Abstract 1356: Therapeutic Targeting RAGE/MT1-MMP/Rac1 Axis in Redox-Sensitive Signaling Pathway in Diabetic Vascular Remodeling
Background: Advanced glycation end products (AGE)/receptor for AGE (RAGE) axis is essential for the pathogenesis of vascular disease complicated with diabetes. Membrane type 1-matrix metalloproteinase (MT1-MMP) functions as not only a proteolytic enzyme but also a signaling molecule. Rho GTPase Rac1, a component of NADPH oxidase, plays a central role in redox-sensitive signaling pathways of vascular disease.
Hypothesis: RAGE and MT1-MMP form a complex and the RAGE/MT1-MMP/Rac1 axis is essential for redox-sensitive signaling of diabetic vascular remodeling. Methods and Results: We added AGEs to cultured rabbit smooth muscle cells (SMCs). AGE induced Rac-1 and p47phox activation within 1 minute as evidenced by membrane translocation and increased the activity of NADPH oxidase within 5 minutes as determined by lucigenin chemiluminescence method, resulting in ROS generation. NF-∈°B phosphorylation was subsequently induced within 15 minutes, resulting in increased expression of MCP-1, MMP-9, PAI-1 and RAGE. This AGE-induced activation of redox-sensitive signaling pathway was blocked by inhibition of RAGE and MT1-MMP by small interfering RNA (siRNA). Statins and angiotensin II receptor blockers inhibited the redox-sensitive pathway including Rac1 activation and RAGE expression. Geranylgeranyltransferase I responsible for initial Rac1 activation of lipid modification was activated by AGEs within 1 minute, which was inhibited by siRNA to MT1-MMP. These data suggest that MT1-MMP is upstream of Rac1 and that RAGE and MT1-MMP may form a complex. Fluorescent immunochemistry demonstrated that MT1-MMP was co-localized in part with RAGE in cultured SMCs. Thus, we performed immunoprecipitation using an antibody to MT1-MMP, indicating that RAGE was found in association with MT1-MMP in the immunoprecipitates recovered from the SMCs. Importantly, RAGE and MT1-MMP also formed a complex with RAGE in the immunoprecipitates from the aortas of spontaneously diabetic Torii rats, a type 2 diabetic animal model.
Conclusions: We provide novel evidence that the RAGE/MT1-MMP/Rac1 axis is essential for AGEs-induced redox-sensitive signaling pathway. This study identifies an attractive therapeutic targeting RAGE/MT1-MMP/Rac1 in diabetic vascular disease.