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Infiltrating monocyte-derived macrophages (MDMs) and resident microglia dominate central nervous system (CNS) injury sites. Differential roles for these cell populations after injury are beginning to be uncovered. Here, we show evidence that MDMs and microglia directly communicate with one another and differentially modulate each other’s functions. Importantly, microglia-mediated phagocytosis and inflammation are suppressed by infiltrating macrophages. In the context of spinal cord injury (SCI), preventing such communication increases microglial activation and worsens functional recovery. We suggest that macrophages entering the CNS provide a regulatory mechanism that controls acute and long-term microglia-mediated inflammation, which may drive damage in a variety of CNS conditions. [ABSTRACT FROM AUTHOR]

Abstract Ceruloplasmin (Cp) is an important extracellular regulator of iron metabolism. We showed previously that it stimulates Reelin proteolytic processing and cell aggregation in cultures of developing neurons. Reelin is a secreted protein required for the correct positioning of neurons in the brain. It is cleaved in vivo into N-terminally-derived 300K and 180K fragments through incompletely known mechanisms. One of Reelin signaling targets is the actin-binding protein cofilin, the phosphorylation of which is diminished in Reelin-deficient mice. This work looked for in vivo evidence of a relationship between Cp, Reelin and neuronal organization during brain development by analyzing wild-type and Cp-null mice. Cp as well as the full-length, 300K and 180K Reelin species appeared together in wild-type brains at embryonic day (E) 12.5 by immunoblotting. In wild-type compared to Cp-null brains, there was more 300K Reelin from E12.5 to E17.5, a period characterized by extensive, radially directed neuronal migration in the cerebral cortex. Immunofluorescence labeling of tissue sections at E16.5 revealed the localization of Cp with radial glia and meningeal cells adjacent to Reelin-producing Cajal-Retzius neurons, underlining the proximity of Cp and Reelin. Cofilin phosphorylation was seen starting at E10.5–E12.5 and lasted longer until postnatal day 7 in wild-type than Cp-null mice. Finally, using CUX1 as a marker revealed defective accumulation of neurons in layers II/III in neonatal and adult Cp-null mice. These results combined with our earlier work point to a potentially new role of Cp in Reelin processing and signaling and neuronal organization in the cerebral cortex in vivo. Highlights • Ceruloplasmin (Cp) and Reelin are expressed together in mouse developing brain. • 300K Reelin and phospho-cofilin are downregulated in developing Cp-null brain. • The cerebral cortex of Cp-null mice shows defect in layers II/III neurons. [ABSTRACT FROM AUTHOR]

The proper formation and maintenance of functional synapses in the central nervous system (CNS) requires communication between neurons and astrocytes and the ability of astrocytes to release neuromodulatory molecules. Previously, we described a novel role for the astrocyte-secreted matricellular protein SPARC (Secreted Protein, Acidic and Rich in Cysteine) in regulating α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) and plasticity at developing synapses. SPARC is highly expressed by astrocytes and microglia during CNS development but its level is reduced in adulthood. Interestingly, SPARC has been shown to be upregulated in CNS injury and disease. However, the role of SPARC upregulation in these contexts is not fully understood. In this study, we investigated the effect of chronic SPARC administration on glutamate receptors on mature hippocampal neuron cultures and following CNS injury. We found that SPARC treatment increased the number of GluA1-containing AMPARs at synapses and enhanced synaptic function. Furthermore, we determined that the increase in synaptic strength induced by SPARC could be inhibited by Philanthotoxin-433, a blocker of homomeric GluA1-containing AMPARs. We then investigated the effect of SPARC treatment on neuronal health in an injury context where SPARC expression is upregulated. We found that SPARC levels are increased in astrocytes and microglia following middle cerebral artery occlusion (MCAO) in vivo and oxygen-glucose deprivation (OGD) in vitro. Remarkably, chronic pre-treatment with SPARC prevented OGD-induced loss of synaptic GluA1. Furthermore, SPARC treatment reduced neuronal death through Philanthotoxin-433 sensitive GluA1 receptors. Taken together, this study suggests a novel role for SPARC and GluA1 in promoting neuronal health and recovery following CNS damage. [ABSTRACT FROM AUTHOR]

