Gd3+ Encapsulation on 2D-g-C3N4 Nanostructure for Spintronics and Ultrasound Assisted Photocatalytic Applications: First-Principles and Experimental Studies.

Atomically thin two-dimensional (2D) semiconductors have high potential in optoelectronics and magneto-optics appliances due to their tunable band structures and physicochemical stability. The work demonstrates that Gd3+ incorporated 2D-g-C3N4 nanosheet (Gd3+/2D-g-C3N4 NS) is synthesized through chemisorption methodology for defect enrichment. The material characterizations reveal that the ion decoration enhances the surface area and defect concentration of the 2D sheet. The experimental observations have been further corroborated with the help of density functional theory (DFT) simulation. Spin asymmetry polarizations near the Fermi level, obtained through the partial density of states (PDOS) analyses, reveal the magnetic nature of the synthesized material, validating the room temperature ferromagnetism obtained through a vibrating-sample magnetometer (VSM). Gd3+/2D-g-C3N4 NS shows significant enhancement in saturation magnetization (Ms) experimentally and computationally compared to the pristine one. The magnetic catalyst shows 98% remediation efficiency for ultrasound-assisted visible-light-driven photodegradation of methyl orange (MO). The synergistic approach of liquid chromatography-mass spectrometry (LC-MS) analyses and DFT studies elucidates reaction intermediates and unveils the degradation mechanism. Post-characterization studies assure the stability of the magnetic catalyst through optical, chemical, magnetic, and microscopic analyses. So, the synthesized material can be proficiently used as a magnetic nanocatalyst in wastewater treatments and spin-electronics applications.

Wastewater-based surveillance of Vibrio cholerae: Molecular insights on biofilm regulatory diguanylate cyclases, virulence factors and antibiotic resistance patterns.

Vibrio cholerae is an inherent inhabitant of aquatic ecosystems. The Indian state of West Bengal, especially the Gangetic delta region is the highest cholera affected region and is considered as the hub of Asiatic cholera. V. cholerae were isolated from publicly accessible wastewater of Midnapore, West Bengal, India. Serotyping determined all isolates to be of non-O1/non-O139 serogroups. Moderate biofilm-forming abilities were noticed in most of the isolates (74.7 %) while, high biofilm formation was recorded for only 6.3 % isolates and 19 % of isolates exhibited low/non-biofilm-forming abilities. PCR-based screening of crucial diguanylate cyclases (DGCs) involved in cyclic-di-GMP-mediated biofilm signaling was performed. cdgH and cdgM were the most abundant DGCs among 93.7 % and 91.5 % of isolates, respectively. Other important DGCs, i.e., cdgK, cdgA, cdgL, and vpvC were present in 84 %, 75.5 %, 72 % and 68 % of isolates, respectively. Besides, the non-O1/non-O139 isolates were screened for the occurrence of virulence factor encoding genes. Moreover, among these non-O1/non-O139 isolates, two strains (3.17 %) harbored both ctxA and ctxB genes, which encode the cholera toxin associated with epidemic cholera. ompU was the most prevalent virulence factor, present in 24.8 % of isolates. Other virulence factors like, zot and st were found in 4.7 % and 9.5 % of isolates. Genes encoding tcp and ace were found to be PCR-negative for the isolates. Additionally, crucial virulence factor regulators, toxT, toxR and hapR were found to be PCR-positive in all the isolates. Antibiotic resistance patterns displayed further vulnerabilities with decreased sensitivity towards commonly used antibiotics with multiple antibiotic resistance index ranging between 0.37 and 0.62. The presence of cholera toxin-encoding multi-drug resistant (MDR) V. cholerae strains in environmental settings is alarming. High occurrence of DGCs are considered to encourage further investigations to use them as alternative therapeutic targets against MDR cholera pathogen due to their unique presence in bacterial systems.

Optimized sperm selection: a highly efficient device for the isolation of progressive motile sperm with low DNA fragmentation index.

