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The reaction of phosphorus (P) between sediments and water in streams strongly affects the surface water P concentrations. A new reactive transport model (SWEMP: Sediment-Water Exchange Model for Phosphorus) was developed to describe redox dependent P sorption in the sediment and vertical diffusive transport of solutes to the overlying stream. The model parameters were independently obtained to first predict P release in ten different sediment-water batch systems and in two flumes. Input parameters are the degree of P saturation of the sediment, its organic matter content, dissolved oxygen (DO) concentration and temperature. The dissolved P concentrations in the overlying waters ranged from 0.02 to 1.2 mg P L -1 in these systems and were correctly predicted by the model within, on average, a factor 1.3 (batch) or 1.1 (flume). The P flux from the sediment towards the overlying water increased with increasing sediment P:Fe ratio and respiration rates, and with decreasing DO and water pH. After validation of the model with experimental data, it was used to predict monthly P concentrations in Flemish rivers using the total P emission data, total discharge, average sediment properties and the monthly averaged water temperatures, DO concentrations and electric conductivity. The monthly average P concentrations oscillate annually between 0.24 and 0.73 mg P L -1 and predictions matched the long-term monitoring data within 10 % using only one adjustable parameter for the entire water system (N > 250,000). The model predicts that summer peaks in P are related to internal loading from the sediment under anoxic conditions rather than to emission-dilution effects, i.e. external input of P and/or its concentration at lower flow rates. This suggests that, surface water P concentrations can be lowered by enhanced DO in the water, the addition of Fe and Al rich binding agents to the sediments and by reducing P emissions., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Toon van Dael reports financial support was provided by Research Foundation Flanders., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Since the onset of the coronavirus disease (COVID-19) pandemic in Belgium, UZ/KU Leuven has played a crucial role as the National Reference Centre (NRC) for respiratory pathogens, to be the first Belgian laboratory to develop and implement laboratory developed diagnostic assays for SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) and later to assess the quality of commercial kits. To meet the growing demand for decentralised testing, both clinical laboratories and government-supported high-throughput platforms were gradually deployed across Belgium. Consequently, the role of the NRC transitioned from a specialised testing laboratory to strengthening capacity and coordinating quality assurance. Here, we outline the measures taken by the NRC, the national public health institute Sciensano and the executing clinical laboratories to ensure effective quality management of molecular testing throughout the initial two years of the pandemic (March 2020 to March 2022)., (© 2024. The Author(s).)

Isolating and characterizing emerging SARS-CoV-2 variants is key to understanding virus pathogenesis. In this study, we isolated samples of the SARS-CoV-2 R.1 lineage, categorized as a variant under monitoring by the World Health Organization, and evaluated their sensitivity to neutralizing antibodies and type I interferons. We used convalescent serum samples from persons in Canada infected either with ancestral virus (wave 1) or the B.1.1.7 (Alpha) variant of concern (wave 3) for testing neutralization sensitivity. The R.1 isolates were potently neutralized by both the wave 1 and wave 3 convalescent serum samples, unlike the B.1.351 (Beta) variant of concern. Of note, the R.1 variant was significantly more resistant to type I interferons (IFN-α/β) than was the ancestral isolate. Our study demonstrates that the R.1 variant retained sensitivity to neutralizing antibodies but evolved resistance to type I interferons. This critical driving force will influence the trajectory of the pandemic.

