The effects of hyaluronate-containing medium on human embryo attachment to endometrial epithelial cells in vitro

Abstract

STUDY QUESTION Does embryo transfer medium containing hyaluronate (HA) promote the attachment phase of human embryo implantation? SUMMARY ANSWER HA-containing medium does not promote human blastocyst attachment to endometrial epithelial cells in vitro. WHAT IS KNOWN ALREADY Embryo transfer media containing high concentrations of HA are being used to increase implantation and live birth rates in IVF treatment, although the mechanism of action is unknown. STUDY DESIGN, SIZE, DURATION Expression of HA-interacting genes in frozen-thawed oocytes/embryos was assessed by microarray analysis (n = 21). Fresh and frozen human blastocysts (n = 98) were co-cultured with human endometrial epithelial Ishikawa cell layers. Blastocyst attachment and the effects of a widely used HA-containing medium were measured. PARTICIPANTS/MATERIALS, SETTING, METHODS Human embryos surplus to treatment requirements were donated with informed consent from several ART centres. Blastocyst-stage embryos were transferred at day 6 to confluent Ishikawa cell layers; some blastocysts were artificially hatched. Blastocyst attachment was monitored from 1 to 48 h, and the effects of blastocyst pre-treatment for 10 min with HA-containing medium were determined. MAIN RESULTS AND THE ROLE OF CHANCE Human embryos expressed the HA receptor genes CD44 and HMMR, hyaluronan synthase genes HAS1–3, and hyaluronidase genes HYAL1–3, at all stages of preimplantation development. Attachment of partially hatched blastocysts to Ishikawa cells at 24 and 48 h was related to trophectoderm grade (P = 0.0004 and 0.007, respectively, n = 34). Blastocysts of varying clinical grades that had been artificially hatched were all attached within 48 h (n = 21). Treatment of artificially hatched blastocysts with HA-containing medium did not significantly affect attachment at early (1–6 h) or late (24 and 48 h) time points, compared with control blastocysts (n = 43). LIMITATIONS, REASONS FOR CAUTION Using an adenocarcinoma-derived cell line to model embryo-endometrium attachment may not fully recapitulate in vivo interactions. The high levels of blastocyst attachment seen with this in vitro model may limit the sensitivity with which the effects of HA can be observed. WIDER IMPLICATIONS OF THE FINDINGS Morphological trophectoderm grade can be correlated with blastocyst attachment in vitro. HA-containing medium may increase pregnancy rates by mechanisms other than promoting blastocyst attachment to endometrium. STUDY FUNDING/COMPETING INTEREST(S) This work was funded by a grant from the Wellbeing of Women, the NIHR Local Comprehensive Research Network and NIHR Manchester Clinical Research Facility, the Department of Health Scientist Practitioner Training Scheme, and the Ministry of Higher Education, The State of Libya. None of the authors has any conflict of interest to declare.

Involvement of uterine natural killer cells in the natural selection of human embryos at implantation

“…57 HYAL2 transcripts are constitutively expressed in human preimplantation embryos. 26 Here, we showed by immunocytochemistry that this glycosylphosphatidylinositol-anchored enzyme localises to both the trophectoderm and inner cell mass of human blastocysts. Importantly, HYAL2 was also detectable in BCM, with levels correlating to both embryo morphology and implantation competence.…”

Clinical efficacy of hyaluronate-containing embryo transfer medium in IVF/ICSI treatment cycles: a cohort study

“…This study was undertaken as the second of a two-part clinical service evaluation of the use of HA-rich medium (EmbryoGlue ® ) for ET ( Ruane et al , 2020 ).…”

“…For EmbryoGlue ® , the 4-fold increase in the concentration of HA relative to 0.125 mg/ml in G 2+ appears to be the critical difference between the two culture media, although the concentration of recombinant HSA is 4-fold lower in EmbryoGlue ® medium compared with 10 mg/ml in G 2+ . Furthermore, we do not know the molecular weight profile of the HA in commercial EmbryoGlue ® , and this in turn has significant implications for our understanding ( Fouladi-Nashta et al , 2017 ; Ruane et al , 2020 ). The impact of other unknown components of these commercial embryo culture media may have significant influence on the mechanism of action of HA; while this remain unknown to us, our aim was to assess the efficacy of specific commercially available embryo culture media containing HA.…”

“…HA affects cellular processes, including adhesion ( Vigetti et al , 2014 ), as well as promoting embryo development and endometrial decidualization ( Urman et al , 2008 ; Block et al , 2009 ), and has been proposed to promote blastocyst attachment to the endometrial epithelium ( Aplin and Kimber, 2004 ). Our recent experimental study described an in vitro model of embryo implantation and showed a trend towards protracted attachment kinetics of blastocysts exposed to HA-rich ET medium (EmbryoGlue ® ) ( Ruane et al , 2020 ). We further manipulated embryo-endometrial interactions in vitro using mouse blastocysts and found that degradation of HA at the embryo-endometrial interface significantly enhanced the kinetics of blastocyst attachment ( Berneau et al , 2019 ).…”

“…Therefore, in order to continually evaluate the use of EmbryoGlue ® in clinical practice, we carried out two parallel studies. The first, a pre-clinical study on human embryos donated to research, showed that EmbryoGlue ® did not increase the attachment of embryos to endometrial epithelium in vitro ( Ruane et al , 2020 ). In the second study, presented here, we evaluated clinical treatment outcomes in a cohort study before and after EmbryoGlue ® was introduced into routine clinical practice.…”