The establishment of methods for reproduction of functional teeth by use of long-term culture

本研究的目的1和2分别是“通过阐明牙齿发育过程中基因的表达，建立颅神经嵴细胞（NCCs）多能相关基因的索引”和“利用上述索引建立多能细胞的生产方法”。但组织切片中的RNA含量太少，不能作为上述指标。因此，我停止了实验。因此，我们主要开展了“建立大鼠全胚胎培养（WEC）复制牙齿、牙周组织和其他颅面器官的方法”的研究，即，3.在上述实验过程中，我们针对牙齿发育过程与脑垂体发育过程的差异进行了研究。我们得到了如下新发现。1.外胚层-内胚层边界和中脑-后脑NCC有助于牙齿的发生。2.外胚层-内胚层边界的非神经元前神经嵴（ANR）产生ACTH产生细胞。3.前索板（PrP）侵入ANR和神经板，参与前索间充质和前脑间充质的形成，并产生垂体内分泌细胞。4. PrP源性细胞的定位分布是牙齿、腺垂体和下丘脑分化所必需的。5. PrP消融小鼠中发生的许多发育缺陷与Shh K. O相似。小鼠我们建立了用于繁殖产生垂体激素的细胞的小生境，例如，上述3项结果表明，在Rathck's囊与间脑的长期培养中，促肾上腺皮质激素（ACTH）、α-GSU、α-Msh、TSHβ和LHβ的含量均明显高于对照组。6.我们显示的域是重要的牙齿，下巴的发育过程中的下巴的鸟类的生殖。7.我们正在利用WEC建立包括与摄食行为有关的下丘脑核团在内的颅面器官的复制方法。

The purposes 1 and 2 of this study was "making an index about multipotent-rerated genes in cranial neural crest cells (NCCs) by the elucidating gene expression during tooth development" and "establishing a method to produce multipotenial cells by use of the index mentioned above", respectively. However, the amount of RNA of NCCs in the histological section was too small to make the index mentioned above. Therefore, I had stopped the experiments. Therefore, we mainly carried out the studies for "establishing methods reproducing the tooth, periodontal tissues and other craniofacial organs by use of rat whole embryo culture (WEC)", i.e., the purposes 3.During above experimens, we focused on the differences between developmental processes of tooth and pituitary, and carried out the studies. We got new findings as follows. 1. The ectoderm-endoderm boundaries and midbrain-hindbrain NCCs contributes to tooth initiation. 2. The non-neuronal anterior neural ridge (ANR) of ectoderm-endoderm boundaries gives rise to ACTH-producing cells. 3. The prechordal plate (PrP) intrudes itself in ANR and neural plate, contributes to prechordal mesenchyme and intraforebrain mesenchyme, and gives rise to endocrine cells in pituitary. 4. The site-specific distribution of PrP-derived cells is required to the differentiation of the tooth, adenohypophysis and hypothalamus. 5. The many developmental defects occur in PrP-ablation mice similarly to in Shh K.O. mice. We established the niche for reproducing the cells producing pituitary hormones, e.g., ACTH,αGSU,αMsh, TSHβ and LHβ, in the above results 3 by the long-term maxillofacial organ culture of Rathck's pouch accompanied with diencephalon. 6. We show the domain that is important for reproduction of tooth, jaw in a development process of a chin of birds. 7. We are establishing the reproduction methods to craniofacial organ, including hypothalamic nucleus relating to feeding behavior, by use of WEC.