Background: Hospital antibiograms guide initial empiric antibiotic treatment selections, but do not directly inform escalation of treatment among nonresponding patients., Methods: Using gram-negative bacteremia as an exemplar condition, we sought to introduce the concept of an escalation antibiogram. Among episodes of gram-negative bacteremia between 2017 and 2020 from 6 hospitals in the Greater Toronto Area, we generated escalation antibiograms for each of 12 commonly used agents. Among organisms resistant to that antibiotic, we calculated the likelihood of susceptibility to each of the other 11 agents. In subgroup analyses, we examined escalation antibiograms across study years, individual hospitals, community versus hospital onset, and pathogen type., Results: Among 6577 gram-negative bacteremia episodes, the likelihood of coverage was ampicillin 31.8%, cefazolin 62.7%, ceftriaxone 67.1%, piperacillin-tazobactam 72.5%, ceftazidime 74.1%, trimethoprim-sulfamethoxazole 74.4%, ciprofloxacin 77.1%, tobramycin 88.3%, gentamicin 88.8%, ertapenem 91.0%, amikacin 97.5%, and meropenem 98.2%. The escalation antibiograms revealed marked shifts in likelihood of coverage by the remaining 11 agents. For example, among ceftriaxone-resistant isolates, piperacillin-tazobactam susceptibility (21.2%) was significantly lower than trimethoprim-sulfamethoxazole (54.2%, P < .0001), ciprofloxacin (63.0%, P < .0001), ertapenem (73.4%, P < .0001), tobramycin (80.1%, P < .0001), gentamicin (82.8%, P < .0001), meropenem (94.3%, P < .0001), and amikacin (97.1%, P < .0001). Trimethoprim-sulfamethoxazole was the second-ranked agent in the meropenem escalation antibiogram (49.6%) and first in the amikacin escalation antibiogram (86.0%). Escalation antibiograms were consistent across 4 study years and 6 hospitals., Conclusions: Escalation antibiograms can be generated to inform empiric treatment changes in nonresponding patients. These tools can yield important insights such as avoiding the common maneuver of escalating from ceftriaxone to piperacillin-tazobactam in suspected gram-negative bacteremia., Competing Interests: Potential conflicts of interest. The authors: No reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest., (© The Author(s) 2022. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Background: Widespread testing for severe acute respiratory coronavirus virus 2 (SARS-CoV-2) is necessary to curb the spread of coronavirus disease 2019 (COVID-19), but testing is undermined when the only option is a nasopharyngeal swab. Self-collected swab techniques can overcome many of the disadvantages of a nasopharyngeal swab, but they require evaluation., Methods: Three self-collected non-nasopharyngeal swab techniques (saline gargle, oral swab and combined oral-anterior nasal swab) were compared to a nasopharyngeal swab for SARS-CoV-2 detection at multiple COVID-19 assessment centers in Toronto, Canada. The performance characteristics of each test were assessed., Results: The adjusted sensitivity of the saline gargle was 0.90 (95% CI 0.86-0.94), the oral swab was 0.82 (95% CI, 0.72-0.89) and the combined oral-anterior nasal swab was 0.87 (95% CI, 0.77-0.93) compared to a nasopharyngeal swab, which demonstrated a sensitivity of ˜90% when all positive tests were the reference standard. The median cycle threshold values for the SARS-CoV-2 E-gene for concordant and discordant saline gargle specimens were 17 and 31 (P < .001), for the oral swabs these values were 17 and 28 (P < .001), and for oral-anterior nasal swabs these values were 18 and 31 (P = .007)., Conclusions: Self-collected saline gargle and an oral-anterior nasal swab have a similar sensitivity to a nasopharyngeal swab for the detection of SARS-CoV-2. These alternative collection techniques are cheap and can eliminate barriers to testing, particularly in underserved populations.

