不同条件下小鼠肺泡巨噬细胞体外培养的实验研究OA北大核心CSTPCD
Experimental study of mouse alveolar macrophages cultured in vitro under different conditions
目的:应用文献报道的肺泡巨噬细胞(AMs)体外培养方法,利用GM-CSF、TGF-β和PPAR-γ激动剂罗格列酮(简称GTR)培养体系,分析不同年龄小鼠、不同来源前体细胞产生AM样细胞或AMs的特征.方法:收取不同周龄小鼠支气管肺泡灌洗液(BALF)、骨髓单个核细胞以及胚胎15~17 d小鼠肝脏单个核细胞,种入12孔板,置于GTR体系中培养.流式细胞术检测产出细胞AM表面标志分子F4/80、CD11b、CD170和CD11c表达.结果:培养后,光镜下BALF来源细胞基本呈圆形,骨髓来源细胞近半呈圆形,胎肝来源细胞85%左右呈圆形.4周龄小鼠BALF-AMs比例接近90%,比例和数量均随小鼠年龄增加而增加,12周龄小鼠BALF-AMs数量接近1×106个.4周龄小鼠骨髓源AM样细胞比例不到20%,8周龄和12周龄小鼠比例为40%~45%,数量随小鼠年龄增加而增加,12周龄小鼠骨髓源AM样细胞数量约为0.8×106个/孔.胎肝源AM样细胞比例约为84%,1个孔产出的AM样细胞约为4×106个.结论:不同年龄小鼠、不同来源前体细胞产生AM样细胞或AMs的纯度和数量有较大差别,应根据具体研究要求选择合适方法.
Obiective:To compare characteristics of alveolar macrophages(AMs)or AM-like cells in vitro cultured from different progenitors in mice of different ages in presence of GM-CSF,TGF-β and PPAR-γ agonist rosiglitazone(in brief,GTR),according to a previously reported optimized culture procedure.Methods:After bronchoalveolar lavage fluid(BALF)and bone marrow were isolated from mice of different ages and fetal livers were isolated from embryonic-day-15~17 mice,mononuclear cells were seeded into 12-well plates and cultured in presence of GTR.Generated cells were subjected to flow cytometry analysis to analyze expressions of AM markers F4/80,CD11b,CD170 and CD11c.Results:After culture,almost all BALF-derived cells were round-shaped under light microscope,about half bone marrow-derived cells appeared round-shaped,and about 85% fetal liver-derived cells showed round-shaped morphology.Frequency of BALF-AMs from 4-week-old mice was nearly 90%.Frequency and number of BALF-AMs increased with age of mice.Number of BALF-AMs from 12-week-old mice was nearly 1×106.Frequency of bone marrow-derived AM-like cells from 4-week-old mice was lower than 20% and increased to about 40%~45% for 8-week-old and 12-week-old mice.Number of bone marrow-derived AM-like cells increased with age of mice,and about 0.8×106 cells/well bone marrow-derived AM-like cells were produced from 12-week-old mice.Frequency of fetal liver-derived AM-like cells was around 84% and their number could reach 4×106 cells/well.Conclusion:Purity and number of AMs or AM-like cells in vitro cultured from different progenitors in mice of different ages vary dramatically.Detailed condition should be chosen according to specific study requirements.
云宏芳;蒋慧;牛春晓;张纪岩
军事医学研究院军事认知与脑科学研究所,北京 100850陆军军医大学西南医院急诊科,重庆 400038
基础医学
肺泡巨噬细胞体外培养肺泡灌洗液骨髓胎肝
Alveolar macrophagesIn vitro cultureBronchoalveolar lavage fluidBone marrowFetal liver
《中国免疫学杂志》 2024 (006)
1121-1125 / 5
国家自然科学基金(81930027,92169207).
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