Department of Cardiovascular, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang, China (mainland).
Department of Gynecology and Obstetrics, The Firts Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang, China (mainland).
Med Sci Monit. 2017 Oct 6;23:4789-4797. doi: 10.12659/msm.903661.
BACKGROUND Raf kinase inhibitor protein (RKIP) regulates growth and differentiation and plays a role in key signal transduction cascades in mammalian cells. Nevertheless, the underlying mechanism for which RKIP regulates cell-cell adhesion remains unknown. Our study investigated the function of the RKIP overexpression on adhesion molecules expression induced by tumor necrosis factor (TNF)-α in cultured mouse vascular smooth muscle cells (MOVACs). MATERIAL AND METHODS The expression levels of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) were detected by ELISA kit, reverse transcription-PCR, and western blot assays. The protein expression of RKIP, p65, and inhibitor of nuclear factor (NF)-κBα (IκBα) were detected by western blot analysis. The activity of NF-kappaB was determined using a Dual-Luciferase Reporter assay. RESULTS The results showed that MOVACs transfected with pCMV5-HA-RKIP significantly inhibited TNF-α induced mRNA and protein expression of ICAM-1 and VCAM-1. The adhesion of THP-1 cells was also detected and inhibited by pCMV5-HA-RKIP in TNF-α-treated MOVACs. RKIP also suppressed the TNF-α-induced activation of NF-kappaB and the protein expression of phosphorylated IκB-α, and promoted the protein expression of IkB-α and nuclear translocation of p65 NF-kappaB. Furthermore, RKIP and the inhibitor of NF-kappaB (BAY11-7082) reduced the upregulation of ICAM-1 and VACM-1 induced by TNF-α. CONCLUSIONS Taken together, these results suggested that RKIP may inhibit the TNF-α-induced expression of adhesion molecules in MOVACs through inactivation of the NF-kappaB pathway.
Raf 激酶抑制剂蛋白（RKIP）调节生长和分化，并在哺乳动物细胞中的关键信号转导级联中发挥作用。然而，RKIP 调节细胞-细胞黏附的潜在机制尚不清楚。本研究探讨了 RKIP 过表达对肿瘤坏死因子（TNF）-α诱导的培养小鼠血管平滑肌细胞（MOVAC）中黏附分子表达的影响。
通过 ELISA 试剂盒、逆转录-PCR 和 Western blot 检测血管细胞黏附分子-1（VCAM-1）和细胞间黏附分子-1（ICAM-1）的表达水平。通过 Western blot 分析检测 RKIP、p65 和核因子（NF）-κB 抑制剂（IκBα）的蛋白表达。使用双荧光素酶报告基因检测法测定 NF-κB 的活性。
结果表明，转染 pCMV5-HA-RKIP 的 MOVAC 显著抑制了 TNF-α诱导的 ICAM-1 和 VCAM-1 的 mRNA 和蛋白表达。在 TNF-α处理的 MOVAC 中，pCMV5-HA-RKIP 还抑制了 THP-1 细胞的黏附。RKIP 还抑制了 TNF-α诱导的 NF-κB 激活和磷酸化 IκB-α的蛋白表达，并促进了 IkB-α的蛋白表达和 p65 NF-κB 的核转位。此外，RKIP 和 NF-κB 抑制剂（BAY11-7082）降低了 TNF-α诱导的 ICAM-1 和 VACM-1 的上调。
综上所述，这些结果表明，RKIP 可能通过失活 NF-κB 途径抑制 TNF-α诱导的 MOVAC 中黏附分子的表达。
