Cutting edge: precursor frequency affects the helper dependence of cytotoxic T cells.

Generation of CTL immunity often depends on the availability of CD4 T cell help. In this report, we show that CTL responses induced by cross-priming can be converted from CD4-dependent to CD4-independent by increasing the frequency of CTL precursors. In the absence of CD4 T cells, high numbers of CTL precursors were able to expand in number and become effector CTL. The ability of high frequencies of CD8 T cells to override help was not due to their ability to signal CD40 via expression of CD154. These findings suggest that when precursor frequencies are high, priming of CD8 T cell responses may not require CD4 T cell help.     

Oxidative metabolism of spironolactone: evidence for the involvement of electrophilic thiosteroid species in drug-mediated destruction of rat hepatic cytochrome P450

In a preliminary paper [Decker et al. (1986) Biochem. Biophys. Res. Commun. 136, 1162] we have shown that the antimineralocorticoid spironolactone (SPL) preferentially inactivates dexamethasone (DEX) inducible rat hepatic cytochrome P450p isozymes in a suicidal manner. These findings are now confirmed, and the kinetic characteristics of such a process are detailed. In an effort to elucidate the mechanism of SPL-mediated inactivation of cytochrome P450, we have examined the metabolism of SPL in vitro. Incubation of [14C]SPL and NADPH with liver microsomes prepared from DEX-pretreated rats results in the formation of several polar metabolites separable by HPLC with UV detection. This process is found to be dependent on NADPH, O2, SPL, and enzyme concentration, as well as temperature. Furthermore, metabolite formation was significantly attenuated by P450 inhibitors CO and n-octylamine. Mass spectral analysis (thermospray LC/MS, FAB/MS, and FAB/MS/MS) of the two most prominent polar metabolites indicated that these compounds had molecular weights that corresponded to the sulfinic and sulfonic acid derivatives of deacetyl-SPL (SPL-SH). These findings document the formation of previously unreported polar metabolites of SPL by rat liver microsomes enriched in cytochrome P450p and implicate a role for this isozyme in the oxidation of the thiol moiety of deacetyl-SPL. The detection of such metabolites also implicates a catalytic trajectory that includes the thiyl radical and/or sulfenic acid species as a plausible protagonist in drug-mediated inactivation of cytochrome P450p.     

Assessing the impact of episodic pollution

The increased tightening of controls on industrial and municipal wastewater discharges has resulted in steady improvements in the quality of many important rivers over recent years. However, episodic pollution, particularly from farm wastes and combined sewer overflows continues to pose a major problem, and is one of the main causes of poor quality rivers today. Despite our acknowledgement of this continuing problem, very little is known of the mechanistic basis of responses and recovery of aquatic organisms and communities exposed to intermittent pulses of common pollutants. The majority of ecotoxicological studies to date have been concerned with the effects of continuous exposure. Although such studies may provide a means of predicting the impact of episodic pollution events, a more appropriate test design would be to assess toxicity under pulsed and fluctuating exposure. Studies should also include a post-exposure observation period and should consider recovery of individuals and communities. This paper reviews the results of reported studies relevant to the investigation of episodic pollution and pays particular attention to the effects of magnitude, duration and frequency of exposure. Results of field investigations using an in situ bioassay technique are also presented to emphasize the importance of field validation of proposed water quality criteria for intermittent pollution events.     

Importance of fungi in the diet of Gammarus pulex and Asellus aquaticus

An important component of the interaction between macroinvertebrates and leaf litter in streams in the extent to which consumers can differentiate between undecomposed and decomposing leaves. The detritivores Gammarus pulex and Asellus aquaticus fed preferentially on conditioned rather on unconditioned leaf material. Growth in A. aquaticus was significantly reduced when unconditioned leaves were provided, but in G. pulex no significant effect of conditioning on growth was observed. The capacity of G. pulex to tolerate reductions in food quality seems to be a consequence of a compensatory system in which respiration rates change to compensate for reductions in food quality. In this way a constant growth rate is maintained. Increases in ingestion rates to compensate for low quality food were not observed.     

