IADR Abstract Archives
Human β-defensin 3 Reduces TNF-α–induced Inflammation In HUVECs
Objectives: Human β-defensin 3 (hBD3) is an antimicrobial peptide with immunoregulatory properties mainly expressed in the oral cavity and it has been shown for its protective role in the pathogenesis of atherosclerosis in our previous study. This study further investigated the role of hBD3 in the initiation stage of this vascular inflammatory disease with human primary umbilical vein endothelial cells (HUVECs) triggered by tumor necrosis factor (TNF)-α and the underlying mechanisms.
Methods: The effects of hBD3 upon TNF-α-induced endothelial injury of HUVECs were evaluated. The inflammatory mediators released by HUVECs were measured with enzyme-linked immunosorbent assay (ELISA). Reactive oxygen species (ROS) induced by TNF-α was measured using DCFH-DA. F-actin staining was evaluated with phalloidine. In addition, protein levels of key molecules regarding cell adhesion, signaling pathways and apoptosis were detected using western blot.
Results: First, hBD3 reduced the production of interleukin-6 (IL-6), IL-8, monocyte chemoattractant protein-1 (MCP-1) and macrophage migration inhibitory factor (MIF) of HUVECs in a dose-dependent manner. In addition, hBD3 significantly prevented intracellular ROS production of HUVECs. Second, western blot analysis demonstrated that hBD3 dose-dependently suppressed the protein levels of intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in TNF-α-induced HUVECs. As a result, hBD3 inhibited monocyte adhesion to TNF-α-treated endothelial cells. Additionally, hBD3 suppressed TNF-α-induced F-actin reorganization in HUVECs. Third, hBD3 markedly inhibited NF-κB activation by the downregulation of phosphorylation of IKK-α/β, IκB and the subunit of p65 within 30 min. Moreover, the phosphorylations of p38 and c-Jun N-terminal protein kinases (JNK) of mitogen-activated protein kinase (MAPK) pathway were also inhibited by hBD3 in HUVECs. Finally, hBD3 inhibited TNF-α-stimulated apoptosis of HUVECs.
Conclusions: hBD3 exerts anti-inflammatory and anti-oxidative effect in endothelial cells induced by TNF-α by inhibition of NF-κB and MAPK signaling.
Methods: The effects of hBD3 upon TNF-α-induced endothelial injury of HUVECs were evaluated. The inflammatory mediators released by HUVECs were measured with enzyme-linked immunosorbent assay (ELISA). Reactive oxygen species (ROS) induced by TNF-α was measured using DCFH-DA. F-actin staining was evaluated with phalloidine. In addition, protein levels of key molecules regarding cell adhesion, signaling pathways and apoptosis were detected using western blot.
Results: First, hBD3 reduced the production of interleukin-6 (IL-6), IL-8, monocyte chemoattractant protein-1 (MCP-1) and macrophage migration inhibitory factor (MIF) of HUVECs in a dose-dependent manner. In addition, hBD3 significantly prevented intracellular ROS production of HUVECs. Second, western blot analysis demonstrated that hBD3 dose-dependently suppressed the protein levels of intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in TNF-α-induced HUVECs. As a result, hBD3 inhibited monocyte adhesion to TNF-α-treated endothelial cells. Additionally, hBD3 suppressed TNF-α-induced F-actin reorganization in HUVECs. Third, hBD3 markedly inhibited NF-κB activation by the downregulation of phosphorylation of IKK-α/β, IκB and the subunit of p65 within 30 min. Moreover, the phosphorylations of p38 and c-Jun N-terminal protein kinases (JNK) of mitogen-activated protein kinase (MAPK) pathway were also inhibited by hBD3 in HUVECs. Finally, hBD3 inhibited TNF-α-stimulated apoptosis of HUVECs.
Conclusions: hBD3 exerts anti-inflammatory and anti-oxidative effect in endothelial cells induced by TNF-α by inhibition of NF-κB and MAPK signaling.