Central Charged Residues in DIIIS4 Regulate Deactivation Gating in Skeletal Muscle Sodium Channels

Summary 1. Mutations in the S4 segment of domain III in the voltage gated skeletal muscle sodium channel hNa_V1.4 were constructed to test the roles of each charged residue in deactivation gating. Mutations comprised charge reversals at K1-R6, charge neutralization, and substitution at R4 and R5. 2. Charge-reversing mutations at R4 and R5 produced the greatest alteration of activation parameters compared to hNa_V1.4. Effects included depolarization of the conductance/voltage (g/V) curve, decreased valence and slowing of kinetics. 3. Reversal of charge at R2 to R4 hyperpolarized, and reversal at R5 or R6 depolarized the h _∞ curve. Most DIIIS4 mutations slowed inactivation from the open state. R4E slowed closed state fast inactivation and R5E inhibited its completion. 4. Deactivation from the open and/or inactivated state was prolonged in mutations reversing charge at R2 to R4 but accelerated by reversal of charge at R5 or R6. Effects were most pronounced at central charges R4 and R5. 5. Charge and structure each contribute to effects of mutations at R4 and R5 on channel gating. Effects of mutations on activation and deactivation at R4 and, to a lesser extent R5, were primarily owing to charge alteration, whereas effects on fast inactivation were charge independent.     

Novel probiotic Bifidobacterium longum subsp. infantis CECT 7210 strain active against rotavirus infections

Rotavirus is the leading cause of severe acute gastroenteritis among children worldwide. It is well known that breast-feeding and vaccination afford infants protection. Since breast-feeding has drastically decreased in developed countries, efforts have been focused on the potential use of probiotics as preventive agents. In this study, a novel Bifidobacterium longum subsp. infantis strain was isolated from infant feces and selected, based on its capacity to inhibit in vitro rotavirus Wa replication (up to 36.05% infectious foci reduction) and also to protect cells from virus infection (up to 48.50% infectious foci reduction) in both MA-104 and HT-29 cell lines. Furthermore, studies using a BALB/c mouse model have proved that this strain provides preliminary in vivo protection against rotavirus infection. The strain has been deposited in the Spanish Type Culture Collection under the accession number CECT 7210. This novel strain has the main properties required of a probiotic, such as resistance to gastrointestinal juices, biliary salts, NaCl, and low pH, as well as adhesion to intestinal mucus and sensitivity to antibiotics. The food safety status has been confirmed by the absence of undesirable metabolite production and in acute ingestion studies of mice. Overall, these results demonstrate that Bifidobacterium longum subsp. infantis CECT 7210 can be considered a probiotic able to inhibit rotavirus infection.     

Chikungunya virus neutralization antigens and direct cell-to-cell transmission are revealed by human antibody-escape mutants

Chikungunya virus (CHIKV) is an alphavirus responsible for numerous epidemics throughout Africa and Asia, causing infectious arthritis and reportedly linked with fatal infections in newborns and elderly. Previous studies in animal models indicate that humoral immunity can protect against CHIKV infection, but despite the potential efficacy of B-cell-driven intervention strategies, there are no virus-specific vaccines or therapies currently available. In addition, CHIKV has been reported to elicit long-lasting virus-specific IgM in humans, and to establish long-term persistence in non-human primates, suggesting that the virus might evade immune defenses to establish chronic infections in man. However, the mechanisms of immune evasion potentially employed by CHIKV remain uncharacterized. We previously described two human monoclonal antibodies that potently neutralize CHIKV infection. In the current report, we have characterized CHIKV mutants that escape antibody-dependent neutralization to identify the CHIKV E2 domain B and fusion loop "groove" as the primary determinants of CHIKV interaction with these antibodies. Furthermore, for the first time, we have also demonstrated direct CHIKV cell-to-cell transmission, as a mechanism that involves the E2 domain A and that is associated with viral resistance to antibody-dependent neutralization. Identification of CHIKV sub-domains that are associated with human protective immunity, will pave the way for the development of CHIKV-specific sub-domain vaccination strategies. Moreover, the clear demonstration of CHIKV cell-to-cell transmission and its possible role in the establishment of CHIKV persistence, will also inform the development of future anti-viral interventions. These data shed new light on CHIKV-host interactions that will help to combat human CHIKV infection and inform future studies of CHIKV pathogenesis.     

