Cellular bases of olfactory circuit assembly revealed by systematic time-lapse imaging.

Neural circuit assembly features simultaneous targeting of numerous neuronal processes from constituent neuron types, yet the dynamics is poorly understood. Here, we use the Drosophila olfactory circuit to investigate dynamic cellular processes by which olfactory receptor neurons (ORNs) target axons precisely to specific glomeruli in the ipsi- and contralateral antennal lobes. Time-lapse imaging of individual axons from 30 ORN types revealed a rich diversity in extension speed, innervation timing, and ipsilateral branch locations and identified that ipsilateral targeting occurs via stabilization of transient interstitial branches. Fast imaging using adaptive optics-corrected lattice light-sheet microscopy showed that upon approaching target, many ORN types exhibiting "exploring branches" consisted of parallel microtubule-based terminal branches emanating from an F-actin-rich hub. Antennal nerve ablations uncovered essential roles for bilateral axons in contralateral target selection and for ORN axons to facilitate dendritic refinement of postsynaptic partner neurons. Altogether, these observations provide cellular bases for wiring specificity establishment.

SPIRO - the automated Petri plate imaging platform designed by biologists, for biologists.

Phenotyping of model organisms grown on Petri plates is often carried out manually, despite the procedures being time-consuming and laborious. The main reason for this is the limited availability of automated phenotyping facilities, whereas constructing a custom automated solution can be a daunting task for biologists. Here, we describe SPIRO, the Smart Plate Imaging Robot, an automated platform that acquires time-lapse photographs of up to four vertically oriented Petri plates in a single experiment, corresponding to 192 seedlings for a typical root growth assay and up to 2500 seeds for a germination assay. SPIRO is catered specifically to biologists' needs, requiring no engineering or programming expertise for assembly and operation. Its small footprint is optimized for standard incubators, the inbuilt green LED enables imaging under dark conditions, and remote control provides access to the data without interfering with sample growth. SPIRO's excellent image quality is suitable for automated image processing, which we demonstrate on the example of seed germination and root growth assays. Furthermore, the robot can be easily customized for specific uses, as all information about SPIRO is released under open-source licenses. Importantly, uninterrupted imaging allows considerably more precise assessment of seed germination parameters and root growth rates compared with manual assays. Moreover, SPIRO enables previously technically challenging assays such as phenotyping in the dark. We illustrate the benefits of SPIRO in proof-of-concept experiments which yielded a novel insight on the interplay between autophagy, nitrogen sensing, and photoblastic response.

Genetically Encoded Tools for Optical Dissection of the Mammalian Cell Cycle.

Eukaryotic cells spend most of their life in interphase of the cell cycle. Understanding the rich diversity of metabolic and genomic regulation that occurs in interphase requires the demarcation of precise phase boundaries in situ. Here, we report the properties of two genetically encoded fluorescence sensors, Fucci(CA) and Fucci(SCA), which enable real-time monitoring of interphase and cell-cycle biology. We re-engineered the Cdt1-based sensor from the original Fucci system to respond to S phase-specific CUL4Ddb1-mediated ubiquitylation alone or in combination with SCFSkp2-mediated ubiquitylation. In cultured cells, Fucci(CA) produced a sharp triple color-distinct separation of G1, S, and G2, while Fucci(SCA) permitted a two-color readout of G1 and S/G2. Fucci(CA) applications included tracking the transient G1 phase of rapidly dividing mouse embryonic stem cells and identifying a window for UV-irradiation damage in S phase. These results show that Fucci(CA) is an essential tool for quantitative studies of interphase cell-cycle regulation.

Considerations for future modification of The Association for the Study of Reproductive Biology embryo grading system incorporating time-lapse observations.

The Association for the Study of Reproductive Biology (ASEBIR) Interest Group in Embryology (in Spanish 'Grupo de Interés de Embriología') reviewed key morphokinetic parameters to assess the contribution of time-lapse technology (TLT) to the ASEBIR grading system. Embryo grading based on morphological characteristics is the most widely used method in human assisted reproduction laboratories. The introduction and implementation of TLT has provided a large amount of information that can be used as a complementary tool for morphological embryo evaluation and selection. As part of IVF treatments, embryologists grade embryos to decide which embryos to transfer or freeze. At the present, the embryo grading system developed by ASEBIR does not consider dynamic events observed through TLT. Laboratories that are using TLT consider those parameters as complementary data for embryo selection. The aim of this review was to evaluate review time-specific morphological changes during embryo development that are not included in the ASEBIR scoring system, and to consider them as candidates to add to the scoring system.

