Wang Hua-Li, Zhang Shu-Lan, Lu Yan-Ming, Jiang Ji-Yong, Zhu Xiao-Yu
Department of Obstetrics and Gynecology, The Second Affiliated Hospital, China Medical University, Shenyang, Liaoning, 110004, P. R. China.
Ai Zheng. 2006 May;25(5):570-5.
BACKGROUND & OBJECTIVE: Hepatocyte growth factor (HGF) specifically binds to its receptor c-met, activates a complex set of intracellular pathways, and plays important roles in regulating tumor invasion, metastasis, and angiogenesis. HGF and c-met are overexpressed in ovarian cancer. This study was designed to investigate the role of HGF in ovarian cancer invasion and its signal transduction pathway.
HGF-induced invasion of ovarian cancer cell line SKOV3 was analyzed with Boyden chamber invasion assay. The expression changes of c-met and matrix metalloproteinase-9 (MMP-9) in SKOV3 cells before and after treatment with HGF and nuclear factor kappaB (NF-kappaB) inhibitor pyrrolidine dithiocarbamate (PDTC) were measured by real-time reverse transcription-polymerase chain reaction (RT-PCR), Western blot, and flow cytometry. The expression of NF-kappaB in SKOV3 cells was evaluated by immunocytochemistry and Western blot.
The invasive cells were significantly more in HGF group than in control group and PDTC plus HGF group (343+/-13 vs. 167+/-11 and 241+/-10, P<0.01). HGF promoted the expression of MMP-9 mRNA by 5.66+/-0.10 folds, but had no effect on c-met mRNA expression; PDTC suppressed the HGF-driven expression of MMP-9 mRNA by 2.75+/-0.80 folds. The positive rates of c-met and MMP-9 were significantly higher in HGF-treated cells than in control cells [(96.6+/-2.0)% vs. (73.3+/-2.4)%, P<0.01; (74.6+/-4.4)% vs. (16.0+/-2.9)%, P<0.01]. The protein levels of c-met and MMP-9 were significantly higher in HGF-treated cells than in control cells (2.84+/-0.18 vs. 1.65+/-0.19, P<0.01; 2.94+/-0.13 vs. 0.54+/-0.18, P<0.01). PDTC significantly suppressed the HGF-driven expression of MMP-9 protein: the positive rate and protein level of MMP-9 were (25.8+/-3.7)% and 0.87+/-0.14 (P<0.05). HGF promoted the expression of NF-kappaB protein in a time-dependent manner. The expression peak appeared 1 h after treatment with HGF (from 0.42+/-0.11 to 1.16+/-0.21, P<0.01). PDTC significantly inhibited the HGF-driven expression of NF-kappaB protein (0.38+/-0.12, P<0.01).
HGF might stimulate the invasion of SKOV3 cells by up-regulating the expression of MMP-9 via NF-kappaB pathway.
肝细胞生长因子（HGF）特异性结合其受体c-met，激活一系列复杂的细胞内信号通路，在调节肿瘤侵袭、转移及血管生成中发挥重要作用。HGF和c-met在卵巢癌中均呈过表达。本研究旨在探讨HGF在卵巢癌侵袭中的作用及其信号转导通路。
采用Boyden小室侵袭实验分析HGF诱导的卵巢癌细胞系SKOV3的侵袭能力。运用实时逆转录-聚合酶链反应（RT-PCR）、蛋白质印迹法及流式细胞术检测HGF及核因子κB（NF-κB）抑制剂吡咯烷二硫代氨基甲酸盐（PDTC）处理前后SKOV3细胞中c-met和基质金属蛋白酶-9（MMP-9）的表达变化。通过免疫细胞化学和蛋白质印迹法评估SKOV3细胞中NF-κB的表达。
HGF组侵袭细胞数显著多于对照组及PDTC+HGF组（343±13 vs. 167±11和241±10，P<0.01）。HGF使MMP-9 mRNA表达升高5.66±0.10倍，但对c-met mRNA表达无影响；PDTC使HGF驱动的MMP-9 mRNA表达降低2.75±0.80倍。HGF处理组细胞中c-met和MMP-9的阳性率显著高于对照组[（96.6±2.0）% vs. （73.3±2.4）%，P<0.01；（74.6±4.4）% vs. （16.0±2.9）%，P<0.01]。HGF处理组细胞中c-met和MMP-9的蛋白水平显著高于对照组（2.84±0.18 vs. 1.65±0.19，P<0.01；2.94±0.13 vs. 0.54±0.18，P<0.01）。PDTC显著抑制HGF驱动的MMP-9蛋白表达：MMP-9的阳性率和蛋白水平分别为（25.8±3.7）%和0.87±0.14（P<0.05）。HGF以时间依赖性方式促进NF-κB蛋白表达。HGF处理1小时后表达达到峰值（从0.42±0.11升至1.16±0.21，P<0.01）。PDTC显著抑制HGF驱动的NF-κB蛋白表达（0.38±0.12，P<0.01）。
HGF可能通过NF-κB途径上调MMP-9表达，从而促进SKOV3细胞的侵袭。
