Effect of sperm DNA fragmentation on blastocyst formation rate and subsequent IVF outcome

Background: Male infertility associated with sperm DNA alteration has raised a new issue in assisted reproduction techniques (ARTs).Methods: It was a retrospective analytical study on 250 cases of routine IVF/ICSI performed at Swagat ART Centre from January 2017 to January 2020. We divided the patient according to the sperm DNA fragmentation index (DFI) as normal DFI≤15%, n=95, a moderate DFI≤30%, n=89, and a high DFI group >30%, n=66. Oocytes of each patient were almost equally divided and fertilization method was adopted as half IVF half ICSI or only ICSI in poor quality (oligo, astheno, teratozoospermia or with two or all three defect and compared the fertilization, cleavage, embryo formation, blastocyst formation, pregnancy and early embryo formation rate among these six groups. Results: Fertilization, cleavage, embryo formation, and clinical pregnancy rates were reported as higher in ≤15% DFI group of both IVF and ICSI-ET (87.3±26.2, 77.7±26.1, 68.2±28.8, 50.8 in IVF and 78.3±17.8, 70.3±31.2, 67.2±28.8, 57.6 respectively). Significant differences (p<0.01) are observed among all six groups. Higher abortion rate is observed in high DFI group of both IVF and ICSI.Conclusions: High sperm DFI causes low blastocyst formation and pregnancy outcome. Higher abortion rate observed in high DFI group indicated need of further study.

Analysis of maturation dynamics and developmental competence of in vitro matured oocytes under time-lapse monitoring

Background To improve the developmental competence of in vitro cultured oocytes, extensive literature focused on maturation rate improvement with different additives in culture medium, while studies investigating the maturation dynamics of oocytes during in vitro maturation (IVM) and the influencing factors on oocyte viability are scarce. Methods The study involved a retrospective observation by time-lapse monitoring of the IVM process of 157 donated GV oocytes from 59 infertile couples receiving ICSI in 2019, in Tongji Hospital, Wuhan, China. The GV oocytes derived from controlled ovarian hyperstimulation (COH) cycles underwent rescue IVM (R-IVM), and the maturation dynamics, including GVBD time (GV-MI), time from GVBD to maturation (MI-MII), maturation time (GV-MII), and MII arrest duration (MII-ICSI), were recorded by time-lapse monitoring. The matured oocytes were inseminated at different MII arrest points and subsequent embryo developments were assessed. The effects of baseline clinical characteristics, oocyte diameters, and maturation dynamics on the developmental competence of the oocytes were also analyzed. Results Totally, 157 GV oocytes were collected. GVBD happened in 111 oocytes, with a median GV-MI duration of 3.7 h. The median MI-MII duration was 15.6 h and the median GV-MII duration was 19.5 h. The maturation rate reached 56.7% at 24 h and 66.9% at 48 h, and the clinical factors, including patient age, FSH level, AMH level, ovarian stimulation protocol, and serum estradiol and progesterone levels on hCG trigger day, showed no effects on the 24-h maturation rate. The normal fertilization rate of oocytes resuming meiosis within 8 h and matured within 24 h was significantly higher than that of oocytes resuming meiosis after 8 h and matured after 24 h. Furthermore, among those oocytes matured within 24 h, the high-quality embryo formation rate of oocytes resuming meiosis within 4.5 h and matured within 19 h was significantly higher. All stated time was measured from the start point of IVM. Additionally, for oocytes from patients with serum progesterone levels less than 1 ng/ml on hCG trigger day, the high-quality embryo formation rate was significantly increased. Conclusion R-IVM technology could increase the available embryos for patients in routine COH cycles, but excessive culture beyond 24 h is not recommended. GV-MI duration of the oocyte, recorded by time-lapse system, and serum progesterone levels of patients on hCG trigger day can significantly affect the developmental potential of the IVM oocytes.

Melatonin and Myo-Inositol: Supporting Reproduction from the Oocyte to Birth

Human pregnancy is a sequence of events finely tuned by several molecular interactions that come with a new birth. The precise interlocking of these events affecting the reproductive system guarantees safe embryo formation and fetal development. In this scenario, melatonin and myo-inositol seem to be pivotal not only in the physiology of the reproduction process, but also in the promotion of positive gestational outcomes. Evidence demonstrates that melatonin, beyond the role of circadian rhythm management, is a key controller of human reproductive functions. Similarly, as the most representative member of the inositol’s family, myo-inositol is essential in ensuring correct advancing of reproductive cellular events. The molecular crosstalk mediated by these two species is directly regulated by their availability in the human body. To date, biological implications of unbalanced amounts of melatonin and myo-inositol in each pregnancy step are growing the idea that these molecules actively contribute to reduce negative outcomes and improve the fertilization rate. Clinical data suggest that melatonin and myo-inositol may constitute an optimal dietary supplementation to sustain safe human gestation and a new potential way to prevent pregnancy-associated pathologies.

