Strong protection induced by an experimental DIVA subunit vaccine against bluetongue virus serotype 8 in cattle

Bluetongue virus (BTV) infections in ruminants pose a permanent agricultural threat since new serotypes are constantly emerging in new locations. Clinical disease is mainly observed in sheep, but cattle were unusually affected during an outbreak of BTV seroype 8 (BTV-8) in Europe. We previously developed an experimental vaccine based on recombinant viral protein 2 (VP2) of BTV-8 and non-structural proteins 1 (NS1) and NS2 of BTV-2, mixed with an immunostimulating complex (ISCOM)–matrix adjuvant. We demonstrated that bovine immune responses induced by this vaccine were as good or superior to those induced by a classic commercial inactivated vaccine. In this study, we evaluated the protective efficacy of the experimental vaccine in cattle and, based on the detection of VP7 antibodies, assessed its DIVA compliancy following virus challenge. Two groups of BTV-seronegative calves were subcutaneously immunized twice at a 3-week interval with the subunit vaccine (n = 6) or with adjuvant alone (n = 6). Following BTV-8 challenge 3 weeks after second immunization, controls developed viremia and fever associated with other mild clinical signs of bluetongue disease, whereas vaccinated animals were clinically and virologically protected. The vaccine-induced protection was likely mediated by high virus-neutralizing antibody titers directed against VP2 and perhaps by cellular responses to NS1 and NS2. T lymphocyte responses were cross-reactive between BTV-2 and BTV-8, suggesting that NS1 and NS2 may provide the basis of an adaptable vaccine that can be varied by using VP2 of different serotypes. The detection of different levels of VP7 antibodies in vaccinated animals and controls after challenge suggested a compliancy between the vaccine and the DIVA companion test. This BTV subunit vaccine is a promising candidate that should be further evaluated and developed to protect against different serotypes.     

Sensitivity to sensitins and tuberculin in Swedish children. II. A study of preschool children

Non-BCG-vaccinated preschool children (4 or 5 years of age) were simultaneously tested on separate arms with a 2 IU PPD RT23 and 0.1 microgram Mycobacterium avium sensitin RS10 or 0.1 microgram Mycobacterium scrofulaceum sensitin RS95. None of the 762 children had any known exposure to tuberculosis. A total of 8.8% reacted with an induration (greater than or equal to 3 mm to PPD RT23 while 2% reacted with greater than or equal to 6 mm. Half the children were tested with M. avium sensitin: 18.9 and 7.8% reacted when 3 and 6 mm cut-off points, respectively, were taken. The remaining children were tested with M. scrofulaceum sensitin: 18.4 and 6.3%, respectively, reacted. In a previous study of schoolchildren aged 8 or 9 years, reactions to sensitins were considerably more frequent. Thus, sensitisation by atypical mycobacteria seems to increase from the preschool to the early school age. This finding probably reflects a continuous exposure of the children to atypical mycobacteria from various sources. The preschool children with a reaction to PPD RT23 greater than or equal to 6 mm were examined and chest X-rays were performed. All children were healthy but one child had enlarged lymph nodes in the mediastinum and abdomen. It cannot be excluded that these pathological findings were caused by atypical mycobacteria.     

Respiratory failure in scoliosis and other thoracic deformities. A survey of patients with home oxygen or ventilator therapy in Sweden.

Registers covering Swedish patients with home ventilator or long-term oxygen therapy were used to study respiratory failure caused by thoracic deformities. In all, 107 patients were studied. Postpolio scoliosis was found in 47 patients. The age of starting therapy varied between 28 and 80 years. Fourteen patients had thoracic deformities other than scoliosis. The mean scoliotic angle was 135 degrees among the patients with scoliosis, and the mean vital capacity was 26% (range, 13-54%) of predicted normal. The yearly demand for home ventilator or oxygen therapy is calculated to three per million inhabitants. No operated patients had respiratory failure, and no patients were found with idiopathic scoliosis and respiratory failure younger than 30 years of age, which may indicate a preventive effect of corrective surgery on the development of respiratory failure.     

