N-methyl-N-nitro-N-nitrosoguanidine-mediated ING4 downregulation contributed to the angiogenesis of transformed human gastric epithelial cells
N-methyl-N-nitro-N-nitrosoguanidine-mediated ING4 downregulation contributed to the angiogenesis of transformed human gastric epithelial cells
复制标题
N-甲基-N-硝基-N-亚硝基胍介导的ING4下调有助于转化人胃上皮细胞的血管生成
DOI：
10.1016/j.lfs.2018.02.034
复制
发表时间：
2018-04-15
期刊：
影响因子：
6.1
通讯作者：
Xu Lichun
中科院分区：
文献类型：
--
作者：
Chen Yansu;Fu Rui;Xu Lichun
目的：血管生成与胃癌的进展和死亡率相关。流行病学证据表明，长期接触n -亚硝基化合物（NOCs）是导致胃癌死亡的主要原因。因此，进一步降低胃癌死亡率，需要探索NOCs诱导血管生成的确切机制。生长蛋白4抑制剂（inhibitor of growth protein 4, ING4）是一种肿瘤抑制基因，在病理性血管生成中起重要作用。在本研究中，我们将研究长期低剂量暴露于n -甲基-n -硝基-n -亚硝基胍（MNNG）后人胃上皮细胞中ING4的表达水平，以及MNNG减少ING4对转化细胞血管生成的病理影响。主要方法：采用软琼脂集落形成法、免疫印迹法、免疫荧光法和创面愈合法评价转化细胞的特性。采用HUVEC生长和成管实验检测其血管生成能力。采用EMSA、荧光素酶报告基因检测、实时荧光定量PCR和Western blotting等方法探讨其确切机制。关键发现：通过慢性低剂量处理建立转化的人胃上皮细胞，在mnng诱导的细胞转化后期，观察到ING4逐渐下调。此外，我们发现MNNG暴露降低了ING4的表达，通过增加NF-kappa B p65的磷酸化水平和随后的DAN结合活性，以及调节NF-kappa B p65下游促血管生成基因MMP-2和MMP-9的表达，显著加剧了血管生成。意义：我们的发现为mnng转化的人胃上皮细胞血管生成的机制提供了重要的见解，并支持了ING4可能是胃癌相关治疗靶点的概念。
Aims: Angiogenesis is associated with the progression and mortality of gastric cancer. Epidemiological evidences indicate that long-term N-nitroso compounds (NOCs) exposure predominantly contributes to the mortality of gastric cancer. Therefore, further reduced mortality of gastric cancer demands to explore the exact mechanisms of NOCs induced angiogenesis. As a tumor suppressor gene, inhibitor of growth protein 4 (ING4) plays an important role in pathological angiogenesis. In this study, we will investigate ING4 expression level in human gastric epithelial cells after the long-term low dose exposure of N-methyl-N-nitro-N-nitrosoguanidine (MNNG) and the pathological impact of MNNG-reduced ING4 on angiogenesis of transformed cells.Main methods: The soft agar colony formation assay, Western blotting, immunofluorescence and wound healing assay were used to evaluate the characteristics of transformed cells. HUVEC growth and tube formation assays were performed to test the angiogenic abilities. EMSA, luciferase reporter gene assay, real-time PCR and Western blotting were used to explore the exact mechanism.Key findings: By establishing transformed human gastric epithelial cells via chronic low dose treatment, a gradually ING4 downregulation was observed in the later-stage of MNNG-induced cell transformation. Moreover, we demonstrated that MNNG exposure-reduced ING4 expression significantly resulted into aggravating angiogenesis through increasing the phosphorylation level of NF-kappa B p65 and subsequently DAN binding activity and regulating the expressions of NF-kappa B p65 downstream pro-angiogenic genes, MMP-2 and MMP-9.Significance: Our findings provided a significant mechanistic insight into angiogenesis of MNNG-transformed human gastric epithelial cell and supported the concept that ING4 may be a relevant therapeutic target for gastric cancer.