Nasopharyngeal swabs are considered the preferential collection method for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnostics. Less invasive and simpler alternative sampling procedures, such as saliva collection, are desirable. We compared saliva specimens and nasopharyngeal (NP) swabs with respect to sensitivity in detecting SARS-CoV-2. A nasopharyngeal and two saliva specimens (collected by spitting or oral swabbing) were obtained from >2500 individuals. All samples were tested by RT-qPCR, detecting RNA of SARS-CoV-2. The test sensitivity was compared on the two saliva collections with the nasopharyngeal specimen for all subjects and stratified by symptom status and viral load. Of the 2850 patients for whom all three samples were available, 105 were positive on NP swab, whereas 32 and 23 were also positive on saliva spitting and saliva swabbing samples, respectively. The sensitivity of the RT-qPCR to detect SARS-CoV-2 among NP-positive patients was 30.5% (95% CI, 1.9%-40.2%) for saliva spitting and 21.9% (95% CI, 14.4%-31.0%) for saliva swabbing. However, when focusing on subjects with medium to high viral load, sensitivity on saliva increased substantially: 93.9% (95% CI, 79.8%-99.3%) and 76.9% (95% CI, 56.4%-91.0%) for spitting and swabbing, respectively, regardless of symptomatic status. Our results suggest that saliva cannot readily replace nasopharyngeal sampling for SARS-CoV-2 diagnostics but may enable identification of the most contagious cases with medium to high viral loads., (Copyright © 2021 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

In this controlled before-after study, wound swabs were only processed for culture, identification, and susceptibility testing if a quality metric, determined by the Q score, was met. Rejection of low-quality wound swabs resulted in a modest decrease in reflexive antibiotic initiation while reducing laboratory workload and generating few clinician requests., (© The Author(s) 2020. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Widely available and easily accessible testing for COVID-19 is a cornerstone of pandemic containment strategies. Nasopharyngeal swabs (NPS) are the currently accepted standard for sample collection but are limited by their need for collection devices and sampling by trained healthcare professionals. The aim of this study was to compare the performance of saliva to NPS in an outpatient setting. This was a prospective study conducted at three centers, which compared the performance of saliva and NPS samples collected at the time of assessment center visit. Samples were tested by real-time reverse transcription polymerase chain reaction and sensitivity and overall agreement determined between saliva and NPS. Clinical data was abstracted by chart review for select study participants. Of the 432 paired samples, 46 were positive for SARS-CoV-2, with seven discordant observed between the two sample types (four individuals testing positive only by NPS and three by saliva only). The observed agreement was 98.4% (kappa coefficient 0.91) and a composite reference standard demonstrated sensitivity of 0.91 and 0.93 for saliva and NPS samples, respectively. On average, the Ct values obtained from saliva as compared to NPS were higher by 2.76. This study demonstrates that saliva performs comparably to NPS for the detection of SARS-CoV-2. Saliva was simple to collect, did not require transport media, and could be tested with equipment readily available at most laboratories. The use of saliva as an acceptable alternative to NPS could support the use of widespread surveillance testing for SARS-CoV-2.

