Abstract
3 min readAbstract Study question Does the cumulus oophorous complex (COC) extracellular matrix (ECM) hold potential as a marker of oocyte quality and developmental competence in human assisted reproduction? Summary answer PTX3, a component of the COC ECM, is a promising marker of follicle development, oocyte maturation, and embryo quality in intracytoplasmic sperm injection (ICSI). What is known already Assisted reproduction technologies (ART), such as in vitro fertilization (IVF) and ICSI, help treat infertility. However, these procedures carry risks, including ovarian hyper-stimulation syndrome (OHSS) and increased chances of premature birth and low birth weight. Research is exploring new markers for embryo selection and hormonal stimulation. COC ECM has been proposed to play a key role in ovarian function. ECM components, like the long pentraxin PTX3, have been associated with female fertility in preclinical and epidemiological studies and hold promise as biomarkers of oocyte maturation and embryo quality. Study design, size, duration Study design: prospective genetic association study. Inclusion criteria: i) age 25-40 y, ii) FSH ≤ 12.5 U/l, iii) AMH ≥ 1 ng/ml, and iv) BMI 17-29 kg/m2. Exclusion criteria: i) PCOS, ii) endometriosis III or IV, iii) sactosalpinx, or iv) poor responders. Aims to study the relationship between the concentration of PTX3 in the COC ECM, single nucleotide polymorphisms (SNPs) in the PTX3 gene, follicular development, oocyte maturity, and embryo quality. Size 78 patients. Duration 36 months. Participants/materials, setting, methods Participants: women undergoing ICSI at the Fertility Center of IRCCS Humanitas Research Hospital Materials and methods cumulus cells (CCs) and ECMs from denuded COCs were separated by centrifugation. ECMs were classified as MII-ECM, MI-ECM, or GV-ECM, based on oocyte grading, and pooled accordingly. The concentration of PTX3 in each pool was measured by ELISA. Venous blood was collected to extract DNA and genotype the rs6788044, rs3816527, rs2305619, and rs1840680 SNPs in the PTX3 gene. Main results and the role of chance 77 MII-, 36 MI-, and 23 GV-ECM pools were generated in the study, and genomic DNA was available from 72 patients. The presence of PTX3 in the isolated ECMs was initially confirmed by Western Blotting analyses using specific anti-human PTX3 polyclonal and monoclonal antibodies. The concentration of PTX3 in the MII-, MI- and GV-ECM was then measured using an in-house-made ELISA kit and found to change with the genotypes of the rs3816527, rs2305619 and rs1840680 SNPs (rs6788044 was monomorphic in the study population) (p < 0.05, one-way ANOVA). Also, the protein’s levels varied across the three grades of the oocytes (with increasing protein concentration from GV and MI to MII) as assessed by multiple regression modeling (p < 0.05). Interestingly, PTX3 concentration negatively correlated with embryo quality (Spearman r = -0.311, p < 0.01). Furthermore, a relationship emerged between SNP genotypes, antral follicle count, number of retrieved COCs, and MII oocytes (p < 0.05, F test). Overall, these findings provide evidence of associations between PTX3 gene variation, protein’s concentration in the COC ECM, and key outcomes of ICSI (follicle maturation and developmental potential of the COC). Limitations, reasons for caution Small sample size (78 patients) and mono-centric patient enrollment are perceived as major limitations of this study. Also, the concentration of PTX3 was measured in pooled ECM samples; in an ideal setting, this should have been dosed in individual COC ECMs. Wider implications of the findings Our findings might have applications in in vitro maturation techniques (by providing insights into COC ECM structure and function). Also, wider implications are envisaged in human pathophysiology with major regard to disorders of folliculogenesis and female fertility of unclear or unknown etiology. Trial registration number No
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