Acquired CCN2 deficiency increases aortic metalloproteinase activity
Matrix metalloproteinases (MMPs) play a key role in aneurysm formation and rupture in several experimental models. To clarify whether CCN2 deletion regulates MMPs levels and/or activity in the aortic wall, a gelatin zymography was performed using protein lysates from the whole aorta. In CCN2-KO mice, MMP-2 and MMP-9 activity was increased compared to WT mice (Figure 5A ). Ang II infusion further increased this activity both in WT and CCN2 deleted mice. MMP-8 levels were also evaluated by western blot. CCN2 deletion increased relative MMP-8 concentration of both latent and active protein, and this was also observed in response to Ang II administration in WT and CCN2-KO mice (Figure 5B ). Additionally, the location of increased MMP activity was assessed in thoracic descending aorta sections by in situ zymography. In aortas of WT mice, the autofluorescence of elastic layers can be distinguished, and no MMP activity was detected. Ang II administration resulted in local positive MMP-fluorescence (assessed by gelatin-degradation). However, in CCN2-KO mice there was lower elastin-fluorescence but increased MMP-fluorescence between the elastic layers, which was further increased by Ang II (Figure 5C ).