Identification of additional uses for existing drugs is a hot topic in drug discovery and a viable alternative to de novo drug development. HAMI3379 is known as an antagonist of the cysteinyl-leukotriene CysLT 2 receptor, and was initially developed to treat cardiovascular and inflammatory disorders. In our study we identified HAMI3379 as an antagonist of the orphan G protein-coupled receptor GPR17. HAMI3379 inhibits signaling of recombinant human, rat, and mouse GPR17 across various cellular backgrounds, and of endogenous GPR17 in primary rodent oligodendrocytes. GPR17 blockade by HAMI3379 enhanced maturation of primary rat and mouse oligodendrocytes, but was without effect in oligodendrocytes from GPR17 knockout mice. In human oligodendrocytes prepared from inducible pluripotent stem cells, GPR17 is expressed and its activation impaired oligodendrocyte differentiation. HAMI3379, conversely, efficiently favored human oligodendrocyte differentiation. We propose that HAMI3379 holds promise for pharmacological exploitation of orphan GPR17 to enhance regenerative strategies for the promotion of remyelination in patients., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Selective leads: In this study, we generated a new series of serotonin 5-HT 7 receptor antagonists. Their synthesis, structure-activity relationships, and selectivity profiles are reported. This series includes 5-HT 7 antagonists with unprecedented high selectivity for the 5-HT 7 receptor, setting the stage for lead optimization of drugs acting on a range of neurological targets., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Lilly/Avid's AV-1451 is one of the most advanced tau PET tracers in the clinic. Although results obtained in Alzheimer's disease patients are compelling, discrimination of tracer uptake in healthy individuals and patients with supranuclear palsy (PSP) is less clear as there is substantial overlap of signal in multiple brain regions. Moreover, accurate quantification of [ 18 F]AV-1451 uptake in Alzheimer's disease may not be possible., Objectives: The aim of the present study was to characterize the in vitro binding of AV-1451 to understand and identify potential off-target binding that could explain the poor discrimination observed in PSP patients., Methods: [ 3 H]AV-1451 and AV-1451 were characterized in in vitro binding assays using recombinant and native proteins/tissues from postmortem samples of controls and Alzheimer's disease and PSP patients., Results: [ 3 H]AV-1451 binds to multiple sites with nanomolar affinities in brain homogenates and to tau fibrils isolated from Alzheimer's disease or PSP patients. [ 3 H]AV-1451 also binds with similarly high affinities in brain homogenates devoid of tau pathology. This unexpected binding was demonstrated to be because of nanomolar affinities of [ 3 H]AV-1451 for monoamine oxidase A and B enzymes., Conclusions: High affinity of AV-1451 for monoamine oxidase proteins may limit its utility as a tau PET tracer in PSP and Alzheimer's disease because of high levels of monoamine oxidase expression in brain regions also affected by tau deposition, especially if monoamine oxidase levels change over time or with a treatment intervention. © 2017 International Parkinson and Movement Disorder Society., (© 2017 International Parkinson and Movement Disorder Society.)

Pairing orphan G protein–coupled receptors (GPCRs) with their cognate endogenous ligands is expected to have a major impact on our understanding of GPCR biology. It follows that the reproducibility of orphan receptor ligand pairs should be of fundamental importance to guide meaningful investigations into the pharmacology and function of individual receptors. GPR17 is an orphan receptor characterized by some as a dualistic uracil nucleotide/cysteinyl leukotriene receptor and by others as inactive toward these stimuli altogether. Whereas regulation of central nervous system myelination by GPR17 is well established, verification of activity of its putative endogenous ligands has proven elusive so far. Herein we report that uracil nucleotides and cysteinyl leukotrienes do not activate human, mouse, or rat GPR17 in various cellular backgrounds, including primary cells, using eight distinct functional assay platforms based on labelfree pathway-unbiased biosensor technologies, as well as canonical second-messenger or biochemical assays. Appraisal of GPR17 activity can neither be accomplished with co-application of both ligand classes, nor with exogenous transfection of partner receptors (nucleotide P2Y12, cysteinyl-leukotriene CysLT1) to reconstitute the elusive pharmacology. Moreover, our study does not support the inhibition of GPR17 by the marketed antiplatelet drugs cangrelor and ticagrelor, previously suggested to antagonize GPR17. Whereas our data do not disagree with a role of GPR17 per se as an orchestrator of central nervous system functions, they challenge the utility of the proposed (ant)agonists as tools to imply direct contribution of GPR17 in complex biologic settings., (Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.)

