ETHANOL MEDIATED OSTEOPOROSIS AND INTERLEUKIN 6

描述（改编自研究者摘要）： 确定慢性酒精暴露可诱发骨质疏松症， 使人衰弱和昂贵的疾病。 然而，分子机制通过 乙醇对骨的影响是未知的。 有证据 乙醇既能增加骨吸收又能抑制骨生成， 导致骨质减少。 促炎细胞因子白细胞介素-6 IL-6是骨上破骨细胞活性的重要介质。 此外，研究人员提供的证据表明，乙醇诱导IL-6 人骨髓基质细胞中IL-6的表达和IL-6启动子活性 并增加小鼠血清IL-6水平。 因此，他们假设 乙醇诱导骨中IL-6基因转录， 最终导致破骨细胞介导的骨吸收的微环境。 为了探索这一假设，研究人员将执行以下操作 具体目标：1. 阐明了乙醇对 IL-6启动子在人骨髓基质细胞中的转录活性 细胞; 2. 乙醇对IL-6影响程度及方法的测定 人骨髓基质细胞中mRNA的稳定性; 3. 证明IL-6 有助于乙醇诱导的小鼠体内骨丢失的机制。 目标1将通过a）评估乙醇对核反应堆的影响来实现。 转录物产生，B）IL-6启动子的突变分析， 乙醇响应性顺式作用元件，和c）测量 乙醇介导的人骨髓转录因子活性变化 基质细胞系和原代人基质细胞。 目标2将是 通过评估a）乙醇诱导的调节mRNA半衰期， B）IL-6 mRNA的3-非翻译区（UTR）的突变分析 以鉴定受乙醇影响的区域，以及c）鉴定 乙醇诱导的人3 -UTR上反式作用因子的变化 骨髓基质细胞系和原代人基质细胞。 目标3将是 通过评估小鼠骨代谢的差异来完成， 不能表达IL-6（IL-6基因敲除）和对照小鼠（其可以表达 IL-6）喂养液体乙醇饮食3或24周。 骨代谢将是 通过各种测量进行评估，包括骨矿物质的变化 含量通过总骨骼灰烬、骨髓破骨细胞产生来评估 通过骨髓培养，通过小窝吸收测定的破骨细胞活性，体内骨 通过尿吡啶啉交联的再吸收和通过 骨组织形态计量学

DESCRIPTION (Adapted from the Investigator's Abstract): It is well established that chronic ethanol exposure can induce osteoporosis, a debilitating and costly disease. However, the molecular mechanisms through which ethanol achieves its influence on bone is unknown. There is evidence that ethanol may both increase bone resorption and inhibit bone production, resulting in osteopenia. The proinflammatory cytokine, interleukin-6 (IL-6), is an important mediator of osteoclast activity on bone. Furthermore, the investigators provide evidence that ethanol induces IL-6 expression and IL-6 promoter activity in a human bone marrow stromal cell line and increases serum IL-6 levels in mice. Accordingly, they hypothesize that ethanol induces IL-6 gene transcription in bone and the bone microenvironment culminating in osteoclast-mediated bone resorption. To explore this hypothesis, the investigators will perform the following specific aims: 1. Elucidate the mechanism of ethanol s influence on transcriptional activity of the IL-6 promoter in human bone marrow stromal cells; 2. Determine the degree and method of ethanol s influence on IL-6 mRNA stability in human bone marrow stromal cells; 3. Demonstrate that IL-6 contributes to the mechanism of ethanol-induced bone loss in vivo in mice. Aim 1 will be accomplished by a) evaluation of ethanol s effect on nuclear transcript production, b) mutational analysis of the IL-6 promoter for ethanol-responsive cis-acting elements, and c) measurement of ethanol-mediated changes in transcription factor activity in a human marrow stromal cell line and primary human stromal cells. Aim 2 will be accomplished by evaluation of a) ethanol induced modulation mRNA half-life, b) mutational analysis of the 3-untranslated region (UTR) of the IL-6 mRNA to identify regions effected by ethanol, and c) identification of ethanol-induced changes in trans-acting factors on the 3 -UTR in human marrow stromal cell line and primary human stromal cells. Aim 3 will be accomplished by evaluating for differences in bone metabolism in mice which cannot express IL-6 (IL-6 gene knockout) and control mice (which can express IL-6) fed a liquid ethanol diet for 3 or 24 weeks. Bone metabolism will be evaluated by a variety of measurements including alterations in bone mineral content as assessed by total skeletal ash, bone marrow osteoclast production by marrow culture, osteoclast activity by pit resorption assay, in vivo bone resorption by urinary pyridinoline crosslinks, and osteoclast activity by bone histomorphometry.