Oscillatory control of Delta-like1 in cell interactions regulates dynamic gene expression and tissue morphogenesis.

Notch signaling regulates tissue morphogenesis through cell-cell interactions. The Notch effectors Hes1 and Hes7 are expressed in an oscillatory manner and regulate developmental processes such as neurogenesis and somitogenesis, respectively. Expression of the mRNA for the mouse Notch ligand Delta-like1 (Dll1) is also oscillatory. However, the dynamics of Dll1 protein expression are controversial, and their functional significance is unknown. Here, we developed a live-imaging system and found that Dll1 protein expression oscillated in neural progenitors and presomitic mesoderm cells. Notably, when Dll1 expression was accelerated or delayed by shortening or elongating the Dll1 gene, Dll1 oscillations became severely dampened or quenched at intermediate levels, as modeled mathematically. Under this condition, Hes1 and Hes7 oscillations were also dampened. In the presomitic mesoderm, steady Dll1 expression led to severe fusion of somites and their derivatives, such as vertebrae and ribs. In the developing brain, steady Dll1 expression inhibited proliferation of neural progenitors and accelerated neurogenesis, whereas optogenetic induction of Dll1 oscillation efficiently maintained neural progenitors. These results indicate that the appropriate timing of Dll1 expression is critical for the oscillatory networks and suggest the functional significance of oscillatory cell-cell interactions in tissue morphogenesis.

Developmental perturbation in human embryos: Clinical and biological significance learned from time-lapse images.

Background: Time-lapse technology (TLT) has gained widespread adoption worldwide. In addition to facilitating the undisturbed culture of embryos, TLT offers the unique capability of continuously monitoring embryos to detect spatiotemporal changes. Although these observed phenomena play a role in optimal embryo selection/deselection, the clinical advantages of introducing TLT remain unclear. However, manual annotation of embryo perturbation could facilitate a comprehensive assessment of developmental competence. This process requires a thorough understanding of embryo observation and the biological significance associated with developmental dogma and variation. This review elucidates the typical behavior and variation of each phenomenon, exploring their clinical significance and research perspectives. Methods: The MEDLINE database was searched using PubMed for peer-reviewed English-language original articles concerning human embryo development. Main findings: TLT allows the observation of consecutive changes in embryo morphology, serving as potential biomarkers for embryo assessment. In assisted reproductive technology laboratories, several phenomena have not revealed their mechanism, posing difficulties such as fertilization deficiency and morula arrest. Conclusion: A profound understanding of the biological mechanisms and significance of each phenomenon is crucial. Further collaborative efforts between the clinical and molecular fields following translational studies are required to advance embryonic outcomes and assessment.

Two coral fluorescent proteins of distinct colors for sharp visualization of cell-cycle progression.

We cloned and characterized two new coral fluorescent proteins: h2-3 and 1-41. h2-3 formed an obligate dimeric complex and exhibited bright green fluorescence. On the other hand, 1-41 formed a highly multimeric complex and exhibited dim red fluorescence. We engineered 1-41 into AzaleaB5, a practically useful red-emitting fluorescent protein for cellular labeling applications. We fused h2-3 and AzaleaB5 to the ubiquitination domains of human Geminin and Cdt1, respectively, to generate a new color variant of Fucci (Fluorescent Ubiquitination-based Cell-Cycle Indicator): Fucci5. We found Fucci5 provided more reliable nuclear labeling for monitoring cell-cycle progression than the 1st and 2nd generations that used mAG/mKO2 and mVenus/mCherry, respectively.Key words: fluorescent protein, cell cycle, time-lapse imaging, flow cytometry.

In-Depth Quantification of Cell Division and Elongation Dynamics at the Tip of Growing Arabidopsis Roots Using 4D Microscopy, AI-Assisted Image Processing and Data Sonification.

