Harikrishnan Keerthi, Cooley Marion A, Sugi Yukiko, Barth Jeremy L, Rasmussen Lars M, Kern Christine B, Argraves Kelley M, Argraves W Scott
Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC 29425, USA.
Department of Clinical Biochemistry and Pharmacology, Odense University Hospital, Odense, Denmark.
Mech Dev. 2015 May;136:123-32. doi: 10.1016/j.mod.2014.12.005. Epub 2015 Jan 6.
Endothelial to mesenchymal transition (EMT) that occurs during cardiac outflow tract (OFT) development is critical for formation of the semilunar valves. Fibulin-1 (Fbln1) is an extracellular matrix protein that is present at several sites of EMT, including the OFT (i.e., E9.5-10.5). The aim of this study was to determine the role of Fbln1 in EMT during the earliest events of OFT development. Examination of proximal OFT cushions in Fbln1 null embryos detected hypercellularity at both E9.5 (93% increase; p = 0.002) and E10.5 (43% increase; p = 0.01) as compared to wild type, suggesting that Fbln1 normally suppresses OFT endocardial cushion EMT. This was supported by studies of proximal OFT cushion explants, which showed that explants from Fbln1 null embryos displayed a 58% increase in cells migrating from the explants as compared to wild type (p = 0.005). We next evaluated the effects of Fbln1 deficiency on the expression of factors that regulate proximal OFT EMT. At E9.5, Fbln1 null proximal OFT endocardium and EMT-derived mesenchyme showed increased TGFβ2 (58% increase; p = 0.01) and increased Snail1-positive nuclei (27% increase; p = 0.0003). Histological examination of OFT cushions in Fbln1 null embryos (E9.5) also detected cells present in the cushion that were determined to be erythrocytes based on round morphology, autofluorescence, and positive staining for hemoglobin. Erythrocytes were also detected in Fbln1 null OFT cushions at E10.5. Together, the findings indicate that Fbln1 normally suppresses proximal OFT EMT preventing proximal cushion hypercellularity and blood cell accumulation.
心脏流出道（OFT）发育过程中发生的内皮向间充质转化（EMT）对于半月瓣的形成至关重要。纤连蛋白-1（Fbln1）是一种细胞外基质蛋白，存在于包括OFT（即E9.5 - 10.5）在内的多个EMT位点。本研究的目的是确定Fbln1在OFT发育最早阶段的EMT过程中的作用。与野生型相比，对Fbln1基因敲除胚胎的近端OFT垫进行检查发现，在E9.5（增加93%；p = 0.002）和E10.5（增加43%；p = 0.01）时细胞增多，这表明Fbln1通常抑制OFT心内膜垫的EMT。近端OFT垫外植体的研究支持了这一点，该研究表明，与野生型相比，来自Fbln1基因敲除胚胎的外植体中从外植体迁移出的细胞增加了58%（p = 0.005）。接下来，我们评估了Fbln1缺乏对调节近端OFT EMT的因子表达的影响。在E9.5时，Fbln1基因敲除的近端OFT心内膜和EMT衍生的间充质显示TGFβ2增加（增加58%；p = 0.01）以及Snail1阳性细胞核增加（增加27%；p = 0.0003）。对Fbln1基因敲除胚胎（E9.5）的OFT垫进行组织学检查还发现垫中存在基于圆形形态、自发荧光和血红蛋白阳性染色确定为红细胞的细胞。在E10.5时，Fbln1基因敲除的OFT垫中也检测到了红细胞。总之，这些发现表明Fbln1通常抑制近端OFT EMT，防止近端垫细胞增多和血细胞积聚。
