A genetically distinct microglial subset promotes myelination

Microglia are brain‐resident macrophages with important, but insufficiently understood functions in development, health, and disease. In a new exciting study, Wlodarczyk and colleagues uncover a transient subset of CD11c⁺ microglia that regulate CNS myelination via IGF‐1 expression. These findings represent not only the first evidence for a microglial role in myelinogenesis, but the first for a functionally distinct, genetically defined subpopulation of microglia.     

Ethanol-Induced TLR4/NLRP3 Neuroinflammatory Response in Microglial Cells Promotes Leukocyte Infiltration Across the BBB

We reported that the ethanol-induced innate immune response by activating TLR4 signaling triggers gliosis and neuroinflammation. Ethanol also activates other immune receptors, such as NOD-like-receptors, and specifically NLRP3-inflammasome in astroglial cells, to stimulate caspase-1 cleavage and IL-1β and IL-18 cytokines production. Yet, whether microglia NLRs are also sensitive to the ethanol effects that contribute to neuroinflammation is uncertain. Using cerebral cortexes of the chronic alcohol-fed WT and TLR4^?/? mice, we demonstrated that chronic ethanol treatment enhanced TLR4 mediated-NLRP3/Caspase-1 complex activation, and up-regulated pro-inflammatory cytokines and chemokines levels. Ethanol-induced NLRP3-inflammasome activation and mitochondria-ROS generation were also observed in cultured microglial cells. The up-regulation of CD45^high/CD11b⁺ cell populations and matrix metalloproteinase-9 levels was also noted in the cortexes of the ethanol-treated WT mice. Notably, elimination of the TLR4 function abolished most ethanol-induced neuroinflammatory effects. Thus, our results demonstrate that ethanol triggers TLR4-mediated NLRP3-inflammasome activation in glial cells, and suggest that microglia stimulation may compromise the permeability of blood–brain barrier events to contribute to ethanol-induced neuroinflammation and brain damage.     

Primary Microglia Isolation from Mixed Glial Cell Cultures of Neonatal Rat Brain Tissue

Microglia account for approximately 12% of the total cellular population in the mammalian brain. While neurons and astrocytes are considered the major cell types of the nervous system, microglia play a significant role in normal brain physiology by monitoring tissue for debris and pathogens and maintaining homeostasis in the parenchyma via phagocytic activity ^1,2. Microglia are activated during a number of injury and disease conditions, including neurodegenerative disease, traumatic brain injury, and nervous system infection ³. Under these activating conditions, microglia increase their phagocytic activity, undergo morpohological and proliferative change, and actively secrete reactive oxygen and nitrogen species, pro-inflammatory chemokines and cytokines, often activating a paracrine or autocrine loop ^4-6. As these microglial responses contribute to disease pathogenesis in neurological conditions, research focused on microglia is warranted.Due to the cellular heterogeneity of the brain, it is technically difficult to obtain sufficient microglial sample material with high purity during in vivo experiments. Current research on the neuroprotective and neurotoxic functions of microglia require a routine technical method to consistently generate pure and healthy microglia with sufficient yield for study. We present, in text and video, a protocol to isolate pure primary microglia from mixed glia cultures for a variety of downstream applications. Briefly, this technique utilizes dissociated brain tissue from neonatal rat pups to produce mixed glial cell cultures. After the mixed glial cultures reach confluency, primary microglia are mechanically isolated from the culture by a brief duration of shaking. The microglia are then plated at high purity for experimental study.The principle and protocol of this methodology have been described in the literature ^7,8. Additionally, alternate methodologies to isolate primary microglia are well described ^9-12. Homogenized brain tissue may be separated by density gradient centrifugation to yield primary microglia ¹². However, the centrifugation is of moderate length (45 min) and may cause cellular damage and activation, as well as, cause enriched microglia and other cellular populations. Another protocol has been utilized to isolate primary microglia in a variety of organisms by prolonged (16 hr) shaking while in culture ^9-11. After shaking, the media supernatant is centrifuged to isolate microglia. This longer two-step isolation method may also perturb microglial function and activation. We chiefly utilize the following microglia isolation protocol in our laboratory for a number of reasons: (1) primary microglia simulate in vivo biology more faithfully than immortalized rodent microglia cell lines, (2) nominal mechanical disruption minimizes potential cellular dysfunction or activation, and (3) sufficient yield can be obtained without passage of the mixed glial cell cultures.It is important to note that this protocol uses brain tissue from neonatal rat pups to isolate microglia and that using older rats to isolate microglia can significantly impact the yield, activation status, and functional properties of isolated microglia. There is evidence that aging is linked with microglia dysfunction, increased neuroinflammation and neurodegenerative pathologies, so previous studies have used ex vivo adult microglia to better understand the role of microglia in neurodegenerative diseases where aging is important parameter. However, ex vivo microglia cannot be kept in culture for prolonged periods of time. Therefore, while this protocol extends the life of primary microglia in culture, it should be noted that the microglia behave differently from adult microglia and in vitro studies should be carefully considered when translated to an in vivo setting.     

