NITRIC OXIDE REGULATION OF OSTEOBLASTS

描述：（改编自研究者摘要）一氧化氮（NO） 最近发现是一种细胞调节分子， 在心血管、神经和免疫系统中的短程作用。 没有 通过涉及NO合酶的酶促反应源自精氨酸 (NOS)已知有3种亚型。 待检验的假设 这种应用是在成骨细胞中产生的NO（cNOS和iNOS）， 它既是成骨细胞增殖、功能和 生存能力和邻近骨细胞类型的旁分泌调节因子。 没有 在骨微环境中产生的是一种重要的调节剂， 成骨功能 第一个具体目标是研究 促炎细胞因子在调节iNOS表达中的作用， 成骨细胞系和突变型成骨细胞来自NOS敲除小鼠， 敏感的分子生物学方法。 调查人员将寻找一个 成骨细胞中的cNOS，纯化NOS酶并表征激活剂， 抑制剂的 通过NOS亚型特异性免疫染色， 将在转基因小鼠的骨细胞中测定NOS蛋白表达 缺乏特定的NOS亚型，并在一个新的小鼠模型， 炎症介导的骨丢失。 第二个具体目标是研究 NO调节成骨细胞的机制，刺激某些功能 在低浓度下，同时变得具有细胞毒性并触发成骨细胞 高浓度下死亡。 调查人员将重点关注 增殖，基质生物合成，生物能量学和凋亡， 寻找对NO作用的假定细胞内靶蛋白的影响 包括核糖核苷酸还原酶，乌头酸酶，鸟苷酸环化酶， 环氧合酶和细胞外基质蛋白，其可以被硝化 活性NO中间体。 最后，NO的细胞-细胞信号传导作用将 使用成骨细胞/破骨细胞共培养物进行测试。 希望这些 研究将确立NO/NOS系统的生理意义 以及NO调节正常激素调节的程度， 骨生理学 来源于成骨细胞的NO可能会引发广泛的 炎性骨疾病中的骨破坏，包括 风湿性关节炎、骨髓炎和牙周病，而 同时阻断正常骨骼用来 保持骨量。 控制成骨细胞NO生成的策略可能 证明在广泛的骨中对药物干预有效 疾病

DESCRIPTION: (Adapted from investigator's abstract) Nitric oxide (NO) was recently discovered to be a cell regulatory molecule with rapid and potent short-range actions in cardiovascular, neuronal and immune systems. NO originates from arginine by an enzymatic reaction involving NO synthase (NOS), for which there are 3 known isoforms. The hypothesis to be tested in this application is that NO (cNOS and iNOS), produced in osteoblasts, is both a direct autocrine regulator of osteoblast proliferation, function and viability, and a paracrine regulator of neighboring bone cell types. NO produced in the bone microenvironment is an important modulator of osteoblastic function. The first Specific Aim is to study the role of pro-inflammatory cytokines in regulating expression of an iNOS, in various osteoblastic cell lines and mutant osteoblasts from NOS knock-out mice using sensitive molecular biological methods. The investigators will search for a cNOS in osteoblasts, purify the NOS enzymes and characterize activators and inhibitors. Through isoform-specific immunostaining of NOS, the pattern of NOS protein expression will be determined in bone cells of transgenic mice deficient in specific NOS isoforms, and in a new mouse model of inflammation-mediated bone loss. The second Specific Aim is to study the mechanisms by which NO regulates osteoblasts, stimulating certain functions at low concentrations, while becoming cytotoxic and triggering osteoblastic death at high concentrations. The investigators will focus on the processes of proliferation, matrix biosynthesis, bioenergetics and apoptosis, searching for effects on putative intracellular target proteins of NO action including ribonucleotide reductase, aconitase, guanyl cyclase, cyclooxygenase and extracellular matrix proteins which can be nitrated by reactive NO intermediates. Finally, cell-cell signaling action of NO will be tested using osteoblast/osteoclast co-cultures. It is hoped that these studies will establish the physiological significance of the NO/NOS system in bone and the extent to which NO modulates normal hormonal regulation of bone physiology. NO originating from osteoblasts may trigger the extensive osteoclastic bone destruction in inflammatory bone diseases, including rheumatoid arthritis, osteomyelitis and periodontal disease, while simultaneously blocking the process of coupling which normal bone uses to maintain bone mass. Strategies to control osteoblastic NO production may prove effective for pharmacologic intervention in a broad range of bone diseases.