Induction of antitumor immunity by indomethacin

Irradiated tumor cells given, together with indomethacin, to syngeneic mice induced an antitumor response and conferred protection against a challenge of a lethal dose of murine mammary (4T1) and lung (3LL) carcinoma cells. Continuous administration of indomethacin was crucial throughout the entire period of immunization and challenge, as no protection was achieved when the drug was given during only one of these procedures. Antitumor immunity was long-lasting and, when tested in the 4T1 model, 48% of mice were resistant to a second challenge of lethal tumor cells. Tumor-free immune mice that were given indomethacin for more than 300 days remained healthy with normal white blood cell counts and normal spleen size. Cells isolated from immune mice were able to kill tumor cells in culture after in vitro activation by interleukin-2, in a manner similar to cells from naive normal control mice. In addition, the mitogenic response of their T cells was as high as that of the control naive mice. While indomethacin was able to induce antitumor immunity to 4T1 and 3LL murine carcinoma cells, both of which contain a high concentration of endogenic prostaglandin E₂ (PGE2), no such immunity was achieved to murine tumor cells with a low concentration of endogenic PGE2. These results suggest a correlation between PGE2 concentration and the ability of indomethacin to induce antitumor immunity. We therefore suggest that an immunotherapy protocol with long-term dispensation of a tolerable dose of an immunomodulator, given together with irradiated autologous tumor cells, may stimulate antitumor responses to tumors containing high concentrations of endogenic PGE2. Received: 12 August 1999 / Accepted: 21 September 1999     

Increasing Exercise Intensity Reduces Heterogeneity of Glucose Uptake in Human Skeletal Muscles

Proper muscle activation is a key feature of survival in different tasks in daily life as well as sports performance, but can be impaired in elderly and in diseases. Therefore it is also clinically important to better understand the phenomenon that can be elucidated in humans non-invasively by positron emission tomography (PET) with measurements of spatial heterogeneity of glucose uptake within and among muscles during exercise. We studied six healthy young men during 35 minutes of cycling at relative intensities of 30% (low), 55% (moderate), and 75% (high) of maximal oxygen consumption on three separate days. Glucose uptake in the quadriceps femoris muscle group (QF), the main force producing muscle group in recreational cycling, and its four individual muscles, was directly measured using PET and 18F-fluoro-deoxy-glucose. Within-muscle heterogeneity was determined by calculating the coefficient of variance (CV) of glucose uptake in PET image voxels within the muscle of interest, and among-muscles heterogeneity of glucose uptake in QF was expressed as CV of the mean glucose uptake values of its separate muscles. With increasing intensity, within-muscle heterogeneity decreased in the entire QF as well as within its all four individual parts. Among-muscles glucose uptake heterogeneity also decreased with increasing intensity. However, mean glucose uptake was consistently lower and heterogeneity higher in rectus femoris muscle that is known to consist of the highest percentage of fast twitch type II fibers, compared to the other three QF muscles. In conclusion, these results show that in addition to increased contribution of distinct muscle parts, with increases in exercise intensity there is also an enhanced recruitment of muscle fibers within all of the four heads of QF, despite established differences in muscle-part specific fiber type distributions. Glucose uptake heterogeneity may serve as a useful non-invasive tool to elucidate muscle activation in aging and diseased populations.     

Age-Related Loss of Lumbar Spinal Lordosis and Mobility – A Study of 323 Asymptomatic Volunteers

Background

The understanding of the individual shape and mobility of the lumbar spine are key factors for the prevention and treatment of low back pain. The influence of age and sex on the total lumbar lordosis and the range of motion as well as on different lumbar sub-regions (lower, middle and upper lordosis) in asymptomatic subjects still merits discussion, since it is essential for patient-specific treatment and evidence-based distinction between painful degenerative pathologies and asymptomatic aging.

Methods and Findings

A novel non-invasive measuring system was used to assess the total and local lumbar shape and its mobility of 323 asymptomatic volunteers (age: 20–75 yrs; BMI <26.0 kg/m²; males/females: 139/184). The lumbar lordosis for standing and the range of motion for maximal upper body flexion (RoF) and extension (RoE) were determined. The total lordosis was significantly reduced by approximately 20%, the RoF by 12% and the RoE by 31% in the oldest (>50 yrs) compared to the youngest age cohort (20–29 yrs). Locally, these decreases mostly occurred in the middle part of the lordosis and less towards the lumbo-sacral and thoraco-lumbar transitions. The sex only affected the RoE.

