McCluskey J, Martin P
Department of Anatomy and Developmental Biology, University College London, United Kingdom.
Dev Biol. 1995 Jul;170(1):102-14. doi: 10.1006/dbio.1995.1199.
The tissue movements of epithelial spreading and mesenchymal contraction play key roles in many aspects of embryonic morphogenesis. One way of studying these movements in a controlled manner is to make an excisional skin wound to an embryo and watch the wound heal. In this paper we report our studies of healing of a simple excisional lesion made to the limb bud stage mouse embryo. The wounded, living embryo is cultured in a roller bottle; under such conditions the wound heals with a highly reproducible time course and is completely closed by 24 hr. During the healing period the environment bathing the wound can be simply manipulated by adding drugs or factors to the culture medium. We have used DiI to label mesenchymal cells exposed at the margin of the initial wound and, by following their fate and measuring the area of mesenchyme remaining exposed at various time points during the healing process, we have quantified both the extent of mesenchymal contraction and the extent of reepithelialisation by movement of epidermis over mesenchyme. We show that the two types of tissue movement contribute almost equally (50:50) to the total wound closure rate. We have gone on to investigate the cell machinery underlying these processes. In adult wounds the epidermis migrates by means of lamellipodial crawling, but we show that reepithelialisation in the embryo is achieved instead by purse-string contraction of a cable of filamentous actin which assembles in the basal layer of cells at the free edge of the epidermis. Addition of cytochalasin D to the culture medium blocks formation of this actin cable and leads to failure of reepithelialisation. Contraction of adult wound connective tissue appears to be driven by conversion of dermal fibroblasts into a specialist smooth muscle-like fibroblast, the myofibroblast. However, using an antibody recognising the alpha-isoform of smooth muscle actin and specific for smooth muscle cells and myofibroblasts, we show that a similar conversion into myofibroblasts does not occur at any stage during the embryonic wound healing process. These observations indicate that both of the tissue movements of embryonic wound healing utilise cell machinery fundamentally different from that driving the analogous tissue movements of adult healing.
上皮扩展和间充质收缩等组织运动在胚胎形态发生的诸多方面发挥着关键作用。以可控方式研究这些运动的一种方法是对胚胎进行切除性皮肤创伤，并观察伤口愈合情况。在本文中，我们报告了对处于肢芽期的小鼠胚胎进行简单切除性损伤后愈合情况的研究。受伤的活体胚胎在滚瓶中培养；在这种条件下，伤口以高度可重复的时间进程愈合，并在24小时内完全闭合。在愈合期间，通过向培养基中添加药物或因子，可以简单地操控伤口周围的环境。我们使用DiI标记在初始伤口边缘暴露的间充质细胞，并通过追踪它们的命运以及测量在愈合过程中各个时间点仍暴露的间充质面积，我们量化了间充质收缩的程度以及表皮在间充质上移动实现再上皮化的程度。我们发现，这两种类型的组织运动对总伤口闭合率的贡献几乎相等（50:50）。我们接着研究了这些过程背后的细胞机制。在成年伤口中，表皮通过片状伪足爬行迁移，但我们发现胚胎中的再上皮化是通过在表皮自由边缘的细胞基底层组装的丝状肌动蛋白索的荷包口收缩来实现的。向培养基中添加细胞松弛素D会阻止这种肌动蛋白索的形成，并导致再上皮化失败。成年伤口结缔组织的收缩似乎是由真皮成纤维细胞转化为一种特殊的平滑肌样成纤维细胞，即肌成纤维细胞所驱动。然而，使用一种识别平滑肌肌动蛋白α异构体且对平滑肌细胞和肌成纤维细胞具有特异性的抗体，我们发现，在胚胎伤口愈合过程的任何阶段都不会发生类似的向肌成纤维细胞的转化。这些观察结果表明，胚胎伤口愈合的两种组织运动所利用的细胞机制与驱动成年伤口类似组织运动的机制根本不同。
