MLC1 is a membrane protein highly expressed in brain perivascular astrocytes and whose mutations account for the rare leukodystrophy (LD) megalencephalic leukoencephalopathy with subcortical cysts ...disease (MLC). MLC is characterized by macrocephaly, brain edema and cysts, myelin vacuolation and astrocyte swelling which cause cognitive and motor dysfunctions and epilepsy. In cultured astrocytes, lack of functional MLC1 disturbs cell volume regulation by affecting anion channel (VRAC) currents and the consequent regulatory volume decrease (RVD) occurring in response to osmotic changes. Moreover, MLC1 represses intracellular signaling molecules (EGFR, ERK1/2, NF-kB) inducing astrocyte activation and swelling following brain insults. Nevertheless, to date, MLC1 proper function and MLC molecular pathogenesis are still elusive. We recently reported that in astrocytes MLC1 phosphorylation by the Ca
/Calmodulin-dependent kinase II (CaMKII) in response to intracellular Ca
release potentiates MLC1 activation of VRAC. These results highlighted the importance of Ca
signaling in the regulation of MLC1 functions, prompting us to further investigate the relationships between intracellular Ca
and MLC1 properties.
We used U251 astrocytoma cells stably expressing wild-type (WT) or mutated MLC1, primary mouse astrocytes and mouse brain tissue, and applied biochemistry, molecular biology, video imaging and electrophysiology techniques.
We revealed that WT but not mutant MLC1 oligomerization and trafficking to the astrocyte plasma membrane is favored by Ca
release from endoplasmic reticulum (ER) but not by capacitive Ca
entry in response to ER depletion. We also clarified the molecular events underlining MLC1 response to cytoplasmic Ca
increase, demonstrating that, following Ca
release, MLC1 binds the Ca
effector protein calmodulin (CaM) at the carboxyl terminal where a CaM binding sequence was identified. Using a CaM inhibitor and generating U251 cells expressing MLC1 with CaM binding site mutations, we found that CaM regulates MLC1 assembly, trafficking and function, being RVD and MLC-linked signaling molecules abnormally regulated in these latter cells.
Overall, we qualified MLC1 as a Ca
sensitive protein involved in the control of volume changes in response to ER Ca
release and astrocyte activation. These findings provide new insights for the comprehension of the molecular mechanisms responsible for the myelin degeneration occurring in MLC and other LD where astrocytes have a primary role in the pathological process.
Objectives The increased incidence of nosocomial infections by multidrug-resistant organisms has motivated the re-introduction of colistin in combination with other antimicrobials in the treatment of ...infections. We describe the clinical and microbiological outcomes of patients infected with multidrug-resistant Acinetobacter baumannii who were treated with a combination of colistin and rifampicin. Patients and methods Critically ill patients with pneumonia and bacteraemia caused by A. baumannii resistant to all antibiotics except colistin in medical and surgical intensive care units were enrolled. Clinical and microbiological responses and safety were evaluated. Results Twenty-nine patients (47 ± 14 years and APACHE II score 17.03 ± 3.68), of whom 19 were cases of nosocomial pneumonia and 10 were cases of bacteraemia, were treated with intravenous colistin sulphomethate sodium (2 million IU three times a day) in addition to intravenous rifampicin (10 mg/kg every 12 h). All A. baumannii isolates were susceptible to colistin. The mean duration of treatment with intravenous colistin and rifampicin was 17.7 (±10.4) days (range 7–36). Clinical and microbiological responses were observed in 22 of 29 cases (76%) and the overall infection-related mortality was 21% (6/29). Three of the 29 evaluated patients (10%) developed nephrotoxicity when treated with colistin, all of whom had previous renal failure. No cases of renal failure were observed among patients with normal baseline renal function. No neurotoxicity was noted. Conclusions Colistin and rifampicin appears to be an effective and safe combination therapy for severe infections due to multidrug-resistant A. baumannii.
We retrospectively studied patients diagnosed with P. aeruginosa bloodstream infections (BSIs) in two Italian university hospitals. Risk factors for the isolation of multidrug-resistant (MDR) or ...non-MDR P. aeruginosa in blood cultures were identified by a case-case-control study, and a cohort study evaluated the clinical outcomes of such infections. We identified 106 patients with P. aeruginosa BSI over the 2-year study period; 40 cases with MDR P. aeruginosa and 66 cases with non-MDR P. aeruginosa were compared to 212 controls. Independent risk factors for the isolation of MDR P. aeruginosa were: presence of central venous catheter (CVC), previous antibiotic therapy, and corticosteroid therapy. Independent risk factors for non-MDR P. aeruginosa were: previous BSI, neutrophil count <500/mm3, urinary catheterization, and presence of CVC. The 21-day mortality rate of all patients was 33·9%. The variables independently associated with 21-day mortality were presentation with septic shock, infection due to MDR P. aeruginosa, and inadequate initial antimicrobial therapy.