Conclusions

During aging, the lower lumbar spine retains its lordosis and mobility, whereas the middle part flattens and becomes less mobile. These findings lay the ground for a better understanding of the incidence of level- and age-dependent spinal disorders, and may have important implications for the clinical long-term success of different surgical interventions.     

Nuclear autoantigenic sperm protein (NASP), a linker histone chaperone that is required for cell proliferation

A multichaperone nucleosome-remodeling complex that contains the H1 linker histone chaperone nuclear autoantigenic sperm protein (NASP) has recently been described. Linker histones (H1) are required for the proper completion of normal development, and NASP transports H1 histones into nuclei and exchanges H1 histones with DNA. Consequently, we investigated whether NASP is required for normal cell cycle progression and development. We now report that without sufficient NASP, HeLa cells and U2OS cells are unable to replicate their DNA and progress through the cell cycle and that the NASP(-/-) null mutation causes embryonic lethality. Although the null mutation NASP(-/-) caused embryonic lethality, null embryos survive until the blastocyst stage, which may be explained by the presence of stored NASP protein in the cytoplasm of oocytes. We conclude from this study that NASP and therefore the linker histones are key players in the assembly of chromatin after DNA replication.     

Functional assessment of cryopreserved pig aortas for pharmaceutical research

Several in vitro studies have demonstrated diminished post-thaw functional activity. Therefore, the aim of this study was to investigate the consequences of thawing and storage method used on the post-thaw functional activity of cryopreserved pig aortas with the aim of adjusting the freezing and thawing protocol so that the vascular segments are preserved in the best possible state, maintaining structure and functionality so that they can later be transplanted with success. In vitro responses of frozen, thawed pig aortas were used to investigate the functional activity after thawing at 15 degrees C and 100 degrees C/min and after storage in gas or liquid phase of liquid nitrogen. Cryopreservation was performed in RPMI 1640 medium + 10% dimethylsulfoxide and the rate of cooling was -1 degrees C/min, until -150 degrees C was reached.After thawing the maximal contractile responses to all the contracting agonists tested (KCl, noradrenaline) were in the ranges of 13-27% compared with the responses in unfrozen pig aortas. Contractile responses were slightly better when thawing was performed at 15 degrees C/min compared with 100 degrees C/min. The endothelium independent relaxant responses to sodium nitroprusside were reduced ( P < 0.05). Cryostorage of pig arteries also resulted in a loss of the endothelium-dependent relaxant response to acetylcholine. The cryopreservation method used provided a limited preservation of pig aorta contractibility, a reduction of the endothelium independent relaxant responses, and no apparent preservation of the endothelium-dependent relaxation. It is possible that further refinements of the cryopreservation protocol might allow better post-thaw functional recovery of pig aortas.     

Efectos del Modelo de Atención Centrado en la Persona en la calidad de vida de personas con deterioro cognitivo de centros gerontológicos

Introduction

The Model of Person Centered Care has attracted increasing interest for use in gerontology centers. Therefore, the contributions about its impact are scarce in our context. The objective of this paper is to establish the impact that the interventions associated with the Model of Person Centered Care in the «Etxean Ondo» Project have on the quality of life of residents with cognitive impairment.

Material and methods

One hundred and ninetten residents with cognitive impairment were selected: 59 in the control group and 60 in the experimental group. Subjects in each group were sorted by cognitive impairment: mild or severe. Changes were implemented in the physical and organizational environments for the promotion of autonomy and wellbeing. Quality of life was assessed before and 6 months after intervention using the Fumat Scales (mild cognitive impairment) and Qualid (severe cognitive impairment). The t-Student test was used for comparison of means.

Results

In intergroup comparisons, significant differences in the Fumat Scale for the control group with mild cognitive impairment were initially identified. These differences were not recorded in the post assessment. The experimental group with severe cognitive impairment was significantly improved in the Qualid Scale post assessment. In intragroup comparisons, significant improvements were evident in the quality of life of experimental subjects, both with severe cognitive impairment (Qualid) and mild (Fumat).

Conclusions

The findings support the effectiveness of the interventions and identify methodological and conceptual issues that have been considered to analyze the Model of Person Centered Care efects.