Regulation and Function of Macrophage CD163

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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.