PURPOSE: To identify the sperm preparation procedure that selects the best sperm population for medically assisted reproduction. METHODS: Prospective observational study comparing the effect of four different sperm selection procedures on various semen parameters. Unused raw semen after routine diagnostic analysis was split in four fractions and processed by four different methods: (1) density gradient centrifugation (DGC), (2) sperm wash (SW), (3) DGC followed by magnetic activated cell sorting (MACS), and (4) using a sperm separation device (SSD). Each fraction was analyzed for progressive motility, morphology, acrosome index (AI), and DNA fragmentation index (DFI). RESULTS: With DGC as standard of care in intraclass correlation coefficient analysis, only SSD was in strong disagreement regarding progressive motility and DFI [0.26, 95%CI (- 0.2, 0.58), and 0.17, 95%CI (- 0.19, 0.45), respectively]. When controlling for abstinence duration, DFI was significantly lower after both MACS and SSD compared to DGC [- 0.27%, 95%CI (- 0.47, - 0.06), p = 0.01, and - 0.6%, 95%CI (- 0.80, - 0.41), p < 0.001, respectively]. Further comparisons between SSD and MACS indicate significantly less apoptotic cells [Median (IQR) 4 (5), 95%CI (4.1, - 6.8) vs Median (IQR) 5 (8), 95%CI (4.9, - 9.2), p < 0.001, respectively] and dead cells [Median (IQR) 9.5 (23.3), 95%CI (13.2, - 22.4) vs Median (IQR) 22 (28), 95%CI (23.1, - 36.8), p < 0.001, respectively] in the SSD group. CONCLUSION: The selection of a population of highly motile spermatozoa with less damaged DNA from unprocessed semen is ideally performed with SSD. Question remains whether this method improves the embryological outcomes in the IVF laboratory.

Identification of Dp140 and α1-syntrophin as novel molecular interactors of the neuronal CaV2.1 channel.

The primary function of dystrophin is to form a link between the cytoskeleton and the extracellular matrix. In addition to this crucial structural function, dystrophin also plays an essential role in clustering and organizing several signaling proteins, including ion channels. Proteomic analysis of the whole rodent brain has stressed the role of some components of the dystrophin-associated glycoprotein complex (DGC) as potential interacting proteins of the voltage-gated Ca2+ channels of the CaV2 subfamily. The interaction of CaV2 with signaling and scaffolding proteins, such as the DGC components, may influence their function, stability, and location in neurons. This work aims to study the interaction between dystrophin and CaV2.1. Our immunoprecipitation data showed the presence of a complex formed by CaV2.1, CaVα2δ-1, CaVß4e, Dp140, and α1-syntrophin in the brain. Furthermore, proximity ligation assays (PLA) showed that CaV2.1 and CaVα2δ-1 interact with dystrophin in the hippocampus and cerebellum. Notably, Dp140 and α1-syntrophin increase CaV2.1 protein stability, half-life, permanence in the plasma membrane, and current density through recombinant CaV2.1 channels. Therefore, we have identified the Dp140 and α1-syntrophin as novel interaction partners of CaV2.1 channels in the mammalian brain. Consistent with previous findings, our work provides evidence of the role of DGC in anchoring and clustering CaV channels in a macromolecular complex.

Silica-based colloid centrifugation enhances sperm quality in cockerel semen.

