Du Tao, Qu Ying, Li Jianfang, Li Hao, Su Liping, Zhou Quan, Yan Min, Li Chen, Zhu Zhenggang, Liu Bingya
Shanghai Key laboratory of Gastric Neoplasms, Shanghai Institute of Digestive Surgery, Department of Surgery, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, No 197 Ruijin er Road, Shanghai 200025, China.
Mol Cancer. 2014 May 4;13:100. doi: 10.1186/1476-4598-13-100.
Elevated MELK expression is featured in multiple tumors and correlated with tumorigenesis and tumor development. This study is aimed to investigate the mechanisms of MELK-mediated development of gastric cancer.
MELK expression levels in human gastric cancer were determined by quantitative-PCR and immunohistochemistry. The effect of MELK on cell activity was explored by knockdown and overexpression experiments. Cell growth was measured using the CCK-8 assay. Apoptosis and cell cycle distributions were analyzed by flow cytometry. Migration and invasion were tested using a transwell migration assay. Cytoskeletal changes were analyzed by immunofluorescence. To explore the molecular mechanism and effect of MELK on migration and invasion, Western blotting was used to analyze the FAK/Paxillin pathway and pull down assays for the activity of small Rho GTPases. In vivo tumorigenicity and peritoneal metastasis experiments were performed by tumor cell engraftment into nude mice.
MELK mRNA and protein expression were both elevated in human gastric cancer, and this was associated with chemoresistance to 5-fluorouracil (5-FU). Knockdown of MELK significantly suppressed cell proliferation, migration and invasion of gastric cancer both in vitro and in vivo, decreased the percentages of cells in the G1/G0 phase and increased those in the G2/M and S phases. Moreover, knockdown of MELK decreased the amount of actin stress fibers and inhibited RhoA activity. Finally, knockdown of MELK decreased the phosphorylation of the FAK and paxillin, and prevented gastrin-stimulated FAK/paxillin phosphorylation. By contrast, MELK overexpression had the opposite effect.
MELK promotes cell migration and invasion via the FAK/Paxillin pathway, and plays an important role in the occurrence and development of gastric cancer. MELK may be a potential target for treatment against gastric cancer.
MELK表达升高在多种肿瘤中均有体现，且与肿瘤发生及发展相关。本研究旨在探究MELK介导胃癌发展的机制。
采用定量PCR和免疫组织化学法检测人胃癌中MELK的表达水平。通过敲低和过表达实验探究MELK对细胞活性的影响。使用CCK-8法检测细胞生长情况。通过流式细胞术分析细胞凋亡和细胞周期分布。采用Transwell迁移实验检测细胞迁移和侵袭能力。通过免疫荧光分析细胞骨架变化。为探究MELK对迁移和侵袭的分子机制及影响，采用蛋白质印迹法分析FAK/Paxillin通路，并通过下拉实验检测小Rho GTPases的活性。通过将肿瘤细胞接种到裸鼠体内进行体内致瘤性和腹膜转移实验。
人胃癌中MELK mRNA和蛋白表达均升高，且这与对5-氟尿嘧啶（5-FU）的化疗耐药相关。敲低MELK可显著抑制胃癌细胞在体外和体内的增殖、迁移和侵袭，降低G1/G0期细胞百分比，增加G2/M期和S期细胞百分比。此外，敲低MELK可减少肌动蛋白应力纤维的数量并抑制RhoA活性。最后，敲低MELK可降低FAK和桩蛋白的磷酸化水平，并阻止胃泌素刺激的FAK/桩蛋白磷酸化。相比之下，MELK过表达则产生相反的效果。
MELK通过FAK/Paxillin通路促进细胞迁移和侵袭，在胃癌的发生和发展中起重要作用。MELK可能是胃癌治疗的潜在靶点。
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