Kikuchi H, Sawada T, Yanagisawa T
Department of Pharmacology, School of Dentistry, Showa University, Tokyo, Japan.
Arch Oral Biol. 1996 Aug-Sep;41(8-9):871-83. doi: 10.1016/s0003-9969(96)00022-2.
In an attempt to study the effects of a three-dimensional agar surface on in vitro dentinogenesis both in the growing end and in incisally cross-cut pulp, the possible expression of odontoblast phenotype was investigated morphologically, autoradiographically and immunohistochemically. Explants were incubated for 8 days. In the growing end, during the last 4 days, mitotic cells differentiated into [3H]-thymidine-labelled, tubular matrix-forming cells. In cross-cut pulp, however, during the first 4 days, mitotic cells differentiated into [3H]-thymidine-labelled, tubular matrix-forming cells. Electron microscopy demonstrated that, in both regions, tubular matrix-forming cells had characteristics similar to those of primary odontoblasts. When agar was incubated alone, exogenous fibronectin was deposited on it rapidly. After 12 h, endogenous fibronectin appeared on explant peripheral cells. Collagen and materials reacting positively to periodic acid-Schiff (PAS) were first interposed between agar and explant after 4 days. After 8 days, an inner immunonegative layer corresponding to materials reacting positively to PAS or toluidine blue and an outer immunopositive layer of fibronectin or collagen were visible adjacent to the rows of elongated columnar cells. In the presence of Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP), a competitive inhibitor of attachment of cells to fibronectin, explants became detached from the agar surface, and no dentinogenesis occurred. These results indicate that, when in contact with an agar surface that becomes modified by fibronectin and/or by a complex of fibronectin with deposited matrix, dental mesenchymal cells progressively differentiate into tubular matrix-forming cells. Possibly the functional agar surface has the important role of providing a foothold for cell attachment, which is the first step towards in vitro odontoblast differentiation. This system of inducing tubular matrix-forming cells constitutes a useful model for the study of in vitro dentinogenesis.
为了研究三维琼脂表面对生长端和切端横断牙髓体外牙本质形成的影响，通过形态学、放射自显影和免疫组织化学方法研究了成牙本质细胞表型的可能表达。外植体培养8天。在生长端，在最后4天，有丝分裂细胞分化为[3H] -胸苷标记的、形成管状基质的细胞。然而，在横断牙髓中，在最初4天，有丝分裂细胞分化为[3H] -胸苷标记的、形成管状基质的细胞。电子显微镜显示，在两个区域，形成管状基质的细胞具有与初级成牙本质细胞相似的特征。单独培养琼脂时，外源性纤连蛋白会迅速沉积在其上。12小时后，内源性纤连蛋白出现在外植体周围细胞上。4天后，胶原蛋白和对过碘酸-希夫（PAS）呈阳性反应的物质首先插入琼脂和外植体之间。8天后，在成排的细长柱状细胞相邻处可见对应于对PAS或甲苯胺蓝呈阳性反应物质的内部免疫阴性层和纤连蛋白或胶原蛋白的外部免疫阳性层。在存在细胞与纤连蛋白附着的竞争性抑制剂甘氨酰-精氨酰-甘氨酰-天冬氨酰-丝氨酸-脯氨酸（GRGDSP）的情况下，外植体从琼脂表面脱离，且未发生牙本质形成。这些结果表明，当与被纤连蛋白和/或纤连蛋白与沉积基质的复合物修饰的琼脂表面接触时，牙间充质细胞逐渐分化为形成管状基质的细胞。功能性琼脂表面可能具有为细胞附着提供立足点的重要作用，这是体外成牙本质细胞分化的第一步。这种诱导形成管状基质细胞的系统构成了研究体外牙本质形成的有用模型。
