Haplotype-aware inference of human chromosome abnormalities.

Extra or missing chromosomes-a phenomenon termed aneuploidy-frequently arise during human meiosis and embryonic mitosis and are the leading cause of pregnancy loss, including in the context of in vitro fertilization (IVF). While meiotic aneuploidies affect all cells and are deleterious, mitotic errors generate mosaicism, which may be compatible with healthy live birth. Large-scale abnormalities such as triploidy and haploidy also contribute to adverse pregnancy outcomes, but remain hidden from standard sequencing-based approaches to preimplantation genetic testing for aneuploidy (PGT-A). The ability to reliably distinguish meiotic and mitotic aneuploidies, as well as abnormalities in genome-wide ploidy, may thus prove valuable for enhancing IVF outcomes. Here, we describe a statistical method for distinguishing these forms of aneuploidy based on analysis of low-coverage whole-genome sequencing data, which is the current standard in the field. Our approach overcomes the sparse nature of the data by leveraging allele frequencies and linkage disequilibrium (LD) measured in a population reference panel. The method, which we term LD-informed PGT-A (LD-PGTA), retains high accuracy down to coverage as low as 0.05 × and at higher coverage can also distinguish between meiosis I and meiosis II errors based on signatures spanning the centromeres. LD-PGTA provides fundamental insight into the origins of human chromosome abnormalities, as well as a practical tool with the potential to improve genetic testing during IVF.

Two clinical case reports of embryonic mosaicism identified with PGT-A persisting during pregnancy as true fetal mosaicism.

The health risks associated with transferring embryos classified as mosaic by preimplantation genetic testing for aneuploidies (PGT-A) are currently unknown. Such embryos produce PGT-A results indicating the presence of both euploid and aneuploid cells and have historically been deselected from transfer and grouped with uniformly aneuploid embryos as 'abnormal'. In recent years, numerous groups have reported the intentional transfer of mosaic embryos in the absence of uniformly euploid embryos, largely observing births of seemingly healthy babies. However, it remains to be understood whether the embryonic mosaicism invariably becomes resolved during the ensuing pregnancy, or whether the placenta and/or fetal tissues retain aneuploid cells, and if so to what potential clinical effect. Here, we report two cases of mosaicism persisting from the embryonic stage to the established pregnancy. Case 1 involved an embryonic low-level segmental mosaic loss in Chromosome (Chr) 1, which was confirmed in amniocentesis as well as in brain tissue of the products of conception. This pregnancy was terminated due to the chromosomal pathologies associated with 1p36 deletion syndrome, such as severe intellectual disability. Case 2 involved a low-level mosaic Chr 21 trisomy, which was confirmed with chorionic villus sampling and amniocentesis. The ensuing pregnancy was terminated after ultrasound identification of severe abnormalities in the placenta and fetus. Together, these two cases should be taken into account for risk-benefit assessments of prospective mosaic embryo transfers.

Effects Induced by a Weak Static Magnetic Field of Different Intensities on HT-1080 Fibrosarcoma Cells.

In this study, we investigated the effects of weak static magnetic fields (SMFs) on HT-1080 human fibrosarcoma cells. Exposures to SMFs for four consecutive days were varied from 0.5 to 600 µT for treated units, while exposures to control units were held at 45 µT. Growth rates were measured by comparing cell counts, whereas membrane potentials, mitochondrial calcium, mitochondrial superoxide (O2 - ), nitric oxide (NO), hydrogen peroxide (H2 O2 ), intercellular pH, and oxidative stress were measured by using fluorescent dyes. The relative cell growth rates vary with the angle of the SMFs. Increases in the magnitude of the SMFs increased concentrations of mitochondrial calcium and membrane potential and decreased intracellular pH. H2 O2 , an important reactive oxygen species (ROS), increases at 100 and 200 µT, decreases at 300 and 400 µT and increases again at 500 and 600 µT. Overall, oxidative stress increases slightly with increasing SMFs, while superoxide and NO concentrations decrease. These results indicate that weak SMFs can accelerate and inhibit cell growth rates and induce alterations in ROS. Changes in ROS and oxidative stress are important for various cell functions. Calcium influx into mitochondria was one of the initial steps into the corresponding changes. Bioelectromagnetics. 2021. © 2021 Bioelectromagnetics Society.