Apolipoproteins L (ApoLs) are Bcl-2-like proteins expressed under inflammatory conditions in myeloid and endothelial cells. We found that Toll-like receptor (TLR) stimuli, particularly the viral mimetic polyinosinic:polycytidylic acid (poly(I:C)), specifically induce ApoLs7/11 subfamilies in murine CD8α(+)  dendritic cells (DCs). This induction requires the TLR3/TRIF (where TRIF is TIR domain containing adapter-inducing interferon β) signaling pathway and is dependent on IFN-β in all ApoLs subfamilies except for ApoL7c. Poly(I:C) treatment of DCs is also associated with induction of both cell death and autophagy. ApoLs expression is related to promotion of DC death by poly(I:C), as ApoLs7/11 knockdown increases DC survival and ApoLs7 are associated with the anti-apoptotic protein Bcl-xL (where Bcl-xL is B-cell lymphoma extra large). Similarly, in human monocyte-derived DCs poly(I:C) induces both cell death and the expression of ApoLs, principally ApoL3. Finally, the BH3-like peptide of ApoLs appears to be involved in the DC death-promoting activity. We would like to propose that ApoLs are involved in cell death linked to activation of DCs by viral stimuli., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Methicillin-resistant Staphylococcus aureus (MRSA) bloodstream infections (BSI) are associated with considerable morbidity and mortality, especially with persistent (PB) or recurrent bacteremia (RB)., Objective: To determine the frequency of PB and RB in patients with MRSA BSI, and to characterize the isolates from these patients., Methods: Surveillance for MRSA BSI was performed for one year in 13 Canadian hospitals. PB was defined as a positive blood culture that persisted for ≥7 days; RB was defined as the recurrence of a positive blood culture ≥14 days following a negative culture. Isolates were typed using pulsed-field gel electrophoresis (PFGE). Vancomycin susceptibility was determined using Etest., Results: A total of 183 patients with MRSA BSI were identified; 14 (7.7%) had PB and five (2.7%) had RB. Ten (5.5%) patients were known to have infective endocarditis, and five of these patients had PB or RB. Initial and subsequent MRSA isolates from patients with PB and RB had the same PFGE type. There were no significant differences in the distribution of PFGE types in patients with PB or RB (37% CMRSA-2/USA100; 37% CMRSA-10/USA300) compared with that in other patients (56% CMRSA-2/USA100; 32% CMRSA-10/USA300). All isolates were susceptible to vancomycin, but patients with PB or RB were more likely to have initial isolates with vancomycin minimum inhibitory concentration = 2.0 μg/mL (26% versus 10%; P=0.06)., Conclusions: Persistent or recurrent MRSA bacteremia occurred in 10.4% of patients with MRSA BSIs. Initial isolates from patients with persistent or recurrent MRSA BSIs were more likely to exhibit reduced susceptibility to vancomcyin, but were not associated with any genotype.

Replacement of the lost myelin sheath is a therapeutic goal for treating demyelinating diseases of the central nervous system (CNS), such as multiple sclerosis (MS). The G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptor (GPCR) GPR17, which is phylogenetically closely related to receptors of the "purinergic cluster," has emerged as a modulator of CNS myelination. However, whether GPR17-mediated signaling positively or negatively regulates this critical process is unresolved. We identified a small-molecule agonist, MDL29,951, that selectively activated GPR17 even in a complex environment of endogenous purinergic receptors in primary oligodendrocytes. MDL29,951-stimulated GPR17 engaged the entire set of intracellular adaptor proteins for GPCRs: G proteins of the Gα(i), Gα(s), and Gα(q) subfamily, as well as β-arrestins. This was visualized as alterations in the concentrations of cyclic adenosine monophosphate and inositol phosphate, increased Ca²⁺ flux, phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2), as well as multifeatured cell activation recorded with label-free dynamic mass redistribution and impedance biosensors. MDL29,951 inhibited the maturation of primary oligodendrocytes from heterozygous but not GPR17 knockout mice in culture, as well as in cerebellar slices from 4-day-old wild-type mice. Because GPCRs are attractive targets for therapeutic intervention, inhibiting GPR17 emerges as therapeutic strategy to relieve the oligodendrocyte maturation block and promote myelin repair in MS.

