Chronic inflammation in rheumatoid arthritis (RA) is mediated by repeatedly activated proinflammatory Th1 cells. In contrast, Th2 cells that might downmodulate the chronic autoimmune response are rarely found in RA. It has been previously documented that RA T cells are severely impaired in their ability to differentiate into Th2 effectors while exerting enhanced Th1 differentiation. The mechanisms underlying this functional abnormality, however, have not been delineated. As IL-4 is a most critical determinant in regulating immune responses by promoting Th2 cell development and inhibiting Th1 cell differentiation, we analyzed the role of single nucleotide polymorphisms (SNPs) in the IL-4 receptor (IL-4R) α-chain, which is critical for binding of IL-4 and for IL-4 signal transduction, in the differentiation of human T cells. Three hundred and sixty-one healthy individuals were genotyped by allele-specific PCR for the two IL-4R α-chain SNPs that are located in functionally important regions of the IL-4R α-chain – the I50V SNP50 and the Q551R SNP551 in the IL-4-binding and STAT6-binding domains, respectively. Naive and memory CD4-positive T cells were isolated from the peripheral blood of individuals who were homozygous for either allele at SNP50 and SNP551, and were primed for 5 days with mAbs to CD28 and/or CD3 in the presence or absence of exogenous IL-4. The phenotype of the resulting differentiated effector cells was then analyzed by flow cytometric analysis of cytoplasmic cytokines. The SNP551 alleles did not affect T-cell differentiation. In contrast, the inhibitory effect of IL-4 on Th1 cell differentiation was significantly diminished in CD4 T cells that were homozygous for the mutated allele at SNP50 (50V) as compared with those with the wild type allele (I50). Likewise, the augmenting effect of IL-4 on Th2 cell differentiation was enhanced on T cells that were homozygous for the wild-type allele as compared with T cells expressing the mutant allele. These data indicate that the mutant allele of the IL-4R α-chain at SNP50 is associated with a decreased T-cell response to IL-4. To delineate a potential mechanism of different responses to IL-4 in the cells expressing different alleles of the IL-4R, T cells form individuals who were homozygous for either the wild-type or the mutant allele at SNP50 were primed with different concentrations of IL-4 and analyzed by flow cytometry for STAT6 and phosphorylated STAT6. Whereas STAT6 concentrations were not different between T-cell expressing I50 or V50, STAT6 phosphorylation in response to IL-4 stimulation was significantly reduced in T cells expressing the V50 allele compared with T cells expressing I50. Thus, the V50 SNP50 allele of the IL-4R α-chain might regulate T-cell differentiation by diminishing T-cell responses to IL-4, resulting in reduced STAT-6 phosphorylation and subsequently in diminished Th2 cell differentiation. The V50 SNP50 allele might thereby contribute to the development of unbalanced Th subset activation, as characteristic for autoimmune diseases, such as RA.
Supported by the Deutsche Forschungsgemeinschaft, the Interdisciplinary Center for Clinical Research at the University of Erlangen–Nuremberg, and the Dr Robert Pfleger Foundation.