Reduction of Deuterium Level Supports Resistance of Neurons to Glucose Deprivation and Hypoxia: Study in Cultures of Neurons and on Animals

The effect of a reduced deuterium (D) content in the incubation medium on the survival of cultured neurons in vitro and under glucose deprivation was studied. In addition, we studied the effect of a decrease in the deuterium content in the rat brain on oxidative processes in the nervous tissue, its antioxidant protection, and training of rats in the T-shaped maze test under hypoxic conditions. For experiments with cultures of neurons, 7–8-day cultures of cerebellar neurons were used. Determination of the rate of neuronal death in cultures was carried out using propidium iodide. Acute hypoxia with hypercapnia was simulated in rats by placing them in sealed vessels with a capacity of 1 L. The effect on oxidative processes in brain tissues was assessed by changes in the level of free radical oxidation and malondialdehyde. The effect on the antioxidant system of the brain was assessed by the activity of catalase. The study in the T-maze was carried out in accordance with the generally accepted methodology, the skill of alternating right-sided and left-sided loops on positive reinforcement was developed. This work has shown that a decrease in the deuterium content in the incubation medium to a level of −357‰ has a neuroprotective effect, increasing the survival rate of cultured neurons under glucose deprivation. When exposed to hypoxia, a preliminary decrease in the deuterium content in the rat brain to −261‰ prevents the development of oxidative stress in their nervous tissue and preserves the learning ability of animals in the T-shaped maze test at the level of the control group. A similar protective effect during the modification of the 2H/1H internal environment of the body by the consumption of DDW can potentially be used for the prevention of pathological conditions associated with the development of oxidative stress with damage to the central nervous system.

P–190 Can pregnancy be predicted according to mean infra-red attenuated total reflectance (MIR-ATR) spectrometry of incubation medium of embryos in IVF?: A proof of concept

Study question Can MIR-ATR spectrometry of embryo incubation medium be used to predict embryo quality and IVF treatment results. Summary answer MIR-ATR spectrometry is able to distinguish between good and poor embryo quality and may improve the prediction of pregnancy based on better embryo selection. What is known already Infra Red (IR) spectra enable to determine if certain chemical functional components are present in a molecule. Attenuated total reflection (ATR) spectroscopy is a well established, simple and rapid technique for objective classification of biological samples. ATR spectrometry was investigated and confirmed to serve as an additional diagnostic tool in oral and gynecologic cancer. Currently, embryo quality is assessed based on morphokinetics parameters from Time-laps incubator and/or final embryo’s grading Study design, size, duration Culture media of 227 embryos on cleavage stage and blastocyst stage were collected and analysed between January 1st, 2018 and December 31st 2020. Participants/materials, setting, methods The incubation medium liquid of embryos cultured in Time-lapse incubator were collected after transferring the embryos to their final destination. Infra-Red (IR) absorbance spectra was measured. Nominal resolution was 4 cm–1, wavenumber range was from 600 cm–1 to 5000 cm–1 and each absorbance spectrum was normalized to the height of amide I band at ∼1650cm–1. Machine learning techniques utilized to build discrimination models for the absorbance data. Results were correlated with clinical and pregnancy results. Main results and the role of chance This preliminary study demonstrates that ATR spectrometry differs in media of : Day 3 embryos compared to day 5 embryos, top quality compared to poor quality embryos and implanting embryos comparing to non-implanting embryos. We found that MIR-ATR spectrometry might predict pregnancy in accuracy rates of 85%. MIR-ATR spectrometry of incubation medium can be used as an additional tool for selection of embryos for transfer. Limitations, reasons for caution Additional study and collection of larger number of culture Media is requested to validate MIR-ATR spectrometry as an additional tool in clinical set up in IVF units and laboratories. Wider implications of the findings: MIR-ATR spectrometry can be used in the future as an additional tool for selection of embryos and prediction of cycles outcomes. Trial registration number NCT03317418

