Tobacco smoking is associated with sex- and plaque-type specific upregulation of CRLF1 in atherosclerotic lesions

Abstract

Tobacco smoking is a known risk factor for atherosclerotic disease, with more elevated risks in women compared to men. We hypothesized that atherosclerotic plaques from smokers show different gene expression patterns compared to non-smokers, in a sex-specific manner. Gene expression data of 625 carotid plaques (151 females and 474 males) were analyzed for differential gene expression between current smokers (n=226) and non-smokers (n=399). All analyses were stratified by sex and by molecular plaque characteristics. Finally, we projected the activity of gene regulatory networks and utilized single-cell transcriptomics from 38 plaques (26 males and 12 females) to interpret the sex- and plaque-type specific signals. We observed higher expression levels of CRLF1 gene in atherosclerotic plaques from smokers compared to non-smokers (log2FC=0.48, FDR=0.012). CRLF1 upregulation was interacting with sex (p=0.01) and was more pronounced in females (log2FC=0.93, p=1.53E-05) compared to males (log2FC=0.35, p=0.0018). Through single-cell RNA-seq analysis, we identified the highest CRLF1 expression within the transitioning and synthetic smooth muscle cell populations. CRLF1 expression was increased in fibro-inflammatory and fibro-cellular plaque types. Gene annotations pointed to increased expression of CRLF1 in networks with extracellular matrix related genes. Atherosclerotic plaques from current smokers show sex-dependent upregulation of smooth muscle cell gene CRLF1. This may explain the different contributions of smoking to cardiovascular risk in females.