Potential involvement of the immune system in the development of endometriosis

This article presents an overview of immunological factors and their role in the development of endometriosis, with emphasis on inflammatory cytokines, growth and adhesion factors. Although retrograde menstruation is a common phenomenon among women of reproductive age, not all women who have retrograde menstruation develop endometriosis. The development of endometriosis is hypothesised to be a complex process, which may be facilitated by several factors, including the quantity and quality of endometrial cells in peritoneal fluid (PF), increased inflammatory activity in PF, increased endometrial-peritoneal adhesion and angiogenesis, reduced immune surveillance and clearance of endometrial cells, and increased production of autoantibodies against endometrial cells. Potential biomarkers like cytokines and autoantibodies upregulated during development of endometriosis may be useful in the development of a non-surgical diagnostic tool. Although endometriosis can be treated using hormonal suppression, there is need for non-hormonal drugs, which can inhibit the development of endometriosis and alleviate pain or infertility without inhibition of ovulation. New molecules that modulate immune function in endometriosis should be the targets for future research

Quantity and quality of retrograde menstruation: a case control study

<p>Abstract</p> <p>Background</p> <p>The purpose of this study was to test the hypothesis that menstruation is associated with a higher concentration of endometrial cells in peritoneal fluid(PF) and with increased white and red blood cell concentration in PF when compared to nonmenstrual phases of the cycle.</p> <p>Methods</p> <p>PF was obtained at laparoscopy from 107 women with endometriosis (n = 59) and controls with a normal pelvis (n = 48) during the luteal (n = 46), follicular (n = 38) or menstrual (n = 23) phase of the cycle. Endometriosis was classified according to the classification of the American Society for Reproductive Medicine (rAFS into minimal (n = 25), mild(n = 20), moderate(n = 6) and severe(n = 8) disease. Cell counts (leucocytes, erythrocytes, thrombocytes) were determined on a cell counter. In a subset of 32 patients (13 controls and 19 women with endometriosis), PF was fixed, processed and thinlayers were prepared and stained with Papanicolaou method and with immunocytochemistry using monoclonal antibodies against cytokeratin 7(CK 7), CK 8/18, Ber-Ep4, vimentin, calretinin and CD68. Ber-Ep4 is a marker for cells with epithelial origin (in some cases for mesothelial cells as well). CD68 is specific for cells from monocyte/macrophage lineage; CK7 and CK8/18 are markers for both endometrial epithelial and mesothelial cells, whereas calretinin and vimentin are markers for both endometrial stromal and mesothelial cells.</p> <p>Results</p> <p>In comparison with the nonmenstrual phase of the cycle, analysis of PF during menstruation showed an increased concentration of leucocytes (3.3 &#215; 10⁹/L vs 0.8 &#215; 10⁹/L, P = 0.03), erythrocytes (0.3 &#215; 10¹²/L vs 0.02 &#215; 10¹²/L, P = 0.006), hematocrit (0.03 L/L vs 0.003 L/L, P = 0.01) and hemoglobin (0.8 g/dL vs 0.1 g/dL, P = 0.01). Mesothelial cells stained positively with CK7, CK8/18, vimentin, and calretinin. Cells positive for Ber-Ep4 were not observed, except in 2 patients with endometriosis investigated during menses. In all patients 50-98% of single cells were strongly positive for both vimentin and CD68.</p> <p>Conclusion</p> <p>When compared to nonmenstrual phases of the cycle, menstruation is associated with an increased concentration of red and white blood cells in PF. However, the presence of EM cells that are detectable by immunohistochemistry in PF is low during all phases of the cycle, including menstruation.</p

Intra- and interobserver analysis in the morphological assessment of early stage embryos during an IVF procedure: a multicentre study

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Genome-wide haplotyping embryos developing from 0PN and 1PN zygotes increases transferrable embryos in PGT-M.