Overexpression of Douglas-Fir LEAFY COTYLEDON1 (PmLEC1) in Arabidopsis Induces Embryonic Programs and Embryo-Like Structures in the lec1-1 Mutant but Not in Wild Type Plants

Somatic embryogenesis (SE) is the most promising method for the quick propagation of desirable plant genotypes. However, application of SE to conifers remains challenging due to our limited knowledge about the genes involved in embryogenesis and the processes that lead to somatic embryo formation. Douglas-fir, an economically important lumber species, possesses a homolog of the angiosperm embryo-regulatory LEC1 gene. In the present study, we analyzed the potential of Douglas-fir PmLEC1 to induce embryonic programs in the vegetative cells of a heterologous host, Arabidopsis thaliana. PmLEC1 complemented the Arabidopsis lec1-1 null mutant and led to a variety of phenotypes ranging from normal morphology to developmental arrest at various stages in the T1 generation. PmLEC1 did not affect the morphology of wild type Arabidopsis T1 plants. More profound results occurred in T2 generations. PmLEC1 expression induced formation of recurrent somatic embryo-like structures in vegetative tissues of the rescued lec1-1 mutant but loss of apical dominance (bushy phenotype) in wild type plants. The activation of embryonic programs in the lec1-1PmLEC1 T2 plants was confirmed by the presence of the embryo-specific transcripts, OLEOSIN and CRUCIFERIN. In contrast, no embryo-like structures, and no OLEOSIN or CRUCIFERIN were observed in PmLEC1-expressing bushy wild type T2 plants.

Blastocyst formation, embryo transfer and breed comparison in the first reported large scale cloning of camels

Cloning, through somatic cell nuclear transfer (SCNT), has the potential for a large expansion of genetically favorable traits in a population in a relatively short term. In the present study we aimed to produce multiple cloned camels from racing, show and dairy exemplars. We compared several parameters including oocyte source, donor cell and breed differences, transfer methods, embryo formation and pregnancy rates and maintenance following SCNT. We successfully achieved 47 pregnancies, 28 births and 19 cloned offspring who are at present healthy and have developed normally. Here we report cloned camels from surgical embryo transfer and correlate blastocyst formation rates with the ability to achieve pregnancies. We found no difference in the parameters affecting production of clones by camel breed, and show clear differences on oocyte source in cloning outcomes. Taken together we demonstrate that large scale cloning of camels is possible and that further improvements can be achieved.

The Impact of Endometriosis on Embryo Quality in in-vitro Fertilization/Intracytoplasmic Sperm Injection: A Systematic Review and Meta-Analysis

Background: The association between endometriosis and embryological outcomes remains uncertain. The meta-analysis aimed to evaluate the impact of endometriosis on embryo quality.Methods: A systematic review and meta-analysis was conducted to investigate the association between the endometriosis and embryo quality. Searches were performed on the three electronic databases: PubMed, EMBASE, and Web of Science. The detailed characteristics and data of the included studies were extracted. The risk ratio with 95% confidence intervals were calculated using the random and fixed effects model. The main outcome measures were high-quality embryo rate, cleavage rate, and embryo formation rate.Results: A total of 22 studies included were analyzed. Compared with the control group, women with endometriosis had a similar high-quality embryo rate (RR = 1.00; 95% CI, 0.94–1.06), a comparable cleavage rate (RR = 1.00; 95% CI, 0.97–1.02), and a similar embryo formation rate (RR = 1.10; 95% CI, 0.97–1.24). In women with stage III-IV endometriosis, there was no statistically significantly difference in high-quality embryo rate (RR = 1.02; 95% CI, 0.94–1.10), cleavage rate (RR = 1.00; 95% CI, 0.98–1.02), and embryo formation rate (RR = 1.05; 95% CI, 0.97–1.14), compared with those without endometriosis. For women with unilateral endometrioma, pooling of results from the affected ovaries did not show a statistically significantly difference in high-quality embryo rate (RR = 0.99; 95% CI, 0.60–1.63) in comparison to the normal contralateral ovaries.Conclusions: Our results seem to indicate that endometriosis does not compromise embryo quality from the perspective of morphology.

Studying the Bulk and Contour Ice Nucleation of Water Droplets via Quartz Crystal Microbalances

Due to the stochastic and time-dependent character of the ice embryo formation and growth (i.e., a process that can be analyzed statistically, but cannot be predicted precisely), the heterogeneous ice nucleation on atmospheric aerosols or macroscopic solid surfaces is still shrouded in mystery, regardless of the extremely active research and exponential progress within this scientific field. For instance, whether the icing appears from outside-in or inside-out is a subject of intense controversy, with practicability in designing passive icephobic coatings or improving the effectiveness of the cryopreservation technologies. Here, we propose an artful technique for quantitative analysis of the different modes of water freezing using super-nonwettable soot-coated quartz crystal microbalances (QCMs). To achieve this goal, a set of 5 MHz QCMs are loaded one at a time with a 50 μL droplet, whose bulk or contour solidification is detected in real-time. The obtained experimental results show that our sensor devices recognize explicitly if the ice nuclei form predominantly at the liquid–solid interface or spread along the droplet’s entire outer shell by triggering individual reproducible responses in terms of the direction of signal evolution in time. Our results may serve as a foundation for the future incorporation of QCM devices in different freezing assays, where gaining information about the ice adhesion forces and ice layer’s thickness is mandatory.

