CELLULAR AND MOLECULAR BIOLOGY OF CHOLESTEATOMA

耳胆脂瘤起源于鼓膜，其后遗症 中耳炎。胆脂瘤是进行性的上皮性病变， 破坏中耳的骨质结构，有时甚至破坏内耳。 胆脂瘤一旦确诊，只能通过手术来强制治疗。 根治及二期中耳再造术。长期的 首席调查员的研究计划的目标是 了解导致发育的细胞机制， 胆脂瘤的进展和骨质破坏。如此卑鄙的- 站立可能导致非手术预防和治疗的策略 这种疾病的管理。 本申请被设计用来调查 两个重要的细胞间信号分子--白介素1 和一氧化氮(NO)，与胆脂瘤破骨细胞性骨侵蚀有关。 本申请的具体目的是：1)确定 白介素族细胞因子(IL-1α、IL-1)的表达 和IL-1ra)在实验性胆脂瘤中的表达 压力诱导沙土鼠破骨细胞的募集和激活 同步骨模型的Bulla模型；2)研究异构体 一氧化氮合酶(NOS I、II、III)在局部调控中的作用 体外和体内破骨细胞。 有两种广泛的假设，每一种都集中在IL-1和IL-1的亚型上 不是。对于特定的Aim1，将检验假设1：IL-1亚型为 在亚成骨细胞和成纤维细胞中转录 胆脂瘤旁上皮和加压大疱处及 代表着招募和激活的早期信号 局部部位的破骨细胞。实验性沙鼠胆脂瘤， 沙土鼠加压水泡，PCS，聚合酶链式反应产物克隆，DNA 测序、定量RT-PCR、骨组织形态计量学和原位RNA 杂交将被用来检验第一个假设。 对于特定的目标2，假设2将被检验：NO是由向上- 骨衬细胞和成纤维细胞内一氧化氮合酶亚型的调节 胆脂瘤旁和加压下的亚上皮 布拉。这导致了NO的本地细化和招募 和激活破骨细胞。小鼠颅骨器官培养 (元素钙释放和45Ca释放)，沙土鼠加压 Bulla，聚合酶链式反应，聚合酶链式反应产物克隆，DNA测序，定量RT- 聚合酶链式反应、骨形态计量学、RNA原位杂交和NADPH- 黄递酶组织化学将被用来检验第二个假说。

Aural cholesteatomas arise from the tympanic membrane as a sequela of otitis media. Cholesteatomas are progressive, epithelial lesions that destroy the bony structures of the middle and, sometimes, inner ear. Once established, cholesteatomas can only be mandated by surgical eradication and secondary middle ear reconstruction. The long-term objective of the Principal Investigator's research program is to understand the cellular mechanisms that lead to the development, progression, and bone destruction of cholesteatomas. Such an under- standing may lead to strategies for the non-surgical prevention and management of this disease. The present application is designed to investigate the relationship of two important intercellular signaling molecules, interleukin-1 (IL-1) and nitric oxide (NO), to osteoclastic bone erosion in cholesteatoma. The specific aims of this application are: 1) to determine the expression of the interleukin family of cytokines (IL-1 alpha, IL-1 beta, and IL-1ra) in experimental cholesteatoma and also during osteoclast recruitment and activation induced by pressure in the gerbil bulla model of synchronous bone modeling; and 2) to investigate isoforms of nitric oxide synthase (NOS I, II, III) in the local control of osteoclasts in vitro and in vivo. There are two broad hypothesis, each focused on the isoforms of IL-1 and NOS. For Specific Aim1, hypothesis 1 will be tested: IL-1 isoforms are transcribed within bone lining cells and fibroblasts of the sub- epithelium subjacent to cholesteatoma and in the pressurized bulla and represent an early signal for the recruitment and activation of osteoclasts at a local site. Experimental gerbilline cholesteatoma, gerbilline pressurized bulla, PCS, cloning of PCR products, DNA sequencing, quantitative RT-PCR, bone histomorphometry, and RNA in situ hybridization will be used to test this first hypothesis. For Specific Aim 2, hypothesis 2 will be tested: NO is produced by up- regulation of NOS isoforms within bone lining cells and fibroblasts of the sub-epithelium subjacent to cholesteatoma and in the pressurized bulla. This leads to the local elaboration of NO and the recruitment and activation of osteoclasts. Mouse calvarial organ culture (elemental calcium release and 45Ca release), gerbilline pressurized bulla, PCR, cloning of PCR products, DNA sequencing, quantitative RT- PCR, bone histomorphometry, RNA in situ hybridization, and NADPH- diaphorase histochemistry will be used to test this second hypothesis.