Pilot study on the therapeutic potential of radiofrequency magnetic fields: growth inhibition of implanted tumours in mice.

The present study investigated possible therapeutic effects of radiofrequency or hypomagnetic fields on the growth rate of two types of implanted tumours. To this end, mice with implanted fibrosarcoma and pancreatic tumours were exposed continuously to a 2 µT, 10 MHz radiofrequency magnetic field (MF) perpendicular to a 45 µT static MF or to a hypomagnetic (~0.4-1 µT) field. The reasoning for a 10 MHz treatment was based on a current theoretical explanation for MF effects, which predicts a resonance phenomenon in this frequency range. Radiofrequency MFs reduced consistently the growth rate of two implanted tumour types (by ~30% in both cases). Also, hypomagnetic field hindered tumour growth in both tumour types, but the observation was not statistically significant with fibrosarcoma tumours. In conclusion, although experiments included a limited number of animals, the results indicate that MFs may offer a novel therapeutic strategy in the treatment of cancer.

Setting Guidelines for Electromagnetic Exposures and Research Needs.

Current limits for exposures to nonionizing electromagnetic fields (EMF) are set, based on relatively short-term exposures. Long-term exposures to weak EMF are not addressed in the current guidelines. Nevertheless, a large and growing amount of evidence indicates that long-term exposure to weak fields can affect biological systems and might have effects on human health. If they do, the public health issues could be important because of the very large fraction of the population worldwide that is exposed. We also discuss research that needs to be done to clarify questions about the effects of weak fields. In addition to the current short-term exposure guidelines, we propose an approach to how weak field exposure guidelines for long-term exposures might be set, in which the responsibility for limiting exposure is divided between the manufacturer, system operator, and individual being exposed. Bioelectromagnetics. © 2020 Bioelectromagnetics Society.

Possible Mechanism for Synchronized Detection of Weak Magnetic Fields by Nerve Cells.

We propose that biological systems may detect static and slowly varying magnetic fields by the modification of the timing of firing of adjacent nerve cells through the local influence of the magnetic field generated by current from one cell's firing on its nearest neighbors. The time delay of an adjacent nerve cell pulse with respect to the initial clock nerve cell pulse could serve as a signal for sensing the magnitude and direction of the magnetic field in a direction perpendicular to the current flows in the cells. It has been shown that changes in static magnetic fields modify concentrations of reactive oxygen species, calcium, pH, the growth rates of fibrosarcoma cells, and membrane potentials. These are linked to changes in membrane potentials that can either inhibit or accelerate the firing rate of pacemaker or clock cells. This mechanism may have applications to animals' use of magnetic fields for navigation or other purposes, possibly in conjunction with other mechanisms. Bioelectromagnetics. © 2020 Bioelectromagnetics Society.

Assessment of aneuploidy concordance between clinical trophectoderm biopsy and blastocyst.