1. PercollTM is one of the most widely used colloid for animal sperm preparation. The aim of this study was to evaluate whether PercollTM colloid centrifugation could be practical to improve cockerel sperm quality, and to compare the effects of PercollTM single layer centrifugation (SLC) and density gradient centrifugation (DGC) in order to obtain the most optimal protocol for cockerel semen.2. In the experiment with PercollTM SLC for fresh semen, an increase of motile sperm was seen after PercollTM 80% SLC and 90% SLC was conducted, at levels of 28.8% and 30.2% respectively (P < 0.01). The increase of progressively motile sperm after PercollTM 80% SLC and 90% SLC was 177.2% and 202.4% respectively (P < 0.01). Meanwhile, for semen stored at 4°C for 24 h, the increase of motile sperm after PercollTM 70% SLC and 80% SLC was 41.2% and 44.0% (P < 0.01), and the increase of progressive sperm after PercollTM 70% SLC and 80% SLC was 71.3% and 83.1% respectively (P < 0.01). Both the percentage of motile sperm and progressive sperm of the fresh and stored cockerel semen after appropriate PercollTM SLC was significantly enhanced.3. Sperm membrane integrity did not show any decrease after PercollTM centrifugation compared with non-centrifuged semen, which suggested that the PercollTM centrifugation treatment in this study did not cause damage to cockerel sperm membranes.4. In the experiment regarding the comparison of PercollTM SLC and DGC with fresh semen, the increase of motile sperm after PercollTM 80% SLC, 90% SLC and 40%/80% DGC was 29.5%, 36.4%, and 25.0% respectively; and the increase of progressive sperm was 44.7%, 58.5%, and 54.7%, respectively. For semen stored at 4°C for 24 h, the increase of motile sperm after PercollTM 70% SLC, 80% SLC and 35%/70% DGC were 41.2%, 44.0%, and 26.4%; and the increase of progressive sperm was 71.3%, 83.1%, and 43.7%, respectively. There were no significant differences between the increase of sperm motility after PercollTM 80%, 90% SLC or PercollTM 40%/80% DGC in fresh cockerel semen. There was no significant difference between PercollTM 70%, 80% SLC and PercollTM 35%/70% in stored cockerel semen. There was a tendency for sperm recovery rates with PercollTM SLC to be higher than PercollTM DGC, although this did not reach statistical significance in this study.5. It was concluded that PercollTM SLC was more suitable for cockerel sperm separation than PercollTM DGC. The results suggested that PercollTM 80% SLC was the most optimal procedure to separate fresh cockerel sperm and PercollTM 70% SLC was the most optimal procedure to separate stored cockerel sperm. PercollTM SLC is more simple, user-friendly and economical and less time-consuming than DGC for cockerel semen processing.

A d-enantiomeric peptide interferes with heteroassociation of amyloid-ß oligomers and prion protein.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder that affects millions of people worldwide. One AD hallmark is the aggregation of ß-amyloid (Aß) into soluble oligomers and insoluble fibrils. Several studies have reported that oligomers rather than fibrils are the most toxic species in AD progression. Aß oligomers bind with high affinity to membrane-associated prion protein (PrP), leading to toxic signaling across the cell membrane, which makes the Aß-PrP interaction an attractive therapeutic target. Here, probing this interaction in more detail, we found that both full-length, soluble human (hu) PrP(23-230) and huPrP(23-144), lacking the globular C-terminal domain, bind to Aß oligomers to form large complexes above the megadalton size range. Following purification by sucrose density-gradient ultracentrifugation, the Aß and huPrP contents in these heteroassemblies were quantified by reversed-phase HPLC. The Aß:PrP molar ratio in these assemblies exhibited some limited variation depending on the molar ratio of the initial mixture. Specifically, a molar ratio of about four Aß to one huPrP in the presence of an excess of huPrP(23-230) or huPrP(23-144) suggested that four Aß units are required to form one huPrP-binding site. Of note, an Aß-binding all-d-enantiomeric peptide, RD2D3, competed with huPrP for Aß oligomers and interfered with Aß-PrP heteroassembly in a concentration-dependent manner. Our results highlight the importance of multivalent epitopes on Aß oligomers for Aß-PrP interactions and have yielded an all-d-peptide-based, therapeutically promising agent that competes with PrP for these interactions.

Spatial distribution and molecular dynamics of dystrophin glycoprotein components at the neuromuscular junction in vivo.

