We report that a functional promoter SNP (rs3806933) of TSLP
that creates a binding site for the transcription factor AP-1 enhances AP-1 binding to the regulatory element (10
). The functional variant also increases the promoter–reporter activity of long-form TSLP
in response to poly(I:C) stimulation in NHBE (10
). In this study, we found that the TSLP
functional polymorphism was associated with both childhood atopic and adult asthma in a Japanese population. A synergistic relationship is evident between viral infection and allergen sensitization and exposure in provoking exacerbations, and the major trigger for exacerbations in both children and adults is viral infection (27
). Clinical studies suggest that patients with asthma are more susceptible to rhinovirus infections than normal individuals, and sustain a longer duration of lower respiratory tract symptoms when infected with rhinoviruses (30
). TSLP appears to be involved in the development of bronchial asthma through functional genetic polymorphisms that may contribute to Th2-polarized immunity through higher TSLP production by bronchial epithelial cells in response to viral respiratory infections.
A recent study showed that the expression of TSLP
is increased in asthmatic airways, and correlates with lung function (3
). The numbers of both epithelial and submucosal cells expressing TSLP
mRNA correlated inversely with FEV1
). We found a significant correlation between the rs2289278 genotype and lung function (FEV1
/FVC). The rs2289278 variant is located in intron 2 of long-form TSLP
and in the 5′ untranslated region of short-form TSLP
. However, the genetic influences of the polymorphism on the function of the TSLP
gene in asthma are unclear. Further investigation of the functional role of the variant needs to be conducted.
A recent study showed that an SNP in TSLP
is associated with cockroach-allergy IgE in Costa Rican girls and with total IgE in girls from two populations (9
). That study showed significant evidence of linkage to IgE produced in response to cockroach allergy on chromosome 5q23 in female subjects (9
is located near the linkage peak, and exerts female-specific effects on lung disease in mice (33
). Moreover, rs2289276 is reportedly associated with reductions in IgE in the response to cockroach and total IgE (9
). In our Japanese population, we could not find an association between total IgE and the rs2289276 SNP among patients with asthma in this study and a female-specific effect on the associations. Although inconsistent associations may reflect sample size, phenotype heterogeneity, or gene–environment interactions in the population, further replication studies of these findings are needed.
We could not replicate our findings in the populations that were examined in two recent GWASs (24
). Differences in minor allele frequencies (MAFs) of the variants were evident among the control populations. The MAFs of the three variants, rs3806932, rs2289276, and rs11466741, in the Japanese population were 0.29, 0.26, and 0.26, respectively (10
). To test for replication at the level of the gene rather than the SNP, further fine mapping around the TSLP
gene and association studies would be needed.
We could not replicate our findings in European-ancestry and African-ancestry populations for which recent GWASs were performed (24
). Differences in MAFs of the variants were evident among the control populations. The MAFs of the three variants, rs3806932, rs2289276, and rs11466741, in the Japanese population were 0.29, 0.26, and 0.26, respectively (10
), compared with 0.43, 0.29, and 0.29 in the Cjo;djppd Asthma Management Program (CAMP) population, 0.63, 0.18, and 0.31 in the African-American population, and 0.66, 0.17, and 0.28 in the African Caribbean population. Failure to observe SNP-for-SNP replication in ethnically diverse populations is not uncommon, and can result from variations in allele frequencies, population admixture, heterogeneity of the phenotype, and environmental factors. The fact that we could not replicate the association at the SNP level is therefore not surprising, and variants other than those tested in this study are likely to be causal. Elsewhere a gene-based rather than SNP-for-SNP approach was argued to provide evidence for genetic analysis at the functional level (25
). Our findings suggest that replication occurs at the level of the gene rather than the SNP, and further fine mapping around the TSLP
gene is warranted.
We previously showed that a functional rs3806933 SNP in the promoter region of long-form TSLP
(−847T) creates a binding site for AP-1 and enhances AP-1 binding to the regulatory element (10
). In this study, we found the binding of AP-2α protein, a possible transcription suppression factor, to the sequences containing the rs2289276 (−82C/T) SNP, and higher binding to −82C (on the protective allele). Both of the SNPs, rs3806933 (−847C/T) and rs2289276 (−82C/T), were significantly associated with susceptibility to asthma. We also identified a common disease-susceptible haplotype, T(−847)-T(−82)-C(1560), in both childhood atopic and adult asthma. The two different transcription factors on the two promoter SNPs may lead to preferential transcription from the susceptible haplotype T-T-C through their cooperative effects in bronchial epithelial cells. In this study, we did not examine the functional effects of polymorphisms rs3806932, rs2289277, rs10073816, and rs11466741, which were in strong LD with the related variants, rs3806933 and rs2289276. The functions of these linked polymorphisms remain to be elucidated.
Combination therapy with LABA and inhaled corticosteroids reduces the frequency of exacerbations in asthma, and it is also efficacious as an interventional therapy in asthma exacerbations (16
). In this study, SAL was able to suppress the production of TSLP in NHBE cells, and we also demonstrated the synergistic suppression of poly(I:C)-induced TSLP production by DEX and SAL in NHBE cells. Respiratory viral infections are associated with the majority of exacerbations in bronchial asthma, and a recent study showed that combined corticosteroid/β2
agonists synergistically suppress rhinovirus-induced neutrophil (CXCL8) and lymphocyte (CCL5 and CXCL10) chemokine production in airway epithelial cells (34
). Glucocorticosteroids inhibit both NF-κB and AP-1 through glucocorticoid-induced leucine zipper protein (35
), and an NF-κB binding site was identified 3.7 kb upstream from the start of TSLP
). Although further investigation of the molecular mechanisms mediating the suppressive effects of DEX and SAL on TSLP and chemokine production is needed, combination therapy may exert an anti-Th2 polarized inflammatory effect during respiratory viral infections through suppressive effects on TSLP and the production of other chemokines in airway epithelial cells.
In conclusion, we identified TSLP as a susceptibility gene for childhood atopic and adult asthma by means of a case-control study with SNPs, and demonstrated that glucocorticoid/LABA treatment synergistically suppressed poly(I:C)-induced TSLP in NHBE. Our data strongly support the important role of TSLP in bronchial asthma and the clinical benefits of combination therapy in asthma management.