Administration of DHA Reduces Endoplasmic Reticulum Stress-Associated Inflammation and Alters Microglial or Macrophage Activation in Traumatic Brain Injury

We investigated the effects of the administration of docosahexaenoic acid (DHA) post-traumatic brain injury (TBI) on reducing neuroinflammation. TBI was induced by cortical contusion injury in Sprague Dawley rats. Either DHA (16 mg/kg in dimethyl sulfoxide) or vehicle dimethyl sulfoxide (1 ml/kg) was administered intraperitonially at 5 min after TBI, followed by a daily dose for 3 to 21 days. TBI triggered activation of microglia or macrophages, detected by an increase of Iba1 positively stained microglia or macrophages in peri-lesion cortical tissues at 3, 7, and 21 days post-TBI. The inflammatory response was further characterized by expression of the proinflammatory marker CD16/32 and the anti-inflammatory marker CD206 in Iba1⁺ microglia or macrophages. DHA-treated brains showed significantly fewer CD16/32⁺ microglia or macrophages, but an increased CD206⁺ phagocytic microglial or macrophage population. Additionally, DHA treatment revealed a shift in microglial or macrophage morphology from the activated, amoeboid-like state into the more permissive, surveillant state. Furthermore, activated Iba1⁺ microglial or macrophages were associated with neurons expressing the endoplasmic reticulum (ER) stress marker CHOP at 3 days post-TBI, and the administration of DHA post-TBI concurrently reduced ER stress and the associated activation of Iba1⁺ microglial or macrophages. There was a decrease in nuclear translocation of activated nuclear factor kappa-light-chain-enhancer of activated B cells protein at 3 days in DHA-treated tissue and reduced neuronal degeneration in DHA-treated brains at 3, 7, and 21 days after TBI. In summary, our study demonstrated that TBI mediated inflammatory responses are associated with increased neuronal ER stress and subsequent activation of microglia or macrophages. DHA administration reduced neuronal ER stress and subsequent association with microglial or macrophage polarization after TBI, demonstrating its therapeutic potential to ameliorate TBI-induced cellular pathology.     

Detection of MicroRNAs in Microglia by Real-time PCR in Normal CNS and During Neuroinflammation

Microglia are cells of the myeloid lineage that reside in the central nervous system (CNS)¹. These cells play an important role in pathologies of many diseases associated with neuroinflammation such as multiple sclerosis (MS)². Microglia in a normal CNS express macrophage marker CD11b and exhibit a resting phenotype by expressing low levels of activation markers such as CD45. During pathological events in the CNS, microglia become activated as determined by upregulation of CD45 and other markers³. The factors that affect microglia phenotype and functions in the CNS are not well studied. MicroRNAs (miRNAs) are a growing family of conserved molecules (~22 nucleotides long) that are involved in many normal physiological processes such as cell growth and differentiation⁴ and pathologies such as inflammation⁵. MiRNAs downregulate the expression of certain target genes by binding complementary sequences of their mRNAs and play an important role in the activation of innate immune cells including macrophages⁶ and microglia⁷. In order to investigate miRNA-mediated pathways that define the microglial phenotype, biological function, and to distinguish microglia from other types of macrophages, it is important to quantitatively assess the expression of particular microRNAs in distinct subsets of CNS-resident microglia. Common methods for measuring the expression of miRNAs in the CNS include quantitative PCR from whole neuronal tissue and in situ hybridization. However, quantitative PCR from whole tissue homogenate does not allow the assessment of the expression of miRNA in microglia, which represent only 5-15% of the cells of neuronal tissue. Hybridization in situ allows the assessment of the expression of microRNA in specific cell types in the tissue sections, but this method is not entirely quantitative. In this report we describe a quantitative and sensitive method for the detection of miRNA by real-time PCR in microglia isolated from normal CNS or during neuroinflammation using experimental autoimmune encephalomyelitis (EAE), a mouse model for MS. The described method will be useful to measure the level of expression of microRNAs in microglia in normal CNS or during neuroinflammation associated with various pathologies including MS, stroke, traumatic injury, Alzheimer''s disease and brain tumors.     

