in mice via PEG2 to induce macrophage to polarize to M2 type.
作者
孙瑶
吕海金
易小猛
郭俊
易慧敏
SUN Yao;LV Hai-jin;YI Xiao-meng;GUO Jun;YI Hui-min(SICU,The Third Affiliated Hospital,Sun Yat-sen University,Guangzhou 510630,China)
出处
《中山大学学报(医学版)》
CAS
CSCD
北大核心
2019年第3期393-400,共8页
Journal of Sun Yat-Sen University:Medical Sciences
基金
广东省科技计划项目(2014A020211010)