STUDY QUESTION: Is a clinical trophectoderm (TE) biopsy a suitable predictor of chromosomal aneuploidy in blastocysts? SUMMARY ANSWER: In the analyzed group of blastocysts, a clinical TE biopsy was an excellent representative of blastocyst karyotype in cases of whole chromosome aneuploidy, but in cases of only segmental (sub-chromosomal) aneuploidy, a TE biopsy was a poor representative of blastocyst karyotype. WHAT IS KNOWN ALREADY: Due to the phenomenon of chromosomal mosaicism, concern has been expressed about the possibility of discarding blastocysts classified as aneuploid by preimplantation genetic testing for aneuploidy (PGT-A) that in fact contain a euploid inner cell mass (ICM). Previously published studies investigating karyotype concordance between TE and ICM have examined small sample sizes and/or have utilized chromosomal analysis technologies superseded by Next Generation Sequencing (NGS). It is also known that blastocysts classified as mosaic by PGT-A can result in healthy births. TE re-biopsy of embryos classified as aneuploid can potentially uncover new instances of mosaicism, but the frequency of such blastocysts is currently unknown. STUDY DESIGN, SIZE, DURATION: For this study, 45 patients donated 100 blastocysts classified as uniform aneuploids (non-mosaic) using PGT-A by NGS (n = 93 whole chromosome aneuploids, n = 7 segmental aneuploids). In addition to the original clinical TE biopsy used for PGT-A, each blastocyst was subjected to an ICM biopsy as well as a second TE biopsy. All biopsies were processed for chromosomal analysis by NGS, and karyotypes were compared to the original TE biopsy. PARTICIPANTS/MATERIALS, SETTING, METHODS: The setting for this study was a single IVF center with an in-house PGT-A program and associated research laboratory. MAIN RESULTS AND THE ROLE OF CHANCE: When one or more whole chromosomes were aneuploid in the clinical TE biopsy, the corresponding ICM was aneuploid in 90 out of 93 blastocysts (96.8%). When the clinical TE biopsy contained only segmental (sub-chromosomal) aneuploidies, the ICM was aneuploid in three out of seven cases (42.9%). Blastocysts showing aneuploidy concordance between clinical TE biopsy and ICM were also aneuploid in a second TE biopsy in 86 out of 88 cases (97.7%). In blastocysts displaying clinical TE-ICM discordance, a second TE biopsy was aneuploid in only two out of six cases (33.3%). LIMITATIONS, REASONS FOR CAUTION: All embryos in this study had an initial classification of 'aneuploid' and not 'euploid' or 'mosaic'. Therefore, the findings of this study refer specifically to a TE biopsy predicting aneuploidy in the remaining blastocyst, and cannot be extrapolated to deduce the ability of a TE biopsy to predict euploidy in the blastocyst. No conclusions should be drawn from this study about the ability of a mosaic TE biopsy to predict the karyotype of the corresponding blastocyst. Caution should be exercised in generalizing the findings of the sample group of this study to the general IVF blastocyst population. The segmental aneuploidy group only contained seven samples. WIDER IMPLICATIONS OF THE FINDINGS: The high rate of intra-blastocyst concordance observed in this study concerning whole chromosome aneuploidy contributes experimental evidence to the validation of PGT-A at the blastocyst stage. Concomitantly, the results suggest potential clinical value in reassessing blastocysts deemed aneuploid by TE re-biopsy in select cases, particularly in instances of segmental aneuploidies. This could impact infertility treatment for patients who only have blastocysts classified as aneuploid by PGT-A available. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the Zouves Foundation for Reproductive Medicine and Zouves Fertility Center. The authors have no competing interest to disclose. TRIAL REGISTRATION NUMBER: Not applicable.

Births from embryos with highly elevated levels of mitochondrial DNA.