Staphylococcus aureus bacteraemia (SAB) is a leading cause of mortality and morbidity in both nosocomial and community settings. The objective of the study is to explore epidemiological ...characteristics and predisposing risk factors associated with healthcare-associated (HCA) and community-acquired (CA) SAB, and to evaluate any differences in mortality and efficacy of initial antimicrobial therapy on treatment outcome. We conducted a two-part analysis. First, a triple case-control study in which groups of HCA SAB with onset ≥48 h after hospital admission (HCA ≥48 h), HCA SAB with onset <48 h of hospital admission (HCA <48 h), and CA SAB were compared with controls. Second, a cohort study including all patients with SAB was performed to identify factors associated with in-hospital mortality. SAB was diagnosed in 165 patients over the study period (January 2007 to December 2007). Five variables were independently associated with HCA ≥48 h SAB: presence of central venous catheter, solid tumour, chronic renal failure, previous hospitalization and previous antibiotic therapy. Significant risk factors for HCA <48 h SAB were: Charlson Comorbidity Index ≥3, previous hospitalization, living in long-term care facilities and corticosteroid therapy. Factors independently associated with CA SAB were: diabetes mellitus, HIV infection and chronic live disease. Patients with HCA <48 h SAB were significantly more likely to receive initial inadequate antimicrobial treatment than patients with CA or HCA ≥48 h SAB (44.8% versus 33.3% and 31.5%, respectively). Logistic-regression analysis identified three variables as independent predictors of mortality: presentation with septic shock, infection with methicillin-resistant S. aureus, and initial inadequate antimicrobial treatment. More than half of patients with SAB have MRSA strains and presentation with septic shock, and inappropriate empirical therapy was associated with increased mortality.
Background and Purpose
Intrathecally (i.t.) administered nociceptin/orphanin FQ (N/OFQ) evokes antinociceptive effects in rodents. Recent studies in monkeys demonstrated that i.t. co‐application of ...N/OFQ and morphine elicits synergistic antinociceptive actions suggesting mixed N/OFQ peptide (NOP) and μ opioid receptor agonists as innovative spinal analgesics. Thus, novel N/OFQ related peptides were synthesized in order to identify and pharmacologically characterize a mixed NOP/ μ opioid receptor agonist.
Experimental Approach
The following in vitro assays were used: calcium mobilization in cells expressing the human NOP or classical opioid receptors and chimeric G proteins, receptor and 35S‐GTPγS binding, 35S‐GTPγS binding in rat spinal cord membranes, guinea pig ileum bioassay. In vivo experiments were performed in monkeys using the tail withdrawal assay.
Key Results
From calcium mobilization studies Dmt1N/OFQ(1–13)‐NH2 was selected as the most potent and least selective compound. The mixed NOP/opioid full agonist activity and high affinity of Dmt1N/OFQ(1–13)‐NH2 was confirmed at human recombinant receptors in receptor binding, calcium mobilization and/or 35S‐GTPγS binding studies, at rat spinal cord receptors in 35S‐GTPγS binding experiments, and at guinea pig receptors inhibiting neurogenic contractions in the ileum. In vivo in the tail withdrawal assay in monkeys i.t. Dmt1N/OFQ(1–13)‐NH2 was able to elicit robust and long‐lasting antinociceptive effects.
Conclusions and Implications
Collectively, these results demonstrate that Dmt1N/OFQ(1–13)‐NH2 behaves as NOP/opioid receptor universal agonist and substantiate the suggestion that such mixed ligands are worthy of development as innovative spinal analgesics.
The Eurasian lynx is of special conservation concern based on the European Union's Habitat Directive and its populations need to be maintained or restored at favourable conservation status. To ...evaluate lynx population status, appropriate monitoring needs to be in place. We modelled the distribution dynamics of lynx in the Alps (200 000 km2) during 1994–2014 at a resolution of 100 km2. Lynx distribution and detection probability varied by year, country, forest cover, elevation and distance to the nearest release site. Occupancy of neighbouring quadrats had a strong positive effect on colonization and persistence rates. Our analyses demonstrate the importance of accounting for imperfect detection: the raw data underestimated the lynx range by 55% on average, depending on country and winter. Over the past 20 years the Alpine lynx range has expanded at an average rate of 4% per year, which was partly due to the lynx translocations to new areas. Our approach to large‐scale distribution modelling and analysing trends using site occupancy models can be applied retrospectively and is useful in many cases where a network of trained people is established to report the presence of target species, for example, in Europe where member states of the European Union have to report conservation status of species of community interest. Hence, dynamic occupancy models are an appealing framework for inference about the large‐scale range dynamics based on opportunistic data and a useful tool for large‐scale management and conservation programmes.
