Department of Obstetrics, Gynecology and Reproductive Sciences, Mount Sinai School of Medicine, New York, NY 10029, USA.
Gynecol Oncol. 2011 Oct;123(1):129-37. doi: 10.1016/j.ygyno.2011.06.006. Epub 2011 Jul 22.
Our previous report has implicated the involvement of VEGF-VEGFR-2 h signaling in LPA-induced EOC invasion. However, the mechanism by which LPA regulates VEGF and VEGFR-2 expression remains to be elucidated. In the present study, we systematically examined the signal transduction pathways activated by LPA and further evaluated whether LPA's effect on VEGF-VEGFR-2 signaling and EOC invasion was mediated by the activation of NF-κB pathway.
Using a signal transduction PathwayFinder PCR array, we examined the expression change of 86 key genes representing 18 signal transduction pathways in DOV13 and SKOV3 cells upon LPA (20 μM) treatment. We also used quantitative PCR, Western blotting and ELISA to evaluate the effect of NF-κB pathway inhibition on VEGF(121), VEGF(165) and VEGFR-2 mRNA and protein expression/secretion with or without the presence of LPA (20 μM) in SKOV3. Cell invasion under various treatment conditions was assessed by Matrigel invasion assay and MMP-2 secretion was detected by gelatin zymography.
Our results showed that in both DOV13 and SKOV3, several of the NF-κB pathway components, such as TNF, are consistently activated by LPA stimulation. In addition, treatment with an NF-κB pathway activation inhibitor, at 10 μM, significantly decreased LPA-induced VEGF(121), VEGF(165) and VEGFR-2 mRNA expression and VEGF secretion, as well as LPA-induced SKOV3 invasion (p<0.05). When combined with an EGFR inhibitor, NF-κB pathway inhibition exhibited a significantly stronger effect than used alone (p<0.05) on reducing LPA-induced VEGF secretion and cell invasion. Additionally, NF-κB inhibition also decreased LPA-induced MMP-2 secretion and MMP-1 expression (p<0.05).
These results suggest that the NF-κB pathway plays an important role in LPA-induced VEGF signaling and EOC invasion and targeting this pathway may reveal potential therapeutic options for metastatic EOC.
我们之前的报告表明，VEGF-VEGFR-2h 信号参与了 LPA 诱导的卵巢癌细胞侵袭。然而，LPA 调节 VEGF 和 VEGFR-2 表达的机制仍有待阐明。在本研究中，我们系统地研究了 LPA 激活的信号转导途径，并进一步评估了 LPA 对 VEGF-VEGFR-2 信号和卵巢癌细胞侵袭的影响是否是通过 NF-κB 途径的激活介导的。
使用信号转导途径 Finder PCR 阵列，我们检测了在 LPA（20μM）处理后 DOV13 和 SKOV3 细胞中代表 18 种信号转导途径的 86 个关键基因的表达变化。我们还使用定量 PCR、Western 印迹和 ELISA 来评估 NF-κB 途径抑制对 VEGF(121)、VEGF(165)和 VEGFR-2mRNA 和蛋白表达/分泌的影响，以及在存在或不存在 LPA（20μM）的情况下在 SKOV3 中的作用。通过 Matrigel 侵袭实验评估各种处理条件下的细胞侵袭，通过明胶酶谱法检测 MMP-2 分泌。
我们的结果表明，在 DOV13 和 SKOV3 中，几种 NF-κB 途径成分，如 TNF，均被 LPA 刺激持续激活。此外，用 NF-κB 途径激活抑制剂（10μM）处理可显著降低 LPA 诱导的 VEGF(121)、VEGF(165)和 VEGFR-2mRNA 表达和 VEGF 分泌，以及 LPA 诱导的 SKOV3 侵袭（p<0.05）。当与 EGFR 抑制剂联合使用时，NF-κB 途径抑制对减少 LPA 诱导的 VEGF 分泌和细胞侵袭的作用比单独使用更强（p<0.05）。此外，NF-κB 抑制还降低了 LPA 诱导的 MMP-2 分泌和 MMP-1 表达（p<0.05）。
这些结果表明，NF-κB 途径在 LPA 诱导的 VEGF 信号和卵巢癌细胞侵袭中起重要作用，靶向该途径可能为转移性卵巢癌提供潜在的治疗选择。
