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Title: Investigating the mechanisms underlying the pathogenesis of alopecia areata
Author: Hamed, Nagy
ISNI:       0000 0004 7425 7255
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2018
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Alopecia areata (AA) is an autoimmune disease of hair follicles (HFs). The exact pathogenesis is unclear, but it is believed to result from T-cell mediated destruction of anagen HF due to the collapse of immune privilege (IP). In this study, we hypothesised that IP collapse is a consequence of an imbalance between regulatory T-cells (Treg) and inflammatory T-cells (Teff), which leads to the upregulation of the expression of Major Histocompatibility Complex (MHC) class I and class II molecules in HF, resulting in IFN-? mediated attack of Teffs against HFs. To test this hypothesis, we compared circulating lymphocytes from AA patients and healthy controls (HCs), using a flow-cytomteric technique. Data analysis showed that CD39 and HLA-DR+ suppressive Tregs were significantly reduced in AA patients indicating impaired function of circulating Tregs. This reduction was accompanied by a marked increase in the circulating Teff proportion, particularly in activated CD8+ T-cells (NKG2D+CD8+ T-cells), Th17 and Th1. These findings were supported by immunofluorescence (IF) staining of AA skin sections, showing a marked reduction of CD39+ Tregs in diseased HF. To further characterise circulating lymphocytes, next generation sequencing (NGS) was performed to analyse T-cell receptors chain ? (TCR?) in patients and controls. Sequencing analysis revealed a predominance of two unique TCR? clones in the total lymphocyte population shared by many AA patients suggesting clonal expansion in response to a specific antigen during the disease process. Interestingly, database searches found that these clones have over 80% amino acid identity to TCR clones from CD8+ T-cells isolated from diseased subjects, and therefore could be pathogenic. There was also predominance of two public TCR clones within the Treg population in the HC group; suggesting that these clones have a protective effect. Our data support the hypothesis of a potential Treg role in preventing AA as their impairment leads to Teff predominance in peripheral blood, therefore specific Treg clones might be used for therapeutic purposes. Epigallocatechin gallate (EGCG) was proposed as a treatment to reestablish Teff-Treg balance, due to its inhibitory effect on the Teff signalling pathway (IFN-?-JAK-STAT) and its stimulatory effect on Treg differentiation. EGCG showed an inhibitory effect on the JAK-STAT pathway by reducing the levels of p-STAT-1 protein in vitro in the Keratinocyte cell line (HaCat) and in the lymphocyte cell line (Jurkat); as well as ex vivo in PBMCs isolated from patient blood. The reduction of p-STAT1 was accompanied by a significant decrease in the expression of MHC class I and II genes in HaCat cells as well as a decrease in the proportion of activated CD8+ T-cells and Th1 cells in peripheral blood of AA patients. This data would support the development of a clinical pilot study to measure the efficacy of EGCG in enhancing hair re-growth in AA patients. To sum up, it has been demonstrated that Teff-Treg imbalance could have a role in IP collapse. Our data brings a new insight into the current understanding of the immunological basis for AA pathogenesis and could be used to facilitate the development of an effective therapeutic intervention.
Supervisor: Tazi-ahnini, Rachid ; McDonagh, Andrew Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available