Detection of five specified menstrual fluid mRNA markers over the uterine cycle

Matrix metallopeptidases (MMPs) 7, 10, and 11 are currently the most commonly employed messenger RNAs (mRNAs) for the identification of menstrual fluid (MF) in forensic analysis. However, no comprehensive study has been carried out to date to explore their time-dependent detection in vaginal samples. This research investigated the detection of MMPs 7, 10, and 11, as well as MMP3 and stanniocalcin 1 (STC1) over the uterine cycle. The aim was to associate relative transcript levels with cycle stages and thus determine which of these transcripts is most suitable for MF identification in a forensic context. Additionally, the effect of hormonal contraceptives (HCs) on their abundance was explored. A total of 300 vaginal swab samples were collected from eight female donors, including a pregnant woman, naturally cycling women, and contraceptive users. Differences among individuals were observed, but these were not consistent within the groups. Only MMP10 and STC1 mRNA abundance appeared to be unaffected by the use of HCs. MMP3, MMP7, and MMP11 transcripts were less abundant in MF samples of some HC users. Overall, MMP3 was most specific to MF, although this transcript was still detected in one of four vaginal material (VM) samples. STC1 was less specific than MMP3 (detected in 39.6 % of VM samples). However, over the days of menstruation, STC1 was more consistently detectable than the MMPs. MMP10 was least specific, with a 78.3 % detection rate in VM samples, but the presence/absence in VM was individual-specific and consistent. MMP10 may therefore be more useful as a VM marker with elevated abundance during menstruation in some individuals. MMP7 and MMP11 were the least reliable mRNAs for MF identification, despite an increased specificity compared to MMP10. Detection rates in MF were lower than those of MMP3 and STC1, whereas detection rates in VM were higher. MMP7 abundance additionally increased approximately 2-5 days after the end of menstruation in all donors except one naturally cycling individual. In view of these results, MMP3 and STC1 were identified as the most useful MF markers for forensic use. Nevertheless, mRNA typing results need to be interpreted with utmost caution.