One of the fundamental questions in plant developmental biology is how cell proliferation and cell expansion coordinately determine organ growth and morphology. An amenable system to address this question is the Arabidopsis root tip, where cell proliferation and elongation occur in spatially separated domains, and cell morphologies can easily be observed using a confocal microscope. While past studies revealed numerous elements of root growth regulation including gene regulatory networks, hormone transport and signaling, cell mechanics and environmental perception, how cells divide and elongate under possible constraints from cell lineages and neighboring cell files has not been analyzed quantitatively. This is mainly due to the technical difficulties in capturing cell division and elongation dynamics at the tip of growing roots, as well as an extremely labor-intensive task of tracing the lineages of frequently dividing cells. Here, we developed a motion-tracking confocal microscope and an Artificial Intelligence (AI)-assisted image-processing pipeline that enables semi-automated quantification of cell division and elongation dynamics at the tip of vertically growing Arabidopsis roots. We also implemented a data sonification tool that facilitates human recognition of cell division synchrony. Using these tools, we revealed previously unnoted lineage-constrained dynamics of cell division and elongation, and their contribution to the root zonation boundaries.

Aging induces aberrant state transition kinetics in murine muscle stem cells.

Murine muscle stem cells (MuSCs) experience a transition from quiescence to activation that is required for regeneration, but it remains unknown if the trajectory and dynamics of activation change with age. Here, we use time-lapse imaging and single cell RNA-seq to measure activation trajectories and rates in young and aged MuSCs. We find that the activation trajectory is conserved in aged cells, and we develop effective machine-learning classifiers for cell age. Using cell-behavior analysis and RNA velocity, we find that activation kinetics are delayed in aged MuSCs, suggesting that changes in stem cell dynamics may contribute to impaired stem cell function with age. Intriguingly, we also find that stem cell activation appears to be a random walk-like process, with frequent reversals, rather than a continuous linear progression. These results support a view of the aged stem cell phenotype as a combination of differences in the location of stable cell states and differences in transition rates between them.

Time-lapse monitoring: An adjunct tool to select embryos for preimplantation genetic testing.

The goal for the present study was to investigate the effect of aneuploidy on embryo morphokinetics events in a time-lapse imaging (TLI) system incubator. This retrospective cohort study was performed in a private university-affiliated in vitro fertilization center, between 2019 March and December 2020. Kinetic data were analyzed in 935 embryos, derived from 316 patients undergoing intracytoplasmic sperm injection cycle with preimplantation genetic testing (PGT) for aneuploidy, individually cultured in a TLI incubator until Day 5 of development. Timing of morphokinetic variables, the incidences of multinucleation, and Known Implantation Data Score (KIDScore)-Day 5 were compared between euploid (n = 352) and aneuploid embryos (n = 583). Aneuploid embryos showed significantly longer timing to complete specific morphokinetic parameters compared to euploidy embryos. Euploidy embryos also showed a significantly higher KIDScore when compared with the aneuploidy ones. Our evidence suggests that TLI monitoring may be an adjunct approach to select embryos for PGT; however, cautious investigation is still needed.

The presence of partial compaction patterns is associated with lower rates of blastocyst formation, sub-optimal morphokinetic parameters and poorer morphologic grade.