STUDY QUESTION Can genome-wide haplotyping increase success following preimplantation genetic testing for a monogenic disorder (PGT-M) by including zygotes with absence of pronuclei (0PN) or the presence of only one pronucleus (1PN)? SUMMARY ANSWER Genome-wide haplotyping 0PNs and 1PNs increases the number of PGT-M cycles reaching embryo transfer (ET) by 81% and the live-birth rate by 75%. WHAT IS KNOWN ALREADY Although a significant subset of 0PN and 1PN zygotes can develop into balanced, diploid and developmentally competent embryos, they are usually discarded because parental diploidy detection is not part of the routine work-up of PGT-M. STUDY DESIGN, SIZE, DURATION This prospective cohort study evaluated the pronuclear number in 2229 zygotes from 2337 injected metaphase II (MII) oocytes in 268 cycles. PGT-M for 0PN and 1PN embryos developing into Day 5/6 blastocysts with adequate quality for vitrification was performed in 42 of the 268 cycles (15.7%). In these 42 cycles, we genome-wide haplotyped 216 good quality embryos corresponding to 49 0PNs, 15 1PNs and 152 2PNs. The reported outcomes include parental contribution to embryonic ploidy, embryonic aneuploidy, genetic diagnosis for the monogenic disorder, cycles reaching ETs, pregnancy and live birth rates (LBR) for unaffected offspring. PARTICIPANTS/MATERIALS, SETTING, METHODS Blastomere DNA was whole-genome amplified and hybridized on the Illumina Human CytoSNP12V2.1.1 BeadChip arrays. Subsequently, genome-wide haplotyping and copy-number profiling was applied to investigate the embryonic genome architecture. Bi-parental, unaffected embryos were transferred regardless of their initial zygotic PN score. MAIN RESULTS AND THE ROLE OF CHANCE A staggering 75.51% of 0PN and 42.86% of 1PN blastocysts are diploid bi-parental allowing accurate genetic diagnosis for the monogenic disorder. In total, 31% (13/42) of the PGT-M cycles reached ET or could repeat ET with an unaffected 0PN or 1PN embryo. The LBR per initiated cycle increased from 9.52 to 16.67%. LIMITATIONS, REASONS FOR CAUTION The clinical efficacy of the routine inclusion of 0PN and 1PN zygotes in PGT-M cycles should be confirmed in larger cohorts from multicenter studies. WIDER IMPLICATIONS OF THE FINDINGS Genome-wide haplotyping allows the inclusion of 0PN and 1PN embryos and subsequently increases the cycles reaching ET following PGT-M and potentially PGT for aneuploidy (PGT-A) and chromosomal structural rearrangements (PGT-SR). Establishing measures of clinical efficacy could lead to an update of the ESHRE guidelines which advise against the use of these zygotes. STUDY FUNDING/COMPETING INTEREST(S) SymBioSys (PFV/10/016 and C1/018 to J.R.V. and T.V.), the Horizon 2020 WIDENLIFE: 692065 to J.R.V., T.V., E.D., A.D. and M.Z.E. M.Z.E., T.V. and J.R.V. co-invented haplarithmisis (‘Haplotyping and copy-number typing using polymorphic variant allelic frequencies’), which has been licensed to Agilent Technologies. H.M. is fully supported by the (FWO) (ZKD1543-ASP/16). The authors have no competing interests to declare

Intra- and inter-observer analysis in the morphological assessment of early-stage embryos

<p>Abstract</p> <p>Background</p> <p>The aim of this study was to determine the intra- and inter-observer variability in the evaluation of embryo quality. Multilevel images of embryos on day 1, day 2 and day 3, were analysed using different morphological parameters.</p> <p>Methods</p> <p>Multilevel images of embryos on day 1, day 2 and day 3, were analysed using a standard scoring system. The kappa coefficient was calculated to measure intra- and inter-observer variability before and after training sessions.</p> <p>Results</p> <p>Good to excellent intra-observer agreement was present for most parameters exceptions being scoring the position of pronuclei and the presence of a cytoplasmic halo on day 1, multinucleation on day 2 and the size of fragments on day 3. Inter-observer agreement was only good to excellent for the number of blastomeres on day 2 and day 3 and the orientation of the cleavage axes on day 2. Training sessions had a positive impact on inter-observer agreement.</p> <p>Conclusion</p> <p>In conclusion, assessment of morphological characteristics of early stage embryos using multilevel images was marked by a high intra-observer and a moderate inter-observer agreement. Training sessions were useful to increase inter-observer agreement.</p

Parental genomes segregate into distinct blastomeres during multipolar zygotic divisions leading to mixoploid and chimeric blastocysts

BACKGROUND: During normal zygotic division, two haploid parental genomes replicate, unite and segregate into two biparental diploid blastomeres. RESULTS: Contrary to this fundamental biological tenet, we demonstrate here that parental genomes can segregate to distinct blastomeres during the zygotic division resulting in haploid or uniparental diploid and polyploid cells, a phenomenon coined heterogoneic division. By mapping the genomic landscape of 82 blastomeres from 25 bovine zygotes, we show that multipolar zygotic division is a tell-tale of whole-genome segregation errors. Based on the haplotypes and live-imaging of zygotic divisions, we demonstrate that various combinations of androgenetic, gynogenetic, diploid, and polyploid blastomeres arise via distinct parental genome segregation errors including the formation of additional paternal, private parental, or tripolar spindles, or by extrusion of paternal genomes. Hence, we provide evidence that private parental spindles, if failing to congress before anaphase, can lead to whole-genome segregation errors. In addition, anuclear blastomeres are common, indicating that cytokinesis can be uncoupled from karyokinesis. Dissociation of blastocyst-stage embryos further demonstrates that whole-genome segregation errors might lead to mixoploid or chimeric development in both human and cow. Yet, following multipolar zygotic division, fewer embryos reach the blastocyst stage and diploidization occurs frequently indicating that alternatively, blastomeres with genome-wide errors resulting from whole-genome segregation errors can be selected against or contribute to embryonic arrest. CONCLUSIONS: Heterogoneic zygotic division provides an overarching paradigm for the development of mixoploid and chimeric individuals and moles and can be an important cause of embryonic and fetal arrest following natural conception or IVF

Effect of day 3 embryo morphometrics and morphokinetics on survival and implantation after slow freezing-thawing and after vitrification-warming: a retrospective cohort study

Morphometric and morphokinetic evaluation of in vitro cultured human embryos allows evaluation without time restriction and reduces intra- and inter-observer variability. Even though these technologies have been reported to improve the quality of cleavage stage embryo evaluation during fresh culture, possible advantages in the evaluation of cryopreserved embryos have been scarcely explored. This study aims to compare morphometric and morphokinetic parameters between slow frozen and vitrified embryos and to determine their relationship to embryo survival and implantation rate (IR) after thawing/warming.status: publishe