Gatekeeper role of brain antigen‐presenting CD11c+ cells in neuroinflammation

Multiple sclerosis is the most frequent chronic inflammatory disease of the CNS. The entry and survival of pathogenic T cells in the CNS are crucial for the initiation and persistence of autoimmune neuroinflammation. In this respect, contradictory evidence exists on the role of the most potent type of antigen‐presenting cells, dendritic cells. Applying intravital two‐photon microscopy, we demonstrate the gatekeeper function of CNS professional antigen‐presenting CD11c⁺ cells, which preferentially interact with Th17 cells. IL‐17 expression correlates with expression of GM‐CSF by T cells and with accumulation of CNS CD11c⁺ cells. These CD11c⁺ cells are organized in perivascular clusters, targeted by T cells, and strongly express the inflammatory chemokines Ccl5, Cxcl9, and Cxcl10. Our findings demonstrate a fundamental role of CNS CD11c⁺ cells in the attraction of pathogenic T cells into and their survival within the CNS. Depletion of CD11c⁺ cells markedly reduced disease severity due to impaired enrichment of pathogenic T cells within the CNS.     

Unique properties of cluster of differentiation 93 in the umbilical cord blood of neonates

It has previously been reported by these authors that cluster of differentiation (CD) 93 is co‐expressed on naive T‐lymphocytes (CD4⁺CD45RA⁺ cells) in neonatal umbilical cord blood cells (UCBCs) but not on normal adult peripheral blood cells (PBCs). In this study, expression of CD93 on other lymphocyte subsets and the concentration of soluble formed CD93 (sCD93) in serum or culture supernatants from neonatal umbilical cord blood (UCB) was examined. It was found that CD93 is also co‐expressed on CD2⁺, CD16⁺, CD56⁺ or CD25⁺ cells in the lymphocyte population of neonatal UCBCs, but not on normal adult PBCs. The concentrations of sCD93 in serum and culture supernatants from neonatal UCB were significantly greater than those from normal adult peripheral blood. The concentrations of sCD93 in culture supernatants from neonatal UCBCs and normal adult PBCs treated with phorbol 12‐myristate 13‐acetate (PMA) were significantly enhanced compared with those without PMA treatment. The degree of enhancement of sCD93 by PMA in culture supernatants from neonatal UCBCs was significantly greater than that of normal adult PBCs and enhancement of sCD93 by PMA in the culture supernatants from neonatal UCBCs and normal adult PBCs was significantly suppressed by PKC inhibitor. Interestingly, the high concentration of serum sCD93 in neonates was significantly decreased in sera from infants at 1 month after birth. Expression of CD93 on the lymphocyte population of PBCs from infants at 1 month after birth was also significantly decreased, compared with that for neonatal UCBCs. These findings indicate that CD93 in neonatal UCB has unique properties as an immunological biomarker.     

HIV receptors within the brain: A study of CD4 and MHC-II on human neurons,astrocytes and microglial cells

We have investigated the level of expression of CD4 and MHC-II antigens on CNS cells and compared it to that on monocytes. MHC-II antigens were expressed spontaneously on cultured astrocytes and monocytes, whereas they were detected only after IFNγ stimulation of microglial cells. In vitro, CD4 receptor was present on monocytes but not on neurons, astrocytes or microglial cells. In normal brain, CD4 antigen was expressed on perivascular microglial cells, a specialized microglia expressing monocytic markers, whereas in HIV1-infected brain, CD4⁺ cells were numerous and scattered throughout the whole parenchyma. These CD4⁺ macrophages may be HIV1-infected monocytes which have crossed the blood-brain barrier after infection, or perivascular microglial cells infected by HIV1-infected blood lymphocytes or free virions.     