Iron accumulation occurs in the CNS in multiple sclerosis (MS) and in experimental autoimmune encephalomyelitis (EAE). However, the mechanisms underlying such iron accumulation are not fully understood. We studied the expression and cellular localization of molecules involved in cellular iron influx, storage, and efflux. This was assessed in two mouse models of EAE: relapsing–remitting (RR-EAE) and chronic (CH-EAE). The expression of molecules involved in iron homeostasis was assessed at the onset, peak, remission/progressive and late stages of the disease. We provide several lines of evidence for iron accumulation in the EAE spinal cord which increases with disease progression and duration, is worse in CH-EAE, and is localized in macrophages and microglia. We also provide evidence that there is a disruption of the iron efflux mechanism in macrophages/microglia that underlie the iron accumulation seen in these cells. Macrophages/microglia also lack expression of the ferroxidases (ceruloplasmin and hephaestin) which have antioxidant effects. In contrast, astrocytes which do not accumulate iron, show robust expression of several iron influx and efflux proteins and the ferroxidase ceruloplasmin which detoxifies ferrous iron. Astrocytes therefore are capable of efficiently recycling iron from sites of EAE lesions likely into the circulation. We also provide evidence of marked dysregulation of mitochondrial function and energy metabolism genes, as well as of NADPH oxidase genes in the EAE spinal cord. This data provides the basis for the selective iron accumulation in macrophage/microglia and further evidence of severe mitochondrial dysfunction in EAE. It may provide insights into processes underling iron accumulation in MS and other neurodegenerative diseases in which iron accumulation occurs. [ABSTRACT FROM AUTHOR]

Summary Macrophages and microglia can be polarized along a continuum toward a detrimental (M1) or a beneficial (M2) state in the injured CNS. Although phagocytosis of myelin in vitro promotes M2 polarization, macrophage/microglia in the injured spinal cord retain a predominantly M1 state that is detrimental to recovery. We have identified two factors that underlie this skewing toward M1 polarization in the injured CNS. We show that TNF prevents phagocytosis-mediated conversion from M1 to M2 cells in vitro and in vivo in spinal cord injury (SCI). Additionally, iron that accumulates in macrophages in SCI increases TNF expression and the appearance of a macrophage population with a proinflammatory mixed M1/M2 phenotype. In addition, transplantation experiments show that increased loading of M2 macrophages with iron induces a rapid switch from M2 to M1 phenotype. The combined effect of this favors predominant and prolonged M1 macrophage polarization that is detrimental to recovery after SCI. [ABSTRACT FROM AUTHOR]

The past decade has revealed much about the complexity of the local inflammatory response after spinal cord injury (SCI). A major challenge is to distinguish between microglia and monocyte-derived macrophages (MDMs) to determine their phenotype and function. Transcriptome studies have revealed microglia-selective genes but are still limited in scope because many markers are downregulated after injury. Additionally, new genetic reporter mice are available to study microglia and MDMs. There is more evidence now for the plasticity and heterogeneity of microglia and MDMs. We also discuss the role of neutrophils that are the first peripheral cells to enter the injured CNS. [ABSTRACT FROM AUTHOR]