Penicillin nonsusceptibility has been demonstrated in group B streptococci (GBS), but there is limited information regarding mechanisms of resistance. We report a case of GBS with reduced susceptibility to penicillin emerging after long-term suppressive oral penicillin therapy for a prosthetic joint infection. Molecular characterization of the isolate before and after long-term penicillin therapy revealed 5 mutations in the ligand-binding regions of PBP1a, -2a, and -2x not previously reported in GBS.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by a selective loss of motor neurones accompanied by intense gliosis in lesioned areas of the brain and spinal cord. Glutamate-mediated excitotoxicity resulting from impaired astroglial uptake constitutes one of the current pathophysiological hypotheses explaining the progression of the disease. In this study, we examined the regulation of glutamate transporters by type 5 metabotropic glutamate receptor (mGluR5) in activated astrocytes derived from transgenic rats carrying an ALS-related mutated human superoxide dismutase 1 (hSOD1(G93A)) transgene. Cells from transgenic animals and wild-type littermates showed similar expression of glutamate-aspartate transporter and glutamate transporter 1 (GLT-1) after in vitro activation, whereas cells carrying the hSOD1 mutation showed a three-fold higher expression of functional mGluR5, as observed in the spinal cord of end-stage animals. In cells from wild-type animals, (S)-3,5-dihydroxyphenylglycine (DHPG) caused an immediate protein kinase C (PKC)-dependent up-regulation of aspartate uptake that reflected the activation of GLT-1. Although this effect was mimicked in both cultures by direct activation of PKC using phorbol myristate acetate, DHPG failed to up-regulate aspartate uptake in cells derived from the transgenic rats. The failure of activated mGluR5 to increase glutamate uptake in astrocytes derived from this animal model of ALS supports the theory of glutamate excitotoxicity in the pathogenesis of the disease.

Excitatory transmission in the CNS necessitates the existence of dynamic controls of the glutamate uptake achieved by astrocytes, both in physiological conditions and under pathological circumstances characterized by gliosis. In this context, this study was aimed at evaluating the involvement of group I metabotropic glutamate receptors (mGluR) in the regulation of glutamate transport in a model of rat astrocytes undergoing in vitro activation using a cocktail of growth factors (G5 supplement). The vast majority of the cells were found to take up aspartate, mainly through the glutamate/aspartate transporter (GLAST), and at least 60% expressed functional mGluR5a. When exposed for 15 s to the selective group I mGluR agonist (S)-3,5-dihydroxyphenylglycine, reactive astrocytes showed a significant increase in their capacity to take up aspartate. This effect was confirmed at the single-cell level, since activation of mGluRs significantly increased the initial slope of aspartate-dependent Na+ entry associated with the activity of glutamate transporters. This up-regulation was inhibited by an antagonist of mGluR5 and, more importantly, was sensitive to a specific glutamate transporter 1 (GLT-1) blocker. The acute influence of mGluR5 on aspartate uptake was phospholipase C- and protein kinase C-dependent, and was mimicked by phorbol esters. We conclude that mGluR5a contributes to a dynamic control of GLT-1 function in activated astrocytes, acting as a glial sensor of the extracellular glutamate concentration in order to acutely regulate the excitatory transmission.

In vitro culture of astroglial progenitors can be obtained from early post-natal brain tissues and several methods have been reported for promoting their maturation into differentiated astrocytes. Hence, a combination of several nutriments/growth factors -- the G5 supplement (insulin, transferrin, selenite, biotin, hydrocortisone, fibroblast growth factor and epidermal growth factor) -- is widely used as a culture additive favouring the growth, differentiation and maturation of primary cultured astrocytes. Considering the key role played by glial cells in the clearance of glutamate in the synapses, cultured astrocytes are frequently used as a model for the study of glutamate transporters. Indeed, it has been shown that when tested separately, growth factors influence the expression and activity of the GLAST and GLT-1. The present study aimed at characterising the functional expression of these transporters during the time course of differentiation of cultured cortical astrocytes exposed to the supplement G5. After a few days, the vast majority of cells exposed to this supplement adopted a typical stellate morphology (fibrous or type II astrocytes) and showed intense expression of the glial fibrillary acidic protein. Both RT-PCR and immunoblotting studies revealed that the expression of both GLAST and GLT-1 rapidly increased in these cells. While this was correlated with a significant increase in specific uptake of radiolabelled aspartate, fluorescence monitoring of the Na+ influx associated with glutamate transporters activity revealed that the exposure to the G5 supplement considerably increased the percentage of cells participating in the uptake. Biochemical and pharmacological studies revealed that this activity did not involve GLT-1 but most likely reflected an increase in GLAST-mediated uptake. Together, these data indicate that the addition of this classical combination of growth factors and nutriments drives the rapid differentiation toward a homogenous culture of fibrous astrocytes expressing functional glutamate transporters.

