Functional consequences of the collagen/elastin switch in vascular remodeling in hyperhomocysteinemic wild-type, eNOS^−/−, and iNOS^−/− mice

Abstract

A decrease in vascular elasticity and an increase in pulse wave velocity in hyperhomocysteinemic (HHcy) cystathionine-beta-synthase heterozygote knockout (CBS(-/+)) mice has been observed. Nitric oxide (NO) is a potential regulator of matrix metalloproteinase (MMP) activity in MMP-NO-tissue inhibitor of metalloproteinase (TIMP) inhibitory tertiary complex. However, the contribution of the nitric oxide synthase (NOS) isoforms eNOS and iNOS in the activation of latent MMP is unclear. We hypothesize that the differential production of NO contributes to oxidative stress and increased oxidative/nitrative activation of MMP, resulting in vascular remodeling in response to HHcy. The overall goal is to elucidate the contribution of the NOS isoforms, endothelial and inducible, in the collagen/elastin switch. Experiments were performed on six groups of animals [wild-type (WT), eNOS(-/-), and iNOS(-/-) with and without homocysteine (Hcy) treatment (0.67 g/l) for 8-12 wk]. In vivo echograph was performed to assess aortic timed flow velocity for indirect compliance measurement. Histological determination of collagen and elastin with trichrome and van Gieson stains, respectively, was performed. In situ measurement of superoxide generation using dihydroethidium was used. Differential expression of eNOS, iNOS, nitrotyrosine, MMP-2 and -9, and elastin were measured by quantitative PCR and Western blot analyses. The 2% gelatin zymography was used to assess MMP activity. The increase in O(2)(-) and robust activity of MMP-9 in eNOS(-/-), WT+Hcy, and eNOS(-/-)+Hcy was accompanied by the gross disorganization and thickening of the ECM along with extensive collagen deposition and elastin degradation (collagen/elastin switch) resulting in a decrease in aortic timed flow velocity. Results show that an increase in iNOS activity is a key contributor to HHcy-mediated collagen/elastin switch and resulting decline in aortic compliance.

miR-29b Participates in Early Aneurysm Development in Marfan Syndrome

“…MMPs are secreted by leukocytes and SMCs, 27,28 and MMP-2 can degrade both elastin and collagen. 29 The absence of aortic wall inflammatory cell infiltration in our model suggests that MMPs are secreted locally. MMP expression has been shown to be upregulated by miR-29b, 15,30 so we believe that the transient increase in miR-29b expression results in enhanced MMP-mediated ELN degradation.…”

NADPH Oxidase-Mediated Triggering of Inflammasome Activation in Mouse Podocytes and Glomeruli During Hyperhomocysteinemia

“…A representative chromatogram is shown in Figure 2A, illustrating the peaks produced from both a standard and typical protein sample when run and separated by the Sepharose 6 column. As depicted in Figure 2B, under control conditions, the specific bands for NALP3 and ASC were located in the lowmolecular-weight fractions (18)(19)(20)(21)(22). Upon stimulation with Hcys for 24 h, the NALP3 and ASC bands markedly shifted to high-molecular-weight fractions (3-7), termed the inflammasome fractions due to complex formation.…”

“…Studies from our laboratory and by others have demonstrated that hHcys induces glomerular injury, affecting glomerular endothelial cells, mesangial cells, and podocytes (27)(28)34). hHcys also induces glomerular damage by inhibition of extracellular matrix degradation and production of local oxidative stress, contributing to the sclerotic process and eventually leading to the loss of renal function (21,30). Despite extensive studies, the early mechanisms triggering the pathogenic actions of hHcys are not yet fully understood.…”

Aberrant expression of FcγRIIIA (CD16) contributes to the development of atherosclerosis

“…Immunofluorescent staining was performed according to the standard procedures described previously (Cairns et al, 2002). The protein level of MMP-9 in aorta was determined by Western blot analysis according to the standard procedure as previously described (Steed et al, 2010). GAPDH was used as a loading control.…”