Th17 transcription factor RORC2 is inversely correlated with FOXP3 expression in the joints of children with juvenile idiopathic arthritis.
OBJECTIVE
To investigate the relationship between interleukin 17 (IL-17) producing T cells (Th17) and CD4+CD25+FOXP3+ regulatory T cells (Tregs) in blood and synovial fluid (SF) of patients with juvenile idiopathic arthritis (JIA).


METHODS
Sixty-five children with JIA (18 males and 47 females, median age 6.2 yrs; 45 with oligoarticular and 20 with polyarticular course) and 75 age- and sex-matched healthy controls were studied. Flow cytometry was used to analyze the forkhead box P3 (FOXP3)-positive Treg cells in peripheral blood (PB) and synovial fluid mononuclear cells (SFMC). FOXP3 and retinoic-acid related orphan receptor C isoform 2 (RORC2) messenger RNA (mRNA) were assessed by real-time polymerase chain reaction analysis. Cytokines (IL-17 and Th1/Th2 related cytokines) were measured in culture supernatants of 11 paired PBMC and SFMC activated with PMA and ionomycin.


RESULTS
FOXP3+ T cells and FOXP3 mRNA amounts were significantly lower in PB of children with JIA as compared with controls (p = 0.0002 and p = 0.001, respectively) and a higher percentage of Treg cells with concomitant higher level of FOXP3 transcript levels were observed in SF when compared with their PB counterparts (both p < 0.0001). SF CD4+FOXP3+ T cells were characterized by higher amounts of FOXP3 protein per cell when compared with peripheral CD4+FOXP3+ T cells, as revealed by the difference in FOXP3 median fluorescence intensity (median +/- SD, arbitrary units, 54 +/- 22.6 vs 19.5 +/- 4.2; p < 0.001). RORC2 transcript levels were higher in JIA joints when compared with matched PB samples (median fold increase 3.9, p < 0.0001) but negatively correlated with FOXP3 mRNA levels (r = -0.623, p = 0.04). Stimulated SFMC displayed an impaired ability to produce IL-17 when compared with PBMC and, interestingly, an inverse relationship between IL-17 levels and the percentage of CD4+CD25+FOXP3+ SF T cells (r = -0.510, p = 0.047) was seen.


CONCLUSIONS
We demonstrated for the first time an increased synovial expression of the transcription factor of Th17, RORC2, in JIA, and its inverse relationship with FOXP3 mRNA. These results extend research on "Th17" and Tregs in JIA.
Olivito
Biagio
B
Department of Paediatrics, Rheumatology and Immunology Unit, University of Florence, Anna Meyer Children's Hospital, Florence, Italy. b.olivito@meyer.it
Simonini
Gabriele
G
Ciullini
Sara
S
Moriondo
Maria
M
Betti
Letizia
L
Gambineri
Eleonora
E
Cantarini
Luca
L
De Martino
Maurizio
M
Azzari
Chiara
C
Cimaz
Rolando
R
D000293:Adolescent; D001171:Arthritis, Juvenile / Q000378:metabolism* / Q000473:pathology; D016022:Case-Control Studies; D002648:Child; D002675:Child, Preschool; D005260:Female; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D006801:Humans; D020381:Interleukin-17 / Q000378:metabolism; D007596:Joints / Q000378:metabolism* / Q000473:pathology; D007963:Leukocytes, Mononuclear / Q000378:metabolism / Q000473:pathology; D008297:Male; D057132:Nuclear Receptor Subfamily 1, Group F, Member 3; D012333:RNA, Messenger / Q000378:metabolism; D018168:Receptors, Retinoic Acid / Q000378:metabolism*; D011988:Receptors, Thyroid Hormone / Q000378:metabolism*; D013582:Synovial Fluid / Q000378:metabolism; D013583:Synovial Membrane / Q000378:metabolism* / Q000473:pathology; D013601:T-Lymphocytes / Q000378:metabolism / Q000473:pathology; D050378:T-Lymphocytes, Regulatory / Q000378:metabolism / Q000473:pathology
C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D020381:Interleukin-17; D057132:Nuclear Receptor Subfamily 1, Group F, Member 3; D012333:RNA, Messenger; C539900:RORC protein, human; D018168:Receptors, Retinoic Acid; D011988:Receptors, Thyroid Hormone