Regulation and Function of Macrophage CD163

描述（由申请人提供）：最近的关键观察表明CD163在炎症调节中起重要作用。CD163仅在单核细胞（Mo）和巨噬细胞（Mo）上表达，去年被证明是血红蛋白-接触珠蛋白（Hb/Hp）复合物的内吞清除受体。值得注意的是，触珠蛋白有两种与CD163具有不同亲和力的等位基因形式，并且触珠蛋白表型是糖尿病肾病和冠状动脉血管成形术后再狭窄的预测因子。此外，CD163在单核细胞上的表达被糖皮质激素和IL-10显著增加，这两种介质在创伤或感染反应中迅速增加，并且具有降低脂多糖（LPS）毒性的良好特征。使用新开发的ELISA，我们发现(i)暴露于低至50 pg/ml的脂多糖（LPS）后，CD163的细胞外结构域迅速从单核细胞脱落；（ii）在体外循环心脏手术期间，血浆中可溶性CD163 （sCD163）迅速显著上升，以及在实验对象中大量注射LPS后。因此，有必要更好地了解CD163的产生和功能。这项RO3应用的目标是生成试剂和试点数据，使PI能够建立一个新的研究项目，专注于了解CD163在调节炎症中的作用。特异性目的旨在阐明糖皮质激素、细胞因子和LPS对CD163表达和脱落的影响的分子机制。此外，新测序的小鼠CD163将在合成、脱落和Hb/Hp摄取方面与人类CD163进行比较，以便为未来的研究建立合适的小鼠模型。新的单克隆抗体（mab）、可溶性CD163融合蛋白和表达人、鼠CD163的转染物也将被开发出来。融合蛋白和单克隆抗体将通过原代Mo培养物、Mo细胞系和转染物测试通过CD163对Hb/Hp复合物摄取的抑制作用。这些试剂还将用于中试实验，探讨表面和可溶性CD163对免疫功能的影响。机制和功能信息的发展，人类和小鼠试剂的产生，以及小鼠模型的潜在验证，应该为更精确地确定CD163在炎症控制中的作用奠定基础。

DESCRIPTION (provided by applicant): Recent key observations suggest that CD163 plays an important role in the regulation of inflammation. Expressed only on monocytes (Mo) and macrophages (MO), CD163 was shown last year to be an endocytic scavenger receptor for hemoglobin-haptoglobin (Hb/Hp) complexes. It is notable that haptoglobin has two allelic forms with different affinities for CD163, and haptoglobin phenotype is a predictor of diabetic nephropathy and of restenosis after coronary angioplasty. Moreover, CD163 expression on monocytes is markedly increased by glucocorticoids and IL-10--two mediators that increase rapidly in response to trauma or infection and are well characterized for reducing lipopolysaccharide (LPS) toxicity. Using a newly developed ELISA we show (i) that the extracellular domain of CD163 is rapidly shed from the monocyte after exposure to as little as 50 pg/ml of lipopolysaccharide (LPS) and (ii) that soluble CD163 (sCD163) in plasma rises rapidly and markedly during cardiac surgery with cardiopulmonary bypass, as well as following a bolus injection of LPS into experimental human subjects. A better understanding of the production and function of CD163 is therefore warranted. The goal of this RO3 application is the generation of reagents and pilot data that will enable the PI to establish a new research program focused on understanding the role that CD163 plays in regulating inflammation. The Specific Aims are designed to elucidate the molecular mechanisms of glucocorticoid, cytokine and LPS effects on CD163 expression and shedding. In addition, the newly sequenced murine CD163 will be compared to the human CD163 in terms of synthesis, shedding and Hb/Hp uptake in order to establish an appropriate murine model for future studies. New monoclonal antibodies (mAbs), soluble CD163 fusion proteins, and transfectants expressing both human and murine CD163 will also be developed. Fusion proteins and mAbs will be tested for inhibition of uptake of Hb/Hp complexes via CD163 using primary Mo cultures, Mo cell lines and transfectants. These reagents will also be used in pilot experiments to probe the effect of surface and soluble CD163 on immunological functions. The development of mechanistic and functional information, the generation of human and mouse reagents, and the potential validation of a mouse model should lay the foundation for an expanded application to determine more precisely the role of CD163 in the control of inflammation.