The role of MAC1 in diesel exhaust particle‐induced microglial activation and loss of dopaminergic neuron function

Increasing reports support that air pollution causes neuroinflammation and is linked to central nervous system (CNS) disease/damage. Diesel exhaust particles (DEP) are a major component of urban air pollution, which has been linked to microglial activation and Parkinson's disease‐like pathology. To begin to address how DEP may exert CNS effects, microglia and neuron‐glia cultures were treated with either nanometer‐sized DEP (< 0.22 μM; 50 μg/mL), ultrafine carbon black (ufCB, 50 μg/mL), or DEP extracts (eDEP; from 50 μg/mL DEP), and the effect of microglial activation and dopaminergic (DA) neuron function was assessed. All three treatments showed enhanced ameboid microglia morphology, increased H₂O₂ production, and decreased DA uptake. Mechanistic inquiry revealed that the scavenger receptor inhibitor fucoidan blocked DEP internalization in microglia, but failed to alter DEP‐induced H₂O₂ production in microglia. However, pre‐treatment with the MAC1/CD11b inhibitor antibody blocked microglial H₂O₂ production in response to DEP. MAC1^?/? mesencephalic neuron‐glia cultures were protected from DEP‐induced loss of DA neuron function, as measured by DA uptake. These findings support that DEP may activate microglia through multiple mechanisms, where scavenger receptors regulate internalization of DEP and the MAC1 receptor is mandatory for both DEP‐induced microglial H₂O₂ production and loss of DA neuron function.     

Suppressor of Cytokine Signaling-2 (SOCS2) Regulates the Microglial Response and Improves Functional Outcome after Traumatic Brain Injury in Mice

Traumatic brain injury (TBI) is frequently characterized by neuronal, axonal and myelin loss, reactive gliosis and neuroinflammation, often associated with functional deficits. Endogenous repair mechanisms include production of new neurons from precursor cells, but usually the new neurons fail to integrate and survive more than a few weeks. This is in part mediated by the toxic and inflammatory environment present in the injured brain which activates precursor cells to proliferate and differentiate but limits survival of the newborn progeny. Therefore, an understanding of mechanisms that regulate production and survival of newborn neurons and the neuroinflammatory response after brain injury may lead to therapeutic options to improve outcomes. Suppressor of Cytokine Signaling 2 (SOCS2) promotes hippocampal neurogenesis and survival of newborn neurons in the adult brain and regulates anti-inflammatory responses in the periphery, suggesting it may be a useful candidate to improve outcomes of TBI. In this study the functional and cellular responses of SOCS2 over-expressing transgenic (SOCS2Tg) mice were compared to wildtype littermates following mild or moderately severe TBI. Unlike wildtype controls, SOCS2Tg mice showed functional improvement on a ladder test, with a smaller lesion volume at 7d post injury and increased numbers of proliferative CD11b⁺ microglia/macrophages at 35d post-injury in the mild injury paradigm. At 7d post-moderately severe injury there was an increase in the area covered by cells expressing an anti-inflammatory M2 phenotype marker (CD206⁺) but no difference in cells with a pro-inflammatory M1 phenotype marker (CD16/32⁺). No effect of SOCS2 overexpression was observed in production or survival of newborn neurons, even in the presence of the neuroprotective agent erythropoietin (EPO). Therefore, SOCS2 may improve outcome of TBI in mice by regulating aspects of the neuroinflammatory response, promoting a more anti-inflammatory environment, although this was not sufficient to enhance survival of newborn cortical neurons.     

