Sperm Factor Initiates Capacitance and Conductance Changes in Mouse Eggs That Are More Similar to Fertilization Than IP3- or Ca2+-induced Changes

AbstractWe used patch clamp electrophysiology and concurrent imaging with the Ca2+-sensitive dye, fura-2, to study the temporal relationship between membrane capacitance and conductance and intracellular free Ca2+ concentration ([Ca2+]i) during mouse egg fertilization. We found an ∼2 pF step increase in egg membrane capacitance and a minor increase in conductance with no change in [Ca2+]i at sperm fusion. This was followed ∼1 min later by a rise in [Ca2+]i that led to larger changes in capacitance and conductance. The most common pattern for these later capacitance changes was an initial capacitance decrease, followed by a larger increase and eventual return to the approximate starting value. There was some variation in this pattern, and sub-μM peak [Ca2+]i favored capacitance decrease, while higher [Ca2+]i favored capacitance increase. The magnitude of accompanying conductance increases was variable and did not correlate well with peak [Ca2+]i. The intracellular introduction of porcine sperm factor reproduced the postfusion capacitance and conductance changes with a similar [Ca2+]i dependence. Raising [Ca2+]i by the intracellular introduction of IP3 initiated fertilization-like capacitance changes, but the conductance changes were slower to activate. Capacitance decrease could be induced when [Ca2+]i was increased modestly by activation of an endogenous Ca2+ current, with little effect on resting conductance. These results suggest that net turnover of the mouse egg surface membrane is sensitive to [Ca2+]i and that sperm and the active component of sperm factor may be doing more than initiating the IP3-mediated release of intracellular Ca2+

Simulations of Intracellular Calcium Release Dynamics in Response to a High-Intensity, Ultrashort Electric Pulse

Numerical simulations for electrically induced, intracellular calcium release from the endoplasmic reticulum are reported. A two-step model is used for self-consistency. Distributed electrical circuit representation coupled with the Smoluchowski equation yields the ER membrane nanoporation for calcium outflow based on a numerical simulation. This is combined with the continuum Li-Rinzel model and drift diffusion for calcium dynamics. Our results are shown to be in agreement with reported calcium release data. A modest increase (rough doubling) of the cellular calcium is predicted in the absence of extra-cellular calcium. In particular, the applied field of 15 kV/cm with 60 ns pulse duration makes for a strong comparison. No oscillations are predicted and the net recovery period of about 5 min are both in agreement with published experimental results. A quantitative explanation for the lack of such oscillatory behavior, based on the density dependent calcium fluxes, is also provided

The Role of Psychological Well-Being in Weight Loss: New Insights from a Comprehensive Lifestyle Intervention

Background/Objective: Although the literature suggested that impaired psychological well-being (PWB) is associated with obesity, evidence on the role of PWB in weight outcomes is limited and inconclusive. This research aimed to investigate the joint role of PWB in achieving clinically significant weight loss (CWL; loss of 5% of the initial weight) through a comprehensive lifestyle intervention for obesity using a broad-based evaluation. Method: This study is a prospective cohort of 96 patients with obesity attending a comprehensive lifestyle intervention for weight loss. Data on weight, lifestyle, PWB, and distress, were collected before and after the intervention. Results: 30.5% of the participants achieved CWL at the end of treatment. A more pronounced increase in autonomy (odds ratio = 0.80 [95% CI: 0.68, 0.93], p ≤.01) and somatization (odds ratio = 0.83 [95% CI: 0.70, 0.98], p ≤.05) from pre- to post-treatment were independently associated with a lower probability of CWL. Conclusions: Unbalanced dimensions of PWB, in particular exceedingly high autonomy, may contribute to a poor weight loss outcome. This study paves the way for the addition of psychotherapeutic strategies geared to euthymia in comprehensive lifestyle intervention

Are HLA-G polymorphisms associated to recurrent pregnancy loss?

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Dietary Supplementation with Boswellia serrata, Verbascum thapsus, and Curcuma longa in Show Jumping Horses: Effects on Serum Proteome, Antioxidant Status, and Anti-Inflammatory Gene Expression

Intense exercise can cause inflammation and oxidative stress due to the production of reactive oxygen species. These pathophysiological processes are interdependent, and each one can induce the other, creating a vicious circle. A placebo-controlled blind study was carried out in show jumping horses (n. 16) to evaluate the effects of a commercial dietary supplement (Dolhorse® N.B.F. Lanes srl, Milan, Italy) containing Verbascum thapsus leaf powder (1.42%), Curcuma longa (14.280 mg/kg), and Boswellia serrata (Roxb ex Colebr) (14.280 mg/kg) extracts. Before and after 10 days of dietary supplementation, blood samples were collected to evaluate the protein levels, antioxidants, and inflammatory responses by proteomic analysis or real-time Reverse Transcriptase-Polymerase Chain Reaction (real-time RT-PCR). A total of 36 protein spots, connected to 29 proteins, were modulated by dietary supplementation, whereas real-time RT-PCR revealed a significant downregulation of proinflammatory cytokines (interleukin 1α (p < 0.05) and interleukin-6 (0.005), toll-like receptor 4 (p < 0.05), and IKBKB (p < 0.05) in supplemented sport horses. Immunoglobulin chains, gelsolin, plasminogen, vitamin D binding protein, apolipoprotein AIV, and filamin B were overexpressed, whereas haptoglobin, α-2-HS-glycoprotein, α2-macroglobulin, afamin, amine oxidase, 60S acidic ribosomal protein, and complement fragments 3, 4, and 7 were reduced. No effect was observed on the antioxidant defense systems. The present results suggest this phytotherapy may reinforce the innate immune responses, thus representing a valid adjuvant to alleviate inflammation, which is a pathophysiological process in sport horses

