Huper Gudrun, Marks Jeffrey R
Duke University Medical Center, Department of Surgery and Institute for Genome Sciences and Policy, Durham, North Carolina, USA.
Cancer Res. 2007 Apr 1;67(7):2990-3001. doi: 10.1158/0008-5472.CAN-06-4065.
Epithelial cells within the normal breast duct seem to be the primary target for neoplastic transformation events that eventually produce breast cancer. Normal epithelial cells are easily isolated and propagated using standard techniques. However, these techniques almost invariably result in populations of cells that are largely basal in character. Because only approximately 20% of human breast cancers exhibit a basal phenotype, our understanding of the disease may be skewed by using these cells as the primary comparator to cancer. Further, because germ line mutations in BRCA1 yield breast cancers that are most often of the basal type, a comparison of normal basal and luminal cells could yield insight into the tissue and cell type specificity of this hereditary cancer susceptibility gene. In this report, we describe a simplified and efficient method for isolating basal and luminal cells from normal human breast tissue. These isogenic cells can be independently propagated and maintain phenotypic markers consistent with their respective lineages. Using these cultured cells, we show that basal and luminal cells exhibit distinct responses to ionizing radiation. Basal cells undergo a rapid but labile cell cycle arrest, whereas luminal cells show a much more durable arrest, primarily at the G(2)-M boundary. Molecular markers, including p53 protein accumulation, p53-activated genes, and BRCA1 nuclear focus formation all correlate with the respective cell cycle responses. Further, we show that short-term cultures of human breast tissue fragments treated with ionizing radiation show a similar phenomenon as indicated by the biphasic accumulation of p53 protein in the basal versus luminal layer. Together, these results indicate that normal basal cells have a transitory cell cycle arrest after DNA damage that may underlie their increased susceptibility to transformation after the loss of functional BRCA1.
正常乳腺导管内的上皮细胞似乎是最终导致乳腺癌的肿瘤转化事件的主要靶点。使用标准技术可以轻松分离和培养正常上皮细胞。然而，这些技术几乎总是产生主要具有基底特征的细胞群体。由于只有约20%的人类乳腺癌表现出基底表型，将这些细胞用作癌症的主要对照可能会使我们对该疾病的理解产生偏差。此外，由于BRCA1的种系突变产生的乳腺癌最常见为基底型，比较正常基底细胞和管腔细胞可能有助于深入了解这种遗传性癌症易感性基因的组织和细胞类型特异性。在本报告中，我们描述了一种从正常人乳腺组织中分离基底细胞和管腔细胞的简化高效方法。这些同基因细胞可以独立培养，并维持与其各自谱系一致的表型标记。使用这些培养的细胞，我们表明基底细胞和管腔细胞对电离辐射表现出不同的反应。基底细胞经历快速但不稳定的细胞周期停滞，而管腔细胞则表现出更持久的停滞，主要在G(2)-M边界。分子标记，包括p53蛋白积累、p53激活基因和BRCA1核灶形成，均与各自的细胞周期反应相关。此外，我们表明，用电离辐射处理的人乳腺组织片段的短期培养显示出类似的现象，基底层与管腔层中p53蛋白的双相积累表明了这一点。总之，这些结果表明，正常基底细胞在DNA损伤后有短暂的细胞周期停滞，这可能是它们在功能性BRCA1缺失后对转化易感性增加的基础。
