Fine morphological assessment of quality of human mature oocytes after slow freezing or vitrification with a closed device: a comparative analysis

BACKGROUND: Human mature oocytes are very susceptible to cryodamage. Several reports demonstrated that vitrification might preserve oocyte better than slow freezing. However, this is still controversial. Thus, larger clinical, biological and experimental trials to confirm this concept are necessary. The aim of the study was to evaluate and compare fine morphological features in human mature oocytes cryopreserved with either slow freezing or vitrification. METHODS: We used 47 supernumerary human mature (metaphase II) oocytes donated by consenting patients, aged 27-32 years, enrolled in an IVF program. Thirtyfive oocytes were cryopreserved using slow freezing with 1.5 M propanediol +0.2 M sucrose concentration (20 oocytes) or a closed vitrification system (CryoTip Irvine Scientific CA) (15 oocytes). Twelve fresh oocytes were used as controls. All samples were prepared for light and transmission electron microscopy evaluation. RESULTS: Control, slow frozen/thawed and vitrified/warmed oocytes (CO, SFO and VO, respectively) were rounded, 90-100 mum in diameter, with normal ooplasm showing uniform distribution of organelles. Mitochondria-smooth endoplasmic reticulum (M-SER) aggregates and small mitochondria-vesicle (MV) complexes were the most numerous structures found in all CO, SFO and VO cultured for 3-4 hours. M-SER aggregates decreased, and large MV complexes increased in those SFO and VO maintained in culture for a prolonged period of time (8-9 hours). A slight to moderate vacuolization was present in the cytoplasm of SFO. Only a slight vacuolization was present in VO, whereas vacuoles were almost completely absent in CO. Amount and density of cortical granules (CG) appeared abnormally reduced in SFO and VO, irrespective of the protocol applied. CONCLUSIONS: Even though, both slow freezing and vitrification ensured a good overall preservation of the oocyte, we found that: 1) prolonged culture activates an intracellular membrane "recycling" that causes the abnormal transformation of the membranes of the small MV complexes and of SER into larger rounded vesicles; 2) vacuolization appears as a recurrent form of cell damage during slow freezing and, at a lesser extent, during vitrification using a closed device; 3) premature CG exocytosis was present in both SFO and VO and may cause zona pellucida hardenin

Mancozeb impairs the ultrastructure of mouse granulosa cells in a dose-dependent manner

Mancozeb, an ethylene bis-dithiocarbamate, is widely used as a fungicide and exerts reproductive toxicity in vivo and in vitro in mouse oocytes by altering spindle morphology and impairing the ability to fertilize. Mancozeb also induces a premalignant status in mouse granulosa cells (GCs) cultured in vitro, as indicated by decreased p53 expression and tenuous oxidative stress. However, the presence and extent of ultrastructural alterations induced by mancozeb on GCs in vitro have not yet been reported. Using an in vitro model of reproductive toxicity, comprising parietal GCs from mouse antral follicles cultured with increasing concentrations of mancozeb (0.001-1 µg/ml), we sought to ascertain the in vitro ultrastructural cell toxicity by means of transmission (TEM) and scanning (SEM) electron microscopy. The results showed a dose-dependent toxicity of mancozeb on mouse GCs. Ultrastructural data showed intercellular contact alterations, nuclear membrane irregularities, and chromatin marginalization at lower concentrations, and showed chromatin condensation, membrane blebbing, and cytoplasmic vacuolization at higher concentrations. Morphometric analysis evidenced a reduction of mitochondrial length in GCs exposed to mancozeb 0.01-1 µg/ml and a dose-dependent increase of vacuole dimension. In conclusion, mancozeb induced dose-dependent toxicity against GCs in vitro, including ultrastructural signs of cell degeneration compatible with apoptosis, likely due to the toxic breakdown product ethylenethiourea. These alterations may represent a major cause of reduced/delayed/missed oocyte maturation in cases of infertility associated with exposure to pesticides