BACKGROUND: Compaction is an important marker of embryonic genome activation and marks a critical step in the development to blastocyst. The objective of our study was to determine whether visualization of the embryonic compaction process through time-lapse imaging (TL) can assist in predicting the kinetics of embryo development as well as the likelihood for blastocyst formation, grade, or ploidy. METHODS: This study is a retrospective review of prospectively collected datafrom a single academic institution. Couples included were thosewho underwent preimplantation genetic testing for aneuploidy (PGT-A) following in vitro fertilization between Januaryand December 2020. Embryos were cultured in the Embrysocope. Embryo morphokinetic data was prospectively collected and analyzed.TL videos werelater reviewed in detail for compaction pattern. Embryo compaction patterns (CP) were categorized as follows: 1) full compaction (CP-F), 2) partial compaction with cell extrusion (P-ext), 3) partial compactionwith cell exclusion (P-exc) and 4) partial compactionwith both cell extrusion and exclusion (P-both). Assessment of embryo decompaction and re-compaction was evaluated. The association between CP, morphokinetic parameters,blastocyst formation, grade and ploidy were then analyzed. RESULTS: A total of 349 embryos were studied. Amongst embryos which progressed to morula (n = 281), the distribution of compaction patterns were: CP-F 45.6%, P-ext12.5%, P-exc29.5% and P-both 12.5%. Embryos exhibiting a CP-F were more likely to proceed to blastocyst compared with those that demonstrated partial compaction patterns (p = 0.006). When compared to CP-F, partial compaction patterns were significantly associated with poorer ICM and TE grades (P < 0.001). Of the 281 morula, 59.8% (n = 168) demonstrated at least one episode of decompaction and re-compaction. Of the 249 blastocysts formed, 200 were cryopreserved for future use after undergoing PGT-A evaluation. Of those, 42.5% were diagnosed as euploid, 39.0% as aneuploid, 9.0% as mosaic and 9.5% had no result. When compared to CP-F, partialCPs exhibited a significantly greater percentage of mosaic embryos (3.6% v. 15.6%, p = 0.032). Additionally, we found that a greater percentage of embryos demonstrating CP-F exhibited morphokinetics that fell into optimal ranges for embryo development when compared to those with partial compaction patterns. CONCLUSION: Time-lapse visualization of compaction patterns identified exclusions and/or extrusions as negative indicators of blastocyst formation and blastocyst grade. When compared to full compaction patterns, partial compaction patterns were associated with delayed embryonic development as well as lower rates of optimal kinetic development.

Using the embryo-uterus statistical model to predict pregnancy chances by using cleavage stage morphokinetics and female age: two centre-specific prediction models and mutual validation.

BACKGROUND: The predictive capability of time-lapse monitoring (TLM) selection algorithms is influenced by patient characteristics, type and quality of data included in the analysis and the used statistical methods. Previous studies excluded DET cycles of which only one embryo implanted, introducing bias into the data. Therefore, we wanted to develop a TLM prediction model that is able to predict pregnancy chances after both single- and double embryo transfer (SET and DET). METHODS: This is a retrospective study of couples (n = 1770) undergoing an in vitro fertilization cycle at the Erasmus MC, University Medical Centre Rotterdam (clinic A) or the Reinier de Graaf Hospital (clinic B). This resulted in 2058 transferred embryos with time-lapse and pregnancy outcome information. For each dataset a prediction model was established by using the Embryo-Uterus statistical model with the number of gestational sacs as the outcome variable. This process was followed by cross-validation. RESULTS: Prediction model A (based on data of clinic A) included female age, t3-t2 and t5-t4, and model B (clinic B) included female age, t2, t3-t2 and t5-t4. Internal validation showed overfitting of model A (calibration slope 0.765 and area under the curve (AUC) 0.60), and minor overfitting of model B (slope 0.915 and AUC 0.65). External validation showed that model A was capable of predicting pregnancy in the dataset of clinic B with an AUC of 0.65 (95% CI: 0.61-0.69; slope 1.223, 95% CI: 0.903-1.561). Model B was less accurate in predicting pregnancy in the dataset of clinic A (AUC 0.60, 95% CI: 0.56-0.65; slope 0.671, 95% CI: 0.422-0.939). CONCLUSION: Our study demonstrates a novel approach to the development of a TLM prediction model by applying the EU statistical model. With further development and validation in clinical practice, our prediction model approach can aid in embryo selection and decision making for SET or DET.

Comparison of euploid blastocyst expansion with subgroups of single chromosome, multiple chromosome, and segmental aneuploids using an AI platform from donor egg embryos.

PURPOSE: This retrospective observational study compares how different classes of blastocyst genotypes from egg donor cycles differentially blastulate and expand using a standard assay. METHODS: Quantitative measurements of expansion utilized a customized neural network that segments all sequential time-lapse images during the first 10 h of expansion. RESULTS: Analyses were performed using two developmental time perspectives using time-lapse imaging. The first was the time to blastocyst formation (tB), which broadly reflects variations in developmental rate. Euploidy peaked at 100-115 h from fertilization. In contrast, aneuploidy peaks flanked this interval bi-modally. These distributions limit ploidy discrimination based upon traditional standard grading features when assessed in real time. In contrast, from the second perspective of progressive blastocyst expansion that is normalized to each individual blastocyst's tB time, euploidy was significantly increased at expansion values > 20,000µ2 across all tB intervals studied. A Cartesian coordinate plot graphically summarizes information useful to rank order blastocysts within cohorts for transfer. Defined aneuploidy subgroups, distinguished by the number and complexity of chromosomes involved, also showed distributive differences from both euploids and from each other. A small subset of clinically significant trisomies did not show discriminating features separating them from other euploids. CONCLUSION: A standard assay of blastocyst expansion normalized to each individual blastocyst's time of blastocyst formation more usefully discriminates euploidy from aneuploidy than real-time expansion comparisons using absolute developmental time from fertilization.