A bimolecular fluorescence complementation (BiFC) approach was used to study the molecular interactions between different components of the postsynaptic protein complex at the neuromuscular junction of living mice. We show that rapsyn forms complex with both α-dystrobrevin and α-syntrophin at the crests of junctional folds. The linkage of rapsyn to α-syntrophin and/or α-dystrobrevin is mediated by utrophin, a protein localized at acetylcholine receptor (AChR)-rich domains. In mice deficient in α-syntrophin, in which utrophin is no longer present at the synapse, rapsyn interaction with α-dystrobrevin was completely abolished. This interaction was completely restored when either utrophin or α-syntrophin was introduced into muscles deficient in α-syntrophin. However, in neuromuscular junctions deficient in α-dystrobrevin, in which utrophin is retained, complex formation between rapsyn and α-syntrophin was unaffected. Using fluorescence recovery after photobleaching, we found that α-syntrophin turnover is 5-7 times faster than that of AChRs, and loss of α-dystrobrevin has no effect on rapsyn and α-syntrophin half-life, whereas the half-life of AChR was significantly altered. Altogether, these results provide new insights into the spatial distribution of dystrophin glycoprotein components and their dynamics in living mice.

Untargeted/Targeted 2D Gas Chromatography/Mass Spectrometry Detection of the Total Volatile Tea Metabolome.

Identifying all analytes in a natural product is a daunting challenge, even if fractionated by volatility. In this study, comprehensive two-dimensional gas chromatography/mass spectrometry (GC×GC-MS) was used to investigate relative distribution of volatiles in green, pu-erh tea from leaves collected at two different elevations (1162 m and 1651 m). A total of 317 high and 280 low elevation compounds were detected, many of them known to have sensory and health beneficial properties. The samples were evaluated by two different software. The first, GC Image, used feature-based detection algorithms to identify spectral patterns and peak-regions, leading to tentative identification of 107 compounds. The software produced a composite map illustrating differences in the samples. The second, Ion Analytics, employed spectral deconvolution algorithms to detect target compounds, then subtracted their spectra from the total ion current chromatogram to reveal untargeted compounds. Compound identities were more easily assigned, since chromatogram complexities were reduced. Of the 317 compounds, for example, 34% were positively identified and 42% were tentatively identified, leaving 24% as unknowns. This study demonstrated the targeted/untargeted approach taken simplifies the analysis time for large data sets, leading to a better understanding of the chemistry behind biological phenomena.

ABC of multifaceted dystrophin glycoprotein complex (DGC).

Dystrophin protein in association with several other cellular proteins and glycoproteins leads to the formation of a large multifaceted protein complex at the cell membrane referred to as dystrophin glycoprotein complex (DGC), that serves distinct functions in cell signaling and maintaining the membrane stability as well as integrity. In accordance with this, several findings suggest exquisite role of DGC in signaling pathways associated with cell development and/or maintenance of homeostasis. In the present review, we summarize the established facts about the various components of this complex with emphasis on recent insights into specific contribution of the DGC in cell signaling at the membrane. We have also discussed the recent advances made in exploring the molecular associations of DGC components within the cells and the functional implications of these interactions. Our review would help to comprehend the composition, role, and functioning of DGC and may lead to a deeper understanding of its role in several human diseases.

Biochemical and Functional Interplay Between Ion Channels and the Components of the Dystrophin-Associated Glycoprotein Complex.

Dystrophin is a cytoskeleton-linked membrane protein that binds to a larger multiprotein assembly called the dystrophin-associated glycoprotein complex (DGC). The deficiency of dystrophin or the components of the DGC results in the loss of connection between the cytoskeleton and the extracellular matrix with significant pathophysiological implications in skeletal and cardiac muscle as well as in the nervous system. Although the DGC plays an important role in maintaining membrane stability, it can also be considered as a versatile and flexible molecular complex that contribute to the cellular organization and dynamics of a variety of proteins at specific locations in the plasma membrane. This review deals with the role of the DGC in transmembrane signaling by forming supramolecular assemblies for regulating ion channel localization and activity. These interactions are relevant for cell homeostasis, and its alterations may play a significant role in the etiology and pathogenesis of various disorders affecting muscle and nerve function.

