Molecular Mechanism of Retinoic Acid-Induced Pattern Duplication During Limb Development

已知肢体的前后模式是由位于肢芽后缘的极化活动区(ZPA)发出的信号决定的。维甲酸被认为是ZPA的形态来源。然而，最近关于肢体发育机制的研究表明，维甲酸是极化活动的诱导剂，但不是直接的ZPA信号。以雏鸡肢芽为模型，研究了维甲酸诱导肢体复制的机制，获得了以下结果。ZPA植入肢芽前部诱导的HoxD表达位于移植物远端和顶端外胚层脊(AER)附近。去除后部的AER会导致HoxD表达下降。因此，除了ZPA因子外，来自AER的信号因子也是诱导和维持HoxD表达所必需的。Msh相关的同源异型盒基因msx1在LIM…的AER和下层间充质中表达。更多的B花蕾。Msx1基因的表达依赖于AER的信号，而与果蝇配对相关的同源盒基因PRX1的中胚层表达与AER无关，这些同源盒基因在肢芽中的表达受到AER和ZPA2信号的差异调控。几种生长因子，包括成纤维细胞生长因子和转化生长因子β，被认为与肢体发育有关，因为这些生长因子在非洲爪哇胚胎发育中起着重要的中胚层诱导和轴向决定作用。(1)已鉴定出4个成纤维细胞生长因子受体基因，并在肢芽中表达。其中，FGFR2显示外胚层表达，而中胚层表达受到中央限制，提示成纤维细胞生长因子家族参与皮肤和骨的形态发生。正如最初在器官培养中观察到的小鼠肢芽中的表达一样，成纤维细胞生长因子-4基因也在鸡的肢芽中表达。利用逆转录病毒载体异位表达成纤维细胞生长因子-4基因，在鸡胚胎中产生了多个数字。因此，成纤维细胞生长因子-4可能是导致肢体生长的AER因素之一。(2)已知的转化生长因子-β家族的I型和II型受体具有细胞内特有的丝氨酸/苏氨酸激酶域。已鉴定出两种鸡激活素受体，受体基因在早期肢芽中弱表达。随后，受体基因在软骨前层、成肌细胞和外胚层中表达。在雏鸡体内发现了I型受体的两个新成员：一个是BMP-2/4，另一个是转化生长因子-β1。肝细胞生长因子(HGF)在鸡胚胎发育过程中具有诱导次级体轴的作用。在萌芽阶段，当产生HGF的MRC-5细胞被移植到小腿芽的前部时，显示出诱导复制肢体的作用。HGF基因在AER下方的肢芽远端间充质中表达，提示HGF在肢体发育过程中是一种AER诱导和/或维持因子。较少

Anterior-posterior pattern of the limb is known to be determined by a signal emanating from the zone of polarizing activity(ZPA) located in the posterior margin of limb bud. Retinoic acid was postulated to be a morphogen of the ZPA.However, recent findings on the mechanism of limb development argue that retinoic acid is an inducer of polarizing activity but not a direct ZPA signal. Mechanism of retinoic acid-induced limb duplication was investigated using chick limb bud, and following results were obtained.1. Expression of HoxD induced by ZPA implantation to the anterior region of the limb bud is distal to the graft and subjacent to the apical ectodermal ridge(AER). Removal of the AER in the posterior region results in the decrease in HoxD expression. Signaling factor from the AER is therefore required for the induction and maintenance of HoxD expression in addition to the ZPA factor. A msh-related homeobox gene, Msx1, is expressed in the AER and underlying distal mesenchyme of the lim … More b bud. Expression of the Msx1 gene depends on the signals from the AER, whereas mesodermal expression of Prx1, a homeobox gene related to Drosophila paired, is independent of the AER.These homeobox genes expressed specifically in the limb bud are differentially regulated by signals from the AER and ZPA.2. Several growth factors, including fibroblast growth factor(FGF) and transforming growth factor beta(TGF-beta), are presumed to be involved in limb development, since the growth factors are known to play important roles for mesoderm induction and axial determination in Xenopus emgbryogenesis.(1) Four FGF receptor genes have been identified, and are expressed in the limb bud. Among them, FGFR2 reveals ectodermal expression and centrally restricted mesodermal expression, suggesting the involvement of the FGF family in skin and bone morphogenesis. The FGF-4 gene is also expressed in the AER of the chick limb bud, as originally observed in the mouse limb bud in organ culture. Ectopic expression of the FGF-4 gene using retrovirus vector results in the supernumeral digits in the chick embryo. Thus, FGF-4 is likely to be one of the AER factors responsible for limb outgrowth.(2) Type I and II receptors for TGF-beta family are known to have characteristic intracellular Ser/Thr kinase domain. Two chicken activin receptors were identified, and the receptor genes are weakly expressed in the early limb bud. Later, the receptor genes are expressed in the prechondrium, myoblasts and ectoderm. Two novel members of the type I receptors were identified in the chick ; one for BMP-2/4 and the other for TGF-beta1. The former is weakly expressed in the prechondrium of the developing limb.(3) Hepatocyte growth factor(HGF) is known to induce secondary body axis in the chick embryo. HGF-producing MRC-5 cells were shown to induce duplicated limb when implanted to the anterior region of the leg bud at budding stages. The HGF gene is expressed in the distal mesenchyme of the limb bud underneath the AER, suggesting HGF as an AER-inducing and/or maintenance factor during limb development. Less