Interleukin-1beta and tumor necrosis factor-alpha are expressed by different subsets of microglia and macrophages after ischemic stroke in mice

<p>Abstract</p> <p>Background</p> <p>Interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) are expressed by microglia and infiltrating macrophages following ischemic stroke. Whereas IL-1β is primarily neurotoxic in ischemic stroke, TNF-α may have neurotoxic and/or neuroprotective effects. We investigated whether IL-1β and TNF-α are synthesized by overlapping or segregated populations of cells after ischemic stroke in mice.</p> <p>Methods</p> <p>We used flow cytometry and immunohistochemistry to examine cellular co-expression of IL-1β and TNF-α at 6, 12 and 24 hours after permanent middle cerebral artery occlusion in mice, validating the results by the use of bone marrow chimeric mice.</p> <p>Results</p> <p>We found that IL-1β and TNF-α were expressed in largely segregated populations of CD11b⁺CD45^dimmicroglia and CD11b⁺CD45^highmacrophages, with cells expressing both cytokines only rarely. The number of Gr1⁺granulocytes producing IL-1β or TNF-α was very low, and we observed no IL-1β- or TNF-α-expressing T cells or astrocytes.</p> <p>Conclusion</p> <p>Taken together, the results show that IL-1β and TNF-α are produced by largely segregated populations of microglia and macrophages after ischemic stroke in mice. Our findings provide evidence of a functional diversity among different subsets of microglia and macrophages that is potentially relevant to future design of anti-inflammatory therapies in stroke.</p

Glutamate-system defects behind psychiatric manifestations in a familial hemiplegic migraine type 2 disease-mutation mouse model

Migraine is a complex brain disorder, and understanding the complexity of this prevalent disease could improve quality of life for millions of people. Familial Hemiplegic Migraine type 2 (FHM2) is a subtype of migraine with aura and co-morbidities like epilepsy/seizures, cognitive impairments and psychiatric manifestations, such as obsessive-compulsive disorder (OCD). FHM2 disease-mutations locate to the ATP1A2 gene encoding the astrocyte-located α(2)-isoform of the sodium-potassium pump (α(2)Na(+)/K(+)-ATPase). We show that knock-in mice heterozygous for the FHM2-associated G301R-mutation (α(2)(+/G301R)) phenocopy several FHM2-relevant disease traits e.g., by mimicking mood depression and OCD. In vitro studies showed impaired glutamate uptake in hippocampal mixed astrocyte-neuron cultures from α(2)(G301R/G301R) E17 embryonic mice, and moreover, induction of cortical spreading depression (CSD) resulted in reduced recovery in α(2)(+/G301R) male mice. Moreover, NMDA-type glutamate receptor antagonists or progestin-only treatment reverted specific α(2)(+/G301R) behavioral phenotypes. Our findings demonstrate that studies of an in vivo relevant FHM2 disease knock-in mouse model provide a link between the female sex hormone cycle and the glutamate system and a link to co-morbid psychiatric manifestations of FHM2

Selectivity, efficacy and toxicity studies of UCCB01-144, a dimeric neuroprotective PSD-95 inhibitor

Inhibition of postsynaptic density protein-95 (PSD-95) decouples N-methyl-d-aspartate (NMDA) receptor downstream signaling and results in neuroprotection after focal cerebral ischemia. We have previously developed UCCB01-144, a dimeric PSD-95 inhibitor, which binds PSD-95 with high affinity and is neuroprotective in experimental stroke. Here, we investigate the selectivity, efficacy and toxicity of UCCB01-144 and compare with the monomeric drug candidate Tat-NR2B9c. Fluorescence polarization using purified proteins and pull-downs of mouse brain lysates showed that UCCB01-144 potently binds all four PSD-95-like membrane-associated guanylate kinases (MAGUKs). In addition, UCCB01-144 affected NMDA receptor signaling pathways in ischemic brain tissue. UCCB01-144 reduced infarct size in young and aged male mice at various doses when administered 30 min after permanent middle cerebral artery occlusion, but UCCB01-144 was not effective in young male mice when administered 1 h post-ischemia or in female mice. Furthermore, UCCB01-144 was neuroprotective in a transient stroke model in rats, and in contrast to Tat-NR2B9c, high dose of UCCB01-144 did not lead to significant changes in mean arterial blood pressure or heart rate. Overall, UCCB01-144 is a potent MAGUK inhibitor that reduces neurotoxic PSD-95-mediated signaling and improves neuronal survival following focal brain ischemia in rodents under various conditions and without causing cardiovascular side effects, which encourages further studies towards clinical stroke trials

Development of a hypoallergenic recombinant parvalbumin for first-in-man subcutaneous immunotherapy of fish allergy.