CDH1 mutation screen in a BRCA1/2-negative familial breast-/ovarian cancer cohort.

PURPOSE: Mutations in the CDH1 gene are linked both to diffuse gastric cancer and invasive lobular carcinoma (ILC). A high mutation rate is found in families fulfilling the diagnostic criteria for hereditary diffuse gastric cancer. Aim of this study was to clarify whether or not there is a significant contribution of CDH1 mutations in hereditary breast-/ovarian cancer (HBOC). METHODS: Ninety-seven unrelated probands fulfilling the diagnostic criteria for HBOC (96 affected, 1 unaffected) but tested negative for pathogenic BRCA1/2 mutations were screened for CDH1 mutations by denaturing high performance liquid chromatography (DHPLC) and subsequent Sanger sequencing of suspicious and positive DHPLC results. RESULTS: In total, we found two potentially pathogenic CDH1 alterations, c.1774G > A, pAla592Thr, and c.2512 A > G, p.Ser838Gly, classified as variants of unknown significance according to ClinVar. In addition, we detected a high number of known CDH1 polymorphisms (n = 62), some of them more frequent in patients with lobular (55%) than in those with invasive ductal carcinoma (27%). CONCLUSION: Although none of the probands studied carried a clearly pathogenic CDH1 mutation, CDH1 could be considered a potential breast cancer gene, esp. for ILC worth including it in the NGS (next generation sequencing) HBOC panel.

A n-3 PUFA depletion applied to rainbow trout fry (Oncorhynchus mykiss) does not modulate its subsequent lipid bioconversion capacity.

Nutritional strategies are currently developed to produce farmed fish rich in n-3 long-chain PUFA (LC-PUFA) whilst replacing fish oil by plant-derived oils in aquafeeds. The optimisation of such strategies requires a thorough understanding of fish lipid metabolism and its nutritional modulation. The present study evaluated the fatty acid bioconversion capacity of rainbow trout (Oncorhynchus mykiss) fry previously depleted in n-3 PUFA through a 60-d pre-experimental feeding period with a sunflower oil-based diet (SO) followed by a 36-d experimental period during which fish were fed either a linseed oil-based diet (LO) (this treatment being called SO/LO) or a fish oil-based diet (FO) (this treatment being called SO/FO). These treatments were compared with fish continuously fed on SO, LO or FO for 96 d. At the end of the 36-d experimental period, SO/LO and SO/FO fish recovered >80 % of the n-3 LC-PUFA reported for LO and FO fish, respectively. Fish fed on LO showed high apparent in vivo elongation and desaturation activities along the n-3 biosynthesis pathway. However, at the end of the experimental period, no impact of the fish n-3 PUFA depletion was observed on apparent in vivo elongation and desaturation activities of SO/LO fish as compared with LO fish. In contrast, the fish n-3 PUFA depletion negatively modulated the n-6 PUFA bioconversion capacity of fish in terms of reduced apparent in vivo elongation and desaturation activities. The effects were similar after 10 or 36 d of the experimental period, indicating the absence of short-term effects.

Hepatocyte Growth Factor and Satellite Cell Activation.

Satellite cells are the "currency" for the muscle growth that is critical to meat production in many species, as well as to phenotypic distinctions in development at the level of species or taxa, and for human muscle growth, function and regeneration. Careful research on the activation and behaviour of satellite cells, the stem cells in skeletal muscle, including cross-species comparisons, has potential to reveal the mechanisms underlying pathological conditions in animals and humans, and to anticipate implications of development, evolution and environmental change on muscle function and animal performance.

