A general view is that initiation of adaptive immune responses to pathogens is divided into two parts, the recognition of 'danger'via the innate immune system and the recognition of foreign antigen by the adaptive immune system. It is widely accepted that a breach in tolerance in the adaptive immune response leads to recognition of self-antigen contributing to autoimmunity. In this model, the source of 'danger'has never been completely defined. We propose a new model whereby the source of 'danger'is actually internal and not external, which we explore in this application. In our model, DNA damage accumulates every day in individuals as a result of environmental exposures, e.g. smoking, UV or ionizing radiation, oxidative stress, chemical exposures, cell replication, inflammatory stress, or normal metabolic activities. Activation of cell cycle checkpoints and the DNA repair machinery repairs DNA damage. However, in rheumatoid arthritis, via intrinsic mechanisms regulated by the presence of HLA-DRB1*04 alleles or other intrinsic pathways, these repair mechanisms;ATR, DNA-PKcs, and ATM, and cell cycle checkpoints, JNK2, p53, p21, p27, CHEK2, RANGAP1 are defective resulting in failure of DNA repair and loss of genomic integrity. Loss of genomic integrity results in chronic NF- B activation and induction of pro-inflammatory cytokines. Since pro- inflammatory cytokines also activate NF- B, this produces a continuous cycle resulting in chronic inflammation. Methotrexate, in cell models and in lymphocytes from subjects with rheumatoid arthritis, corrects expression levels of JNK2, p53, p21, and p27, but not CHEK2 and RANGAP1. Therefore, to further test our hypothesis, we propose longitudinal studies of patients initiating methotrexate therapy to assess restoration of expression of these proteins, of cell cycle checkpoints, of genomic integrity, and restoration of chronic NF-kB activation to basal levels. An alternative model is that chronic inflammation drives excess cytokine production and NF-kB activation and excessive NF-kB activation opposes cell cycle checkpoints and DNA repair machinery producing the above phenotype actually observed in rheumatoid arthritis. To test this second hypothesis, we propose to perform longitudinal studies of patients initiating Enbrel therapy that will reduce excess TNF-a levels and NF- kB activation driven by excessive levels of TNF-a. We will determine if restoration of TNF-? levels to baseline also corrects defective cell cycle checkpoint and DNA repair machinery, thus restoring genomic integrity. The second major test of our hypothesis is that cell cycle checkpoint defects, loss of DNA repair machinery, loss of genomic integrity and excessive NF-kB activation existing in RA lymphocytes are genetically determined, in part, by the major genetic risk factor for RA, HLA-DRB1*04 alleles. To test this hypothesis, we will determine the extent to which this molecular and cellular phenotype is present in HLA-DRB1*04 positive and negative subjects with and without rheumatoid arthritis.
Contributions of defects in cell cycle checkpoints and DNA damage repair, the major genetic risk factor, HLA- DRB1*04, and the major environmental risk factor, smoking, to induction and pathogenesis of rheumatoid arthritis (RA) are not well understood. We propose a unifying hypothesis and a series of experiments to test the validity of our hypothesis. These studies have the potential to vastly improve our understanding of underlying pathogenesis of RA and to identify novel targets to be exploited to develop new and better therapies directed at the underlying molecular defects of RA.
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|Aune, Thomas M; Crooke 3rd, Phillip S; Spurlock 3rd, Charles F (2016) Long noncoding RNAs in T lymphocytes. J Leukoc Biol 99:31-44|
|Spurlock 3rd, Charles F; Gass 4th, Henry M; Bryant, Carson J et al. (2015) Methotrexate-mediated inhibition of nuclear factor ÎºB activation by distinct pathways in T cells and fibroblast-like synoviocytes. Rheumatology (Oxford) 54:178-87|
|Spurlock 3rd, Charles F; Tossberg, John T; Olsen, Nancy J et al. (2015) Cutting Edge: Chronic NF-ÎºB Activation in CD4+ T Cells in Rheumatoid Arthritis Is Genetically Determined by HLA Risk Alleles. J Immunol 195:791-5|
|Spurlock 3rd, Charles F; Tossberg, John T; Matlock, Brittany K et al. (2014) Methotrexate inhibits NF-ÎºB activity via long intergenic (noncoding) RNA-p21 induction. Arthritis Rheumatol 66:2947-57|