Background/Purpose: Juvenile idiopathic arthritis (JIA) is the most common autoimmune arthritis in children. Polyarticular JIA and extended oligoarticular JIA both have genetic associations near genes encoding proteins important for production of T helper (Th) cells. Effector T cells, including Th cells, are generated in a differentiation process controlled by proteins, non-coding RNAs, and epigenetic modifications that drive one lineage while suppressing other lineages. These pathways make unique cell types designed to fight specific pathogens. Naïve T helper (Th0) cells proliferate in response to antigen encounter but are not differentiated. In the presence of polarizing cytokines Th0 cells differentiate into Th1, Th2, and Th17 cells that secrete characteristic cytokines upon stimulation. Th1 cells secrete the pro-inflammatory cytokine IFNγ. This study uniquely assesses Th cell differentiation and cytokine secretion in young children with polyarticular JIA and pediatric healthy controls.
Methods: Peripheral blood mononuclear cells (PBMCs) were collected from 20 JIA patients, 20 age-matched control children, and healthy adults. Children were 2 to 8 years old (average 62 months) at time of sample collection. JIA patients are polyarticular (n=18) or extended oligoarticular (n=2), female (n=16), and ANA positive (n=19). Some samples were collected at diagnosis (n=5). PBMCs were analyzed for naïve and memory T cells by measuring cell surface expression of CD3, CD4, CD8, CD197, and CD45RA with flow cytometry. Th cell differentiation and activity were measured in an ex vivo assay wherein PBMCs are treated with or without cytokines to make Th0, Th1, Th2, and Th17 cell cultures. Cultures are then stimulated, followed by assessment of secreted effector cytokines, cell RNA, and cell protein. Cell proliferation in the ex vivo cultures was measured using a fluorescence-based assay.
Results: PBMC cell surface staining found no difference in the number of CD3+, CD4+, or CD8+ cells between groups. Both JIA and healthy pediatric control PBMCs have significantly more CD4+ and CD8+ naïve (CD197+,CD45RA+), less CD4+ central memory (CD197+,CD45RA-), and less CD8+ effector memory (CD197-,CD45RA-) cells. There is no difference in the proliferation index for CD3+ cells for adult, JIA and child Th0 cell cultures or JIA and child Th1 cell cultures. Th1 cell cultures generated from JIA patients secrete higher levels of the characteristic cytokine IFNγ and exhibit increased levels of IFNγ transcripts than cultures from healthy pediatric controls.
Conclusion: Analysis of pediatric PBMCs revealed an increase in naïve T cells and decrease in memory cells as compared to adults, highlighting important differences in adult versus child biologic samples. JIA Th1 cell cultures exhibit an increase in IFNγ expression at both the transcript and protein levels as compared to pediatric controls. Our evidence does not support a mechanism involving a difference in PBMCs or proliferation of CD3+ cells. The present study suggests that JIA Th1 cell cultures have altered transcriptional regulation of the IFNG locus, resulting in an increase in secretion of the pro-inflammatory cytokine IFNγ.
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