Problems Affecting Labor

Much experimental work has been devoted in comparing the folding behavior of proteins sharing the same fold but different sequence. The recent design of proteins displaying very high sequence identities but different 3D structure allows the unique opportunity to address the protein-folding problem from a complementary perspective. Here we explored by ℙ-value analysis the pathways of folding of three different heteromorphic pairs, displaying increasingly high-sequence identity (namely, 30%, 77%, and 88%), but different structures called G A (a 3-α helix fold) and G B (an α/β fold). The analysis, based on 132 site-directed mutants, is fully consistent with the idea that protein topology is committed very early along the pathway of folding. Furthermore, data reveals that when folding approaches a perfect two-state scenario, as in the case of the G A domains, the structural features of the transition state appear very robust to changes in sequence composition. On the other hand, when folding is more complex and multistate, as for the G Bs, there are alternative nuclei or accessible pathways that can be alternatively stabilized by altering the primary structure. The implications of our results in the light of previous work on the folding of different members belonging to the same protein family are discussed

Tunneling and delocalization in hydrogen bonded systems: a study in position and momentum space

Novel experimental and computational studies have uncovered the proton momentum distribution in hydrogen bonded systems. In this work, we utilize recently developed open path integral Car-Parrinello molecular dynamics methodology in order to study the momentum distribution in phases of high pressure ice. Some of these phases exhibit symmetric hydrogen bonds and quantum tunneling. We find that the symmetric hydrogen bonded phase possesses a narrowed momentum distribution as compared with a covalently bonded phase, in agreement with recent experimental findings. The signatures of tunneling that we observe are a narrowed distribution in the low-to-intermediate momentum region, with a tail that extends to match the result of the covalently bonded state. The transition to tunneling behavior shows similarity to features observed in recent experiments performed on confined water. We corroborate our ice simulations with a study of a particle in a model one-dimensional double well potential that mimics some of the effects observed in bulk simulations. The temperature dependence of the momentum distribution in the one-dimensional model allows for the differentiation between ground state and mixed state tunneling effects.Comment: 14 pages, 13 figure

Efficient multiple time scale molecular dynamics: using colored noise thermostats to stabilize resonances

Multiple time scale molecular dynamics enhances computational efficiency by updating slow motions less frequently than fast motions. However, in practice the largest outer time step possible is limited not by the physical forces but by resonances between the fast and slow modes. In this paper we show that this problem can be alleviated by using a simple colored noise thermostatting scheme which selectively targets the high frequency modes in the system. For two sample problems, flexible water and solvated alanine dipeptide, we demonstrate that this allows the use of large outer time steps while still obtaining accurate sampling and minimizing the perturbation of the dynamics. Furthermore, this approach is shown to be comparable to constraining fast motions, thus providing an alternative to molecular dynamics with constraints.Comment: accepted for publication by the Journal of Chemical Physic

The Mechanism of DNA Sensing by AIM2-Like Receptors

AIM2-Like Receptors are a family of nuclear and cytosolic foreign-DNA sensors consisting of an N-terminal PYD and one or two C-terminal DNA-binding HIN200 domains. AIM2 is a cytosolic sensor which forms a supramolecular structure known as the inflammasome, while IFI16 resides in both the nucleus and cytoplasm, where it can associate with ASC and form its own inflammasome, as well as lead to activation of the interferon pathway by parallel means. Persistent questions in innate immunology remain of how such sensors respond to foreign DNA while remaining silent towards host DNA, as well as their role in activating downstream effectors. Using fluorescence anisotropy, FRET, EMSA, and various mutational studies, it is shown here that the PYD of IFI16 plays a positive, cooperative role in DNA binding, allowing it to oligomerize in a length-dependent manner. Disruption of the non-DNA-binding PYD in turn disrupts DNA binding, demonstrating that oligomerization and DNA binding are coupled events. By fluorescence anisotropy, FRET, EMSA, electron microscopy, and mutagenesis studies, a model which doesn’t invoke the previous autoinhibition is put forth to explain the behavior and activation of AIM2. Results also give clues into how AIM2 may then recruit the next member of the downstream pathway, ASC. Finally, using time-dependent FRET assays as well as competition anisotropy experiments, it is demonstrated that nucleosomes act as effective barriers to IFI16 DNA binding and oligomerization. The length-dependent rates of FRET signal also support a model in which IFI16 uses one-dimensional diffusion as an efficient means of oligomerization. Taken together, these results show that the AIM2-Like Receptor family share many common characteristics that give insight into their roles in host defense and autoimmune disorders

TEST OF PHYSIOLOGICAL PERFORMANCE: RATIONALE AND FEASIBILITY

Rigorous clinical evaluation of the physiological performance is currently performed with complex and long procedures which need expensive technology and skilled operators. In a wide range of situations (frail patients, daily clinical practice, etc.), these approaches are difficult to be applied and simpler tests, with a lack of scientific background, are mandatory. To avoid these problems, we propose a test (test of physiological performance (TOPP)) to evaluate the physiological behavior of a subject, in a really easy and safe clinical setting, measuring only the heart rate. The subject is submitted to an active standing-up test and then two submaximal exercises (with a low power load) on a cycle-ergometer. The heart rate modifications due to each submaximal step are analyzed by exponential interpolation to calculate the ascending and descending time constants and evaluate the way each subject adapts his heart rate to work. The standard deviation of the RR for each stationary phase (warm-up, load, recovery) was calculated as an index of short-term variability. Then a standard Fourier analysis of the stationary periods of the standing-up procedures allows to quickly and easily evaluate the autonomic nervous activation. We tested the protocol on five healthy subjects to verify the feasibility and the acceptance of the procedure. The five subjects demonstrated a good tolerance of the entire procedure. The standing-up showed a behavior of the autonomic system consistent with the physiology (with an increase in sympathetic activation in the passage to standing position). The analysis of the two submaximal steps highlights how younger and trained subjects present lower heart rates (both in the ascending phase and in the recovery) with a quicker adaptation ability (smaller time constants) consistent with what is expected. The short-term variability of heart rate is greater in young and trained subjects, thus confirming how the sympatho-vagal balance, in these subjects, is more dynamic. The proposed test is well tolerated by the subjects and the results, albeit in a small cohort of healthy volunteers, are consistent with what is expected from physiology and is already present in the literature. Our work aims to be a proposal with a feasibility check of a method for evaluating performance. The work to be done for the clinical validation of the TOPP is still long, but we are aware that it can give important results and that the TOPP can become an effective tool for the assessment of the physiological performance even of fragile subjects

