Department of Surgical Oncology and General Surgery, First Hospital of China Medical University, Shenyang City, China.
Oncogene. 2012 Mar 15;31(11):1398-407. doi: 10.1038/onc.2011.340. Epub 2011 Aug 8.
Aberrant expression of miR-335 has been frequently reported in cancer studies, suggesting that there is a close correlation between miR-335 and cancer during its development, progression, metastasis and prognosis. The expression of miR-335 in gastric cancer and its effects are not known. Relative expression of miR-335 in 4 gastric cancer cell lines and in 70 gastric cancer tissues was confirmed by real-time quantitative reverse transcriptase-PCR compared with controls. Transwell cell migration and Matrigel invasion assay in vitro and metastasis formation assay in vivo were used to examine the effects of miR-335 expression on gastric cancer cell invasion and metastasis. The effect of miR-335 expression on gastric cancer cell proliferation was estimated by the 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. Luciferase reporter assay and western blot were used to examine the potential target genes and related pathways. Gene silencing with small-interfering RNA was used to examine the effects of target genes on gastric cancer cell invasion. miR-335 was dramatically downregulated in gastric cancer cell lines than in the normal gastric cell line GES-1. Low expression of miR-335 was significantly associated with lymph-node metastasis, poor pT stage, poor pN stage and invasion of lymphatic vessels. Overexpression of miR-335 suppressed gastric cancer cell invasion and metastasis in vitro and in vivo, but has no significant effects on cell proliferation. Furthermore, miR-335 might suppress gastric cancer invasion and metastasis by targeting Bcl-w and specificity protein 1 (SP1). Taken together, our results provide evidence that miR-335 might function as a metastasis suppressor in gastric cancer by targeting SP1 directly and indirectly through the Bcl-w-induced phosphoinositide 3-kinase-Akt-Sp1 pathway. miR-335 showing altered expression at different stages of gastric cancer could be a target for gastric cancer therapies and could be further developed as a potential prognostic factor.
miR-335 的异常表达在癌症研究中经常被报道，这表明 miR-335 在癌症的发展、进展、转移和预后过程中与癌症密切相关。miR-335 在胃癌中的表达及其作用尚不清楚。通过实时定量逆转录聚合酶链反应（real-time quantitative reverse transcriptase-PCR）与对照组相比，证实了 miR-335 在 4 种胃癌细胞系和 70 种胃癌组织中的相对表达。体外 Transwell 细胞迁移和 Matrigel 侵袭实验以及体内转移形成实验用于检测 miR-335 表达对胃癌细胞侵袭和转移的影响。通过 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide）实验评估 miR-335 表达对胃癌细胞增殖的影响。荧光素酶报告基因实验和 Western blot 用于检测潜在的靶基因和相关途径。用小干扰 RNA 基因沉默实验检测靶基因对胃癌细胞侵袭的影响。miR-335 在胃癌细胞系中的表达明显低于正常胃细胞系 GES-1。miR-335 的低表达与淋巴结转移、不良 pT 分期、不良 pN 分期和淋巴管浸润显著相关。过表达 miR-335 可抑制胃癌细胞在体外和体内的侵袭和转移，但对细胞增殖无显著影响。此外，miR-335 可能通过靶向 Bcl-w 和特异性蛋白 1（SP1）抑制胃癌的侵袭和转移。综上所述，我们的研究结果表明，miR-335 可能通过直接和间接靶向 Bcl-w 诱导的磷脂酰肌醇 3-激酶-Akt-Sp1 通路抑制 SP1 来发挥胃癌转移抑制因子的作用。miR-335 在胃癌不同阶段的表达改变可能成为胃癌治疗的靶点，并可进一步开发为潜在的预后因素。