In ovo electroporation of chicken limb bud ectoderm: Electroporation to chick limb ectoderm: Electroporation to chick limb ectoderm.

BACKGROUND: Deciphering how ectodermal tissues form, and how they maintain their integrity, is crucial for understanding epidermal development and pathogenesis. However, lack of simple and rapid gene manipulation techniques limits genetic studies to elucidate mechanisms underlying these events. RESULTS: Here we describe an easy method for electroporation of chick limb bud ectoderm enabling gene manipulation during ectoderm development and wound healing. Taking advantage of a small parafilm well that constrains DNA plasmids locally and the fact that the limb ectoderm arises from a defined site, we target the limb ectoderm forming region by in ovo electroporation. This approach results in focal and efficient transgenesis of the limb ectodermal cells. Further, using a previously described Msx2 promoter, gene manipulation can be specifically targeted to the apical ectodermal ridge (AER), a signaling center regulating limb development. Using the electroporation technique to deliver a fluorescent marker into the embryonic limb ectoderm, we show its utility in performing time-lapse imaging during wound healing. This analysis revealed previously unrecognized dynamic remodeling of the actin cytoskeleton and lamellipodia formation at the edges of the wound. We find that the lamellipodia formation requires activity of Rac1 GTPase, suggesting its necessity for wound closure. CONCLUSION: Our method is simple and easy. Thus, it would permit high throughput tests for gene function during limb ectodermal development and wound healing.

Transient localization of the Arp2/3 complex initiates neuronal dendrite branching in vivo.

The formation of neuronal dendrite branches is fundamental for the wiring and function of the nervous system. Indeed, dendrite branching enhances the coverage of the neuron's receptive field and modulates the initial processing of incoming stimuli. Complex dendrite patterns are achieved in vivo through a dynamic process of de novo branch formation, branch extension and retraction. The first step towards branch formation is the generation of a dynamic filopodium-like branchlet. The mechanisms underlying the initiation of dendrite branchlets are therefore crucial to the shaping of dendrites. Through in vivo time-lapse imaging of the subcellular localization of actin during the process of branching of Drosophila larva sensory neurons, combined with genetic analysis and electron tomography, we have identified the Actin-related protein (Arp) 2/3 complex as the major actin nucleator involved in the initiation of dendrite branchlet formation, under the control of the activator WAVE and of the small GTPase Rac1. Transient recruitment of an Arp2/3 component marks the site of branchlet initiation in vivo These data position the activation of Arp2/3 as an early hub for the initiation of branchlet formation.

Na+/K+ ATPase α1 and ß3 subunits are localized to the basolateral membrane of trophectoderm cells in human blastocysts.