Ultrastructure of the Flame Bulbs of Urastoma cyprinae (Platyhelminthes, ‘Prolecithophora’, Urastomidae)

The ultrastructure of the flame bulbs of the turbellarian Urastoma cyprinae from Mytilus galloprovincialis in the Mediterranean is described. The nucleus of the terminal cell is located some distance basal to the rootlets of the cilia forming the flame; the cytoplasm contains numerous tubules approximately 54–66 nm in diameter, and vesicles. Thick walled, densely packed rod-like structures coil around each other with a tendency towards longitudinal orientation close to the flame. The rod-like structures tightly surround the basal part of the flame and the distal cytoplasmic tube in the apical part of the flame. Some of them, including the inner predominantly longitudinally directed ones, are continuous with the cytoplasm of the terminal cell, others are continuous with the cytoplasm of the distal cytoplasmic tube. Internal leptotriches arise from the cytoplasm of the terminal cell and intrude between the basal parts of the cilia of the flame. The distal cytoplasmic tube possesses a septate junction. The flame bulb of Urastoma differs distinctly from those known from other Platyhelminthes; implications for the phylogeny of Platyhelminthes are discussed.     

A cladistic analysis of the genera in the subfamily Pudicinae (Nematoda, Trichostrongyloidea, Heligmonellidae).

A parsimony analysis was performed on 37 specific taxa belonging to the subfamily Pudicinae (family Heligmonellidae), which contains parasites mainly from South American caviomorph rodents. Thirteen characters were used from the synlophe (rotation of axis, presence of carene, carene asymmetry, presence of comaretes, single ventral comarete length, ridge discontinuity, ventral ridge numbers, presence of a peculiar posterior synlophe, presence of supernumerary spines) and the male caudal bursa (relative length of rays 9 and 10, caudal bursa type, division of the dorsal ray, divergence of the 10th rays). The cladogram shows a consistency index of 1.0. The subfamily Pudicinae has two synapomorphies. Two suprageneric groups are recognized. Suprageneric group 1 shows one synapomorphy and contains Heligmostrongylus, Fuellebornema, Sciurodendrium and Pseudoheligmosomum; suprageneric group 2 shows two synapomorphies and contains Pudica, Acanthostrongylus, Justinema and Durettestrongylus. Five genera are defined on the basis of synapomorphies. The genera Heligmostrongylus, Sciurodendrium and Pudica which are considered paraphyletic, however, are retained due to lack of knowledge as to their relationships.     

High-dose versus standard-dose vitamin D supplementation in older adults with COVID-19 (COVIT-TRIAL): A multicenter,open-label,randomized controlled superiority trial

BackgroundVitamin D supplementation has been proposed as a treatment for Coronavirus Disease 2019 (COVID-19) based on experimental data and data from small and uncontrolled observational studies. The COvid19 and VITamin d TRIAL (COVIT-TRIAL) study was conducted to test whether a single oral high dose of cholecalciferol (vitamin D3) administered within 72 hours after the diagnosis of COVID-19 improves, compared to standard-dose cholecalciferol, the 14-day overall survival among at-risk older adults infected with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).Methods and findingsThis multicenter, randomized, controlled, open-label, superiority trial involved collaboration of 9 medical centers in France. Patients admitted to the hospital units or living in nursing homes adjacent to the investigator centers were eligible if they were ≥65 years, had SARS-CoV-2 infection of less than 3 days, and at least 1 COVID-19 worsening risk factor (among age ≥75 years, SpO2 ≤94%, or PaO₂/FiO₂ ≤300 mm Hg). Main noninclusion criteria were organ failure requiring ICU, SpO2 ≤92% despite 5 L/min oxygen, life expectancy <3 months, vitamin D supplementation >800 IU/day during the preceding month, and contraindications to vitamin D supplements. Eligible and consenting patients were randomly allocated to either a single oral high-dose (400,000 IU) or standard-dose (50,000 IU) cholecalciferol administered under medical supervision within 72 hours after the diagnosis of COVID-19. Participants and local study staff were not masked to the allocated treatment, but the Steering Committee and the Data and Safety Monitoring Board were masked to the randomization group and outcome data during the trial. The primary outcome was 14-day overall mortality. Between April 15 and December 17, 2020, of 1,207 patients who were assessed for eligibility in the COVIT-TRIAL study, 254 met eligibility criteria and formed the intention-to-treat population. The median age was 88 (IQR, 82 to 92) years, and 148 patients (58%) were women. Overall, 8 (6%) of 127 patients allocated to high-dose cholecalciferol, and 14 (11%) of 127 patients allocated to standard-dose cholecalciferol died within 14 days (adjusted hazard ratio = 0.39 [95% confidence interval [CI], 0.16 to 0.99], P = 0.049, after controlling for randomization strata [i.e., age, oxygen requirement, hospitalization, use of antibiotics, anti-infective drugs, and/or corticosteroids] and baseline imbalances in important prognostic factors [i.e., sex, ongoing cancers, profuse diarrhea, and delirium at baseline]). The number needed to treat for one person to benefit (NNTB) was 21 [NNTB 9 to ∞ to number needed to treat for one person to harm (NNTH) 46]. Apparent benefits were also found on 14-day mortality due to COVID-19 (7 (6%) deaths in high-dose group and 14 (11%) deaths in standard-dose group; adjusted hazard ratio = 0.33 [95% CI, 0.12 to 0.86], P = 0.02). The protective effect of the single oral high-dose administration was not sustained at 28 days (19 (15%) deaths in high-dose group and 21 (17%) deaths in standard-dose group; adjusted hazard ratio = 0.70 [95% CI, 0.36 to 1.36], P = 0.29). High-dose cholecalciferol did not result in more frequent adverse effects compared to the standard dose. The open-label design and limited study power are the main limitations of the study.ConclusionsIn this randomized controlled trial (RCT), we observed that the early administration of high-dose versus standard-dose vitamin D3 to at-risk older patients with COVID-19 improved overall mortality at day 14. The effect was no longer observed after 28 days.Trial registrationClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT04344041","term_id":"NCT04344041"}}NCT04344041.

