Nakatomi Mitsushiro, Quispe-Salcedo Angela, Sakaguchi Masaka, Ida-Yonemochi Hiroko, Okano Hideyuki, Ohshima Hayato
Division of Anatomy and Cell Biology of the Hard Tissue, Department of Tissue Regeneration and Reconstruction, Niigata University Graduate School of Medical and Dental Sciences, 2-5274 Gakkocho-dori, Chuo-ku, Niigata, 951-8514, Japan.
Division of Anatomy, Department of Health Promotion, Kyushu Dental University, 2-6-1 Manazuru, Kokurakita-ku, Kitakyushu, 803-8580, Japan.
Histochem Cell Biol. 2018 Apr;149(4):383-391. doi: 10.1007/s00418-018-1651-3. Epub 2018 Feb 14.
The Nestin gene encodes type VI intermediate filament and is known to be expressed in undifferentiated cells during neurogenesis and myogenesis. To regulate Nestin expression, the first or second intron enhancer is activated in a tissue-dependent manner, for example, the former in mesodermal cells and the latter in neural stem cells. Although Nestin has also been used as a differentiation marker for odontoblasts during tooth development, how Nestin expression is regulated in odontoblasts remains unclear. Therefore, this study aimed to compare the expression patterns of Nestin-GFP (green fluorescent protein) with that of endogenous Nestin in developing teeth of Nestin-EGFP (enhanced GFP) transgenic mice, in which the second intron enhancer is connected with the EGFP domain, at postnatal 7d, 3w, and 8w. Immunohistochemical and in situ hybridization analyses revealed that endogenous Nestin protein and Nestin mRNA were intensely expressed in differentiated odontoblasts, while GFP immunoreactivity, which reflects the activity of Nestin second intron enhancer-mediated transcription, was mainly observed in the subodontoblastic layer. These results indicate that the first intron enhancer may be activated in differentiated odontoblasts. Intriguingly, Nestin-GFP expression in the subodontoblastic layer was found to be restricted to the coronal pulp of molars, which is susceptible to tooth injuries. Because the subodontoblastic layer serves as a reservoir of newly differentiated odontoblast-like cells upon exogenous stimuli to dentin, our findings suggest that the original odontoblasts and regenerated odontoblast-like cells may differently regulate Nestin expression.
巢蛋白基因编码VI型中间丝，已知在神经发生和肌发生过程中，未分化细胞中表达该基因。为了调节巢蛋白的表达，第一或第二内含子增强子以组织依赖的方式被激活，例如，前者在中胚层细胞中被激活，后者在神经干细胞中被激活。尽管巢蛋白也被用作牙齿发育过程中成牙本质细胞的分化标志物，但成牙本质细胞中巢蛋白的表达是如何被调节的仍不清楚。因此，本研究旨在比较巢蛋白-绿色荧光蛋白（GFP）与内源性巢蛋白在出生后7天、3周和8周的巢蛋白-增强绿色荧光蛋白（EGFP）转基因小鼠发育牙齿中的表达模式，其中第二内含子增强子与EGFP结构域相连。免疫组织化学和原位杂交分析显示，内源性巢蛋白和巢蛋白mRNA在分化的成牙本质细胞中强烈表达，而反映巢蛋白第二内含子增强子介导转录活性的GFP免疫反应性主要在成牙本质细胞下层观察到。这些结果表明，第一内含子增强子可能在分化的成牙本质细胞中被激活。有趣的是，发现成牙本质细胞下层的巢蛋白-GFP表达仅限于磨牙的冠髓，冠髓易受牙齿损伤。由于成牙本质细胞下层在牙本质受到外源性刺激时作为新分化的成牙本质样细胞的储存库，我们的研究结果表明，原始成牙本质细胞和再生的成牙本质样细胞可能以不同方式调节巢蛋白的表达。
