Phthalate exposure to adult mice alters non-coding RNAs in cauda epididymosomes

We have previously reported associations between urinary biomarkers of anti-androgenic phthalates and small non-coding RNA (sncRNA) compositions of extracellular vesicles (EVs) in seminal plasma of men undergoing fertility treatment. To complement these findings, we assessed sncRNAs of cauda epididymosomes in mice after exposure to the anti-androgenic phthalates, Di-n-butyl phthalate (DBP), Di(2-ethylhexyl) phthalate (DEHP), and their mixture.Male mice were exposed to 2.5 mg/kg/day of DBP or DEHP and their mixture (2.5 mg/kg/day each of DBP and DEHP) for six weeks. Cauda epididymosomal RNA were isolated and subjected to sequencing to assess their non-coding RNA (ncRNA) content. Mapping of reads to transcriptomes and differential expression of normalized read counts were conducted using counts between the exposed groups relative to controls using STAR alignment tool and EdgeR (Fold change, FC ≥ 1.5 and p < 0.01), respectively.The mapped reads of epididymosomal ncRNAs were mostly piRNAs (47%), followed by tRFs (34%) and miRNAs (6%). Phthalate exposures altered a total of 79 (DBP = 47; DEHP = 11; mixture = 21) epididymosomal sncRNA biotypes (miRNA = 14, tRFs = 5 & piRNA = 60). These ncRNAs were predicted to target genes involved in various functional processes, including tissue development, olfactory perception, and potassium channel activity.This is the first study, to our knowledge, to show that exposure to phthalates in adult male mice disrupts the ncRNA profiles of cauda epididymosomes. These data suggest that cauda epididymosomes ncRNAs are responsive to environmental exposures and may have implication in the health and development of the next generation.