Cell density during differentiation can alter the phenotype of bone marrow-derived macrophages.
Cell density during differentiation can alter the phenotype of bone marrow-derived macrophages.
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DOI：
10.1186/2045-3701-3-30
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发表时间：
2013
影响因子：
7.5
通讯作者：
Hu J
中科院分区：
文献类型：
--
作者：
Lee CM;Hu J
骨髓来源的巨噬细胞(BMDM)是研究巨噬细胞功能的主要细胞。然而，尽管目前有许多可用的方案，但在生成生物计量吸入器方面缺乏显著的共识，可能会模糊比较不同研究或实验室的结果的可靠性。在这项研究中，我们讨论了细胞密度对由此产生的巨噬细胞数量的影响。参照已发表的方法，将野生型C57BL/6小鼠骨髓细胞接种于每10 cm 4 × 10 5或5 × 10 6细胞，在20%L细胞条件培养液中培养7天后，分析细胞表面标志、促炎细胞因子的产生和对极化信号的反应性。低密度组CD11b+F4/80+巨噬细胞(97.28 ± 0.52%)，以Ly-6C-Ly-6G-和c-fms+为主；高密度组CD11b+F4/80+CD11b+F4/80+CD11c+(68.72 ± 2.52%)和Ly-6C-Ly-6G+(71.10 ± 0.90%)。培养密度越高的骨髓基质细胞分泌的促炎细胞因子IL-6、IL-12和肿瘤坏死因子-α也越少，吞噬能力也越弱，在内毒素和IL-4刺激下，M1和M2相关基因的表达模式也不同。总体而言，我们的发现表明，在BMDM分化过程中改变细胞密度可以产生不同的巨噬细胞群，这可能会改变功能研究的结果。
Bone marrow-derived macrophages (BMDMs) are widely used primary cells for studying macrophage function. However, despite numerous protocols that are currently available, lack of a notable consensus on generating BMDMs may obscure the reliability in comparing findings from different studies or laboratories. In this study, we addressed the effect of cell density on the resulting macrophage population. With reference to previously published methods, bone marrow cells from wild type C57BL/6 mice were plated at either 4 × 105 cells or 5 × 106 cells per 10 cm and cultured in 20% L-cell conditioned media for 7 days, after which they were analyzed for cell surface markers, production of proinflammatory cytokines, and responsiveness to polarizing signals. Reproducibly, cells plated at lower density gave a pure population of CD11b+F4/80+ macrophages (97.28 ± 0.52%) with majority being Ly-6C-Ly-6G- and c-Fms+, while those plated at higher density produced less CD11b+F4/80+ cells and a considerably higher proportion of CD11b+F4/80+CD11c+ (68.72 ± 2.52%) and Ly-6C-Ly-6G+ (71.10 ± 0.90%) cells. BMDMs derived from higher plating density also secreted less proinflammatory cytokines such as IL-6, IL-12 and TNF-α and were less phagocytic, and had a different pattern of expression for M1- and M2-related genes upon LPS or IL-4 stimulation. Overall, our findings indicate that altering cell density during BMDM differentiation can give rise to distinct macrophage populations that could vary the outcome of a functional study.
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影响因子：
3.2
作者：
Francke, Alexander;Herold, Joerg;Braun-Dullaeus, Ruediger C.
通讯作者：
Braun-Dullaeus, Ruediger C.
DOI：
10.1038/nri3073
发表时间：
2011-10-14
期刊：
Nature reviews. Immunology
影响因子：
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作者：
通讯作者：
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DOI：
10.1101/pdb.prot5080
发表时间：
2008-12-01
期刊：
CSH protocols
影响因子：
--
作者：
Weischenfeldt, Joachim;Porse, Bo
通讯作者：
Porse, Bo
DOI：
10.1007/978-1-61779-527-5_12
发表时间：
2012-01-01
期刊：
LEUCOCYTES: METHODS AND PROTOCOLS
影响因子：
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作者：
Manzanero, Silvia
通讯作者：
Manzanero, Silvia
DOI：
10.1038/nri2506
发表时间：
2009-03
期刊：
Nature reviews. Immunology
影响因子：
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作者：
通讯作者：
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