Rountree C Bart, Ding Wei, He Lina, Stiles Bangyan
Department of Pediatrics, College of Medicine, The Pennsylvania State University, Hershey, Pennsylvania 17033-0850, USA.
Stem Cells. 2009 Feb;27(2):290-9. doi: 10.1634/stemcells.2008-0332.
PTEN (phosphatase and tensin homolog deleted on chromosome 10) is a lipid phosphatase that regulates mitogenic signaling pathways, and deficiency of PTEN results in cell proliferation, survival, and malignancy. Murine liver-specific Pten deletion models develop liver malignancy by 12 months of age. Using this model, we describe a population of CD133+ liver cancer stem cells isolated during the chronic injury phase of disease progression and before primary carcinoma formation. We performed immunohistochemistry and flow cytometry isolation using livers from 3- and 6-month-old Pten(loxP/loxP); Alb-Cre+ mice (mutants) and controls. CD133+CD45- nonparenchymal (NP) cells were analyzed for gene expression profile and protein levels. Single CD133+CD45- oval cells were isolated for clonal expansion and tumor analysis. Cultured and freshly isolated liver CD133+CD45- and CD133-CD45- NP cells were injected into immune-deficient and immune-competent mice. In mutant mice, the NP fraction increased in CD133+CD45- cells in 3- and 6-month-old Pten-deleted animals compared with controls. Clone lines expanded from single CD133+CD45- cells demonstrated consistent liver progenitor cell phenotype, with bilineage gene expression of hepatocyte and cholangiocyte markers. CD133+ cells from expanded clone lines formed robust tumors in immune-deficient and immune-competent mice. Furthermore, freshly isolated CD133+CD45- NP liver cells from 6-month-old mutants formed tumors in vivo, and CD133-CD45- NP cells did not. Consistent with a cancer stem cell phenotype, CD133+ cells demonstrate resistance to chemotherapy agents compared with CD133- cells. CD133+CD45- nonparenchymal cells from chronic injury Pten(loxP/loxP); Alb-Cre+ mice represent a bipotent liver progenitor cell population with cancer stem cell phenotype.
PTEN（第10号染色体缺失的磷酸酶及张力蛋白同源物）是一种脂质磷酸酶，可调节有丝分裂信号通路，PTEN的缺失会导致细胞增殖、存活及恶变。小鼠肝脏特异性Pten缺失模型在12月龄时会发生肝脏恶变。利用该模型，我们描述了一群在疾病进展的慢性损伤阶段且在原发性癌形成之前分离出的CD133⁺肝癌干细胞。我们使用3月龄和6月龄的Pten(loxP/loxP); Alb-Cre⁺小鼠（突变体）及对照小鼠的肝脏进行免疫组织化学和流式细胞术分离。对CD133⁺CD45⁻非实质（NP）细胞进行基因表达谱和蛋白水平分析。分离单个CD133⁺CD45⁻卵圆细胞进行克隆扩增和肿瘤分析。将培养的以及新鲜分离的肝脏CD133⁺CD45⁻和CD133⁻CD45⁻NP细胞注射到免疫缺陷和免疫健全的小鼠体内。在突变小鼠中，与对照相比，3月龄和6月龄Pten缺失动物的CD133⁺CD45⁻细胞中的NP组分增加。从单个CD133⁺CD45⁻细胞扩增出的克隆系表现出一致的肝祖细胞表型，具有肝细胞和胆管细胞标志物的双谱系基因表达。来自扩增克隆系的CD133⁺细胞在免疫缺陷和免疫健全的小鼠中形成了强大的肿瘤。此外，从6月龄突变体新鲜分离的CD133⁺CD45⁻NP肝细胞在体内形成了肿瘤，而CD133⁻CD45⁻NP细胞则没有。与癌症干细胞表型一致，与CD133⁻细胞相比，CD133⁺细胞对化疗药物具有抗性。来自慢性损伤Pten(loxP/loxP); Alb-Cre⁺小鼠的CD133⁺CD45⁻非实质细胞代表了具有癌症干细胞表型的双能肝祖细胞群体。
