New pathways in the pathogenesis of rheumatoid arthritis

Elmesmari, Aziza Atiya (2014) New pathways in the pathogenesis of rheumatoid arthritis. PhD thesis, University of Glasgow.

Abstract

Rheumatoid Arthritis (RA) is a common chronic autoimmune disease that is characterized by synovial tissue inflammation eventually leading to joint destruction with severe functional deterioration and increased mortality – the underlying pathogenesis of RA remains unsolved. The principle of new therapeutic development is to define and characterize a molecular pathway both in terms of its basic biology and also its context-dependent effects in the synovial compartment. A hallmark pathological feature of RA is a rapid influx and accumulation of immune cells such as monocytes/macrophages into the synovium. Monocytes/ macrophages are major effector cells in RA synovitis, principally acting by releasing TNF-α, IL-6 and other inflammatory cytokines and chemokines. The recruitment of effectors cells is an important step in RA progression and is mediated by chemokines and their receptors. Two pathways will be studied in this project, both with potential relevance to the accumulation and activation of inflammatory leukocytes to the synovium, namely microRNAs and sphingolipid enzymes.
MicroRNAs are a recently discovered class of post-transcriptional regulators that induce mRNA target degradation or translation inhibition. They have been shown to be involved in the regulation of the immune response and the development of autoimmunity. Of particular interest in the context of RA is miR-155, which is upregulated in RA synovial macrophages where it regulates cytokine expression such as TNF-α. Until now little was known about the role of miR-155 in regulating monocyte migration. Therefore, we sought to focus on the functional contribution of miR-155 in monocyte migration by the modulation of the expression of chemokines and their reciprocal chemokine receptors. Firstly the absolute copy numbers of miR- 155 transcripts in peripheral blood (PB) and synovial fluid (SF) monocytes of RA and healthy controls were assessed. To examine the role of miR-155 in monocyte migration and retention in the joint space, I overexpressed miR-155 in PB CD14+ monocytes of healthy controls and RA patients and examined the expression of chemokines and chemokine receptors mRNA levels by taqman low-density array (TLDA) and quantified the production of these chemokines in culture supernatant by multiplex assay. The role of miR-155 was investigated further using bone marrow monocytes (BMMO) from miR-155−/− and wild type (WT) mice. RA PB and SF CD14+ monocytes expressed higher copy numbers of miR-155 compared with healthy controls. RA SF monocytes exhibited the highest expression levels of miR- 155. The copy number of miR-155 expression was significantly increased in anti- citrullinated protein antibody (ACPA) positive RA compared with ACPA negative RA. The RA PB monocyte miR-155 copy number correlated positively and significantly with DAS28. Overexpression of miR-155 in PB monocytes led to an increased production of chemokines (CCL3/MIP-1α, CCL4/MIP-1β, CCL5/ RANTES, and CCL8/MCP2) and a reduction in expression of inflammatory chemokine receptor CCR2 while homeostatic CCR7 was up regulated. Commensurate with this, these receptors were expressed in an opposite direction inBMMO from miR-155 deficient cells; CCR7 was significantly down regulated and CCR2 expression level was increased. These observations suggest that CCR2 and CCR7 were under the tight control of miR-155 and that this regulation is preserved across the species; together suggesting that miR-155 can act as an important regulator of these receptors. We conclude that deregulation of miR- 155 in RA monocytes contributes to monocyte retention at sites of inflammation due to induction of chemokine production and down-regulation of inflammatory chemokine receptors. Furthermore, these data imply that miR-155 levels may reflect RA disease activity and could be a potential diagnostic or clinical disease activity biomarker for RA.
Sphingosine kinases (SPHKs), SPHK1 and SPHK2, are isozymes that phosphorylate sphingosine into sphingosine-1- phosphate (S1P). S1P, a pleiotropic lipid mediator of inflammation, subsequently binds with any of the five G-protein coupled protein S1P receptors (S1PR1-5) and stimulates an array of cellular responses. Defects in
S1P/S1PRs signalling have been shown to be associated with various pathologies. Until now however, no comprehensive analysis of expression of its components in RA has been performed. My data show that S1P concentrations were significantly elevated in the serum of RA patients with active disease compared to RA patients in remission and in healthy controls. Moreover, S1P1, S1P3, S1P5 and SPHK1 were differentially regulated in RA immune cell subsets, such as neutrophils, monocytes (CD14+), lymphocytes (CD4+ and CD8+) compared to healthy controls. In addition, compared with osteoarthritis (OA) pathological control, RA synovial tissues were strongly positive for the SPHK1, S1P1, and S1P3. Interestingly, RA patients treated with biological DMARDs had attenuated levels of SPHK1, S1P3 and S1P5, but not S1P1, when compared with patients treated with conventional DMARDs. Therefore, my study warrants further investigation of the clinical significance of S1P as a biomarker for disease activity and to explore the utility of novel therapeutic tools available to modulate the SPHK/S1PR/S1P axis in RA with a view to defining new therapeutic possibilities.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Rheumatoid Arthritis, MicroRNAs, miR-155, monocyte migration, chemokine, chemokine receptor, Sphingosine kinases and S1P receptors.
• Subjects: Q Science > QR Microbiology > QR180 Immunology; R Medicine > R Medicine (General); R Medicine > RC Internal medicine
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Infection & Immunity
• Supervisor's Name: McInnes, Professor Iain
• Date of Award: 2014
• Depositing User: Dr Aziza Elmesmari
• Unique ID: glathesis:2014-5706
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 31 Oct 2014 16:43
• Last Modified: 16 Oct 2017 08:34
• URI: https://theses.gla.ac.uk/id/eprint/5706