Abstract 291: Fibulin-1 is Required to Prevent Hypoplastic Semilunar Valve Formation
Fibulin-1 (Fbln1), an extracellular matrix protein, is prominently expressed during outflow tract (OFT) morphogenesis as a component of both the endocardial cushions and OFT valves. Here we sought to investigate the role of Fbln1 in the process of OFT valvulogenesis. Since Fbln1 null mice die perinatally, we first evaluated the effects of Fbln1 deficiency on OFT valve morphology in hearts of embryos at E17.5. Morphometric analysis of 3D reconstructions revealed that in Fbln1 nulls the volume of the anterior leaflet of the pulmonary valve, and the left and non-coronary leaflets of the aortic valve were significantly reduced as compared to wild type (WT) (30-40% reduction; WT n=6, null n=8; p<0.05). The reduced leaflet size correlated with decreased leaflet cell numbers (45%, 30% and 30% respectively; p<0.05). TUNEL and BrdU analysis showed no difference between E17.5 WT and Fbln1 null OFT valves. To determine when abnormalities of OFT valve leaflet cell numbers first occurs, OFT endocardial cushions and prevalvular mesenchyme from E10.5-14.5 null and WT embryos were analyzed. At E10.5, the proximal OFT cushions of Fbln1 nulls were hypercellular (34% increase; p<0.05; n=5 for each genotype) whereas cellularity of distal cushions was not different from WT (p= 0.49). The fact that the ratio of proximal OFT cushion mesenchymal cells to endothelial cells was increased in E10.5 Fbln1 null cushions suggests that alteration in EMT might contribute to the hypercellularity. At E12.5, cellularity of the proximal OFT cushions was observed to decrease by 35% (p<0.05; WT n=4; null n=4). This was accompanied by a 172% increase in apoptosis, with no alteration in the level of proliferation (p<0.05; WT n=3; null n=4). Decreased cellularity and increased apoptosis persisted in the OFT prevalvular mesenchyme of Fbln1 nulls at E14.5 (p<0.05; WT n=2; null n=2). Taken together, Fbln1 specifically regulates cellularization of the proximal endocardial cushion and prevalvular mesenchyme. Mechanistically, Fbln1 may act as a suppressor of cushion EMT and indirectly or directly as an inhibitor of apoptosis.
- © 2013 by American Heart Association, Inc.