PAR2 plays a prominent role in airway inflammation and asthma [5, 16, 7]. This is the first study exploring associations between F2RL1 gene SNPs and clinical and immunological characteristics of asthma. We genotyped three SNPs (rs1529505, rs2242991 and rs631465) in F2RL1 gene since they are not linked to each other, and previous studies have shown that these three SNPs are associated with monocytes [14] and asthma [9]. We had hypothesized that F2RL1 SNP genotypes that increase whole blood PAR2 expression would be associated with increased inflammatory markers in blood and increased asthma severity, but the data do not support this hypothesis. In fact, rs1529505 (CT and TT), was associated with a higher frequency of eosinophils and Th2 cells), but decreased PAR2 mRNA expression in peripheral blood. In addition, the genotype had no association with disease severity or lung function. These results suggest that the relationship between PAR2 expression and characteristics of asthma is quite complex. On the other hand, the CC genotype of rs631465 was associated with decreased PAR2 mRNA expression, but rs2242991 was not associated with any of the parameters we analyzed here.
Our observation that carries of the heterozygote and minor rs1529505 genotype (CT and TT) had less PAR2 mRNA in peripheral blood is concordant with data on whole blood and lung found in the Genotype-Tissue Expression (GTEx) portal. However, this association is not consistent across all tissues. For example, in other tissues the opposite has been observed; the major genotype rs1529505 CC is associated with reduced PAR-2 in the gastrocnemius muscle (GTEx) and in synovial tissue (21). Based on in silico prediction software (AliBaba2.1), the rs1529505 SNP “CC” genotype may affect binding sites for Sp1 and AP-2alpha transcription factors, emphasizing a potential role for this SNP in modulating PAR2 mRNA expression. Therefore, differential association of the CC genotype with PAR-2 mRNA expression in various tissues may be the result of differential expression of transcription factors across tissues or across the dominant PAR2-expressing cell in peripheral blood and/or certain tissue. Both Sp1 [17, 18] and AP-2alpha [19, 20] have been associated with asthma and regulation of asthma related factors.
While the rs1529505 SNP was associated with whole blood PAR2 mRNA expression in our cohort, there was no association between this SNP and surface PAR2 protein expression on peripheral blood monocytes. However, a large proportion of PAR2 expressed by monocytes is intracellular [21], which indicates that surface PAR2 expression on monocytes may have little to do with mRNA abundance. Alternatively, the significant association observed between the CC genotype of rs1529505 and increased PAR2 mRNA expression in whole blood may be the result of increased PAR2 mRNA expression in other blood PAR2-expressing cells, such as eosinophils, T lymphocytes or dendritic cells [22–25].
One reason for the seemingly opposite finding of the association of rs1529505 CC genotype with increased PAR2 expression, yet lower blood eosinophil and Th2 cell counts, could be enhanced infiltration to the lung. Indeed, PAR2 expression in the airways promotes eosinophil infiltration in murine models [26]. If the CC genotype of rs1529505 also increases PAR2 expression in the lungs, it may lead to increased inflammatory cell migration from peripheral blood into the airways resulting in decreased numbers of eosinophils and Th2 cells circulating in the blood. Future studies should explore associations between the rs1529505 SNP, airway PAR2 expression and airway eosinophil counts to test this hypothesis.
The rs1529505 SNP was not associated with asthma severity in this study, nor with any other clinical characteristic, or serum cytokine levels. The reason for this lack of association despite association with higher numbers of eosinophils and Th2 cells in peripheral blood, is not clear. Larger cohort studies and analysis of more immune and clinical markers associated with asthma severity may clarify this point.
The CC genotype of rs631465 SNP was associated with increased PAR2 mRNA expression compared to CT genotype in this study cohort. However, this finding is contradictory to a previously published observation in Korean children where the expression of PAR2 mRNA in PBMC of subjects with CT or TT genotype was higher than in those with the CC genotype of rs631465 [9]. This may be the result of the different ethnic backgrounds of our population.
The rs2242991 was not associated with any parameters studied. It is however interesting that even though we had a small number of individuals with CT genotype we identified a trend for association of the rs631465 SNPs with obesity, as has been shown before [10]. Ultimately, larger studies are needed to explore the association of the rs2242991 and rs631465 SNPs with asthma, which might also depend on the ethnicity of the individuals in the studied cohort.