Methylome signature of bronchoalveolar cells from Multiple Sclerosis patients in relation to smoking

Background Despite compelling evidence that cigarette smoking impacts the risk of developing Multiple Sclerosis (MS), little is known about smoking-associated changes in the primary exposed cells in the lung of patients. We aimed to examine molecular changes occurring in pulmonary immune cells from MS patients in relation to smoking and in comparison to healthy controls (HC). Methods We profiled genome-wide DNA methylation (5mC) and hydroxymethylation (5hmC) in bronchoalveolar lavage (BAL) cells collected from female MS patients (9 smokers, 8 non-smokers) and healthy volunteers (10 smokers, 12 non-smokers), using Illumina Infinium EPIC arrays on conventional (BS) and oxidative (oxBS) bisulfite DNA. We profiled gene expression using RNA-sequencing in non-smoker MS patients and controls. Findings The most prominent changes were found in relation to smoking, with 1376 BS-differentially methylated positions (DMPs), 131 5mC-DMPs and 4 5hmC-DMP (adjusted P < 0.05) differing between MS smokers and non-smokers. Approximately 30% of the affected genes overlapped with smoking-associated changes in HC, leading to a strong common smoking signature in both MS and HC after gene ontology analysis. Smoking in MS patients resulted in additional discrete changes related to neuronal processes. To further explore MS-specific signature, we performed RNA sequencing in BAL cells from non-smoker MS patients and controls. Overall, methylome and transcriptome analyses in non-smokers suggest that BAL cells from MS patients display very subtle but concordant changes associated to genes connected to reduced transcriptional/translational processes and enhanced cellular adhesion and migration, further corroborating findings from animal studies. Interpretation Our study provides new insights into the impact of smoking on lung inflammation in the immunopathogenesis of MS. - 39 bronchoalveolar lavage samples (mainly alveolar macrophages) from healthy (H) individuals and multiple sclerosis (MS) patients; smokers (S) and nonsmokers (NS). Includes both BS-treated and oxBS-treated samples for 5mC and 5hmC interrogation.