Follicle size indicates oocyte maturity and blastocyst formation but not blastocyst euploidy following controlled ovarian hyperstimulation of oocyte donors.

STUDY QUESTION: Is there is an association between follicle size and the quality of oocytes retrieved from them as judged by ability to achieve the blastocyst stage, blastocyst grades and blastocyst ploidy? SUMMARY ANSWER: Although follicle size is a valuable predictor of oocyte maturity and is a significant predictor of the ability of a fertilized oocyte to become a quality blastocyst, the ploidy of each quality blastocyst is not related to the size of the follicle from which its oocyte was retrieved. WHAT IS KNOWN ALREADY: It is unclear whether the oocytes within larger follicles are the best oocytes of the cohort. Although there have been studies examining follicle size in relation to embryo quality, there has been no study relating the incidence of euploidy in embryos to follicle size. STUDY DESIGN, SIZE, DURATION: The purpose of this study was to examine follicle sizes and the oocytes from those follicles (and the embryos that result from those oocytes) to see if there is an association between follicle size and the quality of oocytes as judged by ability to achieve the blastocyst stage, blastocyst grades and blastocyst ploidy. Follicle sizes for oocytes were assessed both as diameters (mm) and as Z values (expressed as their size relative to the mean and standard deviation of that donor's follicular cohort). Comparisons were made using cumulative histograms, rolling averages and receiver operator characteristic (ROC) curves and its AUC. PARTICIPANTS/MATERIALS, SETTING, METHODS: Twenty-two oocyte donors (ages: 24.5 ± 3.5 years) whose recipients would use ICSI for insemination were enrolled in this study. Follicles were aspirated one-at-a-time to be certain that the aspirated oocyte was from the same follicle measured. The follicle measurement (size) was noted in the embryology records. Oocytes were cultured individually throughout their time in the embryology laboratory so that follicle sizes could be uniquely associated with each oocyte. Oocytes and embryos were analyzed according to the size of the follicle from which the oocyte was retrieved. MAIN RESULTS AND THE ROLE OF CHANCE: Three hundred seventeen oocytes (96.1%) had an associated follicle size. Of the oocytes with follicle sizes, 255 (80.4%) had a polar body (MII), and 60 (18.9%) were immature: 31 (9.8%) with a visible germinal vesicle (GV stage) and 29 (9.1%) with neither a polar body nor a visible germinal vesicle (MI). The incidence of MII oocytes was significantly associated with larger follicle size using either mm (ROC's AUC = 0.87; P < 0.0001) or Z values (ROC's AUC = 0.86; P < 0.0001). Among MII oocytes there was no association with follicle size for the appearance of 228 oocytes with two pronuclei (2 PN). Among 2 PN's, the development of 94 quality blastocysts that underwent trophectoderm biopsy (TE Bx) exhibited a significant association with larger follicles using either mm (ROC's AUC = 0.59; P = 0.01) or Z values (ROC's AUC = 0.57; P = 0.01). The use of follicle diameter as a feature to distinguish between fertilized oocytes that would ultimately become blastocysts versus those that would not become blastocysts resulted in an enrichment for blastocyst formation from 20 to 40%. Of the 94 quality blastocysts, 51 were determined by next generation sequencing (NGS) to be euploid.Although oocyte maturity and the incidence of blastocyst formation were associated with follicle size, the incidence of euploidy among biopsied blastocysts was not. Follicles measured by two different methods (mm or Z values) led to predominantly the same conclusions. LIMITATIONS, REASONS FOR CAUTION: This study investigated the relationship between follicle size and measures of oocyte/embryo quality when donors were treated similarly. Therefore, this study does not investigate the effects of triggering and retrieving oocytes when the follicle cohorts are of different sizes or lead follicles are of different sizes. Although no association was found between follicle size and euploid blastocysts, the fact that blastocyst ploidy is not entirely dependent upon oocyte ploidy (e.g. aneuploidies derived from mitotic errors or from the fertilizing sperm) makes it difficult to infer the relationship between follicle diameter and oocyte ploidy. WIDER IMPLICATIONS OF THE FINDINGS: It is confirmed that follicle diameter is predictive of oocyte maturity. However, once oocyte maturity is known, the diameter of the follicle from which the oocyte was retrieved is not instructive. Embryos generated through fertilization and development of the mature oocytes from any observed follicle diameter were equally likely to become euploid blastocysts. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by ReproART: Georgian American Center for Reproductive Medicine. None of the authors declare any actual conflicts of interest. D.H.M. received compensation from ReproART, Biogenetics Corporation and the Sperm and Embryo Bank of New York and honoraria and travel funding from Ferring Pharmaceuticals and from Granata Bio. S.M. received compensation from Cooper Genomics and an honorarium and travel funding from Ferring Pharmaceuticals. L.C. is the founder of LTD Ovamedi, the organization that represents Cooper Genomics in Georgia, and received travel funding from the European Society for Human Reproduction and Embryology. TRIAL REGISTRATION NUMBER: N/A.

Adaptive Foxp3+ regulatory T cell-dependent and -independent control of allergic inflammation.

