Ming Mei, Yu Jie-ping, Zhou Yan-hong, Cao Jun, Song Wen-chong, Yu Hong-gang, Luo He-sheng
Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, China.
Zhonghua Zhong Liu Za Zhi. 2007 Jan;29(1):4-8.
To study the effect of gastrin on the mRNA and protein expression of urokinase-type plasminogen activator (u-PA) in human colon cancer cells and detect the role of p38 MAPK in this process.
Lipofectin method was used to transfect pCR3. 1/CCK2R vector expressing gastrin receptor into a colon cancer cell line colo320. Gastrin and gastrin antagonist were used to up-regulate and down-regulate the signaling pathway, respectively. Human colon cancer colo320 cells and colo320/ CCK2,R cells were cultured and then stimulated with gastrin for different time; SB203580 was added into culture medium to prevent p38 kinase pathway before incubating with gastrin; Western blot and RT-PCR were used to examine the u-PA expression. Western blot was employed to detect p38 kinase phosphorylation.
Gastrin increased evidently the mRNA and protein expressions of u-PA and induced p38 kinase phosphorylation in colo320/CCK,R cells time-dependently. However, the extent of enhancement of u-PA and p38 MAPK expression in colo320 cells was much less than that in colo320/CCK2R cells. The gastrin antagonist L-365, 260 showed an effect of competitive inhibition on gastrin-induced u-PA expression and p38 kinase phosphorylation. The inhibitor SB203580 could sufficiently suppress gastrin-induced p38 kinase phosphorylation and significantly attenuate gastrin-induced u-PA mRNA and protein expressions in colo320/ CCK2 R cells in a dose-dependent manner.
Gastrin-gastrin receptor signal transduction pathway can obviously induce u-PA expression in human colon cancer cells via activating the phosphorylation of p38 kinase.
研究胃泌素对人结肠癌细胞中尿激酶型纤溶酶原激活剂（u-PA）mRNA和蛋白表达的影响，并检测p38丝裂原活化蛋白激酶（p38 MAPK）在此过程中的作用。
采用脂质体法将表达胃泌素受体的pCR3.1/CCK2R载体转染至结肠癌细胞系colo320。分别用胃泌素和胃泌素拮抗剂上调和下调该信号通路。培养人结肠癌colo320细胞和colo320/CCK2,R细胞，然后用胃泌素刺激不同时间；在与胃泌素孵育前，向培养基中加入SB203580以阻断p38激酶通路；采用蛋白质免疫印迹法（Western blot）和逆转录-聚合酶链反应（RT-PCR）检测u-PA表达。用Western blot检测p38激酶磷酸化水平。
胃泌素能明显上调colo320/CCK,R细胞中u-PA的mRNA和蛋白表达，并呈时间依赖性诱导p38激酶磷酸化。然而，colo320细胞中u-PA和p38 MAPK表达的增强程度远低于colo320/CCK2R细胞。胃泌素拮抗剂L-365,260对胃泌素诱导的u-PA表达和p38激酶磷酸化具有竞争性抑制作用。抑制剂SB203580能充分抑制胃泌素诱导的p38激酶磷酸化，并以剂量依赖性方式显著减弱胃泌素诱导的colo320/CCK2 R细胞中u-PA的mRNA和蛋白表达。
胃泌素-胃泌素受体信号转导通路可通过激活p38激酶的磷酸化明显诱导人结肠癌细胞中u-PA的表达。