To access publisher's full text version of this article, please click on the hyperlink in Additional Links field or click on the hyperlink at the top of the page marked Files. This article is open access.The FAST (food allergy-specific immunotherapy) project aims at developing safe and effective subcutaneous immunotherapy for fish allergy, using recombinant hypoallergenic carp parvalbumin, Cyp c 1.Preclinical characterization and good manufacturing practice (GMP) production of mutant Cyp (mCyp) c 1.Escherichia coli-produced mCyp c 1 was purified using standard chromatographic techniques. Physicochemical properties were investigated by gel electrophoresis, size exclusion chromatography, circular dichroism spectroscopy, reverse-phase high-performance liquid chromatography and mass spectrometry. Allergenicity was assessed by ImmunoCAP inhibition and basophil histamine release assay, immunogenicity by immunization of laboratory animals and stimulation of patients' peripheral blood mononuclear cells (PBMCs). Reference molecules were purified wild-type Cyp c 1 (natural and/or recombinant). GMP-compliant alum-adsorbed mCyp c 1 was tested for acute toxicity in mice and rabbits and for repeated-dose toxicity in mice. Accelerated and real-time protocols were used to evaluate stability of mCyp c 1 as drug substance and drug product.Purified mCyp c 1 behaves as a folded and stable molecule. Using sera of 26 double-blind placebo-controlled food-challenge-proven fish-allergic patients, reduction in allergenic activity ranged from 10- to 5,000-fold (1,000-fold on average), but with retained immunogenicity (immunization in mice/rabbits) and potency to stimulate human PBMCs. Toxicity studies revealed no toxic effects and real-time stability studies on the Al(OH)3-adsorbed drug product demonstrated at least 20 months of stability.The GMP drug product developed for treatment of fish allergy has the characteristics targeted for in FAST: i.e. hypoallergenicity with retained immunogenicity. These results have warranted first-in-man immunotherapy studies to evaluate the safety of this innovative vaccine.info:eu-repo/grantAgreement/EC/FP7/20187

Differences in Origin of Reactive Microglia in Bone Marrow Chimeric Mouse and Rat After Transient Global Ischemia.

Current understanding of microglial involvement in disease is influenced by the observation that recruited bone marrow (BM)-derived cells contribute to reactive microgliosis in BM-chimeric mice. In contrast, a similar phenomenon has not been reported for BM-chimeric rats. We investigated the recruitment and microglial transformation of BM-derived cells in radiation BM-chimeric mice and rats after transientglobal cerebral ischemia, which elicits a characteristic microglialreaction. Both species displayed microglial hyperplasia and rod cell transformation in the hippocampal CA1 region. In mice, a subpopulation of lesion-reactive microglia originated from transformed BM-derived cells. By contrast, no recruitment or microglial transformation of BM-derived cells was observed in BM-chimeric rats. These results suggest that reactive microglia in rats originate from resident microglia, whereas they have a mixed BM-derived and resident origin in mice, depending on the severity of ischemic tissue damage

Cortical morphogenesis during embryonic development is regulated by miR-34c and miR-204

The porcine brain closely resembles the human brain in aspects such as development and morphology. Temporal miRNA profiling in the developing embryonic porcine cortex revealed a distinct set of miRNAs, including miR-34c and miR-204, which exhibited a highly specific expression profile across the time of cortical folding. These miRNAs were found to target Doublecortin (DCX), known to be involved in neuron migration during cortical folding of gyrencephalic brains. In vivo modulation of miRNA expression in mouse embryos confirmed that miR-34c and miR-204 can control neuronal migration and cortical morphogenesis, presumably by posttranscriptional regulation of DCX