Flow Cytometric Characterization of T Cell Subsets and Microglia After Repetitive Mild Traumatic Brain Injury in Rats

Although, there is growing awareness in the progressive neurodegeneration of chronic traumatic encephalopathy, changes of immune reactions remain equivocal at best. Thus, in a clinically relevant rat repetitive mild traumatic brain injury (rmTBI) model, some immunologic cells (T cell subsets, microglia) in the injured brain and peripheral blood were analyzed by flow cytometry and immunofluorescence. In the injured brain, CD3⁺ T cells showed a bimodal increase during 42 days post-injury (dpi). CD3⁺CD4⁺ T cells firstly increased and then decreased, while CD3⁺CD8⁺ T cells had reversed tendency. CD86⁺/CD11b⁺ M1-like microglia increased at 42 dpi and CD206⁺/CD11b⁺ M2-like microglia peaked at 7 dpi. In addition, peripheral immune suppression was implicated in the chronic phase after rmTBI. Taken together, the study provided useful information on long-term dynamic changes of some immune cells after rmTBI in rats.     

Homeostatic and injury‐induced microglia behavior in the aging brain

Microglia cells are essential for brain homeostasis and have essential roles in neurodegenerative diseases. Aging is the main risk factor for most neurodegenerative diseases, and age‐related changes in microglia may contribute to the susceptibility of the aging brain to dysfunction and neurodegeneration. We have analyzed morphology and dynamic behavior of neocortical microglia in their physiological environment in young adult (3‐month‐old), adult (11‐ to 12‐month‐old), and aged (26‐ to 27‐month‐old) C57BL/6J‐Iba1‐eGFP mice using in vivo 2‐photon microscopy. Results show that surveying microglial cells in the neocortex exhibit age‐related soma volume increase, shortening of processes, and loss of homogeneous tissue distribution. Furthermore, microglial process speed significantly decreased with age. While only a small population of microglia showed soma movement in adult mice, the microglia population with soma movement was increased in aged mice. However, in response to tissue injury, the dynamic microglial response was age‐dependently diminished. These results provide novel insights into microglial behavior and indicate that microglial dysfunction in the aging brain may contribute to age‐related cognitive decline and neurodegenerative diseases.     

Response of the mouse lung transcriptome to welding fume: effects of stainless and mild steel fumes on lung gene expression in A/J and C57BL/6J mice

Background

Nitrogen dioxide (NO₂) is an air pollutant associated with poor respiratory health, asthma exacerbation, and an increased likelihood of inhalational allergies. NO₂ is also produced endogenously in the lung during acute inflammatory responses. NO₂ can function as an adjuvant, allowing for allergic sensitization to an innocuous inhaled antigen and the generation of an antigen-specific Th2 immune response manifesting in an allergic asthma phenotype. As CD11c⁺ antigen presenting cells are considered critical for naïve T cell activation, we investigated the role of CD11c⁺ cells in NO₂-promoted allergic sensitization.

Methods

We systemically depleted CD11c⁺ cells from transgenic mice expressing a simian diphtheria toxin (DT) receptor under of control of the CD11c promoter by administration of DT. Mice were then exposed to 15 ppm NO₂ followed by aerosolized ovalbumin to promote allergic sensitization to ovalbumin and were studied after subsequent inhaled ovalbumin challenges for manifestation of allergic airway disease. In addition, pulmonary CD11c⁺ cells from wildtype mice were studied after exposure to NO₂ and ovalbumin for cellular phenotype by flow cytometry and in vitro cytokine production.

Results

Transient depletion of CD11c⁺ cells during sensitization attenuated airway eosinophilia during allergen challenge and reduced Th2 and Th17 cytokine production. Lung CD11c⁺ cells from wildtype mice exhibited a significant increase in MHCII, CD40, and OX40L expression 2 hours following NO₂ exposure. By 48 hours, CD11c⁺MHCII⁺ DCs within the mediastinal lymph node (MLN) expressed maturation markers, including CD80, CD86, and OX40L. CD11c⁺CD11b^- and CD11c⁺CD11b⁺ pulmonary cells exposed to NO₂ in vivo increased uptake of antigen 2 hours post exposure, with increased ova-Alexa 647⁺ CD11c⁺MHCII⁺ DCs present in MLN from NO₂-exposed mice by 48 hours. Co-cultures of ova-specific CD4⁺ T cells from naïve mice and CD11c⁺ pulmonary cells from NO₂-exposed mice produced IL-1, IL-12p70, and IL-6 in vitro and augmented antigen-induced IL-5 production.