Electrophysiological Activity and Survival Rate of Rats Nervous Tissue Cells Depends on D/H Isotopic Composition of Medium

The deuterium content modification in an organism has a neuroprotective effect during the hypoxia model, affecting anxiety, memory and stress resistance. The aim of this work was to elucidate the possible mechanisms of the medium D/H composition modification on nerve cells. We studied the effect of an incubation medium with a 50 ppm deuterium content compared to a medium with 150 ppm on: (1) the activity of Wistar rats’ hippocampus CA1 field neurons, (2) the level of cultured cerebellar neuron death during glucose deprivation and temperature stress, (3) mitochondrial membrane potential (MMP) and the generation of reactive oxygen species in cultures of cerebellar neurons. The results of the analysis showed that the incubation of hippocampal sections in a medium with a 50 ppm deuterium reduced the amplitude of the pop-spike. The restoration of neuron activity was observed when sections were returned to the incubation medium with a 150 ppm deuterium content. An environment with a 50 ppm deuterium did not significantly affect the level of reactive oxygen species in neuron cultures, while MMP decreased by 16–20%. In experiments with glucose deprivation and temperature stress, the medium with 50 ppm increased the death of neurons. Thus, a short exposure of nerve cells in the medium with 50 ppm deuterium acts as an additional stressful factor, which is possibly associated with the violation of the cell energy balance. The decrease in the mitochondrial membrane potential, which is known to be associated with ATP synthesis, indicates that this effect may be associated with the cell energy imbalance. The decrease in the activity of the CA1 field hippocampal neurons may reflect reversible adaptive changes in the operation of fast-reacting ion channels.

Mathematical model of transmembrane potential dynamics of loach early embryogenesis

Heavy metals in the water environment are known to have a negative effect on the viability of fish in early development. We have discussed the influence of environmental factors on early embryo development from the viewpoint of the correlation adaptometry method. The analysis of time series with the subsequent construction of a mathematical model was used to determine the change in the greatest effect of certain types of ions on the values of the transmembrane potential for prognostic purposes. The membrane potential is accepted as an integral indicator of the state of the embryos. Structures of five elements of the same type were constructed for the time shifts from 0 to 180 minutes. Each element in the system characterizes the value of the transmembrane potential that was measured in a cell incubated in one of the five solutions during early embryo development. Mathematical models describing the cell membrane potential dynamics have been created and studied. It was noted that the transmembrane potential dynamics of embryo cells is dependent on a change in the value of the correlation coefficient between elements of the system. A decrease in the sum of the correlations between individual elements of the system with an increase in the magnitude of the time shift is established. The results of the numerical solutions of the system equations indicated the sequence of changes in the greatest effect of the incubation medium on the value of the membrane potential in cells. The study of the membrane potentials’ dynamics, using the total values of the strength of correlation, confirmed the influence of heavy metals in the incubation medium on the membrane potential of embryo cell in early development.

Effect of sodium orthovanadate on ovulation isolated ovarian follicles in Siberian Sturgeon (Acipenser baerii) In Vitro

Effects of sodium orthovanadate on oocyte ovulation were examined during in vitro culture of Siberian sturgeon ovarian follicles from hibernating fish. It was shown that sodium orthovanadate stimulates ovulation of Siberian sturgeon oocytes in a dose-dependent manner. The stimulating or inhibitory effect of vanadate depends on the time of addition to the incubation medium. It was also shown that the stimulating effects of orthovanadate depend on the physiological status of hibernating females whose oocytes were isolated

The Release of a Soluble Glycosylated Protein from Glycogen by Recombinant Lysosomal α-Glucosidase (rhGAA) In Vitro and Its Presence in Serum In Vivo

