Department of Obstetrics and Gynecology, Shimane University School of Medicine, Izumo, Japan.
Oncology. 2010;79(3-4):238-46. doi: 10.1159/000322644. Epub 2011 Mar 3.
In the current study, we investigated the mechanism relating downregulation of mitogen-activated protein kinase kinase-4 (MKK4) expression to the development of endometrial cancer.
MKK4 expression in endometrial cancer was assessed by immunohistochemistry using 87 paraffin-embedded tissue specimens, and clinical data was collected via a retrospective chart review. MKK4 gene knockdown using silencing RNA and an MKK4 gene transfection system was used to assess MKK4 function in tissue samples of endometrial cancer.
Lower expression of MKK4 immunointensity was observed in 63.2% (55/87) of the analyzed tumors. High-grade endometrioid adenocarcinoma (G2 and G3) (p = 0.024), postmenopausal status (p = 0.018), and patient age (≥ 60) (p = 0.012) were significantly correlated with lower MKK4 expression. Patients with lower MKK4 expression in endometrial cancer tissues tended to have a shorter overall survival (p = 0.197). Using cell growth and anchorage-independent assays, we determined that both the growth and colony-forming ability of MKK4-transfected HEC1B cells, a line with a low endogenous expression of MKK4, were significantly reduced compared to control vector-transfected cells. Overexpression of the MKK4 gene in HEC1B cells resulted in reduced cell migration activity in a simulated wound healing assay. To confirm that MKK4 expression is related to tumor suppressor function, we used 2 independent but complementary approaches. MKK4 gene knockdown in JHEM1 cells, which overexpressed MKK4, increased proliferation activity. Additionally, the engineered expression of MKK4 in Ishikawa cells, a line with low endogenous MKK4 expression, produced a phenotype similar to that of HEC1B. Similar results were produced in tumor xenografts in nude mice.
These results indicate that MKK4 acts as a tumor suppressor, and reduced expression of MKK4 may contribute to the development of endometrial cancer.
本研究旨在探讨丝裂原活化蛋白激酶激酶 4（MKK4）表达下调与子宫内膜癌发生发展之间的关系。
采用免疫组化法检测 87 例石蜡包埋组织标本中的 MKK4 表达情况，并通过回顾性病历分析收集临床资料。采用沉默 RNA 技术和 MKK4 基因转染系统下调 MKK4 基因表达，评估 MKK4 基因在子宫内膜癌组织中的功能。
在 87 例分析的肿瘤中，有 63.2%（55/87）的肿瘤 MKK4 免疫强度表达较低。高级别子宫内膜样腺癌（G2 和 G3）（p=0.024）、绝经后状态（p=0.018）和患者年龄（≥60 岁）（p=0.012）与较低的 MKK4 表达显著相关。在子宫内膜癌组织中表达较低的 MKK4 的患者总生存时间更短（p=0.197）。通过细胞生长和非锚定依赖性测定，我们发现与转染对照载体的细胞相比，MKK4 低表达的 HEC1B 细胞的生长和集落形成能力显著降低。在 HEC1B 细胞中转染 MKK4 基因后，在模拟伤口愈合试验中细胞迁移活性降低。为了证实 MKK4 表达与肿瘤抑制功能有关，我们采用了 2 种独立但互补的方法。在过表达 MKK4 的 JHEM1 细胞中敲低 MKK4 基因可增加增殖活性。此外，在 Ishikawa 细胞（一种内源性 MKK4 表达较低的细胞系）中构建 MKK4 的表达，产生了与 HEC1B 相似的表型。在裸鼠肿瘤异种移植中也得到了类似的结果。
这些结果表明 MKK4 作为一种肿瘤抑制因子发挥作用，MKK4 表达下调可能导致子宫内膜癌的发生。