In a randomized trial, Cedric Annweiler and colleagues evaluate whether a single high dose of vitamin D3 improves survival among older adults in France with SARS-CoV-2 infection.     

The Role of Microsite Conditions in Restoring Trembling Aspen (Populus tremuloides Michx) from Seed

Encouraging natural regeneration of Populus tremuloides Michx (trembling aspen) from seed is a largely unexplored means for reintroducing the species into reclamation areas. We evaluated the effects of microsite (surface contour and substrate type) on aspen seedling establishment and growth on a reclaimed coal mine. The 4.6 ha study site was divided into six 48 m‐wide strips that had 15 or 40 cm capping material salvaged from a nearby forest floor added to the mine surface. We surveyed 126 m long transects located in the center of each strip for microsite conditions, and the presence and height of aspen seedlings. We found that aspen seedlings generally preferred mineral‐organic substrates and concave microsites. To facilitate the regeneration of aspen by seed, we suggest that land managers increase small‐scale roughness and microtopographic diversity on reclaimed sites .     

Apposite insulin-like growth factor (IGF) receptor glycosylation is critical to the maintenance of vascular smooth muscle phenotype in the presence of factors promoting osteogenic differentiation and mineralization

Vascular calcification is strongly linked with increased morbidity and mortality from cardiovascular disease. Vascular calcification is an active cell-mediated process that involves the differentiation of vascular smooth muscle cells (VSMCs) to an osteoblast-like phenotype. Several inhibitors of this process have been identified, including insulin-like growth factor-I (IGF-I). In this study, we examined the role of the IGF receptor (IGFR) and the importance of IGFR glycosylation in the maintenance of the VSMC phenotype in the face of factors known to promote osteogenic conversion. IGF-I (25 ng/ml) significantly protected VSMCs from β-glycerophosphate-induced osteogenic differentiation (p < 0.005) and mineral deposition (p < 0.01). Mevalonic acid depletion (induced by 100 nm cerivastatin) significantly inhibited these IGF protective effects (p < 0.01). Mevalonic acid depletion impaired IGFR processing, decreased the expression of mature IGFRs at the cell surface, and inhibited the downstream activation of Akt and MAPK. Inhibitors of N-linked glycosylation (tunicamycin, deoxymannojirimycin, and deoxynojirimycin) also markedly attenuated the inhibitory effect of IGF-I on β-glycerophosphate-induced mineralization (p < 0.05) and activation of Akt and MAPK. These results demonstrate that alterations in the glycosylation of the IGFR disrupt the ability of IGF-I to protect against the osteogenic differentiation and mineralization of VSMCs by several interrelated mechanisms: decreased IGFR processing, reduced IGFR cell-surface expression, and reduced downstream signaling via the Akt and MAPK pathways. IGF-I thus occupies a critical position in the maintenance of normal VSMC phenotype and protection from factors known to stimulate vascular calcification.