We analysed range dynamics of a reintroduced large carnivore, the Eurasian lynx, in the Alps (200 000 km2) over 20 years, combining a cutting edge occupancy model with citizen science. Lynx distribution and detection probability varied by distance to the nearest release site, year, forest cover, elevation and country. Occupancy of neighbouring quadrats had a strong positive effect on colonization and persistence rates. Our analyses demonstrate the importance of accounting for imperfect detection: the raw data underestimated the lynx range by 55% on average. Over the past 20 years the Alpine lynx range expanded at an average rate of 4% per year, which was partly due to the lynx translocations to new areas. Our approach to large‐scale distribution modelling and analysing trends using site occupancy models can be applied retrospectively and is useful in many cases where a network of trained people is established to report the presence of target species.
Inferring the distribution and abundance of a species from field records must deal with false‐negative and false‐positive errors. False‐negative errors occur if a species present goes undetected, ...while false‐positive errors are typically a consequence of species misidentification. False‐positive observations in studies of rare species may cause an overestimation of the distribution or abundance of the species and distort trend indices. We illustrate this issue with the monitoring of the Eurasian lynx in the Alps. We developed a three‐level classification of field records according to their reliability as inferred from whether they were validated or not. The first category (C1) represents ‘hard fact’ data (e.g. dead lynx); the second category (C2) includes confirmed data (e.g. tracks verified by an expert); and the third category (C3) are unconfirmed data (e.g. any kind of direct visual observation). For lynx, which is a comparatively well‐known species in the Alps, we use site‐occupancy modelling to estimate its distribution and show that the inferred lynx distribution is highly sensitive to presence sign category: it is larger if based on C3 records compared with the more reliable C1 and C2 records. We believe that the reason for this is a fairly high frequency of false‐positive errors among C3 records. This suggests that distribution records for many lesser‐known species may be similarly unreliable, because they are mostly or exclusively based on unconfirmed and thus soft data. Nevertheless, such soft data form a considerable part of species assessments as presented, for example in the International Union for Conservation of Nature Red List. However, C3 records can often not be discarded because they may be the only information available. When inferring the distribution of rare carnivores, especially for species with an expanding or shrinking range, we recommend a rigorous discrimination between fully reliable and un‐ or only partly reliable data, in order to identify possible methodological problems in the distribution maps related to false‐positive records.
Highlights • We used baculovirus and inactivated FMDV (iFMDV) to formulate vaccines. • We tested such vaccines in the murine model. • Co-inoculation of Bv with iFMDV protects against viral challenge ...at 2 dpv. • Such protection can be maintained using iFMDV oil vaccine up to 14 dpv.
Supplementing kindergarten children during a cold season with a prebiotic inulin-type fructans product with shorter and longer fructan chains has been shown to reduce febrile episodes requiring ...medical attention and to lower the incidence of sinusitis. These beneficial effects may be connected to the specific modulation of children’s gut microbiota. By applying quantitative and qualitative microbiota analysis this study aimed at characterising the gut microbiota composition and at exploring effects of prebiotic intervention on the gut microbiota during a 24-weeks intervention and during antibiotic treatment in healthy children. The study was a randomised, placebo-controlled trial with 258 healthy children aged 3 to 6 years consuming 6 g/day prebiotic inulin-type fructans or maltodextrin. During the course of the study, faecal samples were collected and subject to targeted qPCR analysis and phylogenetic profiling by multiplexed high throughput sequencing of the prokaryotic 16S rRNA gene PCR amplicons. The microbiota composition of the cohort could be clustered into three distinct constellations (enterotypes). Prebiotic intake resulted in a selective modulation of the gut microbiota composition. Relative abundance of Bifidobacterium was significantly higher in the prebiotic group (n=104) compared to control group (n=105) and this effect was found for all three enterotypes. Antibiotic administration decreased the relative abundance of Bifidobacterium in both groups. Nonetheless, children of the prebiotic group receiving antibiotic treatment displayed significantly higher levels of Bifidobacterium than children receiving the placebo control. Prebiotic supplementation induced specific changes in the gut microbiota composition of children aged 3 to 6 years. Moreover, it attenuated antibiotic-induced disturbances in the gut microbiota composition as shown by higher relative abundance of bifidobacteria at the end of the antibiotic treatment in the prebiotic group. With the previously reported benefits on immune function, the study contributes to the evidence on the immune-modulating effects of prebiotics through gut microbiota modifications. The study was registered as NCT03241355 (https://clinicaltrials.gov/show/NCT03241355).
PDF
![[Dmt1]N/OFQ(1–13)‐NH2: a po...](http://api.summon.serialssolutions.com/2.0.0/image/issn/XR4PS5YY4K/0007-1188_1476-5381/small "[Dmt1]N/OFQ(1–13)‐NH2: a potent nociceptin/orphanin FQ and opioid receptor universal agonist")