Nandrolone decanoate has long-term effects on dominance in a competitive situation in male rats

The aim of the present study was to examine possible long-term effects of the anabolic androgenic steroid (AAS), nandrolone decanoate (ND), on dominance in a provoking and competitive situation in sexually matured male rats. The experimental group (n=10) received daily injections of ND [15 mg/kg in a volume of 1 ml/kg subcutaneous (s.c.) injection for 14 days]. During the corresponding period, the controls (n=10) were given daily injections of an oil vehicle (1 ml/kg s.c.). All animals were tested in a competitive situation at four occasions after the end of the treatment period (week 5, 8, 11 and 14). Water-deprived pairs of rats, consisting of one ND-treated rat and one control, had to compete for access to water. The results showed that the ND-treated rats approached the water spout significantly more often compared to the controls. During the competition tests, the ND-treated rats spent more time drinking, an effect that was prominent for 11 weeks after the end of the treatment period. The ND-treated rats also displayed more frequently piloerection than the controls. The results indicate that ND has long-term effect on dominance in a provoking and competitive situation.     

Ventilatory response to CO2 re-breathing before and after nocturnal nasal intermittent positive pressure ventilation in patients with chronic alveolar hypoventilation

Long-term nocturnal nasal intermittent positive pressure ventilation (NIPPV) has beneficial effects on daytime PaCO2 in patients with chronic alveolar hypoventilation. Our aim was to investigate if these beneficial effects are related to improved respiratory drive as measured by ventilatory response to CO2. In 17 hypoventilated patients (mean age 62 years) we obtained daytime arterial blood gases, nocturnal transcutaneous oxygen saturation, nocturnal transcutaneous PaCO2 ventilatory response to CO2 re-breathing, spirometry and indices of respiratory muscle strength before and after 9 months of NIPPV. Patients served as their own controls. After 9 months of NIPPV day-time PaCO2 decreased from 7.1 kPa to 6.3 kPa, (P<0.001) and PaO2 increased from 8.1 kPa to 9.3 kPa, (P<0.01). The changes in morning and daytime PaCO2 and in nocturnal transcutaneous oxygen saturation were significantly correlated to the changes in several variables derived from the ventilatory response to CO2 re-breathing. In patients with substantial improvement in daytime PaCO2 we found significant improvements in ventilatory response to CO2 re-breathing. The present study confirms the beneficial effect of long-term NIPPV on daytime arterial blood gases. The results are consistent with the hypothesis that the improvement of daytime PaCO2 is related to improved respiratory drive observed after NIPPV.     

An otopetrin family proton channel promotes cellular acid efflux critical for biomineralization in a marine calcifier

Otopetrins comprise a family of proton-selective channels that are critically important for the mineralization of otoliths and statoconia in vertebrates but whose underlying cellular mechanisms remain largely unknown. Here, we demonstrate that otopetrins are critically involved in the calcification process by providing an exit route for protons liberated by the formation of CaCO₃. Using the sea urchin larva, we examined the otopetrin ortholog otop2l, which is exclusively expressed in the calcifying primary mesenchymal cells (PMCs) that generate the calcitic larval skeleton. otop2l expression is stimulated during skeletogenesis, and knockdown of otop2l impairs spicule formation. Intracellular pH measurements demonstrated Zn²⁺-sensitive H⁺ fluxes in PMCs that regulate intracellular pH in a Na⁺/HCO₃⁻-independent manner, while Otop2l knockdown reduced membrane proton permeability. Furthermore, Otop2l displays unique features, including strong activation by high extracellular pH (>8.0) and check-valve–like outwardly rectifying H⁺ flux properties, making it into a cellular proton extrusion machine adapted to oceanic living conditions. Our results provide evidence that otopetrin family proton channels are a central component of the cellular pH regulatory machinery in biomineralizing cells. Their ubiquitous occurrence in calcifying systems across the animal kingdom suggest a conserved physiological function by mediating pH at the site of mineralization. This important role of otopetrin family proton channels has strong implications for our view on the cellular mechanisms of biomineralization and their response to changes in oceanic pH.

