GENES REGULATED BY PHOSPHATE-INDUCED PARATHYROID CELL PROLIFERATION IN ORGAN CULTURE : ANALYSIS USING OLIGO-DNA MICROARRAY

1例低磷酸盐血症性骨软化症患者接受高磷酸盐治疗后，发生了第三性甲状旁腺功能亢进。四种甲状旁腺机能亢进的基因分析揭示了几种基因突变，表明高浓度磷酸盐刺激甲状旁腺生长，伴随甲状旁腺的单克隆增殖（参考文献（1））。为了阐明磷酸盐诱导的甲状旁腺细胞增殖，我们建立了原发性（I °-）或继发性甲状旁腺机能亢进（II °-HPT）患者甲状旁腺组织的器官培养。在该器官培养物中，完整的PTH活跃分泌超过1个月。当甲状旁腺组织用高磷酸盐培养基培养时，完整的PTH分泌增加，细胞增殖也被刺激，从而再现了II °-HPT患者甲状旁腺的临床特征。基因分析的寡聚DNA微阵列，可以分析整个人类基因在一个单一的运行（41000基因）显示，甲状旁腺细胞表达丰富的胶原蛋白，蛋白质细胞粘附和形态发生，信号转导，多巴胺受体D4，朊病毒等。此外，甲状旁腺特异性基因，如甲状旁腺激素，嗜铬粒蛋白-A，钙敏感受体，维生素D受体也显着表达。当甲状旁腺细胞用高磷酸盐培养基培养时，包括PTH在内的100多个基因增加超过100%，Ⅱ °-HPT患者的甲状旁腺细胞增殖程度高于Ⅰ °-HPT患者。据报道参与甲状旁腺细胞增殖的生长因子，如FGF-2、IGF-1或TGF-α，在高磷酸盐培养基中没有增加，表明甲状旁腺细胞通过未知途径增殖。通过分析微阵列的早期点，我们正在研究什么信号是第一个刺激甲状旁腺细胞增殖的反应高磷酸盐浓度。

A patient with hypophosphatemic osteomalacia, who was treated with high phosphate, developed tertiary hyperparathyroidism. Gene analysis of the four hyperparathyroid glands revealed several gene mutations, suggesting that high phosphate concentration stimulated parathyroid growth, accompanied by monoclonal proliferation of parathyroid (Reference(1)).To clarify phosphate-induced cell proliferation of parathyroid gland, we established organ culture of parathyroid tissue obtained from patient with primary (I°-)or secondary hyperparathyroidism (II°-HPT). In this organ culture, intact PTH was actively secreted for more than 1 month. When parathyroid tissue were cultured with high phosphate medium, intact PTH secretion was increased and cell proliferation was also stimulated, thus reproducing clinical features of parathyroid glands in patients with II°-HPT。Gene analyses by oligo-DNA microarray, which can analyze whole human genes at a single run (41000genes) revealed that parathyroid cells express abundantly collagens, proteins for cell adhesion and morphogenesis, signal transductions, dopamine receptor D4, prion, etc. Furthermore, parathyroid-specific genes such as PTH, chromogranin-A, calcium-sensing receptor, and vitamin D receptor were also significantly expressed. When parathyroid cells were cultured with high-phosphate medium, more than 100 genes including PTH were increased more than 100%., and parathyroid cells obtained from patients with II°-HPT proliferated to a greater extent than those from I°-HPT. Growth factors which are reported to be involved in parathyroid cell proliferation, such as FGF-2,IGF-1,or TGF-α, did not increase in response to high-phosphate medium, suggesting parathyroid cells proliferate via unknown-pathway. By analyzing microarray at earlier points, we are investigating what signals are the first to stimulate proliferation of parathyroid cells in response to high phosphate concentrations.