Role of the redox responsive transcription factor, NRF2 in immune cell function

Hamdam, Junnat Role of the redox responsive transcription factor, NRF2 in immune cell function. Doctor of Philosophy thesis, University of Liverpool.

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Abstract

Dendritic cells (DCs) are potent innate antigen presenting cells which are able to sense and engulf pathogens from sites of infection, which are then processed and presented to adaptive T lymphocytes in secondary lymphoid organs. Therefore, they are critical for the initiation and modulation of primary antigen-specific adaptive immune responses. They also play a critical role in the maintenance of T cell tolerance. T cells play vital roles in mediating both cellular and humoral-specific adaptive immune responses. There are various types of T cells present within the immune system including the cytotoxic CD8 T cells and T helper CD4 cells. CD4 T cells can be further subdivided into various subtypes examples of which include T helper 1 cells (Th1), Th2, Th17 and T regulatory cells. Each subset has its own distinctive function, transcriptional regulation and effector cytokine profile. It is established that appropriate DC and T cell immune function is highly dependent on their intracellular redox status. Cellular redox homeostasis is maintained through a balance between oxidising agents e.g. reactive oxygen species (ROS) and anti-oxidant or reducing agents e.g. glutathione (GSH). Excessive ROS production resulting in oxidative stress is extremely deleterious to the cell and if left unimpeded can result in cell necrosis, tissue damage and the onset of disease. As a result, mammalian cells have evolved an inducible adaptive defence system which provides protection against such oxidative or chemical insult. The functionality of this cellular defence system is principally governed by the activity of the redox-sensitive transcription factor Nrf2. In response to oxidative stress, Nrf2 induces the transcription of a battery of cytoprotective and antioxidant genes involved in GSH synthesis, detoxification of xenobiotics and their reactive metabolites and the maintenance of cellular redox homeostasis. It is now emerging that Nrf2 plays a pivotal role in immunity. However, its precise role in DC and T cell function is unclear. Using immature bone marrow-derived DCs (iDCs) from Nrf2+/+ and Nrf2-/- mice, the work presented in this thesis demonstrates that Nrf2 deficiency in iDCs resulted in lowered GSH levels, enhanced iDCs co-stimulatory receptor expression, impaired endocytic and phagocytic capacity, and increased iDC-mediated antigen-specific CD8 T cell stimulatory capacity in response to both an antigenic and self-peptide. Furthermore, artificially lowering GSH levels in the iDCs did not recapitulate the Nrf2 deficient iDC phenotype. Moreover, Nrf2-/- DCs exhibited an enhanced capacity to present cell-associated peptide antigens to antigen-specific CD8 T cells, resulting in increased CD8 T cell effector function. Loss of Nrf2 in LPS-stimulated DCs results in a lowered Th1 cytokine profile. These results have implications for Nrf2 in DC-mediated CD8 T-cell immunity, peripheral CD8 T cell tolerance and CD4 effector differentiation. The role of Nrf2 in T cell function is poorly understood. Using thymocytes, splenocytes and lymph-node derived T cells from Nrf2+/+ and Nrf2-/- mice, we demonstrate that loss of Nrf2 did not affect the development of CD4+CD25+ natural occurring Treg, mature CD4 and CD8 T cell populations within the thymus. Furthermore, Nrf2 deficiency did not alter the composition of CD4 and CD8 T cell populations within secondary lymphoid organs. It was observed that splenic Nrf2-/- naïve T cells exhibited enhanced ROS generation, accompanied by low level increases in T cell activation markers. However, the marginal augmentation of Nrf2-/- naïve T cell activation status did not result in increased T cell receptor (TCR)-triggered T cell proliferation. In contrast, Nrf2 deficient effector T cells exhibited enhanced TCR/CD3-triggered proliferation, associated with increased Th1 and decreased Th2 effector function. Importantly, we demonstrated that Nrf2-/- effector T cells secreted increased levels of IL-17A and IL-22, a signature cytokine profile indicative of the more recently identified Th17 cell lineage. This was also observed under Th17 polarising conditions, further suggesting that loss of Nrf2 predisposes effector Th17 development. The implications of the latter findings are significant given the pivotal role that Th17 cells play in the pathogenesis of a variety of autoimmune diseases including multiple sclerosis (MS) and Systemic lupus erythematosus (SLE). Nrf2 plays a critical role in the detoxification of xenobiotics in the liver, which as the primary drug-metabolising organ, is subjected to an array of xenobiotics and their respective metabolites. Individuals vary in their responses to xenobiotic exposure from adaptation to severe adverse drug reactions. However, it is unknown whether this human disparity in drug response is a consequence of inter-individual variation in the Nrf2 adaptive defence system to xenobiotic stress. In light of this, we aimed to firstly investigate whether variation in the Nrf2 adaptive system was present within individuals in response to a chemical inducer of Nrf2, CDDO-Me. To address this issue, basal and induced Nrf2 protein levels and downstream NQO1 expression were measured in activated human T cell blasts, in response to increasing concentrations of CDDO-Me. Examination of various donor-derived T cells, demonstrated that humans vary in their Nrf2 response to CDDO-Me, with respect to nuclear Nrf2 and NQO1 mRNA expression. Therefore we concluded that inter-individual variation does exist in the human’s Nrf2 adaptive system in response to a known Nrf2 probe. Overall, the experimental results obtained in this PhD programme have provided detail on the role of Nrf2 in immune cells and highlights the potential for therapeutically targeting Nrf2 in immune-mediated disease.

Item Type:	Thesis (Doctor of Philosophy)
Additional Information:	Date: 2013-08 (completed)
Subjects:	?? RM ??
Divisions:	Faculty of Health and Life Sciences
Depositing User:	Symplectic Admin
Date Deposited:	11 Feb 2014 16:01
Last Modified:	16 Dec 2022 04:41
DOI:	10.17638/00014133
Supervisors:	Sathish, Jean Goldring, Christopher
URI:	https://livrepository.liverpool.ac.uk/id/eprint/14133