Otopetrins are a recently discovered family of proton channels (1, 2) that play a key role in the acid-sensing taste receptors (3, 4) and were demonstrated to be critically involved in statoconia and otolith formation of the mammalian vestibular organ and the fish inner ear (5, 6). Mutations and knockdown of otop1 resulted in agenesis of statoconia (5) and otoliths (6), while the physiological function of this proton channel in the CaCO₃ mineralization process remains obscure. Otopetrins are an evolutionary conserved family of proton channels found in animals ranging from basal metazoans to vertebrates and humans (1). In vertebrates, three otopetrin family members, Otop1 through Otop3, were identified that have a dimeric architecture with each subunit consisting of 12 transmembrane domains. Electrophysiological characterization of these proteins in heterologous expression systems demonstrated highly selective zinc-sensitive proton conductance (1).During the formation of CaCO₃, protons are liberated that need to be removed from the calcification front to protect cellular pH homeostasis and to promote further mineral precipitation (7–9). In this way, biomineralization and pH homeostasis are intrinsically linked processes that require efficient proton transport strategies. In the sea urchin larva, which develops an elaborate calcitic skeleton, amorphous CaCO₃ (ACC) is initially formed within intracellular compartments of primary mesenchyme cells (PMCs) that is then exocytosed to the growing skeleton (10, 11). During this process, 1.6 moles of protons are liberated per mole of CaCO₃ precipitated, leading to a substantial cellular acid load during the rapid early formation of the larval skeleton (8). To prevent an intracellular acidosis, PMCs must have outward directed proton fluxes that are comparable with those found in acid-secreting cells, including osteoclasts, renal tubular cells, or gastric parietal cells (9). Previous studies addressing pH regulatory mechanisms in PMCs demonstrated that the Slc4 family transporter Sp-Slc4a10 is critically involved in the cellular accumulation of dissolved inorganic carbon (i.e., HCO₃⁻) (12). In PMCs, bicarbonate has a dual function by serving as a calcification substrate and acting as a proton buffer in the cytosol (12). During active calcification, intracellular pH and HCO₃⁻ levels are increased, accompanied by an up-regulation of Sp-Slc4a10 on the protein and messenger RNA (mRNA) level (13). Interestingly, PMCs reduce Na⁺/H⁺ exchange capacities during this phase of active mineralization, suggesting an alternative route for the exit of proton across the plasma membrane in place. Although the involvement of vesicular proton accumulation by a V-type H⁺ ATPase has been suggested by pharmacological approaches, the direct export of protons from the cytosol across the plasma membrane remained obscure (13). A genome-wide analysis of the skeletogenic gene regulatory network demonstrated specific expression of the otop2l gene in PMCs of the sea urchin embryo (14). Otop2l is the single member of the otopetrin family in the sea urchin located downstream of ALX1 and Ets1 in the PMC gene regulatory network (14). Given their deep phylogenetic origin and their ubiquitous association with calcifying systems, the physiological characterization of otopetrins in mineralizing cells can provide fundamental knowledge about the mechanisms of biomineralization.In this work, we address the cellular expression and localization of Otop2l in PMCs by a suite of molecular and biochemical techniques. Using Otop2l morphants, we test the contribution of this channel to the calcification process. The physiological function of Otop2l in PMCs was investigated using intracellular pH recordings to characterize cellular proton fluxes in combination with proton channel inhibitors and knockdown of otop2l. Finally, heterologous expression and recalcification experiments are used to demonstrate that otop2l encodes a proton channel that supports the mineralization processes, depending on transmembrane proton gradients.     

Qualify of life and palliation predict survival in patients with chronic alveolar hypoventilation and nocturnal ventilatory support

Objectives  Non-invasive positive pressure ventilation (NPPV) improves health-related quality of life (HRQL) in patients with chronic alveolar hypoventilation (CAH). We studied the prognostic impact of HRQL on survival in relation to clinical factors. Patients  Forty-four patients with CAH due to post-polio (12), scoliosis (11), post-tb (17) or other diagnoses (4) who received nocturnal NPPV were prospectively studied during 6–10 years. Measurements  Blood gases and HRQL were analysed at baseline and after 9 months and after 8 years. HRQL was evaluated with measures of functioning (SIP), emotional well-being (HADS and MACL), and global QL. Results  Blood gases and HRQL measures improved during NPPV. The overall 5-year survival rate was 73%. In multivariate survival analysis, a diagnosis of post-polio and low baseline SIP physical index scores, indicating low levels of physical dysfunction, predicted longer survival (P = 0.02, respectively). Similarly, palliation of physical dysfunction and preserved or improved global QL by 9 months were associated with longer overall survival (P = 0.009 and P = 0.001, respectively; multivariate Cox regression). Conclusion  Seventy-three percent of patients treated for CAH with NPPV survived more than 5 years. Diagnosis and self-rated physical functioning at pre-treatment were related to survival, as were major improvements in physical functioning and global QL during NPPV.