Division of Pathology, School of Molecular Medical Sciences, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom.
PLoS One. 2010 May 19;5(5):e10714. doi: 10.1371/journal.pone.0010714.
CD133 is a membrane molecule that has been, controversially, reported as a CSC marker in colorectal cancer (CRC). In this study, we sought to clarify the expression and role of CD133 in CRC. Initially the size of the CD133-expressing (CD133+) population in eight well-described CRC cell lines was measured by flow cytometry and was found to range from 0% to >95%. The cell line HT29 has a CD133+ population of >95% and was chosen for functional evaluation of CD133 after gene knockdown by RNA interference. A time course assay showed that CD133 inhibition had no significant effect on cell proliferation or apoptosis. However, CD133 knockdown did result in greater susceptibility to staurosporine-induced apoptosis (p = 0.01) and reduction in cell motility (p<0.04). Since gene knockdown may cause "off-target" effects, the cell line SW480 (which has a CD133+ population of 40%) was sorted into pure CD133+ and CD133- populations to allow functional comparison of isogenic populations separated only by CD133 expression. In concordance with the knockdown experiments, a time course assay showed no significant proliferative differences between the CD133+/CD133- populations. Also greater resistance to staurosporine-induced apoptosis (p = 0.008), greater cell motility (p = 0.03) and greater colony forming efficiency was seen in the CD133+ population than the CD133- population in both 2D and 3D culture (p<0.0001 and p<0.003 respectively). Finally, the plasticity of CD133 expression in tumour cells was tested. Quantitative PCR analysis showed there was transcriptional repression in the CD133- population of SW480. Prolonged culture of a pure CD133- population resulted in re-emergence of CD133+ cells. We conclude that CD133 expression in CRCs is associated with some features attributable to stemness and that there is plasticity of CD133 expression. Further studies are necessary to delineate the mechanistic basis of these features.
CD133 是一种膜分子，其作为结直肠癌（CRC）中的 CSC 标志物存在争议。在这项研究中，我们试图阐明 CD133 在 CRC 中的表达和作用。最初，通过流式细胞术测量了八个描述良好的 CRC 细胞系中 CD133 表达（CD133+）群体的大小，发现范围从 0%到>95%。细胞系 HT29 的 CD133+群体>95%，并选择其进行 RNA 干扰基因敲低后的 CD133 功能评估。时程测定表明，CD133 抑制对细胞增殖或凋亡没有明显影响。然而，CD133 敲低确实导致对 staurosporine 诱导的凋亡更加敏感（p=0.01），并且细胞迁移能力降低（p<0.04）。由于基因敲低可能导致“脱靶”效应，因此将细胞系 SW480（CD133+群体为 40%）分选为纯 CD133+和 CD133-群体，以允许仅通过 CD133 表达分离的同基因群体进行功能比较。与敲低实验一致，时程测定表明 CD133+/CD133-群体之间没有明显的增殖差异。在 2D 和 3D 培养中，与 CD133-群体相比，CD133+群体中 staurosporine 诱导的凋亡（p=0.008）、细胞迁移能力（p=0.03）和集落形成效率更高（p<0.0001 和 p<0.003）。最后，测试了肿瘤细胞中 CD133 表达的可塑性。定量 PCR 分析显示，SW480 的 CD133-群体存在转录抑制。纯 CD133-群体的延长培养导致 CD133+细胞的重新出现。我们得出结论，CRC 中的 CD133 表达与一些归因于干性的特征相关，并且 CD133 表达具有可塑性。需要进一步的研究来描绘这些特征的机制基础。
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