The possibility to isolate stem cells from the adult central nervous system and to maintain and propagate these cells in vitro has raised a general interest with regards to their use in cell replacement therapy for degenerative brain diseases. Considering the critical role played by astrocytes in the control of glutamate homeostasis, we have characterised the expression of functional glutamate transporters in neural stem cells exposed to selected culture conditions favouring their differentiation into astrocytes. Commonly, neural stem cells proliferate in suspension as neurospheres in serum-free medium. The addition of serum or a supplement of growth factors (G5) to the culture medium was found to trigger cell adhesion on coated surfaces and to favour their differentiation. Indeed, after 7 days in these conditions, the vast majority of the cells adopted markedly distinct morphologies corresponding to protoplasmic (with serum) or fibrous (with G5 supplement) astrocytes and approximately 35-40% acquired the expression of the glial fibrillary acidic protein (GFAP). Immunocytochemical analysis also revealed that the treatments with serum or with the G5 supplement triggered the expression of the glial glutamate transporters GLT-1 (35 and 21%, respectively) and GLAST (29 and 69%, respectively). This effect was correlated with a robust increase in the Na+ -dependent [3H]-d-aspartate uptake, which was partially inhibited by dihydrokainate, a selective blocker of GLT-1. Together, these results indicate that in vitro differentiation of cultured neural stem cells can give rise to distinct populations of astrocytes expressing functional glutamate transporters.

Adult bone marrow mesenchymal stem cells are multipotent cells that can differentiate into a variety of mesodermal tissues. Recent studies have reported on their ability to also evolve into non-mesodermal cells, especially neural cells. While most of these studies revealed that manipulating these cells triggers the expression of typical neurone markers, less is known about the induction of neuronal- or glial-related physiological properties. The present study focused on the characterisation of glutamate transporters expression and activity in rat mesenchymal stem cells grown in culture conditions favouring their differentiation into astroglial cells. Ten days exposure of the cells to the culture supplement G5 was found to increase the expression of nestin (neuro-epithelial stem cell intermediate filament), an intermediate filament protein expressed by neural stem cells. Simultaneously, a robust induction of the high-affinity glutamate transporter GLT-1 (and GLAST) expression was detected by RT-PCR and immunocytochemistry. This expression was correlated with a highly significant increase in the Na+-dependent [3H]D-aspartate uptake. Finally, while glial fibrillary acidic protein immunoreactivity could not be detected, the induced expression of the astrocytic enzyme glutamine synthetase was demonstrated. These results indicate that in vitro differentiation of adult mesenchymal stem cells in neural precursors coincides with the induction of functional glutamate transport systems. Although the astrocytic nature of these cells remains to be confirmed, this observation gives support to the study of mesenchymal stem cells as a promising tool for the treatment of neurological diseases involving glutamate excitoxicity.

We report the first characterization of the in vivo porphyrin scavenging abilities of two components of a newly discovered heme scavenging system involving iron-regulated surface determinant (Isd) proteins. These proteins are present within the cell envelope of the Gram-positive human pathogen Staphylococcus aureus. IsdC and IsdE, when expressed heterologously in Escherichia coli, efficiently scavenged intracellular heme and resulted in de novo heme synthesis in excess of 100-fold above background. Magnetic circular dichroism analyses showed that the heme-binding properties of the two proteins differ significantly from one another. IsdC bound almost exclusively free-base protoporphyrin IX, whereas the IsdE protein was associated with low spin Fe(III) and Fe(II) heme. These properties provide important insight into the possible mechanisms of iron scavenging from bound heme by Isd proteins.