Ros M A, Lyons G E, Mackem S, Fallon J F
Departmento de Anatomia y Biologia Celular, Universidad de Cantabria, Santander, Spain.
Dev Biol. 1994 Nov;166(1):59-72. doi: 10.1006/dbio.1994.1296.
Recombinant limb buds constructed of an ectodermal jacket containing dissociated anterior wing bud mesoderm cells form limb-like structures lacking anteroposterior polarity. Polarity can be restored by implanting a small piece of polarizing region into the recombinant limb. We used recombinant limbs to study expression of Msx1 and Msx2 and 5' Hoxd genes in limb development. Detectable mesodermal expression of Msx1 and Msx2 was lost during preparation of the recombinants. When expression was restored both genes were expressed in a symmetrical fashion in the apical mesoderm whether or not the recombinant was polarized. The normal anterior pattern of Msx1 and Msx2 was never reestablished. The roles Msx1 and Msx2 play in limb patterning are at present not clear. In nonpolarized recombinant limbs the 5' Hoxd genes were sequentially and uniformly expressed in the subridge mesoderm across the anteroposterior axis. They exhibited nested domains of expression showing exclusively distal restriction. In polarized recombinants the 5' Hoxd genes were expressed progressively in an asymmetric, nested fashion adjacent to the implanted polarizing region; there was also a distal mesodermal region further from the implant that did not express the most 5' Hoxd genes. Both of these observations are similar to the normal limb. Our results demonstrate that the polarizing region is not necessary for the activation or maintenance of 5' Hoxd gene expression but that its presence is required in order for the normal pattern to be established. We propose that the expression of the Hoxd genes is an intrinsic property of rapidly growing limb cells and that their expression is related to the limb bud proximodistal outgrowth. Our data also support the influence of the polarizing region upon Hoxd gene expression in two ways. First there is a short-range action responsible for the normal asymmetric expression and second a long-range effect suppressing the expression of 5'-Hoxd genes by the anterior cells.
由含有解离的前翅芽中胚层细胞的外胚层套构建的重组肢芽形成缺乏前后极性的肢状结构。通过将一小片极化区域植入重组肢中可以恢复极性。我们利用重组肢来研究Msx1和Msx2以及5'Hoxd基因在肢体发育中的表达。在重组体的制备过程中，Msx1和Msx2可检测到的中胚层表达丧失。当表达恢复时，无论重组体是否极化，这两个基因都以对称方式在顶端中胚层中表达。Msx1和Msx2的正常前部模式从未重新建立。目前尚不清楚Msx1和Msx2在肢体模式形成中所起的作用。在非极化的重组肢中，5'Hoxd基因在前-后轴上的亚嵴中胚层中依次且均匀地表达。它们表现出仅显示远端限制的嵌套表达域。在极化的重组体中，5'Hoxd基因在与植入的极化区域相邻处以不对称的嵌套方式逐渐表达；在离植入物更远的远端中胚层区域也有一个不表达最5'端Hoxd基因的区域。这两个观察结果都与正常肢体相似。我们的结果表明，极化区域对于5'Hoxd基因表达的激活或维持不是必需的，但它的存在是建立正常模式所必需的。我们提出，Hoxd基因的表达是快速生长的肢体细胞的一种内在特性，并且它们的表达与肢芽的近-远侧生长有关。我们的数据还从两个方面支持了极化区域对Hoxd基因表达的影响。首先，有一种短程作用负责正常的不对称表达，其次，有一种长程效应抑制前部细胞对5'-Hoxd基因的表达。
