Original article Cell-free nucleic acids and melatonin levels in human follicular fluid predict embryo quality in patients undergoing in-vitro fertilization treatment

摘要

Despite many advances in assisted reproductive technology (ART), the most viable embryo selection remains a challenge for infertility treatment. This study was designed to investigate whether intra-follicular circulating cell-free DNA (cfDNA) fragments and Melatonin levels predict embryo quality in patients undergoing IVF treatment. A total of eight hundred and ninety-five follicular fluid (ff) samples were collected from 325 infertile patients undergoing IVF treatment. Patients were enrolled from August 2017 to December 2018 in the infertility center of a tertiary care hospital. A clear non-hematic follicular fluid was aspirated after the removal of eggs from the dominant follicles (18?mm) of each patient. Melatonin and E 2 levels in each follicular sample were estimated by immune-chemiluminescence using commercially available kits. ALU-qPCR evaluated cfDNA levels in individual follicular fluid samples. Our study presented a significant and negative relationship between intra-follicular cfDNA and melatonin concentration (-0.541: P??0.001). Each individual follicle contains measurable copy number of cfDNA [mean: 1.85?±?2.98?ng/μl (median; 1.86?ng/μl (95% Cl: 0.96–2.87)]. In pregnant women cfDNA copy number was significantly decreased in follicular fluid samples(ff) aspirated from matured oocytes than in immature ones [p??0.01; β = -0.42?±?0.49; median; 1.45?ng/ml (95% Cl: 0.36–2.97) vs. 3.57?ng/μl (95% Cl: 0.37–4.01) respectively. While melatonin concentration in ff samples corresponding to mature oocytes was significantly higher than in ff samples related to immature oocytes (p??0.001). Moreover, in pregnant women cfDNA level was significantly lower in ff samples related to oocytes which produces top-quality embryos versus low quality embryos [p??0.001; β?=?1.81?±?0.91; median; 1.25?ng/μl (95% Cl: 0.35–1.97)] vs. [(median; 3.65?ng/ml (95% Cl: 1.23–6.36)] respectively. Likewise, in non-pregnant women melatonin levels were significantly decreased in ff samples related to embryos with high fragmentation rate (≥25%) than embryos with low fragmentation rate (25%; p??0.001). Conclusively, this study indicates that Intra-follicular cfDNA and melatonin concentration possibly a new supplemental tool that supports to establish an advanced non-invasive early prognostic test for the patients undergoing IVF/ICSI procedure.