Drews Ulrich, Drews Ute
Abteilung für Klinische Morphologie des Zentrums für Klinische Grundlagenforschung der Universität Ulm, Deutschland.
Wilhelm Roux Arch Entwickl Mech Org. 1973 Sep;173(3):208-227. doi: 10.1007/BF00573115.
From our previous work we have put forward the hypothesis that cholinesterase activity in embryonic cells is related to morphogenetic movements. Therefore, the locomotory behavior of mesenchymal cells differentiating into cartilage by passing through a phase of Cholinesterase activity was analysedin vitro.Mesenchymal cores of chick limb buds stage 23/24 were partially disaggregated and cultured in plastic tissue culture dishes (Fig. 1). Within 31/2 to 5 days aggregates of mesenchymal cells differentiated into cartilage nodules surrounded by myoblasts (Figs. 2, 3 and 5). The cartilaginous nature of the nodules was confirmed by electron microscopy (Figs. 6 and 7). During the culture period serial photographs (24×24 mm) were taken (Tables 1-3). After formalin fixation the histochemical Cholinesterase reaction was carried out inside the culture dishes. Positive and negative cells were identified in the live serial photographs and their locomotory behavior was analysed.Initially the cells behaved like fibroblasts. Movements were regulated by contact inhibition, resulting in radial outward migration within the mesenchymal aggregates. In this first phase of development there was no cholinesterase activity. After 12 to 48 hours in culture however ChE-positive cells could be detected. Positive cells, appearing within a monolayer, detached from the bottom of the culture dish and crawled onto neighboring cells (Figs. 8a and b). In the periphery of the aggregates radial outward migration slowed down considerably. In the center short non-directional movements of positive cells could be observed, frequently leading to overlayering of cell bodies.In the third stage of development the ChE-positive cells stopped moving and transformed into cartilage cells (Fig. 9a and b). Finally, ChE-activity disappeared from the differentiated cartilage cells.From the difference in locomotory behaviour of negative and positive cells it is concluded that the appearance of Cholinesterase is accompanied by a change in the adhesive properties of the cells. An increase in cell adhesiveness enables the ChE-positive cells to detach from the bottom of the culture dish and to establish a new equilibrium of contact inhibition inside the cellular aggregates. This seems to be a prerequisite for the secretion of extracellular matrix and development of firm cell contacts. In vivo cartilage differentiation presumably also starts with an increase in cell adhesiveness in the presumptive cartilage cells. This provokes pseudopodial rearrangements leading to the condensation and demarkation of the cartilage anlagen. The change in adhesiveness is accompanied by Cholinesterase activity.
从我们之前的工作中，我们提出了一个假设，即胚胎细胞中的胆碱酯酶活性与形态发生运动有关。因此，我们在体外分析了通过胆碱酯酶活性阶段分化为软骨的间充质细胞的运动行为。将处于23/24期的鸡胚肢芽的间充质核心部分解离，并在塑料组织培养皿中培养（图1）。在3.5至5天内，间充质细胞聚集体分化为被成肌细胞包围的软骨结节（图2、3和5）。通过电子显微镜证实了结节的软骨性质（图6和7）。在培养期间拍摄了系列照片（24×24毫米）（表1 - 3）。福尔马林固定后，在培养皿内进行组织化学胆碱酯酶反应。在活体系列照片中识别出阳性和阴性细胞，并分析它们的运动行为。
最初，细胞表现得像成纤维细胞。运动受接触抑制调节，导致在间充质聚集体内呈放射状向外迁移。在发育的第一阶段没有胆碱酯酶活性。然而，培养12至48小时后，可以检测到胆碱酯酶阳性细胞。出现在单层内的阳性细胞从培养皿底部脱离，并爬到相邻细胞上（图8a和b）。在聚集体的周边，放射状向外迁移显著减慢。在中心，可以观察到阳性细胞的短程无定向运动，经常导致细胞体重叠。
在发育的第三阶段，胆碱酯酶阳性细胞停止运动并转化为软骨细胞（图9a和b）。最后，分化的软骨细胞中胆碱酯酶活性消失。
从阴性和阳性细胞运动行为的差异可以得出结论，胆碱酯酶的出现伴随着细胞黏附特性的改变。细胞黏附性的增加使胆碱酯酶阳性细胞能够从培养皿底部脱离，并在细胞聚集体内建立新的接触抑制平衡。这似乎是细胞外基质分泌和牢固细胞接触形成的先决条件。在体内，软骨分化可能也始于假定软骨细胞中细胞黏附性的增加。这引发伪足重排，导致软骨原基的凝聚和分界。黏附性的改变伴随着胆碱酯酶活性。