In studies on the degradation of glycogen by rhGAA, a glycosylated protein core material was found which consists of about 5–6% of the total starting glycogen. There was an additional 25% of the glycogen unaccounted for based on glucose released. After incubation of glycogen with rhGAA until no more glucose was released, no other carbohydrate was detected on HPAEC-PAD. Several oligosaccharides are then detectable if the medium is first boiled in 0.1 N HCl or incubated with trypsin. It is present in serum either in an HCl extract or in a trypsin digest. The characteristics of the in vivo serum material are identical to the material in the in vitro incubation medium. One oligosaccharide cannot be further degraded by rhGAA, from the incubation medium as well as from serum co-elute on HPAEC-PAD. Several masked oligosaccharides in serum contain m-inositol, e-inositol, and sorbitol as the major carbohydrates. The presence of this glycosylated protein in serum is a fraction of glycogen that is degraded outside the lysosome and the cell. The glycosylated protein in the serum is not present in the serum of Pompe mice not on ERT, but it is present in the serum of Pompe disease patients who are on ERT, so it is a biomarker of GAA degradation of lysosomal glycogen.

Structural Diversity in Early-Stage Biofilm Formation on Microplastics Depends on Environmental Medium and Polymer Properties

Plastics entering the environment can not only undergo physical degradation and fragmentation processes, but they also tend to be colonized by microorganisms. Microbial colonization and the subsequent biofilm formation on plastics can alter their palatability to organisms and result in a higher ingestion as compared to pristine plastics. To date, the early stage of biofilm formation on plastic materials has not been investigated in context of the environmental medium and polymer properties. We explored the early-stage biofilm formation on polyamide (PA), polyethylene terephthalate (PET), and polyvinyl chloride (PVC) after incubation in freshwater and artificial seawater and categorized the structural diversity on images obtained via scanning electron microscopy. Furthermore, by the measurement of the initial ζ-potential of the plastic materials, we found that PA with the highest negative ζ-potential tended to have the highest structural diversity, followed by PET and PVC after incubation in freshwater. However, PVC with the lowest negative ζ-potential showed the highest structural diversity after incubation in seawater, indicating that the structural diversity is additionally dependent on the incubation medium. Our results give insights into how the incubation medium and polymer properties can influence the early-stage biofilm formation of just recently environmentally exposed microplastics. These differences are responsible for whether organisms may ingest microplastic particles with their food or not.

Transient coating of γ-Fe2O3 nanoparticles with glutamate for its delivery to and removal from brain nerve terminals

Glutamate is the main excitatory neurotransmitter in the central nervous system and excessive extracellular glutamate concentration is a characteristic feature of stroke, brain trauma, and epilepsy. Also, glutamate is a potential tumor growth factor. Using radiolabeled ʟ-[14C]glutamate and magnetic fields, we developed an approach for monitoring the biomolecular coating (biocoating) with glutamate of the surface of maghemite (γ-Fe2O3) nanoparticles. The nanoparticles decreased the initial rate of ʟ-[14C]glutamate uptake, and increased the ambient level of ʟ-[14C]glutamate in isolated cortex nerve terminals (synaptosomes). The nanoparticles exhibit a high capability to adsorb glutamate/ʟ-[14C]glutamate in water. Some components of the incubation medium of nerve terminals, that is, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) and NaH2PO4, decreased the ability of γ-Fe2O3 nanoparticles to form a glutamate biocoating by about 50% and 90%, respectively. Only 15% of the amount of glutamate biocoating obtained in water was obtained in blood plasma. Albumin did not prevent the formation of a glutamate biocoating. It was shown that the glutamate biocoating is a temporal dynamic structure at the surface of γ-Fe2O3 nanoparticles. Also, components of the nerve terminal incubation medium and physiological fluids responsible for the desorption of glutamate were identified. Glutamate-coated γ-Fe2O3 nanoparticles can be used for glutamate delivery to the nervous system or for glutamate adsorption (but with lower effectiveness) in stroke, brain trauma, epilepsy, and cancer treatment following by its subsequent removal using a magnetic field. γ-Fe2O3 nanoparticles with transient glutamate biocoating can be useful for multifunctional theranostics.