The effect of discontinuous gas exchange on respiratory water loss in grasshoppers (Orthoptera: Acrididae) varies across an aridity gradient.

The significance of discontinuous gas-exchange cycles (DGC) in reducing respiratory water loss (RWL) in insects is contentious. Results from single-species studies are equivocal in their support of the classic 'hygric hypothesis' for the evolution of DGC, whereas comparative analyses generally support a link between DGC and water balance. In this study, we investigated DGC prevalence and characteristics and RWL in three grasshopper species (Acrididae, subfamily Pamphaginae) across an aridity gradient in Israel. In order to determine whether DGC contributes to a reduction in RWL, we compared the DGC characteristics and RWL associated with CO2 release (transpiration ratio, i.e. the molar ratio of RWL to CO2 emission rates) among these species. Transpiration ratios of DGC and continuous breathers were also compared intraspecifically. Our data show that DGC characteristics, DGC prevalence and the transpiration ratios correlate well with habitat aridity. The xeric-adapted Tmethis pulchripennis exhibited a significantly shorter burst period and lower transpiration ratio compared with the other two mesic species, Ocneropsis bethlemita and Ocneropsis lividipes. However, DGC resulted in significant water savings compared with continuous exchange in T. pulchripennis only. These unique DGC characteristics for T. pulchripennis were correlated with its significantly higher mass-specific tracheal volume. Our data suggest that the origin of DGC may not be adaptive, but rather that evolved modulation of cycle characteristics confers a fitness advantage under stressful conditions. This modulation may result from morphological and/or physiological modifications.

A computational model of insect discontinuous gas exchange: A two-sensor, control systems approach.

The insect gas exchange system is characterised by branching air-filled tubes (tracheae/tracheoles) and valve-like structures in their outer integument (spiracles) which allow for a periodic gas exchange pattern known as the discontinuous gas exchange cycle (DGC). The DGC facilitates the temporal decoupling of whole animal gas exchange from cellular respiration rates and may confer several physiological benefits, which are nevertheless highly controversial (primarily reduction of cellular oxidative damage and/or respiratory water saving). The intrinsic and extrinsic factors influencing DGCs are the focus of extensive ongoing research and little consensus has been reached on the evolutionary genesis or mechanistic costs and benefits of the pattern. Despite several hypotheses and much experimental and evolutionary biology research, a mechanistic physical model, which captures various key elements of the DGC pattern, is currently lacking. Here, we present a biologically realistic computational, two-sensor DGC model (pH/carbon dioxide and oxygen setpoints) for an Orthopteran gas exchange system, and show computationally for the first time that a control system of two interacting feedback loops is capable of generating a full DGC pattern with outputs which are physiologically realistic, quantitatively matching experimental results found in this taxonomic model elsewhere. A finite-element mathematical approach is employed and various trigger sets are considered. Parameter sensitivity analyses suggest that various aspects of insect DGC are adequately captured in this model. In particular, with physiologically relevant input parameters, the full DGC pattern is induced; and the phase durations, endotracheal carbon dioxide partial pressure ranges, and pH fluctuations which arise are physically realistic. The model results support the emergent property hypothesis for the existence of DGC, and indicate that asymmetric loading and off-loading (hysteresis) in one of the sensor feedback loops, which is a novel inclusion here, is a critical aspect of the insect spiracle-tracheal gas exchange system.

Regional and genotypic differences in intrinsic electrophysiological properties of cerebellar Purkinje neurons from wild-type and dystrophin-deficient mdx mice.