The Epigenetics of the Endocannabinoid System

The endocannabinoid system (ES) is a cell-signalling system widely distributed in biological tissues that includes endogenous ligands, receptors, and biosynthetic and hydrolysing machineries. The impairment of the ES has been associated to several pathological conditions like behavioural, neurological, or metabolic disorders and infertility, suggesting that the modulation of this system may be critical for the maintenance of health status and disease treatment. Lifestyle and environmental factors can exert long-term effects on gene expression without any change in the nucleotide sequence of DNA, affecting health maintenance and influencing both disease load and resistance. This potentially reversible "epigenetic" modulation of gene expression occurs through the chemical modification of DNA and histone protein tails or the specific production of regulatory non-coding RNA (ncRNA). Recent findings demonstrate the epigenetic modulation of the ES in biological tissues; in the same way, endocannabinoids, phytocannabinoids, and cannabinoid receptor agonists and antagonists induce widespread or gene-specific epigenetic changes with the possibility of trans-generational epigenetic inheritance in the offspring explained by the transmission of deregulated epigenetic marks in the gametes. Therefore, this review provides an update on the epigenetics of the ES, with particular attention on the emerging role in reproduction and fertility

Differential effects on membrane permeability and viability of human keratinocyte cells undergoing very low intensity megasonic fields

Among different therapeutic applications of Ultrasound (US), transient membrane sonoporation (SP) - a temporary, non-lethal porosity, mechanically induced in cell membranes through US exposure - represents a compelling opportunity towards an efficient and safe drug delivery. Nevertheless, progresses in this field have been limited by an insufficient understanding of the potential cytotoxic effects of US related to the failure of the cellular repair and to the possible activation of inflammatory pathway. In this framework we studied the in vitro effects of very low-intensity US on a human keratinocyte cell line, which represents an ideal model system of skin protective barrier cells which are the first to be involved during medical US treatments. Bioeffects linked to US application at 1 MHz varying the exposure parameters were investigated by fluorescence microscopy and fluorescence activated cell sorting. Our results indicate that keratinocytes undergoing low US doses can uptake drug model molecules with size and efficiency which depend on exposure parameters. According to sub-cavitation SP models, we have identified the range of doses triggering transient membrane SP, actually with negligible biological damage. By increasing US doses we observed a reduced cells viability and an inflammatory gene overexpression enlightening novel healthy relevant strategies

Ultrasound delivery of Surface Enhanced InfraRed Absorption active gold-nanoprobes into fibroblast cells: a biological study via Synchrotron-based InfraRed microanalysis at single cell level

Ultrasound (US) induced transient membrane permeabilisation has emerged as a hugely promising tool for the delivery of exogenous vectors through the cytoplasmic membrane, paving the way to the design of novel anticancer strategies by targeting functional nanomaterials to specific biological sites. An essential step towards this end is the detailed recognition of suitably marked nanoparticles in sonoporated cells and the investigation of the potential related biological effects. By taking advantage of Synchrotron Radiation fourier transform infrared micro-spectroscopy (SR-microftiR) in providing highly sensitive analysis at the single cell level, we studied the internalisation of a nanoprobe within fibroblasts (NIH-3T3) promoted by low-intensity US. To this aim we employed 20 nm gold nanoparticles conjugated with the IR marker 4-aminothiophenol. The significant Surface Enhanced Infrared Absorption provided by the nanoprobes, with an absorbance increase up to two orders of magnitude, allowed us to efficiently recognise their inclusion within cells. Notably, the selective and stable SR- microftiR detection from single cells that have internalised the nanoprobe exhibited clear changes in both shape and intensity of the spectral profile, highlighting the occurrence of biological effects. Flow cytometry, immunofluorescence and murine cytokinesis-block micronucleus assays confirmed the presence of slight but significant cytotoxic and genotoxic events associated with the US-nanoprobe combined treatments. our results can provide novel hints towards US and nanomedicine combined strategies for cell spectral imaging as well as drug delivery-based therapies

Spatiotopic selectivity of adaptation-based compression of event duration

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Unequal access to testing and vaccination services for the homeless and undocumented population during the COVID-19 pandemic

Objectives: To furnish a model to ensure access and use of healthcare services to the undocumented and homeless population. Methods: Between March 2020 and October 2021, public and third sector actors in Rome implemented an accessible COVID-19 screening service and vaccination program targeting the homeless and undocumented population. Results: 95.6% of the patients tested negative to both rapid and molecular tests. 0.9% tested positive to both. 0.7% were false negatives, while 2.8% were false positives. None of the participants refused the diagnostic treatment. From July to October 2021, 1384 people received a complete cycle of the COVID-19 vaccine through the program. 632 (45.6%) also agreed to perform the antibodies testing before inoculation. 318 (50.31%) of these were positive at the time of vaccination. Conclusion: We present a cost-effective model for reducing structural barriers to access diagnostic and preventive services for the homeless and undocumented population that can be applied to different public health settings