Optimization of microRNA Acquirement from Seminal Plasma and Identification of Diminished Seminal microRNA-34b as Indicator of Low Semen Concentration

International Journal of Molecular Sciences

Marie-LouiseP. van der Hoorn¹  Tess Meuleman²  Michael Eikmans³  Jacqueline D. H. Anholts³  Els van Beelen³  Laura Blijleven³  FransH. J. Claas³ 

[1] Department of Gynecology and Obstetrics, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands;Department of Gynecology and Obstetrics, Radboud University Medical Center, 6525 GA Nijmegen, The Netherlands;Department of Immunohematology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands;

关键词: semen;    seminal plasma;    microRNA;    optimization;    oligozoospermia;    asthenozoospermia;   

DOI  :  10.3390/ijms21114089

来源: DOAJ

【 摘 要 】

About 10–15% of couples who want to conceive suffer from subfertility, while in 30% of these cases, a male factor plays a role. Levels of particular microRNAs in seminal plasma, including those involved in spermatogenesis, may serve as an indicative parameter for subfertility. We first optimized a protocol for acquiring microRNAs from seminal plasma. Next, using a test-validation strategy in a male cohort, we aimed to identify microRNAs of which the levels are related to semen motility and concentration. By qPCR, 742 microRNAs were profiled in three normozoospermic samples, three seminal samples with a low semen motility (asthenozoospermia), and three with a low semen concentration (oligozoospermia). MicroRNAs showing significant differences between groups were further validated in a second cohort consisting of 40 samples with normozoospermia (control group), 47 samples with asthenozoospermia, and 19 samples with oligozoospermia (of which 74% also low motility). Highest microRNA yields were obtained with the Biofluids RNA extraction kit, with inclusion of MS2 RNA carrier and proteinase K treatment to the protocol, and when 50 µL of seminal plasma was used as input. Exosome isolation prior to RNA extraction did not lead to enhanced yields. In the test cohort, 236 microRNAs could be detected, of which 54 microRNAs showed a difference between groups. Five microRNAs were analyzed in the validation cohort. MiR-34b-5p levels in the control group were significantly higher compared to the asthenozoospermia group (p < 0.05) and compared to the oligozoospermia group (p < 0.001). We optimized microRNA acquirement from seminal plasma and identified microRNA levels in relation to semen concentration and motility. As recent human and mouse studies show that the miR-34 family is a marker of low semen concentration and is crucial in spermatogenesis, seminal plasma miR-34b-5p may represent a suitable candidate to study further as a marker of male subfertility.

【 授权许可】

Unknown