Follicular fluid from infertile women with mild endometriosis may compromise the meiotic spindles of bovine metaphase II oocytes

Abstract

What is the potential impact of follicular fluid (FF) from infertile women with mild endometriosis (ME) on oocyte quality, especially on nuclear maturation and the meiotic spindle? FF from infertile women with ME may compromise nuclear maturation and the meiotic spindles of in vitro matured bovine oocytes. Controversial studies have suggested that impaired oocyte quality may be involved in the pathogenesis of endometriosis-related infertility. Moreover, some studies have demonstrated alterations in the composition of FF from infertile women with endometriosis. However, to date no study has evaluated the effect of FF from infertile women with ME on the genesis of meiotic oocyte anomalies. We performed an experimental study. Samples of FF were obtained from February 2009 to February 2011 from 22 infertile women, 11 with ME and 11 with tubal or male factors of infertility (control group), who underwent ovarian stimulation for ICSI at our university IVF Unit. From March 2011 to February 2012 we performed in vitro maturation (IVM) experiments using immature bovine oocytes as described below. FF free of blood and containing a mature oocyte was obtained from 22 infertile women during oocyte retrieval for ICSI. Immature bovine oocytes underwent IVM in the absence of FF (No-FF) and in the presence of four concentrations (1, 5, 10 and15%) of FF from infertile women without endometriosis (C-FF) and with ME (ME-FF). Eleven replicates were performed, each one using FF from a control patient and a patient with ME. Each FF sample was used in only one experiment. After 22-24 h of IVM, oocytes were denuded, fixed and immunostained for morphological visualization of microtubules and chromatin by confocal microscopy. A total of 1324 cumulus-oocyte complexes were matured in vitro. Of these, 1128 were fixed and 1048 were analyzed by confocal microscopy. The percentage of meiotically normal oocytes was significantly higher for oocytes that underwent IVM in the absence of FF (No-FF; 76.5%) and in the presence of 1% (80.9%), 5% (76.6%), 10% (75%) and 15% (76.2%) C-FF than in oocytes that underwent IVM in the presence of 1% (44.4%), 5% (36.7%), 10% (45.5%) and 15% (51.2%) ME-FF (P < 0.01). No differences were observed among FF concentrations within each group. When the four concentrations from each group were pooled, the number of oocytes in metaphase I stage was significantly higher in the ME-FF (50 oocytes) than in the C-FF (29 oocytes) group and the percentage of meiotic abnormalities was significantly higher when oocytes were matured with ME-FF (55.8%) than with C-FF (23.1%), P < 0.01. Owing to the strict selection criteria for FF donors, this study had a small sample size (11 cases and 11 controls), and thus further investigations using a large cohort of patients are needed to confirm these results. In addition, data obtained from studies using animal models may not necessarily be extrapolated to humans and studies evaluating in vivo matured oocytes from infertile women with ME are important to confirm our results. Our results open new insights into the pathogenic mechanisms of infertility related to mild endometriosis, suggesting that FF from infertile women with mild endometriosis may be involved in the worsening of oocyte quality of these women. This study was supported by the National Council for Scientific and Technological Development (CNPq), Brazil. The authors declare no conflicts of interest.