A Role of Osteoimmunology in Leptin-Regulated Bone Metabolism

描述（由申请人提供）：骨免疫学被定义为免疫系统和骨代谢之间的相互作用。这一领域是从认识到在炎症和应激时期，免疫细胞在骨骼中产生的分泌因子可以深刻地影响骨骼调节而发展起来的。瘦素是一种由含脂肪细胞产生的16kda脂肪细胞因子。瘦素与下丘脑中央长形受体的结合通过降低成骨细胞功能刺激骨质流失。体外实验也证明，瘦素与骨髓基质细胞（BMSCs）的直接结合可以促进它们向成骨谱系分化。我们假设并产生了初步数据，表明瘦素介导的骨调节存在新的第三种机制。据推测，瘦素通过与巨噬细胞等免疫细胞结合，通过继发性改变骨髓间充质干细胞分化和成骨细胞功能，间接调节骨量，在骨髓外周发挥作用。我们将探索这些骨免疫相互作用，以更好地确定骨髓间充质干细胞的生态位，并确定我们的免疫系统在出生后骨骼维护中可能发挥的作用。为了在瘦素的背景下探索我们的假设，我们将采用体外和体内技术。瘦素和瘦素受体缺陷小鼠模型将用于收获原代巨噬细胞和骨髓间充质干细胞。巨噬细胞分泌矿化和分化调节细胞因子的能力将通过珠状多通道酶联免疫吸附试验进行分析，Bio-Rad公司将其命名为BioPlex。在对照或刺激巨噬细胞及其产物存在的情况下，也将监测骨髓间充质干细胞的分化。最后，通过将收获的骨髓间充质干细胞皮下移植到明胶泡沫支架中，探讨瘦素对体内骨形成的局部调节。所得到的听骨将使用显微ct和免疫组织学技术进行定性分析。骨免疫学的这一概念暗示了骨髓腔中形成和影响的循环，对炎症相关的骨骼疾病（包括骨质疏松症、骨转移癌、关节炎、骨髓炎和牙周病）的研究具有重要意义。

DESCRIPTION (provided by applicant): Osteoimmunology is defined as the interplay between the immune system and bone metabolism. This field has developed from the realization that during times of inflammation and stress, immune cells generated in the bone secrete factors that can profoundly affect bone regulation. Leptin is a 16 kDa adipocytokine produced by fat-containing cells. Binding of leptin to central long-form receptors in the hypothalamus stimulates bone loss by decreasing osteoblast function. It has also been demonstrated in vitro that direct binding of leptin to bone marrow stromal cells (BMSCs) can promote their differentiation toward an osteogenic lineage. We hypothesize and have generated preliminary data to suggest that a novel third mechanism of leptin-mediated bone regulation exists. It is hypothesized that leptin acts peripherally in the bone marrow by binding to immune cells such as macrophages to indirectly regulate bone mass through secondary changes in BMSC differentiation and osteoblast function. We will explore these osteoimmunologic interactions to better define the niche of the BMSC and to determine the role our immune system may play in post-natal bone maintenance. To explore our hypothesis in the context of leptin we will employ both in vitro and in vivo techniques. Leptin and leptin-receptor deficient mouse models will be utilized for harvest of primary macrophages and BMSCs. Macrophage cells will be analyzed for their ability to secrete mineralization and differentiation-regulating cytokines through the use of bead-based multichannel ELISA, marketed as BioPlex by Bio-Rad. Differentiation of BMSCs will also be monitored in the presence of control or stimulated macrophages and their products. Finally, local leptin regulation of in vivo bone formation will be explored by re-implanting harvested BMSCs subcutaneously in gelfoam scaffolds. The resulting ossicles will be qualitatively analyzed using micro-CT and immunohistological techniques. This concept of osteoimmunology implies a circle of creation and influence in the marrow cavity and has important implications for the study of inflammation-linked bone disorders including osteoporosis, cancer metastasis to the bone, arthritis, osteomyelitis and periodontal disease.