Cerebellar subregions are recognized as having specialized roles, with lateral cerebellum considered crucial for cognitive processing, whereas vermal cerebellum is more strongly associated with motor control. In human Duchenne muscular dystrophy, loss of the cytoskeletal protein dystrophin is thought to cause impairments in cognition, including learning and memory. Previous studies demonstrate that loss of dystrophin causes dysfunctional signaling at γ-aminobutyric acid (GABA) synapses on Purkinje neurons, presumably by destabilization of GABAA receptors. However, potential differences in the intrinsic electrophysiological properties of Purkinje neurons, including membrane potential and action potential firing rates, have not been investigated. Here, using a 2×2 analysis of variance (ANOVA) experimental design, we employed patch clamp analysis to compare membrane properties and action potentials generated by acutely dissociated Purkinje neurons from vermal and lateral cerebellum in wild-type (WT) mice and mdx dystrophin-deficient mice. Compared to Purkinje neurons from WT mice, neurons from mdx mice exhibited more irregular action potential firing and a hyperpolarization of the membrane potential. Firing frequency was also lower in Purkinje neurons from the lateral cerebellum of mdx mice relative to those from WT mice. Several action potential waveform parameters differed between vermal and lateral Purkinje neurons, irrespective of dystrophin status, including action potential amplitude, slope (both larger in the vermal region), and duration (shorter in the vermal region). Moreover, the membrane potential of Purkinje neurons from the vermal region of WT mice exhibited a significant hyperpolarization and concurrent reduction in the frequency of spontaneous action potentials compared to Purkinje neurons from the lateral region. This regional hyperpolarization and reduction in spontaneous action potential frequency was abolished in mdx mice. These results from mice demonstrate the presence of differential electrophysiological properties between Purkinje neurons from different regions of the WT mouse cerebellum and altered intrinsic membrane properties in the absence of dystrophin. These findings provide a possible mechanism for the observations that absence of cerebellar dystrophin contributes to deficits in mental function observed in humans and mouse models of muscular dystrophy. Moreover, these results highlight the importance of distinguishing functional zones of the cerebellum in future work characterizing Purkinje neuron electrophysiology and studies using the model of dissociated Purkinje neurons from mice.

Temperature-dependent variation in gas exchange patterns and spiracular control in Rhodnius prolixus.

Insects display an array of respiratory behaviors, including the use of discontinuous gas exchange. This pattern is characterized by periods of spiracular closure, micro-openings (flutter), and complete openings during which the majority of gas exchange takes place. A current model of insect spiracular control suggests that spiracles are controlled by two interacting feedback loops, which produce the discontinuous pattern. The flutter period is thought to be initiated by a critically low partial pressure of oxygen, while the open period is initiated by a critically high CO2 threshold. The goal of our study was to test this control model under conditions of feeding-induced or temperature-induced changes in metabolic rate. We manipulated the metabolic rate of the insect Rhodnius prolixus using two discrete mechanisms: (1) feeding the insects a bloodmeal or (2) exposing them to a range of temperatures (18-38°C). Examining the variation in the gas exchange patterns produced by insects in each of these treatments allowed us to determine whether spiracular control is sensitive to metabolic rate and/or temperature. We found that increases in temperature caused significant decreases in open phase burst volumes and premature abandonment of discontinuous gas exchange cycles. These effects were not observed in fed individuals maintained at a single temperature despite their higher metabolic rates. Our results indicate that some part of the spiracular control mechanism is temperature sensitive, suggesting a possible role for pH in CO2 sensing.

Oxygen diffusion limitation triggers ventilatory movements during spiracle closure when insects breathe discontinuously.

During discontinuous gas exchange cycles in insects, spiracular opening follows a typical prolonged period of spiracle closure. Gas exchange with the environment occurs mostly during the period of full spiracular opening. In this study we tested the hypothesis that recently reported ventilatory movements during the spiracle closure period serve to mix the tracheal system gaseous contents, and support diffusive exchanges with the tissues. Using heliox (21% O2, 79% He), we found that by increasing oxygen diffusivity in the gas phase, ventilatory movements of Schistocerca gregaria were significantly delayed compared with normoxic conditions. Exposure to hyperoxic conditions (40% O2, 60% N2) resulted in a similar delay in forced ventilation. Together, these results indicate that limits to oxygen diffusion to the tissues during spiracle closure trigger ventilatory movements, which in turn support tissue demands. These findings contribute to our understanding of the mechanistic basis of respiratory gas exchange between insect tissues and the environment.

