Horiguchi Kotaro, Nakakura Takashi, Yoshida Saishu, Tsukada Takehiro, Kanno Naoko, Hasegawa Rumi, Takigami Shu, Ohsako Shunji, Kato Takako, Kato Yukio
Laboratory of Anatomy and Cell Biology, Department of Health Sciences, Kyorin University, Mitaka, Tokyo, 181-8612, Japan; Institute of Endocrinology, Meiji University, Kawasaki, Kanagawa, 214-8571, Japan.
Department of Anatomy, Graduate School of Medicine, Teikyo University, Tokyo, 173-8605, Japan.
Biochem Biophys Res Commun. 2016 Nov 11;480(2):273-279. doi: 10.1016/j.bbrc.2016.10.049. Epub 2016 Oct 17.
Contact-dependent (juxtacrine) signaling is important for local cell-to-cell interaction and has received attention in recent years regarding its role in pituitary function, differentiation, and development. This study investigated one of the juxtacrine-related molecules, thymocyte differentiation antigen 1 (THY1), in the anterior lobe of the rat pituitary gland. Western blot analysis revealed expression of the THY1 protein in the adult rat anterior lobe. We also found that the THY1 ligand, integrin-β2 (ITGB2), is also expressed in the pituitary gland. In situ hybridization and immunohistochemical analyses showed that both THY1 mRNA and protein were present in almost, if not all, thyroid-stimulating hormone (TSH)-immunopositive cells (thyrotropes) and that ITGB2 was co-expressed in these cells. As THY1 appeared to represent a novel marker for thyrotropes, we then attempted to isolate these cells from various anterior lobe cells by the use of a THY1 antibody and a pluriBead-cascade cell isolation system. This technology allowed the isolation of thyrotropes with 83% purity at about 17-fold enrichment. Furthermore, the isolated THY1-immunopositive cells had higher Tsh mRNA levels compared with THY1-immunonegative cells and released TSH in response to thyrotropin-releasing hormone. These findings indicated that THY1 represents a potent thyrotrope marker and that the thyrotrope isolation method using the THY1 antibody may serve as a powerful tool to analyze their function including juxtacrine regulation through THY1/ITGB2 interaction.
接触依赖性（旁分泌）信号传导对于局部细胞间相互作用很重要，近年来其在垂体功能、分化和发育中的作用受到了关注。本研究调查了大鼠垂体前叶中一种与旁分泌相关的分子——胸腺细胞分化抗原1（THY1）。蛋白质印迹分析显示THY1蛋白在成年大鼠垂体前叶中有表达。我们还发现THY1配体整合素-β2（ITGB2）也在垂体中表达。原位杂交和免疫组织化学分析表明，几乎所有促甲状腺激素（TSH）免疫阳性细胞（促甲状腺细胞）中都存在THY1 mRNA和蛋白，并且ITGB2在这些细胞中共同表达。由于THY1似乎是促甲状腺细胞的一种新型标志物，我们随后尝试使用THY1抗体和多珠级联细胞分离系统从各种垂体前叶细胞中分离这些细胞。该技术能够以约17倍的富集率分离出纯度为83%的促甲状腺细胞。此外，与THY1免疫阴性细胞相比，分离出的THY1免疫阳性细胞具有更高的Tsh mRNA水平，并能对促甲状腺激素释放激素作出反应释放TSH。这些发现表明THY1是一种有效的促甲状腺细胞标志物，并且使用THY1抗体的促甲状腺细胞分离方法可能是分析其功能（包括通过THY1/ITGB2相互作用进行旁分泌调节）的有力工具。
