Successful growth and characterization of mouse pancreatic ductal cells:: Functional properties of the Ki-RASG12V oncogene
Successful growth and characterization of mouse pancreatic ductal cells:: Functional properties of the Ki-RASG12V oncogene
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DOI：
10.1053/j.gastro.2004.03.058
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发表时间：
2004-07-01
期刊：
影响因子：
29.4
通讯作者：
Rustgi, AK
中科院分区：
文献类型：
--
作者：
Schreiber, FS;Deramaudt, TB;Rustgi, AK
背景与目的：Ki-RAS癌基因在胰腺导管肿瘤中发生改变。从细胞角蛋白19（K：19）-Ki-RAS（G12 V）转基因小鼠和对照同窝仔中纯化胰腺导管细胞（PDC），以鉴定Ki-Ras活化在细胞类型特异性背景下的性质。由于Ki-RAS突变在接受放射治疗的患者中具有预后意义，因此我们研究了Ki-RAS状态对放射生存率的影响。方法：培养Ki-RAS（G12 V）突变或野生型（WT）-Ki-RAS小鼠的胰腺导管片段。生长曲线，电子显微镜，流式细胞仪，信号和细胞周期蛋白的分析建立。法尼基转移酶抑制剂（FTI）在照射前后用R115777处理以确定Ki-Ras法尼基化对细胞存活的影响。结果如下：来自WT和K19-Ki-RAS（G12 V）小鼠的PDCs具有导管细胞的特征，在胶原蛋白上形成三维结构，在形态、生长和细胞周期分布上没有差异。这可能是由于Ki-RAS（G12 V）PDCs中p16 INK 4和P27（Kip 1）的上调以及丝裂原活化蛋白激酶途径的过度刺激的缺乏。未观察到Ki-RAS（G12 V）PDC和WT PDC之间的辐射存活率差异。然而，与用P53（V143 A）转导的WT PDC相比，表达突变体p53（V143 A）的Ki-RASG 12 V PDC具有增强的存活。R115777处理致敏Ki-RAS（G12 V）PDC和Ki-RAS（G12 V）/p53（V143 A）PDC，但不致敏WT PDC。结论：描述了鼠WT PDC和Ki-RAS（G12 V）PDC的新表征。细胞周期调节因子的诱导和丝裂原活化蛋白激酶过度刺激的缺乏可能是抑制激活的Ki-RAS（G12 V）介导的增殖的原因。由于其激活是FTI致敏所必需的，因此R115777可能对表达致癌Ki-Ras的胰腺肿瘤有用。
Background & Aims: The Ki-RAS oncogene is altered in pancreatic ductal neoplasms. Pancreatic ductal cells (PDCs) were purified from cytokeratin 19 (K:19)-Ki-RAS(G12V) transgenic mice and control littermates to identify properties of Ki-Ras activation in a cell-type-specific context. Because Ki-RAS mutation has prognostic significance in patients treated with radiation, we studied the influence of Ki-RAS status on radiation survival. Methods: Pancreatic ductal fragments from mice with Ki-RAS(G12V) mutation or wild-type (WT)-Ki-RAS were cultured. Growth curves, electron microscopy, flow cytometry, and analysis of signaling and cell-cycle proteins were established. Farnesyltransferase inhibitor (FTI) treatment with R115777 before and after irradiation was used to determine the effect of Ki-Ras farnesylation on cell survival. Results: PDCs from WT and K19-Ki-RAS(G12V) mice had features of ductal cells with formation of 3-dimensional structures on Collagen without differences in morphology, growth, and cell-cycle distribution. This may result from up-regulation of p16INK4 and P27(Kip1) and lack of hyperstimulation of the mitogen-activated protein kinase pathway in Ki-RAS(G12V) PDCs. No differences in radiation survival between Ki-RAS(G12V) PDCs and WT PDCs were observed. However, Ki-RASG12V PDCs expressing mutant p53(V143A) had enhanced survival compared with WT PDCs transduced with P53(V143A). R115777 treatment sensitized Ki-RAS(G12V) PDCs and Ki-RAS(G12V)/p53(V143A) PDCs, but not WT PDCs. Conclusions: Novel characterization of murine WT PDCs and Ki-RAS(G12V) PDCs is described. Induction of cell-cycle regulators and lack of mitogen-activated protein kinase hyperstimulation likely are responsible for constraining activated Ki-RAS(G12V)-mediated proliferation. Because its activation was required for sensitization by an FTI, R115777 may be useful against pancreatic tumors expressing oncogenic Ki-Ras.