Ground deformation analysis at Campi Flegrei (Southern Italy) by CGPS and tide-gauge network

Campi Flegrei caldera is located 15 km west of the city of Naples, within the central-southern sector of a large graben called Campanian Plain. It is an active volcanic area marked by a quasi-circular caldera depression, formed by a huge ignimbritic eruption occurred about 37000 years ago. This caldera was generated by several collapses produced by strong explosive eruptions (the last eruption, occurred in 1538, built an about 130 m spatter cone called Mt. Nuovo). Campi Flegrei area periodically experiences significant deformation episodes, with uplift phenomena up to more than 3.5 m in 15 years (from 1970 to 1984), which caused during 1983-84 the temporary evacuation of about 40000 people from the ancient part of Pozzuoli town. The deformation field obtainable by CGPS and tidegauge stations plays an important role for the modelling and interpretation of volcanic phenomena, as well as for forecasting purposes. The structural complexity of the Campi Flegrei area, together with the evidence of a strong interaction between magmatic chamber and shallow geothermal system, calls for a detailed characterization of the substructure and of magma-water interaction processes. The incoming experiment of deep drilling, down to about 4 km, will give detailed structural and physical constraints able to resolve the intrinsic ambiguities of geophysical data and in particular geodetic ones. In this poster we describe the recent ground deformations at Campi Flegrei area by means of GPS technique and tide gauge stations, discussing the possible interpretations also in light of further constraints likely coming from the next CFDDP (Campi Flegrei Deep Drilling) deep drilling experiment

Structure analysis and site-directed mutagenesis of defined key residues and motives for pilus-related sortase C1 in group B Streptococcus

In group B Streptococcus (GBS), 3 structurally distinct types of pili have been discovered as potential virulence factors and vaccine candidates. The pilus-forming proteins are assembled into high-molecular-weight polymers via a transpeptidation mechanism mediated by specific class C sortases. Using a multidisciplinary approach including bioinformatics, structural and biochemical studies, and in vivo mutagenesis, we performed a broad characterization of GBS sortase C1 of pilus island 2a. The high-resolution X-ray structure of the enzyme revealed that the active site, into the \u3b2-barrel core of the enzyme, is made of the catalytic triad His157-Cys219-Arg228 and covered by a loop, known as the \u201clid.\u201d We show that the catalytic triad and the predicted N- and C-terminal transmembrane regions are required for the enzyme activity. Interestingly, by in vivo complementation mutagenesis studies, we found that the deletion of the entire lid loop or mutations in specific lid key residues had no effect on catalytic activity of the enzyme. In addition, kinetic characterizations of recombinant enzymes indicate that the lid mutants can still recognize and cleave the substrate-mimicking peptide at least as well as the wild-type protein.\u2014Cozzi, R., Malito, E., Nuccitelli, A., D\u2019Onofrio, M., Martinelli, M., Ferlenghi, I., Grandi, G., Telford, J. L., Maione, D., Rinaudo, C. D. Structure analysis and site-directed mutagenesis of defined key residues and motives for pilus-related sortase C1 in group B Streptococcus

Bioelectric Effects of Intense Nanosecond Pulses

Electrical models for biological cells predict that reducing the duration of applied electrical pulses to values below the charging time of the outer cell membrane (which is on the order of 100 ns for mammalian cells) causes a strong increase in the probability of electric field interactions with intracellular structures due to displacement currents. For electric field amplitudes exceeding MV/m, such pulses are also expected to allow access to the cell interior through conduction currents flowing through the permeabilized plasma membrane. In both cases, limiting the duration of the electrical pulses to nanoseconds ensures only nonthermal interactions of the electric field with subcellular structures. This intracellular access allows the manipulation of cell functions. Experimental studies, in which human cells were exposed to pulsed electric fields of up to 300 kY/cm amplitude with durations as short as 3 ns, have confirmed this hypothesis and have shown that it is possible to selectively alter the behavior and/or survival of cells. Observed nanosecond pulsed effects at moderate electric fields include intracellular release of calcium and enhanced gene expression, which could have long term implications on cell behavior and function. At increased electric fields, the application of nanosecond pulses induces a type of programmed cell death, apoptosis, in biological cells. Cell survival studies with 10 ns pulses have shown that the viability of the cells scales inversely with the electrical energy density, which is similar to the ‘dose’ effect caused by ionizing radiation. On the other hand, there is experimental evidence that, for pulses of varying durations, the onset of a range of observed biological effects is determined by the electrical charge that is transferred to the cell membrane during pulsing. This leads to an empirical similarity law for nanosecond pulse effects, with the product of electric field intensity, pulse duration, and the square root of the number of pulses as the similarity parameter. The similarity law allows one not only to predict cell viability based on pulse parameters, but has also been shown to be applicable for inducing platelet aggregation, an effect which is triggered by internal calcium release. Applications for nanosecond pulse effects cover a wide range: from a rather simple use as preventing biofouling in cooling water systems, to advanced medical applications, such as gene therapy and tumor treatment. Results of this continuing research are leading to the development of wound healing and skin cancer treatments, which are discussed in some detail