Expression of genes encoding vascular endothelial growth factor and its Flk-1 receptor in the chick embryonic heart
Expression of genes encoding vascular endothelial growth factor and its Flk-1 receptor in the chick embryonic heart
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DOI：
10.1006/jmcc.2000.1141
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发表时间：
2000-06-01
影响因子：
5
通讯作者：
Nagai, R
中科院分区：
文献类型：
--
作者：
Sugishita, Y;Takahashi, T;Nagai, R
血管内皮生长因子（VEGF）在胚胎血管发育中起重要作用。心脏是产生VEGF的主要器官之一，但胚胎心肌细胞中VEGF基因的表达调控机制尚不清楚。因此，我们从培养的10日龄鸡胚心室肌细胞（CEVM）中克隆了编码VEGF及其受体（KDR/flk-1或Quek-1同源物）的cDNA。逆转录-聚合酶链反应结果显示，这些chide VEGF mRNA至少由四种不同的种类组成，分别对应于190、166、146和122个氨基酸的亚型。在胚胎心脏和CEVM中，166和122氨基酸的异构体占优势。北方印迹分析检测到丰富的VEGF mRNA在胚胎心脏和CEVM，即使在基础状态。在CEVM中，毛喉素（100 μ M）或佛波醇12-肉豆蔻酸酯，13-乙酸酯（200 nM）以时间依赖性方式显著增加CEVM中VEGF mRNA的水平。与此相反，VEGF mRNA的基础水平减弱染料木素（100 μ M），但不是由H89（100 μ M）或双吲哚马来酰亚胺（75 μ M）。北方印迹分析也检测到鸡flk-1 mRNA在胚胎心脏的丰度，并在CEVM的程度要小得多。VEGF和flk-1 mRNA在6、8和10日龄鸡胚心脏中持续高表达。在10天龄的胚胎心脏中，原位杂交证实编码VEGF的mRNA主要在心室肌细胞中表达。与此相反，flk-1 mRNA的检测微血管内皮细胞，并在较小程度上在心室肌细胞。这些数据表明，VEGF产生于胚胎心室肌细胞，即使在基础状态，VEGF mRNA的水平可能受到不同的蛋白激酶的调节。鸡胚心脏产生的VEGF可能通过作用于周围的内皮细胞，并可能作用于心室肌细胞本身，在胚胎心血管发育中发挥重要作用。(C)北京大学出版社.
Vascular endothelial growth factor (VEGF) is known to play an essential role in embryonic vascular development. The heart is one of the main organs that produce VEGF, but it is still unknown how expression of VEGF gene is regulated in embryonic cardiac myocytes. Thus, we cloned cDNAs encoding VEGF and its receptor (a KDR/flk-1 or Quek-1 homologue) from cultured 10-day-old chick embryonic ventricular myocytes (CEVM). Reverse transcription-polymerase chain reaction revealed that the chide VEGF mRNAs consisted of at least four different species corresponding to the isoforms of 190, 166, 146 and 122 amino acids. In the embryonic heart and CEVM, the isoforms of 166 and 122 amino acids were dominant. Northern blot analysis detected an abundance of VEGF mRNA in both the embryonic heart and CEVM, even at the basal state. The levels of VEGF mRNA in CEVM were significantly augmented by forskolin (100 mu M), or phorbol 12-myristate, 13-acetate (200 nM) in a time-dependent manner in CEVM. In contrast, the basal levels of VEGF mRNA were attenuated by genistein (100 mu M), but not by H89 (100 mu M) or bisindolylmaleimide (75 mu M). Northern blot analysis also detected the chick flk-1 mRNA in abundance in the embryonic heart, and to a much lesser extent in CEVM. The expression levels of VEGF and flk-1 mRNA species were continuously high in the 6, 8 and 10-day-old chick embryonic hearts. In the 10-day-old embryonic hearts, in situ hybridization confirmed that mRNA encoding VEGF was mainly expressed in ventricular myocytes. In contrast, the flk-1 mRNA was detected in the microvascular endothelial cells, and to a lesser extent in the ventricular myocytes. These data suggest that VEGF is produced in embryonic ventricular myocytes, even at the basal state, and that the levels of VEGF mRNA may be differently regulated by various protein kinases. VEGF produced by the chick embryonic heart may play important roles in embryonic cardiovascular development by acting on surrounding endothelial cells and, possibly, on ventricular myocytes themselves. (C) 2000 Academic Press.