Does the H67D HFE genotype impact macrophage phenotype and disease process?
Open Access
- Author:
- Nixon, Anne Marie
- Graduate Program:
- Anatomy
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- July 25, 2017
- Committee Members:
- James R. Connor, Dissertation Advisor/Co-Advisor
James R. Connor, Committee Chair/Co-Chair
Gregory M. Holmes, Committee Member
Patricia J. McLaughlin, Committee Member
Robert F. Paulson, Outside Member - Keywords:
- Iron
HFE
Macrophages
Parkinson's Disease
Paraquat
Neurodegeneration - Abstract:
- Iron homeostasis is tightly regulated to prevent disease resulting from too little or too much iron within the body. The HFE protein is involved in iron regulation and when mutated it is associated with several iron-related disorders. In particular, the H63D HFE mutant variant is the most prevalent HFE mutation in the Caucasian population. Although HFE is known to modulate iron homeostasis through its interaction with transferrin receptor (TfR), its precise role in iron metabolism and cellular function remain unclear. One cell type in the body whose function may be especially impacted by the HFE mutation is the macrophage. Macrophages play critical roles in iron regulation through scavenging and recycling of iron, providing a mechanism to both reduce and reallocate excess iron. The effects of perturbed iron regulation in macrophages in neurodegenerative disease progression is not yet established. Here, this thesis aims to investigate the role of HFE in macrophage function, and determine how HFE mutant macrophages contribute to neurodegenerative diseases, specifically Parkinson’s Disease. We hypothesize that the H67D macrophages would have altered iron handling and inflammatory response, promoting Parkinson’s progression. To investigate the role of H67D genotype on macrophage phenotype, bone marrow macrophages from H67D mutant mice were generated and compared to wildtype derived macrophages for the status of iron related proteins, inflammatory cytokine secretion, and macrophage-related cell functions (i.e. migration and phagocytosis). The H67D macrophages had increased levels of L-ferritin, but no significant differences in TfR or ferroportin, indicating differences in iron handling specifically in the resting state. In addition, H67D macrophages functionality was altered as they displayed decreased migration, and increased phagocytosis iii activity compared to WT. The H67D macrophages also expressed increased levels of chemoattractant proteins and decreased inflammatory cytokines. The alterations in macrophage function with the HFE mutation could impact the role of these cells in neurodegenerative disease. Therefore, we generated a Parkinsonism mouse model using paraquat to selectively target and induce the degeneration of dopaminergic neurons within the nigrostriatal pathway. In this model, we found WT mice had impaired motor function, and decreased number of tyrosine hydroxylase (TH) neurons, whereas the H67D mice showed no motor impairment following exposure to paraquat. Surprisingly, the mice with the H67D mutation had a significant reduction in TH neurons in both treated and untreated mice compared to WT. Thus, the HFE mutation was associated with significant loss of TH neurons but this loss was not increased by paraquat. The loss of these neurons could be the result of a developmental flaw or due to an initial stressor that led to neuroprotection; which is discussed in the final chapter. The third chapter used the same paraquat model to determine if there was a genotype effect of cholesterol or statin exposure. Cholesterol and statin have been proposed as agents that progress Parkinson’s Disease. Moreover, the H63D/H67D mutation is associated with disruption in cholesterol metabolism. The results showed that the cholesterol diet negatively affected motor function of WT mice when injected with paraquat, however there was no effect on motor function with a statin diet. The H67D mice have no change in motor function when on either a cholesterol or statin diet indicating the H67D genotype is resistant to the toxic effects of cholesterol. In addition, a statin diet does not exacerbate the results of paraquat induced neurotoxicity. iv v Our results show that the H67D genotype plays a role in iron and immune response of macrophages but also can be neuroprotective in a Parkinsonism disease model possibly due to the initial stress exerted by the macrophages. Overall, these findings encompass a complete role of H67D from periphery to a disease model. The H67D mouse model can further be used to interrogate the mechanism for the neuroprotection against exposure to paraquat and to cholesterol plus paraquat. These data further support the concept that the HFF genotype is a disease modifier.