Sperm Preparation with Microfluidic Sperm Sorting Chip May Improve Intracytoplasmic Sperm Injection Outcomes Compared to Density Gradient Centrifugation.

Does sperm preparation using the FERTILE PLUS™ Sperm Sorting Chip improve fertilization rates, blastocyst formation, utilization, and euploidy rates in patients undergoing intracytoplasmic sperm injection (ICSI), compared with density gradient centrifugation (DGC)? A single-cohort, retrospective data review including data from 53 couples who underwent ICSI cycles within a 12-month period. For each couple, the two closest, consecutive cycles were identified, where one used the standard technique of sperm preparation (DGC) and the subsequent used FERTILE PLUS™, therefore, couples acted as their own controls. Paired samples t-test was used to compare means for the outcomes (fertilization, blastocyst formation, utilization, and euploidy rates). Binary logistic regression analysis assessed the relationship between female age, the presence of male factor infertility, and euploidy rates. Blastocyst, utilization, and euploidy rates were significantly higher for cycles using FERTILE PLUS™ compared to DGC (76% vs 56%, p = 0.002; 60% vs 41%, p = 0.005, and 40% vs 20%, p = 0.001, respectively). Although there was an increase in fertilization rates for cycles using FERTILE PLUS™, this was not significant (72% vs 68%, p = 0.449). The euploidy rates of females ≤ 35 years were significantly increased when the FERTILE PLUS™ sperm preparation method was used, compared to the older age group (OR 2.31, p = 0.007). No significant association was found between the presence or absence of male factor infertility and euploidy rates between the two cycles. This study provides tentative evidence that the FERTILE PLUS™ microfluidic sorting device for sperm selection can improve blastocyst formation, utilization, and euploidy rates following ICSI in comparison to the DGC method.

Pre-freezing selection of Holstein bull semen with the BoviPure colloid as double- or single-layer centrifugation improves the post-thawing quality.

Artificial insemination (AI) is critical for breeding in the dairy industry. High-merit bulls can present low freezability, hampering genetic dissemination. Thawed semen can be improved using density gradient centrifugation (DGC) with colloids, but little information deals with the pre-freezing application. Thus, the BoviPure colloid (optimized for bull spermatozoa) was tested for pre-freezing application as the usual double-layer (DLC) versus single-layer (SLC, quick and economical). Semen from twelve Holstein-Friesian bulls was extended with OPTIXcell extender, frozen (Control), or processed by SLC or DLC and frozen. Sperm were assessed pre-freezing for motility and viability and post-thawing (directly and after 4 h 38 °C) for apoptosis, capacitation status, acrosomal damage, mitochondrial activity, cytoplasmic and mitochondrial reactive oxygen species (ROS), and chromatin status (SCSA for DNA fragmentation and chromatin compaction and monobromobimane, mBBr, for disulfide bridges evaluation). The DGC improved parameters post-thawing (e.g., 57.5%±10.1 motility vs. control 53.3% ± 11.2) at the cost of sperm loss (sperm recovery of DGC 14.4% ± 2.5 and SLC 17.4% ± 2.5). DNA fragmentation (%DFI) decreased (0.21% ± 0.53 vs. control 1.30% ± 0.10), and SLC reduced chromatin compaction. A clustering procedure separated lesser (LF) and greater freezability (GF) bulls. LF samples were especially benefited by DGC, with SLC providing better post-thawing results for this group. In conclusion, pre-freezing DGC improved sperm parameters post-thawing, potentially improving the cryopreservation of low-freezability semen from high-merit bulls. SLC, quicker and economical, would be preferable since it showed similar or higher performance than DLC.