Depressurization-Induced Nucleation in the “Polylactide-Carbon Dioxide” System: Self-Similarity of the Bubble Embryos Expansion

Self-similar expansion of bubble embryos in a plasticized polymer under quasi-isothermal depressurization is examined using the experimental data on expansion rates of embryos in the CO2-plasticized d,l-polylactide and modeling the results. The CO2 initial pressure varied from 5 to 14 MPa, and the depressurization rate was 5 × 10−3 MPa/s. The constant temperature in experiments was in a range from 310 to 338 K. The initial rate of embryos expansion varied from ≈0.1 to ≈10 µm/s, with a decrease in the current external pressure. While modeling, a non-linear behavior of CO2 isotherms near the critical point was taken into account. The modeled data agree satisfactorily with the experimental results. The effect of a remarkable increase in the expansion rate at a decreasing external pressure is interpreted in terms of competing effects, including a decrease in the internal pressure, an increase in the polymer viscosity, and an increase in the embryo radius at the time of embryo formation. The vanishing probability of finding the steadily expanding embryos for external pressures around the CO2 critical pressure is interpreted in terms of a joint influence of the quasi-adiabatic cooling and high compressibility of CO2 in the embryos.

In vitro Regeneration of Clematis Plants in the Nikita Botanical Garden via Somatic Embryogenesis and Organogenesis

The effects of growth regulators, namely, 6-benzylaminopurine (BAP) and thidiazuron (TDZ), on the morphogenic capacity of 13 cultivars of clematis plants, in terms of their morphological structure formation, shoot regeneration, and somatic embryo development, are presented. The clematis cultivars ‘Alpinist,’ ‘Ay-Nor,’ ‘Bal Tsvetov,’ ‘Crimson Star,’ ‘Crystal Fountain,’ ‘Kosmicheskaya Melodiya,’ ‘Lesnaya Opera,’ ‘Madame Julia Correvon,’ ‘Nevesta,’ ‘Nikitsky Rosovyi,’ ‘Nikolay Rubtsov,’ ‘Serenada Kryma,’ and ‘Vechniy Zov’ were taken in collection plots of the Nikita Botanical Gardens for use in study. After explant sterilization with 70% ethanol (1 min), 0.3–0.4% Cl2 (15 min), and 1% thimerosal (10 min), 1-cm long segments with a single node were introduced to an in vitro culture. The explants were established on the basal MS medium supplemented with BAP (2.20–8.90 μM) and 0.049 μM NAA, or TDZ (3.0; 6.0, and 9.0 μM) with 30 g/L sucrose and 9 g/L agar. The medium with 0.89 μM BAP served as the control. Culture vessels and test tubes with the explants were maintained in plant growth chamber-controlled conditions: with a 16-h photoperiod, under cool-white light fluorescent lamps with a light intensity of 37.5 μmol m–2 s–1, at a temperature of 24 ± 1°C. Histological analysis demonstrated that adventitious bud and somatic embryo formation in studied clematis cultivars occurred at numerous areas of active meristematic cell zones. The main role of plant growth regulators and its concentrations were demonstrated. It was determined that maximum adventitious microshoot regeneration without any morphological abnormalities formed on the media supplemented with BAP or TDZ. 4.40 μM BAP, or 6.0 μM TDZ were optimal cytokinin concentrations for micropropagation. The explants of ‘Alpinist,’ ‘Ay-Nor,’ ‘Crimson Star,’ ‘Crystal Fountain,’ ‘Nevesta,’ and ‘Serenada Kryma’ cultivars displayed high morphogenetic capacity under in vitro culturing. During indirect somatic embryogenesis, light intensity 37.5 μmol m–2 s–1 stimulated a higher-number somatic embryo formation and a temperature of 26°C affected somatic embryo development. Active formation of primary and secondary somatic embryos was also demonstrated. 2.20 μM BAP with 0.09 μM IBA affected the high-number somatic embryo formation for eight cultivars. Secondary somatic embryogenesis by the same concentration of BAP was induced. The frequency of secondary somatic embryogenesis was higher in ‘Crystal Fountain’ (100%), ‘Crimson Star’ (100%), ‘Nevesta’ (97%), and ‘Ay-Nor’ (92%) cultivars. Based on these results, the methodology for direct somatic embryogenesis and organogenesis of studied clematis cultivars has been developed.