Cyclic response of masonry piers retrofitted with timber frames and boards

The quasi-static in-plane cyclic response of two single-leaf calcium silicate unreinforced masonry piers was investigated to show the effectiveness of an innovative timber retrofit solution. The aim of the intervention is to increase the pier in-plane and out-of-plane strength and displacement capacity, thus reducing the seismic vulnerability of this typology of unreinforced masonry construction with a light, cost-effective, sustainable and reversible approach. The retrofit technique consists of a timber frame mechanically connected by means of steel fasteners to the masonry pier and building floors. Oriented strand timber boards are then nailed to the frame. In-plane quasi-static shear-compression cyclic tests were performed on two single-leaf calcium silicate brick piers with identical geometry and masonry mechanical properties: one was tested unstrengthened while the other was tested in the retrofitted configuration. The experimental results showed evident improvements in the lateral force-displacement response of the retrofitted specimen. More specifically, compared with the bare masonry pier, the retrofitted pier exhibited slightly higher stiffness, larger strength and significantly greater displacement capacity

Brief communication: Towards a universal formula for the probability of tornadoes

A methodological approach is proposed to provide an analytical (exponential-like) expression for the probability of occurrence of tornadoes as a function of the convective available potential energy and the wind shear (or, alternatively, the storm relative helicity). The resulting expression allows the probability of tornado occurrence to be calculated using variables that are computed by weather prediction and climate models, thus compensating for the lack of resolution needed to resolve these phenomena in numerical simulations

Simulation of a flash-flood event over the Adriatic Sea with a high-resolution atmosphere–ocean–wave coupled system

On the morning of September 26, 2007, a heavy precipitation event (HPE) affected the Venice lagoon and the neighbouring coastal zone of the Adriatic Sea, with 6-h accumulated rainfall summing up to about 360 mm in the area between the Venetian mainland, Padua and Chioggia. The event was triggered and maintained by the uplift over a convergence line between northeasterly flow from the Alps and southeasterly winds from the Adriatic Sea. Hindcast modelling experiments, using standalone atmospheric models, failed to capture the spatial distribution, maximum intensity and timing of the HPE. Here we analyze the event by means of an atmosphere-wave-ocean coupled numerical approach. The combined use of convection permitting models with grid spacing of 1 km, high-resolution sea surface temperature (SST) fields, and the consistent treatment of marine boundary layer fluxes in all the numerical model components are crucial to provide a realistic simulation of the event. Inaccurate representations of the SST affect the wind magnitude and, through this, the intensity, location and time evolution of the convergence zone, thus affecting the HPE prediction. © 2021, The Author(s)

IoT technologies for wine supply chain traceability: Potential application in the Southern Apulia Region (Italy)

The high value and volume of Italian wine production determines a strong stimulus for counterfeiting, which generates negative consequences for grape growers, winemakers and consumers. In this context, IoT technologies and the blockchain can serve as tools to ensure traceability, transparency and efficiency along the whole wine supply chain. Using primary data collected through interviews to the main grape growers and wineries involved in the wine supply chain in the Southern Apulia Region and secondary data, acquired from previous scientific literature, the study proposes a framework for the traceability and efficiency of the wine supply chain based on a combination of blockchain, Radio-Frequency Identification (RFID) and Near Field Communication (NFC) tags, Serial Shipping Container Codes (SSCC) and Quick Response (QR) codes. The developed framework allows for the systematic storage of information about commodities and processes throughout the supply chain, from grape growers to wine consumption and packaging disposal and/or reuse (forward and reverse flows). In addition, it ensures the transparency, safety, and security of all processes involved within the wine supply chain, serving as a quality information management tool. The information collected along the wine supply chain is entered into the management system by farmers, winemakers and bottlers and is accessible to all of them, while the distributors, consumers and the bottle reverse logistics operators, can only consult all of the information stored on the blockchain in order to know the origin, the quality, the processing and the authenticity of wines, without being able to enter data and/or modify the existent information

