Sophora Flavescens Alkaloid-Rich Fraction Induction of IL-10 Production and Prevention of Dexamethasone Suppression of Asthma Patient PBMC IL-10 Production Is Associated with Altered DNA Methylation at foxp3 Gene Promoter

Abstract

S A T U R D A Y 234 IFNg and Foxp3 Methylation, Expression in Buccal Mucosa in Inner-City Children with Allergic Asthma Emily Happy Miller, Hanjie Zhang, Mariangels De Planell Saguer, Stephanie Lovinsky-Desir, Matthew S. Perzanowski, PhD, Wanda Phipatanakul, MD, MS, Elizabeth C. Matsui, MD, MHS, Rachel Miller, MD; Department of Medicine, Columbia University, New York, NY, Division of Pulmonary, Allergy and Critical Care Medicine, Department of Medicine, Columbia University, New York, NY, Division of Pulmonary, Department of Pediatrics, Columbia University, New York, NY, Department of Environmental Health Sciences, Columbia University, New York, NY, Division of Pediatric Allergy/Immunology, Boston Children’s Hospital, Harvard University School of Medicine, Boston, MA, Division of Pediatric Allergy/Immunology, Johns Hopkins School of Medicine, Baltimore, MD, Division of Allergy, Immunology, and Rheumatology and Division of Pulmonary, Department of Pediatrics, Columbia University, New York, NY. RATIONALE: DNA methylation of IFN and Foxp3 has been associated with environmental exposures and asthma-related outcomes. However, it is not known whether epigenetic regulation and expression of these genes in the buccal mucosa can be used as a biomarker for allergic disease in children. METHODS: Buccal mucosa cells from mouse sensitized asthmatic children (ages 5-17) who were recruited to participate in the Mouse Allergen and Asthma Intervention Trial (MAAIT) were collected at baseline and 6 months after intervention or control. DNA and total RNA were extracted. Pyrosequencing of CpG sites of IFNg (promoter) and Foxp3 (promoter, upstream enhancer) and real time PCR was conducted. RESULTS: At baseline, in this mixed cell population, negative correlations between expression and DNA methylation at each gene were not found. Instead, IFN and Foxp3 buccal gene expression positively correlated (N 5 113, r 5 0.59, p<0.0001). Unexpectedly, IFN promoter methylation (CpG-54) positively correlated with Foxp3 gene expression (N5113, r 5 0.20, p5 0.03). Methylation at CpG sites in the Foxp3 promoter (CpG-207) negatively correlated with methylation at Foxp3 enhancer site (eg. CpG 4575; N 5 125, r 5 -0.64, p <0.0001). The same trends repeated 6 months later. CONCLUSIONS: Several patterns of regulatory gene methylation and expression in buccal cells were exhibited in this cohort of mousesensitized, allergic asthmatic children. Future work examining internal validity and reliability of these markers, associations with allergen levels, and associations with allergy and asthma-related outcomes following intervention may yield information on potentially novel biomarkers for pediatric allergic asthma.