STUDY QUESTION: Is there a relation between specific Na+/K+ ATPase isoform expression and localization in human blastocysts and the developmental behavior of the embryo? SUMMARY ANSWER: Na+/K+ ATPase α1, ß1 and ß3 are the main isoforms expressed in human blastocysts and no association was found between the expression level of their respective mRNAs and the rate of blastocyst expansion. WHAT IS KNOWN ALREADY: In mouse embryos, Na+/K+ ATPase α1 and ß1 are expressed in the basolateral membrane of trophectoderm (TE) cells and are believed to be involved in blastocoel formation (cavitation). STUDY DESIGN, SIZE, DURATION: A total of 20 surplus embryos from 11 patients who underwent IVF and embryo transfer at a university hospital between 2009 and 2018 were analyzed. PARTICIPANTS/MATERIALS, SETTING, METHODS: After freezing and thawing Day 5 human blastocysts, their developmental behavior was observed for 24 h using time-lapse imaging, and the expression of Na+/K+ ATPase isoforms was examined using quantitative RT-PCR (RT-qPCR). The expressed isoforms were then localized in blastocysts using fluorescent immunostaining. MAIN RESULTS AND THE ROLE OF CHANCE: RT-qPCR results demonstrated the expression of Na+/K+ ATPase α1, ß1 and ß3 isoforms in human blastocysts. Isoforms α1 and ß3 were localized to the basolateral membrane of TE cells, and ß1 was localized between TE cells. A high level of ß3 mRNA expression correlated with easier hatching (P = 0.0261). LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The expression of mRNA and the localization of proteins of interest were verified, but we have not been able to perform functional analysis. WIDER IMPLICATIONS OF THE FINDINGS: Of the various Na+/K+ ATPase isoforms, expression levels of the α1, ß1 and ß3 mRNAs were clearly higher than other isoforms in human blastocysts. Since α1 and ß3 were localized to the basolateral membrane via fluorescent immunostaining, we believe that these subunits contribute to the dilation of the blastocoel. The ß1 isoform is localized between TE cells and may be involved in tight junction formation, as previously reported in mouse embryos. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the JSPS KAKENHI (https://www.jsps.go.jp/english/index.html), grant number 17K11215. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The authors have no conflicts of interest.

Longitudinal surface measurements of human blastocysts show that the dynamics of blastocoel expansion are associated with fertilization method and ongoing pregnancy.

BACKGROUND: Despite all research efforts during this era of novel time-lapse morphokinetic parameters, a morphological grading system is still routinely being used for embryo selection at the blastocyst stage. The blastocyst expansion grade, as evaluated during morphological assessment, is associated with clinical pregnancy. However, this assessment is performed without taking the dynamics of blastocoel expansion into account. Here, we studied the dynamics of blastocoel expansion by comparing longitudinal blastocoel surface measurements using time-lapse embryo culture. Our aim was to first assess if this is impacted by fertilization method and second, to study if an association exists between these measurement and ongoing pregnancy. METHODS: This was a retrospective cohort study including 225 couples undergoing 225 cycles of in vitro fertilization (IVF) treatment with time-lapse embryo culture. The fertilization method was either conventional IVF, intracytoplasmic sperm injection (ICSI) with ejaculated sperm or ICSI with sperm derived from testicular sperm extraction (TESE-ICSI). This resulted in 289 IVF embryos, 218 ICSI embryos and 259 TESE-ICSI embryos that reached at least the full blastocyst stage. Blastocoel surface measurements were performed on time-lapse images every hour, starting from full blastocyst formation (tB). Linear mixed model analysis was performed to study the association between blastocoel expansion, the calculated expansion rate (µm2/hour) and both fertilization method and ongoing pregnancy. RESULTS: The blastocoel of both ICSI embryos and TESE-ICSI embryos was significantly smaller than the blastocoel of IVF embryos (beta -1121.6 µm2; 95% CI: -1606.1 to -637.1, beta -646.8 µm2; 95% CI: -1118.7 to 174.8, respectively). Still, the blastocoel of transferred embryos resulting in an ongoing pregnancy was significantly larger (beta 795.4 µm2; 95% CI: 15.4 to 1575.4) and expanded significantly faster (beta 100.9 µm2/hour; 95% CI: 5.7 to 196.2) than the blastocoel of transferred embryos that did not, regardless of the fertilization method. CONCLUSION: Longitudinal blastocyst surface measurements and expansion rates are promising non-invasive quantitative markers that can aid embryo selection for transfer and cryopreservation. TRIAL REGISTRATION: Our study is a retrospective observational study, therefore trial registration is not applicable.

Similar fertilization rates and preimplantation embryo development among testosterone-treated transgender men and cisgender women.

