Increased regulatory T-cell numbers are associated with farm milk exposure and lower atopic sensitization and asthma in childhood.

BACKGROUND: European cross-sectional studies have suggested that prenatal and postnatal farm exposure decreases the risk of allergic diseases in childhood. Underlying immunologic mechanisms are still not understood but might be modulated by immune-regulatory cells early in life, such as regulatory T (Treg) cells. OBJECTIVE: We sought to assess whether Treg cells from 4.5-year-old children from the Protection against Allergy: Study in Rural Environments birth cohort study are critical in the atopy and asthma-protective effect of farm exposure and which specific exposures might be relevant. METHODS: From 1133 children, 298 children were included in this study (149 farm and 149 reference children). Detailed questionnaires until 4 years of age assessed farming exposures over time. Treg cells were characterized as upper 20% CD4(+)CD25(+) forkhead box protein 3 (FOXP3)(+) (intracellular) in PBMCs before and after stimulation (with phorbol 12-myristate 13-acetate/ionomycin or LPS), and FOXP3 demethylation was assessed. Atopic sensitization was defined by specific IgE measurements; asthma was defined by a doctor's diagnosis. RESULTS: Treg cells were significantly increased in farm-exposed children after phorbol 12-myristate 13-acetate/ionomycin and LPS stimulation. Exposure to farm milk was defined as a relevant independent farm-related exposure supported by higher FOXP3 demethylation. Treg cell (upper 20% CD4(+)CD25(+), FOXP3(+) T cells) numbers were significantly negatively associated with doctor-diagnosed asthma (LPS stimulated: adjusted odds ratio, 0.26; 95% CI, 0.08-0.88) and perennial IgE (unstimulated: adjusted odds ratio, 0.21; 95% CI, 0.08-0.59). Protection against asthma by farm milk exposure was partially mediated by Treg cells. CONCLUSIONS: Farm milk exposure was associated with increased Treg cell numbers on stimulation in 4.5-year-old children and might induce a regulatory phenotype early in life, potentially contributing to a protective effect for the development of childhood allergic diseases.

TBX21 and HLX1 polymorphisms influence cytokine secretion at birth.

BACKGROUND: TBX21 (T cell specific T-box transcription factor) and HLX1 (H.20-like homeobox 1) are crucial transcription factors of T(H)1-cells, inducing their differentiation and suppressing T(H)2 commitment, particularly important for early life immune development. This study investigated the influence of TBX21 and HLX1 single nucleotide polymorphisms (SNPs), which have previously been shown to be associated with asthma, on T(H)1/T(H)2 lineage cytokines at birth. METHODS AND FINDINGS: Cord blood mononuclear cells (CBMCs) of 200 neonates were genotyped for two TBX21 and three HLX1 SNPs. CBMCs were stimulated with innate (Lipid A, LpA; Peptidoglycan, Ppg), adaptive stimuli (house dust mite Dermatophagoides pteronyssinus 1, Derp1) or mitogen (phytohemagglutinin, PHA). Cytokines, T-cells and mRNA expression of T(H)1/T(H)2-related genes were assessed. Atopic diseases during the first 3 years of life were assessed by questionnaire answered by the parents. Carriers of TBX21 promoter SNP rs17250932 and HLX1 promoter SNP rs2738751 showed reduced or trendwise reduced (p≤0.07) IL-5, IL-13 and TNF-α secretion after LpA-stimulation. Carriers of HLX1 SNP rs2738751 had lower IL-13 levels following Ppg-stimulation (p = 0.08). Carriers of HLX1 exon 1 SNP rs12141189 showed increased IL-5 (LpA, p = 0.007; Ppg, p = 0.10), trendwise increased IL-13 (LpA), higher GM-CSF (LpA/Ppg, p≤0.05) and trendwise decreased IFN-γ secretion (Derp1+LpA-stimulation, p = 0.1). Homozygous carriers of HLX1 promoter SNP rs3806325 showed increased IL-13 and IL-6 (unstimulated, p≤0.03). In carriers of TBX21 intron 3 SNP rs11079788 no differences in cytokine secretion were observed. mRNA expression of T(H)1/T(H)2-related genes partly correlated with cytokines at protein level. TBX21 SNP rs11079788 carriers developed less symptoms of atopic dermatitis at 3 years of age (p = 0.03). CONCLUSIONS: Polymorphisms in TBX21 and HLX1 influenced primarily IL-5 and IL-13 secretion after LpA-stimulation in cord blood suggesting that genetic variations in the transcription factors essential for the T(H)1-pathway may contribute to modified T(H)2-immune responses already early in life. Further follow-up of the cohort is required to study the polymorphisms' relevance for immune-mediated diseases such as childhood asthma.