טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentGuetta Julia
SubjectRegulation of the Macrophage Atherogenecity by Different
Protein Products of Haptoglobin Gene
DepartmentDepartment of Medicine
Supervisor Professor Andrew Peter Levy
Full Thesis textFull thesis text - English Version


Abstract

We have recently demonstrated in multiple studies that the haptoglobin 2-2 genotype is associated with an increased risk for diabetic cardiovascular disease. The major function of haptoglobin (Hp) is to bind to hemoglobin and thereby prevent hemoglobin-induced oxidative tissue damage.

We hypothesized that there may be diabetes- and haptoglobin genotype-dependent differences in the atherogenicity of macrophages, that play a central role in atherosclerosis progression, and that these differences are due to the release of active iron from hemoglobin.

 First, we demonstrated increased oxidative stress in tissue culture cells exposed to haptoglobin 2-2-hemoglobin complexes as opposed to haptoglobin 1-1-hemoglobin complexes and showed that this oxidative stress is inhibited by desferrioxamine. Second, we demonstrated marked diabetes-dependent differences in the oxidative stress in peritoneal macrophages of mice, genetically modified to express the mouse homolog of the Hp 2 allele, as compared with the wild type mouse Hp 1 allele. Taken together these data implicate active iron in the increased susceptibility of individuals with the Hp 2 allele to diabetic vascular disease.

In the second part of the work, we sought to determine if the protein products of the two haptoglobin alleles differed in their ability to modulate the cytokine profile produced by macrophages in response to the haptoglobin-hemoglobin complex. Peripheral blood mononuclear cells were isolated from normal human volunteers and cultured in the presence of complexes formed by the protein products of the two different haptoglobin alleles with hemoglobin. The release of specific cytokines in the conditioned media of these cells was assessed by ELISA. We found that the haptoglobin 1-hemoglobin complex stimulated the secretion of significantly more Il-6 and Il-10 than the haptoglobin 2 -hemoglobin complex and that the release of these cytokines is dependent on the binding of the haptoglobin-hemoglobin complex to the CD163 receptor. In addition, we found a highly significant increase in plasma levels of Il-10 in Hp 1 mice after myocardial ischemia and reperfusion as compared to Hp 2 mice.

We hypothesize that haptoglobin genotype modulates the balance of inflammatory (Th1) and anti-inflammatory (Th2) cytokines produced by macrophages exposed to haptoglobin-hemoglobin complex.

Taken together, our findings may have implications in understanding inter-individual differences in the inflammatory response to hemorrhage and in developing a model to test different therapeutic agents to reduce mortality and morbidity from diabetic cardiovascular complications attributed to the Hp polymorphism.