Sustainable vehicle routing based on firefly algorithm and TOPSIS methodology

In a sustainable management of logistics, transportation plays a crucial role. Traditionally, the main purpose was to solve the Vehicle Routing Problem minimizing the cost associated with the travelled distances. Nowadays, the economic profit cannot be the only driver for achieving sustainability and environmental issues have to be also considered. In this paper, to satisfy the intricate limits involved in real vehicle routing problem, the study has been structured considering different types of vehicles in terms of maximum capacity, velocity and emissions, asymmetric paths, vehicle-client constraints and delivery time windows. The firefly algorithm has been implemented to solve the vehicle routing problem and the TOPSIS technique has been applied to integrate economic and environmental factors. Finally, to prove the effectiveness of the proposed approach, a numerical example has been proposed using data provided by a logistic company located in Sicily

The regulatory G4 motif of the Kirsten ras (KRAS) gene is sensitive to guanine oxidation: Implications on transcription

KRAS is one of the most mutated genes in human cancer. It is controlled by a G4 motif located upstream of the transcription start site. In this paper, we demonstrate that 8-oxoguanine (8-oxoG), being more abundant in G4 than in non-G4 regions, is a new player in the regulation of this oncogene. We designed oligonucleotides mimicking the KRAS G4-motif and found that 8-oxoG impacts folding and stability of the G-quadruplex. Dimethylsulphate-footprinting showed that the G-run carrying 8-oxoG is excluded from the G-tetrads and replaced by a redundant G-run in the KRAS G4-motif. Chromatin immunoprecipitation revealed that the base-excision repair protein OGG1 is recruited to the KRAS promoter when the level of 8-oxoG in the G4 region is raised by H2O2. Polyacrylamide gel electrophoresis evidenced that OGG1 removes 8-oxoG from the G4-motif in duplex, but when folded it binds to the G-quadruplex in a non-productive way. We also found that 8-oxoG enhances the recruitment to the KRAS promoter of MAZ and hnRNP A1, two nuclear factors essential for transcription. All this suggests that 8-oxoG in the promoter G4 region could have an epigenetic potential for the control of gene expression

Radiogenomics in clear cell renal cell carcinoma: correlations between advanced CT imaging (texture analysis) and microRNAs expression

Purpose: A relevant challenge for the improvement of clear cell renal cell carcinoma management could derive from the identification of novel molecular biomarkers that could greatly improve the diagnosis, prognosis, and treatment choice of these neoplasms. In this study, we investigate whether quantitative parameters obtained from computed tomography texture analysis may correlate with the expression of selected oncogenic microRNAs. Methods: In a retrospective single-center study, multiphasic computed tomography examination (with arterial, portal, and urographic phases) was performed on 20 patients with clear cell renal cell carcinoma and computed tomography texture analysis parameters such as entropy, kurtosis, skewness, mean, and standard deviation of pixel distribution were measured using multiple filter settings. These quantitative data were correlated with the expression of selected microRNAs (miR-21-5p, miR-210-3p, miR-185-5p, miR-221-3p, miR-145-5p). Both the evaluations (microRNAs and computed tomography texture analysis) were performed on matched tumor and normal corticomedullar tissues of the same patients cohort. Results: In this pilot study, we evidenced that computed tomography texture analysis has robust parameters (eg, entropy, mean, standard deviation) to distinguish normal from pathological tissues. Moreover, a higher coefficient of determination between entropy and miR-21-5p expression was evidenced in tumor versus normal tissue. Interestingly, entropy and miR-21-5p show promising correlation in clear cell renal cell carcinoma opening to a radiogenomic strategy to improve clear cell renal cell carcinoma management. Conclusion: In this pilot study, a promising correlation between microRNAs and computed tomography texture analysis has been found in clear cell renal cell carcinoma. A clear cell renal cell carcinoma can benefit from noninvasive evaluation of texture parameters in adjunction to biopsy results. In particular, a promising correlation between entropy and miR-21-5p was found