RESEARCH QUESTION: Conflicting data exist on the utility of quantification of mitochondrial DNA (mtDNA) levels as a predictor of blastocyst implantation in the IVF clinic. The current study determined whether blastocysts with highly elevated mtDNA levels could result in healthy pregnancies and births, and whether mitochondrial functional output might be a readout of cell stress in the embryo. DESIGN: mtDNA levels were determined in 109 blastocysts used in clinical transfers into 100 women, noting their clinical outcomes. In a separate set of embryos, mitochondrial function was quantified in a model of embryo stress, aneuploidy. Measurement of mtDNA levels made use of surplus material from the process of preimplantation genetic testing for aneuploidies, and followed recently proposed unifying guidelines for mtDNA quantification. RESULTS: Unusually high mtDNA levels did not preclude blastocyst implantation and healthy births. An analysis of 109 blastocysts showed no significant difference between mtDNA levels in implanted (n = 55) versus non-implanted (n = 54) blastocysts. No obvious differences in the degree of mitochondrial functional output were detected in a model of embryo stress. CONCLUSIONS: Measurement of mtDNA copy number might not provide any advantage in embryo prioritization and could lead to a deselection of blastocysts that would result in healthy pregnancies and births. Furthermore, the quantification of mitochondrial functional output in a model of cellular stress might suggest that mitochondria are not clear targets for biomarker identification as it relates to blastocyst viability. Any suggested link between mtDNA levels, mitochondria or their output with blastocyst transfer outcome requires further validation.

Shock Tube/Laser Absorption and Kinetic Modeling Study of Triethyl Phosphate Combustion.

Pyrolysis and oxidation of triethyl phosphate (TEP) were performed in the reflected shock region at temperatures of 1462-1673 K and 1213-1508 K, respectively, and at pressures near 1.3 atm. CO concentration time histories during the experiments were measured using laser absorption spectroscopy at 4580.4 nm. Experimental CO yields were compared with model predictions using the detailed organophosphorus compounds (OPC) incineration mechanism from the Lawrence Livermore National Lab (LLNL). The mechanism significantly underpredicts CO yield in TEP pyrolysis. During TEP oxidation, predicted rate of CO formation was significantly slower than the experimental results. Therefore, a new improved kinetic model for TEP combustion was developed, which was built upon the AramcoMech2.0 mechanism for C0-C2 chemistry and the existing LLNL submechanism for phosphorus chemistry. Thermochemical data of 40 phosphorus (P)-containing species were reevaluated, either using recently published group values for P-containing species or by quantum chemical calculations (CBS-QB3). The new improved model is in better agreement with the experimental CO time histories within the temperature and pressure conditions tested in this study. Sensitivity analysis was used to identify important reactions affecting CO formation, and future experimental/theoretical studies on kinetic parameters of these reactions were suggested to further improve the model. To the best of our knowledge, this is the first study of TEP kinetics in a shock tube under these conditions and the first time-resolved laser-based species time history data during its pyrolysis and oxidation.

Role of radical pairs and feedback in weak radio frequency field effects on biological systems.

Radio frequency electromagnetic fields (RF) have been shown to modify the concentrations of the radical O2-, H2O2 and cancer cell growth rates at exposure levels below those that cause significant heating. Reactive oxygen species (ROS) are both signaling molecules and species that can do damage, depending on timing, location and concentrations. We briefly look at some mechanisms by which electromagnetic fields can modify the concentrations of ROS and some of the feedback and repair processes that lead to variable biological effects. Of particular interest are the role of radical pairs and their spins, which have received considerable attention recently, and the role of feedback in biological systems, to which less attention has been paid.

Is mitochondrial DNA quantitation in blastocyst trophectoderm cells predictive of developmental competence and outcome in clinical IVF?

Behind every successful IVF embryo transfer, there is a great game of chance. Methods seeking to tilt the balance and increase the likelihood of implantation have been proposed and implemented with varying results, including embryo morphology, FISH-PGS, comprehensive chromosomal screening (CCS), morphokinetics, endometrial receptivity testing. It has been suggested that mitochondrial DNA (mtDNA) copy number could serve as a biomarker for embryo viability, but this concept was recently challenged. The world of IVF is left with unanswered questions: Why are there discrepancies in the reports? Should mtDNA copy number be considered to rank embryos for transfer? And in a broader sense, how well must a technique be validated before its implementation in the IVF clinic? Here, we explore these questions attempting to piece together the published data and suggest future directions to help unravel the subject matter.

The effects of weak magnetic fields on radical pairs.

