Thursday, May 2, 2024
5/2/2024
PROJECT SUMMARY/ABSTRACT Formation of disease-associated autoantibodies (Abs), including anti-cyclic citrullinated peptide (CCP) and rheumatoid factor (RF), is an early step in the pathogenesis of rheumatoid arthritis (RA). In fact, RA-Abs can develop several years before the onset of joint inflammation during a preclinical phase of RA development. Our preliminary data found that the presence of RA-Abs in sputum during preclinical RA is strongly associated with development of classified RA in 3 years. Yet, there is a major gap in our understanding of the cellular mecha- nisms leading to RA-Ab generation in the lung in preclinical RA. The overall hypothesis of the proposed project is that CD4+PDhiCXCR5- T peripheral helper (Tph) cells and CD14+CD16+ intermediate monocytes in the lung contribute to the sputum RA-Ab generation that predicts transition from preclinical to classifiable RA. This hy- pothesis is based on our published and preliminary data including our finding that Tph cells and CD14+CD16+ monocytes are increased in the peripheral blood of individuals At-Risk for RA (i.e., in the preclinical phase of RA based on serum anti-CCP positivity) compared to controls. Tph cells are known to promote B cell differenti- ation and antibody production in inflamed non-lymphoid tissues, such as the lung. Moreover, we found in pre- liminary data that sputum RA-Ab levels strongly correlated with sputum IL-6 levels, which can promote B cell differentiation and antibody production and can be produced at high levels by CD14+CD16+ monocytes. The proposed ancillary studies will leverage induced sputum samples already being collected as part of an ongoing parent R01 study. This 1-year study will include individuals At-Risk for RA, individuals with classified RA and healthy controls. The study will quantify the proportion of T and B cell subsets, including Tph cells, in sputum using flow cytometry. In At-Risk subjects, the % Tph cells will be correlated with levels of sputum RA-Abs and Tph-associated cytokines (IL-21, CXCL13). T cell subsets will also be sorted by FACS and undergo bulk RNA sequencing. It is expected that sputum Tph cells will be increased in individuals At-Risk for RA, correlate with sputum RA-Abs and have a distinct transcriptional profile reflecting a role in B cell differentiation and antibody production. The proposed project will also quantify the proportion of monocyte and macrophage subsets in sputum using flow cytometry. In At-Risk subjects, the % CD14+CD16+ monocytes in sputum will be correlated with levels of sputum RA-Abs and IL-6. Bulk RNA sequencing will be performed on monocyte and macro- phage subsets isolated by FACS. It is expected that sputum CD14+CD16+ monocytes will be increased in indi- viduals At-Risk for RA, correlate with sputum RA-Abs and have a distinct transcriptional profile associated with IL-6 production. Altogether, this proposal will further the strategic vision of the NIH by investigating novel cellu- lar mechanisms in the lung that contribute to the RA-Ab generation associated with transition from preclinical to classified RA and could serve as novel cellular targets for disease prevention in RA.
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