Calvert Richard J, Tepper Shirley, Kammouni Wafa, Anderson Lucy M, Kritchevsky David
Division of Research and Applied Technology, Office of Nutritional Products, Labeling, and Dietary Supplements, Center for Food Safety and Applied Nutrition, US Food and Drug Administration, College Park, MD 20740, USA.
Biochem Pharmacol. 2006 Dec 15;72(12):1749-55. doi: 10.1016/j.bcp.2006.08.026. Epub 2006 Sep 3.
Our previous work established that hypocholesterolemic agents altered K-ras intracellular localization in lung. Here, we examined K-ras activity to define further its potential importance in lung carcinogenesis. K-ras activity in lungs from male A/J, Swiss and C57BL/6 mice was examined. For 3 weeks, mice consumed either 2 or 4% cholestyramine (CS), 1% niacin, 5% konjac mannan (KM), or were injected with lovastatin 25mg/kg three or five times weekly (Lov-3X and Lov-5X). A pair-fed (PF) group was fed the same quantity of diet consumed by the Lov-5X mice to control for lower body weights in Lov-5X mice. After 3 weeks, serum cholesterol was assayed with a commercial kit. Activated K-ras protein from lung was affinity precipitated with a Raf-1 ras binding domain-glutathione-S-transferase fusion protein bound to glutathione-agarose beads, followed by Western blotting, K-ras antibody treatment, and chemiluminescent detection. Only KM reduced serum cholesterol (in two of three mouse strains). In C56BL/6 mice treated with Lov-3X, lung K-ras activity increased 1.8-fold versus control (p=0.009). In normal lung with wild-type K-ras, this would be expected to be associated with maintenance of differentiation. In A/J mice fed 4% CS, K-ras activity increased 2.1-fold (p=0.02), which might be responsible for the reported enhancement of carcinogenesis in carcinogen-treated rats fed CS. KM feeding and PF treatment had no significant effects on K-ras activity. These data are consistent with the concept that K-ras in lung has an oncogenic function when mutated, but may act as a tumor suppressor when wild-type.
我们之前的研究表明，降胆固醇药物会改变肺组织中K-ras的细胞内定位。在此，我们检测了K-ras的活性，以进一步明确其在肺癌发生中的潜在重要性。我们检测了雄性A/J、瑞士和C57BL/6小鼠肺组织中的K-ras活性。连续3周，小鼠分别食用2%或4%的消胆胺（CS）、1%的烟酸、5%的魔芋甘露聚糖（KM），或者每周注射3次或5次25mg/kg的洛伐他汀（Lov-3X和Lov-5X）。设立一对进食量相同的对照组（PF），该组小鼠的进食量与Lov-5X组小鼠相同，以控制Lov-5X组小鼠因体重下降产生的影响。3周后，使用商业试剂盒检测血清胆固醇水平。用与谷胱甘肽琼脂糖珠结合的Raf-1 ras结合域-谷胱甘肽-S-转移酶融合蛋白亲和沉淀肺组织中的活化K-ras蛋白，随后进行蛋白质免疫印迹、K-ras抗体处理和化学发光检测。只有KM降低了血清胆固醇水平（在三种小鼠品系中的两种中）。在接受Lov-3X治疗的C56BL/6小鼠中，肺组织K-ras活性相较于对照组增加了1.8倍（p=0.009）。在具有野生型K-ras的正常肺组织中，这有望与维持细胞分化相关。在喂食4% CS的A/J小鼠中，K-ras活性增加了2.1倍（p=0.02），这可能是之前报道的在喂食CS的致癌物处理大鼠中致癌作用增强的原因。喂食KM和PF处理对K-ras活性没有显著影响。这些数据与以下概念一致：肺组织中的K-ras在发生突变时具有致癌功能，但在野生型时可能作为肿瘤抑制因子发挥作用。