It is proposed that radical concentrations can be modified by combinations of weak, steady and alternating magnetic fields that modify the population distribution of the nuclear and electronic spin state, the energy levels and the alignment of the magnetic moments of the components of the radical pairs. In low external magnetic fields, the electronic and nuclear angular momentum vectors are coupled by internal forces that outweigh the external fields' interactions and are characterized in the Hamiltonian by the total quantum number F. Radical pairs form with their unpaired electrons in singlet (S) or triplet (T) states with respect to each other. At frequencies corresponding to the energy separation between the various states in the external magnetic fields, transitions can occur that change the populations of both electron and nuclear states. In addition, the coupling between the nuclei, nuclei and electrons, and Zeeman shifts in the electron and nuclear energy levels can lead to transitions with resonances spanning frequencies from a few Hertz into the megahertz region. For nuclear energy levels with narrow absorption line widths, this can lead to amplitude and frequency windows. Changes in the pair recombination rates can change radical concentrations and modify biological processes. The overall conclusion is that the application of magnetic fields at frequencies ranging from a few Hertz to microwaves at the absorption frequencies observed in electron and nuclear resonance spectroscopy for radicals can lead to changes in free radical concentrations and have the potential to lead to biologically significant changes.

Inhibition of cellular proliferation and enhancement of hydrogen peroxide production in fibrosarcoma cell line by weak radio frequency magnetic fields.

This study presents experimental data for the effects of weak radio frequency (RF) magnetic fields on hydrogen peroxide (H2O2) production and cellular growth rates of fibrosarcoma HT1080 cells in vitro. Cells were exposed either to 45 µT static magnetic fields (SMFs)-oriented vertical to the plane of growth or to SMFs combined with weak 5 and 10 MHz RF magnetic fields of 10 µTRMS intensity perpendicular to the static field. Cell numbers were reduced up to 30% on Day 2 for the cells exposed to the combination of SMF and a 10 MHz RF magnetic field compared with the SMF control cells. In addition, cells exposed to 10 MHz RF magnetic fields for 8 h increased H2O2 production by 55%. The results demonstrate an overall magnetic field-induced biological effect that shows elevated H2O2 levels with accompanying decrease in cellular growth rates.

Inhomogeneous background magnetic field in biological incubators is a potential confounder for experimental variability and reproducibility.

This report shows that the background magnetic field in biological incubators can vary by orders of magnitude within and between incubators. These variations can be observed within the same incubator in locations that are centimeters apart from each other as well as between incubators that are identical and located in the same laboratory. Additionally, the values measured were frequently outside the range of magnitudes found naturally on the Earth's surface or ordinary habitation spaces. Exposure to such altered magnetic field environments has been experimentally shown to be sufficient to cause numerous effects in cell cultures. Examples of the effects reported span from differential generation of free radicals and heat shock proteins to differences in cellular proliferation, differentiation, and death. Although the effects are not well established and the molecular mechanism of action is currently under debate, these observations alone support the notion that the inhomogeneity of the background magnetic field in incubators is a potential confounding source of the variability and reproducibility for studies performed on cell cultures. In this regard, it is recommended that special measures be adopted to control the background magnetic fields in incubators when investigating the biological effects of exposure to magnetic fields of comparable characteristics as the ones measured in this study, or when studying small biological effects in general.

Impact of weak radiofrequency and static magnetic fields on key signaling molecules, intracellular pH, membrane potential, and cell growth in HT-1080 fibrosarcoma cells.

There are substantial concerns that extended exposures to weak radiofrequency (RF) fields can lead to adverse health effects. In this study, HT-1080 fibrosarcoma cells were simultaneously exposed to a static magnetic flux density between 10 [Formula: see text] and 300 [Formula: see text] and RF magnetic fields with amplitudes ranging from 1 nT to 1.5 µT in the frequency range from 1.8 to 7.2 MHz for four days. Cell growth rates, intracellular pH, hydrogen peroxide, peroxynitrite, membrane potential and mitochondrial calcium were measured. Results were dependent on carrier frequency and the magnitude of the RF magnetic field, modulation frequencies and the background static magnetic field (SMF). Iron sulphur (Fe-S) clusters are essential for the generation of reactive oxygen species and reactive nitrogen species (ROS and RNS). We believe the observed changes are associated with hyperfine couplings between the chemically active electrons and nuclear spins. Controlling external magnetic fields may have important clinical implications on aging, cancer, arthritis, and Alzheimer's.

