Alcohol-induced Osteoimmunopathies: A Unifying Explanatory Mechanism

描述(申请人提供)：酗酒促进骨质疏松的发展。据估计，酗酒者的骨质疏松症患病率为28-52%，而所有美国成年人的患病率约为10%。酗酒还抑制先天和后天免疫功能，导致更高的感染率和更高的发病率和死亡率。由于酒精对破骨细胞和髓系树突状细胞(MDCs)的起源--破骨细胞-树突状细胞(ODC)前体细胞的影响，这些明显无关的酗酒结果错综复杂地联系在一起。由于其在骨骼维护和免疫细胞产生中的作用，ODC是骨骼和免疫系统的关键。这种相互依赖使骨免疫系统特别容易受到长期饮酒对ODC的影响。RANK/c-fos和Wnt/FrizzledFZD/2-catenin通路分别是ODC向破骨细胞和MDCs分化的两个主要信号系统。这两条信号通路的整合是通过Noch信号发生的。Notch激活通过抑制RANK转录而抑制破骨细胞的生成，同时促进FZD(WnT受体)基因表达，促进树突状细胞的生成。长期饮酒抑制了恒河猴骨髓细胞的缺口活性。此外，长期饮酒通过表观遗传机制(RANK启动子CpG低甲基化和FZD启动子CpG高甲基化)增加RANK的表达，降低FZD受体基因的表达。关于这些前体细胞信号通路之间的相互作用的信息很少，酒精对这些信号和表观遗传机制的影响仍有待研究。我们的假设是，长期饮酒扰乱了控制ODC祖细胞分化平衡的关键信号机制，从而导致破骨细胞生成增强和树突状细胞生成受损。针对三个特定目标设计的实验将检验以下假说：(1)通过上调RANK信号促进破骨细胞的生成；(2)通过抑制FZD/2-catenin信号通路损害树突状细胞的生成；以及(3)改变RANK和FZD基因的启动子CpG甲基化，进一步失衡ODC前体细胞的分化。基因组甲基化技术，以及慢病毒载体的生产和相关的协议将在K99期间掌握。这些技术将被用于在R00期间完成拟议的研究。在R00阶段，资助计划书的撰写和指导职责将补充实验室研究。拟议的实验方法将采用恒河猴慢性酒精消费的临床相关模型和一些尖端技术。这项调查还将确定开发治疗干预措施以治疗酗酒者骨免疫功能障碍的关键目标。

DESCRIPTION (provided by applicant): Alcohol abuse promotes the development of osteoporosis. The prevalence of osteoporosis in alcohol abusers is estimated to be 28-52% compared to approximately 10% prevalence for all adult Americans. Alcohol abuse also suppresses both innate and acquired immune function leading to a higher incidence of infections with increased morbidity and mortality. These apparently unrelated outcomes of alcohol abuse are intricately linked through the effects of alcohol on the cell that is the origin of both osteoclasts and myeloid dendritic cells (mDCs): the osteoclast-dendritic cell (ODC) progenitor. Because of its role in bone maintenance and immune cell production, the ODC is a linchpin of the skeletal and immune systems. This interdependence makes the osteoimmune system particularly vulnerable to the effects of chronic alcohol consumption on the ODC. RANK/c-FOS and Wnt/Frizzled (FZD)/2-catenin pathways are two major signaling systems involved in ODC differentiation to osteoclasts and mDCs, respectively. Integration of these two signaling pathways occurs through NOTCH signaling. NOTCH activation suppresses osteoclastogenesis through inhibiting RANK transcription while promoting FZD (Wnt receptor) gene expression, enhancing dendropoiesis. Chronic alcohol consumption suppressed NOTCH activity in bone marrow cells of rhesus macaques. Furthermore, chronic alcohol consumption increased RANK expression and decreased FZD receptor gene expression through epigenetic mechanisms (RANK promoter CpG hypomethylation and FZD promoter CpG hypermethylation). Little information is available regarding the interplay among these progenitor cell signaling pathways, and the impact of alcohol on these signaling and epigenetic mechanisms remains to be examined. Our hypothesis is that chronic alcohol consumption dysregulates key signaling mechanisms controlling the balance of ODC progenitor lineage differentiation, which leads to enhanced osteoclastogenesis and impaired dendropoiesis. Experiments designed to address three Specific Aims will test the hypotheses that chronic alcohol consumption: (1) promotes osteoclastogenesis through upregulation of RANK signaling; (2) impairs dendropoiesis by inhibiting the FZD/2-catenin signaling pathway; and (3) alters promoter CpG methylation in RANK and FZD genes, further unbalancing ODC progenitor cell differentiation. Genome methylation techniques, as well as lentiviral vector production and the associated protocols will be mastered during the K99 period. These techniques will be employed to complete the proposed research during the R00 period. Grant proposal writing and mentoring duties will complement laboratory research during the R00 phase. The proposed experimental approach will employ a clinically relevant model of chronic alcohol consumption in rhesus macaques and a number of cutting edge technologies. This investigation will also identify key targets for developing therapeutic interventions to treat osteoimmune dysfunction in alcohol abusers.