e0083 Membrane type 1 matrix metalloproteinase activation is enhanced by low shear stress through integrin pathway in ApoE-/- mice

Objective Low shear stress and matrix metalloproteinase are involved in atherogenesis and plaque stability. The aim of this study wad to explore the relation and possible mechanism of membrane type-1 matrix metalloproteinase (MT1-MMP) and low shear stress (LSS). Methods 80 male apoE-/- mice were fed a high-fat diet, and lesions in the left carotid artery were induced by perivascular placement of constrictive collars. Results Ultrasound-determined shear stress was significantly lower in the left carotid artery than in the right artery (5.61±0.72 vs 9.76±1.48 N/m2, p<0.01). Confocal microscopy and dual staining revealed higher MT1-MMP expression in left common carotid artery. Cultured human umbilical vein endothelial cells (HUVECs) exposed to 0.4 N/cm2 shear stress (LSS) and 1.2 N/cm2 shear stress (PSS). HUVECs subjected to LSS showed a time-dependent increase in MT1-MMP mRNA level. MT1-MMP mRNA level was downregulated under PSS at 3 and 5 h. MT1-MMP protein showed no change in expression at 1-h LSS, but 3-, 5-, 8- and 16-h treatment produced significantly increased expression and activity. NF-κB DNA-binding activity was stronger at 30-min and 1-h LSS exposure than with control treatment. Pretreatment with 18 μM SN50 efficiently inhibited NF-κB DNA binding activity. In the presence of SN50, MT1-MMP mRNA expression, protein level and activity were significantly attenuated (all p<0.01). MT1-MMP mRNA expression, protein level and activity were inhibited by PD98059 (all p<0.01). To determine the relation between ERK1/2 and NF-κB with MT1-MMP induced by LSS, HUVECs were incubated with PD98059 for 2 h before LSS. The level of phosphor-IKKα/β was reduced, whereas that of phosphor-IκBα was increased as compared with no PD98059 treatment. As well, NF-κB DNA-binding activity was decreased. HUVECs were preincubated for 48 h with FAK siRNA and then treated with LSS. The shear stress-induced increase in MT1-MMP mRNA and protein level and activity was significantly inhibited (all p<0.01). As well, LSS-induced ERK1/2 activation was inhibited with 5-min LSS exposure (p<0.01). To examine NF-κB DNA-binding activity, HUVECs were pretreated for 48 h with FAK siRNA before being exposed to LSS. NF-κB DNA-binding activity was inhibited with FAK siRNA pretreatment. HUVECs were preincubated for 48 h with integrin β1 siRNA and then underwent LSS. The shear stress-induced increase in MT1-MMP mRNA expression, protein level and activity was significantly inhibited (all p<0.01). As well, LSS-induced FAK and ERK1/2 activation was inhibited at 5-min exposure (p<0.01). Next, we examined NF-κB DNA-binding activity with 48-h integrin β1 siRNA pretreatment before exposure to LSS; NF-κB DNA binding activity was inhibited by siRNA integrin β1 pretreatment. Conclusions MT1-MMP induced by LSS is involved in atherosclerotic plaque, via the integrin β1-FAK-ERK1/2-NF-κB pathway.