Adaptive Foxp3(+) regulatory T (Treg) cells develop during induction of mucosal tolerance and after immunization. Large numbers of Foxp3(+) T cells have been found in inflamed tissues. We investigated the role of adaptive Foxp3(+) Treg cells in mucosal tolerance and in chronic allergic lung inflammation. We used two strains of mice that are devoid of naturally occurring Treg cells; one is capable of generating adaptive Foxp3(+) Treg cells upon exposure to antigen, whereas the other is deficient in both naturally occurring and adaptive Foxp3(+) Treg cells. We found that adaptive Foxp3(+) Treg cells were essential for establishing mucosal tolerance and for suppressing IL-4 production and lymphoid neogenesis in chronic inflammation, whereas IL-5 production and eosinophilia could be controlled by Foxp3-independent, IFN-gamma-dependent mechanisms. Thus, whereas adaptive Foxp3(+) Treg cells regulate sensitization to allergens and the severity of chronic inflammation, IFN-gamma-producing cells can play a beneficial role in inflammatory conditions involving eosinophils.

Management of Recurrent Implantation Failure and Hereditary Thrombophilia: A Case Report.

Recurrent implantation failure (RIF) is one of the core problems for assisted reproductive technology (ART). High-quality, euploid embryos and synchronization between the embryonic stage and the uterine endometrial lining are crucial for positive outcomes. Molecular biology techniques have significantly transformed assisted reproductive technology (ART). Numerous couples facing infertility issues have successfully achieved the birth of healthy infants through the application of molecular biology methods: preimplantation genetic testing for aneuploidy (PGT-A) and endometrial receptivity analysis (ERA). Exploring the impact of age on endometrial assays like the endometrial receptivity assay (ERA) yields valuable insights, including the determination of the implantation window and the development of personalized strategies. The authors present the case of a 42-year-old woman who has experienced RIF with euploid embryos, coupled with a hereditary thrombophilia homozygous mutation in the MTHFR genes: A1298C and C677T.

Interleukin 2 signaling is required for CD4(+) regulatory T cell function.

Mice deficient in interleukin (IL)-2 production or the IL-2 receptor alpha or beta chains develop a lethal autoimmune syndrome. CD4(+) regulatory T cells have been shown to prevent autoimmune diseases, allograft rejection, and to down-regulate antibody responses against foreign antigens. To assess the role of IL-2 in the generation and function of regulatory T cells, we transferred CD4(+) T cells from mice genetically deficient in IL-2 or IL-2R(alpha) (CD25) expression. A small number of splenic or thymic CD4(+) T cells from IL-2 knockout mice can protect mice from spontaneous experimental autoimmune encephalomyelitis (EAE). In contrast, splenic or thymic CD4(+) T cells from CD25 knockout donor mice conferred little or no protection. We conclude that T cells with regulatory potential can develop, undergo thymic selection, and migrate to the peripheral lymphoid organs in the absence of IL-2, and do not protect from disease by means of IL-2 secretion. However, IL-2 signaling in regulatory T cells is essential for their protective function. Altogether, our results favor a model whereby IL-2 induces regulatory T cell activity.

Oral tolerance in the absence of naturally occurring Tregs.

Mucosal tolerance prevents pathological reactions against environmental and food antigens, and its failure results in exacerbated inflammation typical of allergies and asthma. One of the proposed mechanisms of oral tolerance is the induction of Tregs. Using a mouse model of hyper-IgE and asthma, we found that oral tolerance could be effectively induced in the absence of naturally occurring thymus-derived Tregs. Oral antigen administration prior to i.p. immunization prevented effector/memory Th2 cell development, germinal center formation, class switching to IgE, and lung inflammation. Oral exposure to antigen induced development of antigen-specific CD4CD25Foxp3CD45RB cells that were anergic and displayed suppressive activity in vivo and in vitro. Oral tolerance to the Th2 allergic response was in large part dependent on TGF-beta and independent of IL-10. Interestingly, Tregs were also induced by single i.p. immunization with antigen and adjuvant. However, unlike oral administration of antigen, which induced Tregs but not effector T cells, i.p. immunization led to the simultaneous induction of Tregs and effector Th2 cells displaying the same antigen specificity.

Unique maturation program of the IgE response in vivo.

A key event in the pathogenesis of asthma and allergies is the production of IgE antibodies. We show here that IgE(+) cells were exceptional because they were largely found outside germinal centers and expressed, from very early on, a genetic program of plasma cells. In spite of their extragerminal center localization, IgE(+) cells showed signs of somatic hypermutation and affinity maturation. We demonstrated that high-affinity IgE(+) cells could be generated through a unique differentiation program that involved two phases: a pre-IgE phase in which somatic hypermutation and affinity maturation take place in IgG1(+) cells, and a post-IgE-switching phase in which IgE cells differentiate swiftly into plasma cells. Our results have implications for the understanding of IgE memory responses in allergy.

CD25- T cells generate CD25+Foxp3+ regulatory T cells by peripheral expansion.

Naturally occurring CD4(+) regulatory T cells are generally identified through their expression of CD25. However, in several experimental systems considerable T(reg) activity has been observed in the CD4(+)CD25(-) fraction. Upon adoptive transfer, the expression of CD25 in donor-derived cells is not stable, with CD4(+)CD25(+) cells appearing in CD4(+)CD25(-) T cell-injected animals and vice versa. We show in this study that CD25(+) cells arising from donor CD25(-) cells upon homeostatic proliferation in recipient mice express markers of freshly isolated T(reg) cells, display an anergic state, and suppress the proliferation of other cells in vitro. The maintenance of CD25 expression by CD4(+)CD25(+) cells depends on IL-2 secreted by cotransferred CD4(+)CD25(-) or by Ag-stimulated T cells in peripheral lymphoid organs.