Ceruloplasmin (Cp) is a ferroxidase that also plays a role in iron efflux from cells. It can thus help to regulate cellular iron homeostasis. In the CNS, Cp is expressed as a membrane-anchored form by astrocytes. Here, we assessed the role of Cp in permanent middle cerebral artery occlusion (pMCAO) comparing wildtype and Cp null mice. Our studies show that the lesion size is larger and functional recovery impaired in Cp null mice compared to wildtype mice. Expression of Cp increased ninefold at 72 h after pMCAO and remained elevated about twofold at day 14. We also assessed changes in mRNA and protein expression of molecules involved in iron homeostasis. As expected there was a reduction in ferroportin in Cp null mice at 72 h. There was also a remarkable increase in DMT1 protein in both genotypes at 72 h, being much higher in wildtype mice (19.5-fold), that then remained elevated about twofold at 14 days. No difference was seen in transferrin receptor 1 (TfR1) expression, except a small reduction in wildtype mice at 72 h, suggesting that the increase in DMT1 may underlie iron uptake independent of TfR1-endosomal uptake. There was also an increase of ferritin light chain in both genotypes. Iron histochemistry showed increased iron accumulation after pMCAO, initially along the lesion border and later throughout the lesion. Immunofluorescence labeling for ferritin (a surrogate marker for iron) and GFAP or CD11b showed increased ferritin in GFAP+ astrocytes along the lesion border in Cp null mice, while CD11b+ macrophages expressed ferritin equally in both genotypes. Increased lipid peroxidation assessed by 4HNE staining was increased threefold in Cp null mice at 72 h after pMCAO; and 3-nitrotyrosine labeling showed a similar trend. Three key pro-inflammatory cytokines (IL-1β, TNFα, and IL-6) were markedly increased at 24 h after pMCAO equally in both genotypes, and remained elevated at lower levels later, indicating that the lack of Cp does not alter key inflammatory cytokine expression after pMCAO. These data indicate that Cp expression is rapidly upregulated after pMCAO, and loss of Cp results in dysregulation of iron homeostasis, increased oxidative damage, greater lesion size and impaired recovery of function.

A critical aspect in all models is the assessment of the final outcome of the modelling procedure. In the case of a focal ischaemic brain injury, apart from the determination of the size of the lesion, another valuable tool is the evaluation of the final functional deficit. Indeed, ischaemic damage leads to the appearance of different degrees of sensoriomotor and cognitive impairments, which may yield useful information on location and size of the lesion and on the efficacy of neuroprotective treatments after the acute injury. In addition, the magnitude of these impairments may also be useful to predict final outcome and to evaluate neuro-restorative therapies in a long-term scenario. To this aim, a wide range of tests has been developed which allow the quantification of all these neurological symptoms. This review intends to compile the most useful behavioural tests designed to assess neurological symptoms in studies of focal experimental cerebral ischemia in rodents induced by middle cerebral artery occlusion, the most commonly used model of ischaemic stroke.

Objective: Findings from several studies have revealed that major depression is associated with an increased cardiovascular risk. The physiopathologic mechanisms of this association remain unclear, although recently, it has been hypothesized that a decreased production of nitric oxide could be a potential contributor to vascular dysfunction in depressive patients. The objective of this study was to evaluate nitric oxide production and vascular endothelial function in treatment-naive young healthy adults with a first episode of major depression., Methods: A case-control study in 50 treatment-naive young adults with a first episode of major depression and 50 healthy control subjects was conducted. Plasma levels of nitric oxide metabolites (nitrates/nitrites) were determined using a colorimetric assay based on Griess reaction. Endothelial function was assessed by flow-mediated vasodilation measurements after reactive hyperemia., Results: The mean age of the depressed patients was 22.6 (standard deviation [SD], 4.6) years, whereas the controls were 23.4 (SD, 4.8) years. Sixteen men (32%) and 34 women (68%) were included in each group. The plasma nitrite/nitrate concentrations were significantly lower in depressive subjects compared with healthy controls (17.5 [SD, 4.9] μmol/L versus 21.6 [SD, 7.0] μmol/L, p < .001); however, flow-mediated vasodilation values were similar in both groups (13.1% [SD, 4.3%] versus 12.1% [SD, 5.0%], p = .10)., Conclusions: Decreased plasma concentrations of nitric oxide metabolites are not associated with vascular endothelial dysfunction in young subjects with a first episode of major depression. Reduced nitrate/nitrite levels could reflect a decreased nitric oxide production in the central nervous system of depressed subjects. Further studies are needed to confirm this hypothesis.