Luo Jiing-Chyuan, Chi Chin-Wen, Lin Hsiao-Yi, Chang Full-Young, Lu Ching-Liang, Chen Chih-Yen, Lee Shou-Dong
Division of Gastroenterology, Department of Medicine, Taipei Veterans General Hospital, National Yang-Ming University, Taipei, Taiwan 11217.
J Pharmacol Exp Ther. 2007 Feb;320(2):687-94. doi: 10.1124/jpet.106.113035. Epub 2006 Oct 31.
Epidermal growth factor (EGF) is essential to heal gastric ulcers, whereas glucocorticoid delays rat gastric ulcer healing. We found that dexamethasone inhibited EGF-stimulated rat gastric epithelial cell (RGM-1) proliferation by cell count and DNA synthesis analysis of flow cytometry and attempted to elucidate the possible mechanistic pathway via Western blot analysis. EGF (10 ng/ml) treatment for 24 h significantly increased RGM-1 cell proliferation, and dexamethasone (10(-8) and 10(-6) M) markedly suppressed EGF-stimulated cell proliferation. Western blotting results demonstrated that the phosphorylated extracellular signal-regulated kinase (pERK) (pERK1/pERK2) significantly increased at 10 min after EGF treatment. This was followed by increase of cyclooxygenase (COX)-2 expression at 3 h after EGF treatment. The continued increase of COX-2 (up to 18 h) resulted in increased intracellular prostaglandin E(2) and cyclin D1 expression significantly after 8 and 12 h of EGF treatment. Dexamethasone substantially reduced EGF-stimulated COX-2 expression at 3 and 6 h and cyclin D1 expression at 8 and 12 h. Pretreatment of RGM-1 cells with dexamethasone or 2'-amino-3'-methoxyflavone (PD98059)-mitogen-activated protein kinase kinase inhibitor (5 x 10(-5) M) significantly reduced EGF-stimulated pERK1/pERK2 expression. Simultaneous treatment of RGM-1 cells with PD98059 and EGF also markedly decreased EGF-stimulated COX-2 expression at 6 h. These findings indicate that dexamethasone significantly suppresses EGF-stimulated gastric epithelial cell proliferation, and one of the pathways involved is via inhibiting activation of ERK1/ERK2, followed by inhibition of COX-2, cyclin D1 expression, and finally DNA synthesis.
表皮生长因子（EGF）对胃溃疡的愈合至关重要，而糖皮质激素会延迟大鼠胃溃疡的愈合。我们通过流式细胞术的细胞计数和DNA合成分析发现，地塞米松抑制了EGF刺激的大鼠胃上皮细胞（RGM-1）增殖，并试图通过蛋白质印迹分析阐明可能的机制途径。用EGF（10 ng/ml）处理24小时可显著增加RGM-1细胞增殖，而地塞米松（10⁻⁸和10⁻⁶ M）则明显抑制EGF刺激的细胞增殖。蛋白质印迹结果表明，EGF处理10分钟后，磷酸化细胞外信号调节激酶（pERK）（pERK1/pERK2）显著增加。随后，EGF处理3小时后环氧合酶（COX）-2表达增加。COX-2的持续增加（直至18小时）导致EGF处理8小时和12小时后细胞内前列腺素E₂和细胞周期蛋白D1表达显著增加。地塞米松在3小时和6小时时显著降低了EGF刺激的COX-2表达，在8小时和12小时时降低了细胞周期蛋白D1表达。用地塞米松或2'-氨基-3'-甲氧基黄酮（PD98059）——丝裂原活化蛋白激酶激酶抑制剂（5×10⁻⁵ M）预处理RGM-1细胞可显著降低EGF刺激的pERK1/pERK2表达。用PD98059和EGF同时处理RGM-1细胞也显著降低了6小时时EGF刺激的COX-2表达。这些发现表明，地塞米松显著抑制EGF刺激的胃上皮细胞增殖，其中一条涉及的途径是通过抑制ERK1/ERK2的激活，随后抑制COX-2、细胞周期蛋白D1表达，最终抑制DNA合成。