RESEARCH QUESTION: What are the effects of testosterone treatment on oocyte fertilization and preimplantation embryo development among transgender men who have undergone fertility preservation? DESIGN: A retrospective study was undertaken in a university-affiliated tertiary hospital between April 2016 and November 2021. Embryos were divided into three groups by source: 210 embryos from 7 testosterone-exposed transgender men, 135 from 10 cisgender women who cryopreserved embryos, and 276 from 24 cisgender women who underwent fertility treatment. Statistical analyses compared assisted reproductive technology outcomes between the group of transgender men and both groups of cisgender women. Morphokinetic and morphological parameters were compared between the embryos derived from these three groups. RESULTS: The transgender men (30.2 ± 3.5 years of age) were significantly younger than the cisgender women who cryopreserved embryos (35.1 ± 1.8 years; P = 0.005) and the cisgender women who underwent fertility treatment (33.8 ± 3.2 years; P = 0.017). After adjusting for participant age, the fertilization rate was comparable between the transgender men and both groups of cisgender women (P = 0.391 and 0.659). There were no significant differences between the transgender men and the cisgender women who preserved fertility in terms of number of cryopreserved embryos (7.2 ± 5.1 and 3.5 ± 2.6; P = 0.473) or the distribution of embryo age at cryopreservation (P = 0.576). All morphokinetic parameters evaluated by time-lapse imaging, as well as the morphological characteristics, were comparable for the embryos in all three groups. CONCLUSIONS: Testosterone exposure among transgender men has no adverse impact upon fertilization rates or preimplantation embryo development and quality.

Improved embryonic development and utilization rates with EmbryoScope: a within-subject comparison versus a benchtop incubator.

The objective of this study was to investigate whether, in consecutive intracytoplasmic sperm injection (ICSI) cycles, embryonic development in an incubator with a time-lapse imaging (TLI) system is better than the previous one obtained in a benchtop incubator (G-185) with similar cultivation characteristics. The study was of a retrospective within-subject design, in which each cycle served as its own control. Data were obtained via the chart review of patients undergoing ICSI in a private university-affiliated in vitro fertilization (IVF) centre who fulfilled the following criteria: second ICSI attempt in which embryos were cultured in a TLI incubator system (TLI group, n = 71), preceded by a first ICSI attempt in which embryos were cultured in a benchtop incubator (Control group, n = 71). Embryonic development up to the fifth day of development, oocyte utilization rate (OUR; transferred embryos plus frozen embryos per total number of retrieved oocytes) and embryo utilization rate (EUR; transferred embryos plus frozen embryos per normally fertilized oocyte) were compared between the groups. There were significant differences in the day 2 non-cleavage rate, day 5 embryo rate, blastocyst development rate, frozen blastocyst rate, OUR, and EUR, in favour of the TLI group. Embryonic development, frozen blastocyst rate, OUR and EUR in the second ICSI cycle were significantly improved when the culture was performed in the EmbryoScope, compared with those rates obtained with culture in a G-185 in the first ICSI cycle of the same patients. The results may also lead to higher cumulative pregnancy outcomes following embryo thawing and transfer.

The KIDScore™ D3 scoring system contributes to the prediction of embryonic development potential: A promising tool for screening high-quality embryos.

Using the KIDScoreTM D3 (KID3) scoring system, day 3 embryos observed by time-lapse imaging (TLI) were scored to explore the predictive value of the KID scoring system on the developmental potential of embryos. The kinetic parameters of 477 normal fertilized embryos from 77 patients who underwent TLI in our hospital from January 2019 to June 2020 were evaluated by KID3, and the embryos were divided into five groups according to the scores for retrospective analysis of blastocyst formation. Additionally, the high-quality blastocyst formation rate, pregnancy rate and early abortion rate were analyzed via KID3 and traditional morphological assessments, and comparisons of differences among different ages were also performed. In the KID3 estimate, the blastocyst or high-quality blastocyst formation rate in the score 5 group was markedly higher than that in the score 1-4 groups. Blastocyst or high-quality blastocyst formation rates in the A group (the results of two evaluation tools indicated they were excellent embryos) and the B group (KID3: excellent embryos, traditional evaluation: not excellent embryos) were evidently increased in comparison with the C or D group (KID3: not excellent embryos, traditional evaluation: excellent embryo or not, respectively). Furthermore, the percentages of score 5 embryos, blastocyst and high-quality blastocyst formation rates for patients ≥ 35 years old were markedly decreased compared with those for patients < 34 years old, while the trends of nondiploid cleavage, multinucleation and asymmetric division were the opposite. Collectively, the KID3 scoring system may be a promising predictive tool for screening embryos with better developmental potential.