Non-coding RNAs (ncRNA) are regulators of cell functions and circulating ncRNAs from the majority of RNA classes, such as miRNA, tRNA, piRNAs, lncRNA, snoRNA, snRNA and miscRNAs, are potential non-invasive biomarkers. Understanding how non-disease traits influence ncRNA expression is essential for assessing their biomarker potential. We studied associations of common traits (sex, age, smoking, body mass, physical activity, and technical factors such as sample storage and processing) with serum ncRNAs. We used RNAseq data from 526 donors from the Janus Serum Bank and traits from health examination surveys. We identified associations between all RNA classes and traits. Ageing showed the strongest association with ncRNA expression, both in terms of statistical significance and number of RNAs, regardless of RNA class. Serum processing modifications and storage times significantly altered expression levels of a number of ncRNAs. Interestingly, smoking cessation generally restored RNA expression to non-smoking levels, although for some isomiRs, mRNA fragments and tRNAs smoking-related expression levels persisted. Our results show that common traits influence circulating ncRNA expression. Therefore it is clear that ncRNA biomarker analyses should be adjusted for age and sex. In addition, for specific ncRNAs identified in our study, analyses should also be adjusted for body mass, smoking, physical activity and serum processing and storage.

Circulating small non-coding RNAs associated with age, sex, smoking, body mass and physical activity