טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentStrauss Merav
SubjectThe Molecular Mechanisms Responsible for the Divergent
Regulation of Cytokine Secretion by Macrophages
after Binding Hb-Hp1 and Hb-Hp2 to CD163
DepartmentDepartment of Medicine
Supervisor Professor Andrew Peter Levy
Full Thesis textFull thesis text - English Version


Abstract

It has been recently demonstrated in multiple independent population-based longitudinal and cross sectional analyses that the haptoglobin (Hp) 2-2 genotype is associated with an increased risk for diabetic cardiovascular disease. The major function of Hp is to bind to hemoglobin (Hb) and thereby prevent Hb-induced oxidative tissue damage. This antioxidant function of Hp is mediated in part by the ability of Hp to prevent the release of iron from bound Hb. In addition to serving as an antioxidant it has been proposed that Hp, as mediated through the CD163 receptor, may also have an immunomodulatory function.

In this work, we first sought to determine if the protein products of the two Hp alleles differed in their ability to modulate the cytokine profile produced by macrophages in response to the Hp/Hb complex. Peripheral blood mononuclear cells (PBMCs) were isolated from normal human volunteers and cultured in the presence of complexes formed by the protein products of the two different Hp alleles with Hb. The release of specific cytokines in the conditioned media of these cells was assessed by ELISA. We found that the Hp1-1-Hb complex stimulated the secretion of significantly more IL-6 and IL-10 than the Hp2-2-Hb complex. We demonstrate that the release of these cytokines is dependent on the binding of the Hp-Hb complex to the CD163 receptor.

Studies using specific protein kinase inhibitors have revealed that both CK2 and protein kinase C (PKC) are involved in the CD163 signaling mechanism that leads to increased cytokine production. This study investigated whether the amount of CK2 associated with CD163, or the activity of CK2 associated with CD163, differs following binding of the different Hp-Hb complexes to CD163. We found no significant difference in the amount of CK2 (neither the alpha nor the beta subunit) associated with CD163 following stimulation with Hp-Hb complexes. However, we found that the activity of CK2 is differentially modulated following treatment of PMBCs with Hp-Hb complexes. Specifically, binding of Hp1-1-Hb results in a significant increase in CD163- associated CK2 activity compared to Hp2-2-Hb complexes. We hypothesize that Hp1-1-Hb increased cytokine production may be due to increased CK2 activity. Therefore, strategies designed to mimic the increased CD163-associated CK2 activity may have a protective effect in atherosclerosis.