Weak Radiofrequency Field Effects on Chemical Parameters That Characterize Oxidative Stress in Human Fibrosarcoma and Fibroblast Cells.

In the last few decades, evidence has surfaced that weak radiofrequency (RF) fields can influence biological systems. This work aims to improve our understanding of how externally applied weak RF fields alter concentrations of chemical parameters that characterize oxidative stress. We conducted a series of experiments to investigate the effects of applying weak RF magnetic fields within the 3-5 MHz region on mitochondrial respiration in both human fibrosarcoma and fibroblast cells over a period of four days. Our experimental data show that RF fields between 3 and 5 MHz were able to change the modulation of mitochondrial signaling by changing the cell growth, mitochondrial mass, and oxidative stress. Exposure to RF fields at 4.2 MHz significantly increased the mitochondrial mass and oxidative stress in fibrosarcoma cells. There are substantial concerns that extended exposure to weak RF fields can lead to health effects. The ability to control these parameters by external magnetic fields may have important clinical implications.

Chromosomal, gestational, and neonatal outcomes of embryos classified as a mosaic by preimplantation genetic testing for aneuploidy.

OBJECTIVE: To understand the clinical risks associated with the transfer of embryos classified as a mosaic using preimplantation genetic testing for aneuploidy. DESIGN: Analysis of data collected between 2017 and 2023. SETTING: Multicenter. PATIENTS: Patients of infertility treatment. INTERVENTION: Comparison of pregnancies resulting from embryos classified as euploid or mosaic using the 20%-80% interval in chromosomal intermediate copy numbers to define a mosaic result. MAIN OUTCOME MEASURES: Rates of spontaneous abortion, birth weight, length of gestation, incidence of birth defects, and chromosomal status during gestation. RESULTS: Implanted euploid embryos had a significantly lower risk of spontaneous abortion compared with mosaic embryos (8.9% [n = 8,672; 95% confidence interval {CI95} 8.3, 9.5] vs. 22.2% [n = 914; CI95 19.6, 25.0]). Embryos with mosaicism affecting whole chromosomes (not segmental) had the highest risk of spontaneous abortion (27.6% [n = 395; CI95 23.2, 32.3]). Infants born from euploid, mosaic, and whole chromosome mosaic embryos had average birth weights and lengths of gestation that were not statistically different (3,118 g and 267 days [n = 488; CI95 3,067, 3,169, and 266, 268], 3052 g and 265 days [n = 488; CI95 2,993, 3,112, and 264,267], 3,159 g and 268 days [n = 194; CI95 3,070, 3,249, and 266,270], respectively). Out of 488 infants from mosaic embryo transfers (ETs), one had overt gross abnormalities as defined by the Centers for Disease Control and Prevention. Most prenatal tests performed on pregnancies from mosaic ETs had normal results, and only three pregnancies produced prenatal test results reflecting the mosaicism detected at the embryonic stage (3 out of 250, 1.2%; CI95 0.25, 3.5). CONCLUSION: Although embryos classified as mosaic experience higher rates of miscarriage than euploid embryos (with a particularly high frequency shortly after implantation), infants born of mosaic ETs are similar to infants of euploid ETs. Prenatal testing indicates that mosaicism resolves during most pregnancies, although this process is not perfectly efficient. In a small percentage of cases, the mosaicism persists through gestation. These findings can serve as risk-benefit considerations for mosaic ETs in the fertility clinic.