DesRochers Teresa M, Kuo Ivana Y, Kimmerling Erica P, Ehrlich Barbara E, Kaplan David L
Department of Biomedical Engineering, Tufts University, Medford, Massachusetts, United States of America.
Departments of Pharmacology and of Cellular and Molecular Physiology, School of Medicine, Yale University, New Haven, Connecticut, United States of America.
PLoS One. 2015 Mar 20;10(3):e0120097. doi: 10.1371/journal.pone.0120097. eCollection 2015.
Mycoplasma contamination of cell cultures is a pervasive, often undiagnosed and ignored problem in many laboratories that can result in reduced cell proliferation and changes in gene expression. Unless contamination is specifically suspected, it is often undetected in two dimensional (2D) cultures and the resulting effects of mycoplasma contamination are rarely appreciated and can lead to incorrect conclusions. Three dimensional (3D) tissue cultures are increasingly utilized to explore tissue development and phenotype. However, 3D cultures are more complex than 2D cell cultures and require a more controlled cellular environment in order to generate structures necessary to mimic in vivo responses and are often maintained for longer time periods. Changes to the microenvironment are assumed to have a more extreme effect upon the success of 3D tissue cultures than 2D cell cultures, but the effects of mycoplasma have not been studied. To test this hypothesis, we grew 2D cell cultures and 3D tissues from pig kidney epithelial cells (LLC-PK1) that were contaminated with mycoplasma and the same stock of cells after mycoplasma removal. We did not observe an effect of mycoplasma contamination on proliferation in 2D monolayer cell culture. However, cyst formation in 3D tissues was altered, with effects upon the number, size and structure of cysts formed. These data serve to reinforce the necessity of testing cell stocks for mycoplasma contamination.
细胞培养中的支原体污染是许多实验室普遍存在的问题，常常未被诊断和忽视，它会导致细胞增殖减少和基因表达变化。除非特别怀疑有污染，否则在二维（2D）培养中通常检测不到支原体，支原体污染产生的影响也很少得到重视，可能会导致错误的结论。三维（3D）组织培养越来越多地用于探索组织发育和表型。然而，3D培养比2D细胞培养更复杂，需要更可控的细胞环境才能生成模拟体内反应所需的结构，并且通常要维持更长时间。人们认为微环境的变化对3D组织培养成功的影响比对2D细胞培养的影响更大，但支原体的影响尚未得到研究。为了验证这一假设，我们用受支原体污染的猪肾上皮细胞（LLC-PK1）以及去除支原体后的同一批细胞培养了2D细胞培养物和3D组织。我们没有观察到支原体污染对2D单层细胞培养中细胞增殖的影响。然而，3D组织中的囊肿形成发生了改变，对形成的囊肿数量、大小和结构产生了影响。这些数据进一步证明了检测细胞株是否受到支原体污染的必要性。