Conclusions

CD11c⁺ cells are critical for NO₂-promoted allergic sensitization. NO₂ exposure causes pulmonary CD11c⁺ cells to acquire a phenotype capable of increased antigen uptake, migration to the draining lymph node, expression of MHCII and co-stimulatory molecules required to activate naïve T cells, and secretion of polarizing cytokines to shape a Th2/Th17 response.     

CD11c+ Cells Partially Mediate the Renoprotective Effect Induced by Bone Marrow-Derived Mesenchymal Stem Cells

Previous studies have shown that induction of immune tolerance by mesenchymal stem cells (MSCs) is partially mediated via monocytes or dendritic cells (DCs). The purpose of this study was to determine the role of CD11c⁺ cells in MSC-induced effects on ischemia/reperfusion injury (IRI). IRI was induced in wildtype (WT) mice and CD11c⁺-depleted mice following pretreatment with or without MSCs. In the in-vitro experiments, the MSC-treated CD11c⁺ cells acquired regulatory phenotype with increased intracellular IL-10 production. Although splenocytes cocultured with MSCs showed reduced T cell proliferation and expansion of CD4⁺FoxP3⁺ regulatory T cells (Tregs), depletion of CD11c⁺ cells was associated with partial loss of MSCs effect on T cells. In in-vivo experiment, MSCs’ renoprotective effect was also associated with induction of more immature CD11c⁺ cells and increased FoxP3 expression in I/R kidneys. However all these effects induced by the MSCs were partially abrogated when CD11c⁺ cells were depleted in the CD11c⁺-DTR transgenic mice. In addition, the observation that adoptive transfer of WT CD11c⁺ cells partially restored the beneficial effect of the MSCs, while transferring IL-10 deficient CD11c⁺ cells did not, strongly suggest the important contribution of IL-10 producing CD11c⁺ cells in attenuating kidney injury by MSCs. Our results suggest that the CD11c⁺ cell-Tregs play critical role in mediating renoprotective effect of MSCs.     

Microglial calcium signal acts as a rapid sensor of single neuron damage in vivo

In the healthy adult brain microglia, the main immune-competent cells of the CNS, have a distinct (so-called resting or surveying) phenotype. Resting microglia can only be studied in vivo since any isolation of brain tissue inevitably triggers microglial activation. Here we used in vivo two-photon imaging to obtain a first insight into Ca²⁺ signaling in resting cortical microglia. The majority (80%) of microglial cells showed no spontaneous Ca²⁺ transients at rest and in conditions of strong neuronal activity. However, they reliably responded with large, generalized Ca²⁺ transients to damage of an individual neuron. These damage-induced responses had a short latency (0.4-4 s) and were localized to the immediate vicinity of the damaged neuron (< 50 μm cell body-to-cell body distance). They were occluded by the application of ATPγS as well as UDP and 2-MeSADP, the agonists of metabotropic P2Y receptors, and they required Ca²⁺ release from the intracellular Ca²⁺ stores. Thus, our in vivo data suggest that microglial Ca²⁺ signals occur mostly under pathological conditions and identify a Ca²⁺ store-operated signal, which represents a very sensitive, rapid, and highly localized response of microglial cells to brain damage. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.     

Prion protein participates in the regulation of classical and alternative activation of BV2 microglia

The cellular prion protein (PrP^C) is a glycoprotein anchored by glycosylphosphatidylinositol (GPI) to the cell surface and is abundantly expressed in the central nervous system. Numerous studies have suggested a protective function for PrP^C, including protection from ischemic and excitotoxic lesions and several apoptotic insults, and recent reports have shown that PrP^C has a context‐dependent neuroprotective function. In this study, we investigated the effect of PPNP down‐regulation on various forms of microglial activation. We first examined the mRNA expression of PRNP upon exposure to IFN‐γ, IL‐4, or IL‐10 in BV2 microglia. We then analyzed the effect of si‐RNA‐mediated disruption of PRNP on different parameters of microglial activation in IFN‐γ‐, IL‐4‐, or IL‐10‐stimulated microglia. The results showed that PRNP mRNA expression was invariably down‐regulated in microglia upon exposure to IFN‐γ, IL‐4, or IL‐10. PRNP silencing prior to cytokines treatment reduced the responsiveness of microglia to INF‐γ treatment, significantly altered IL‐4‐induced microglial activation phenotype, and had no effect on IL‐10‐induced microglial activation. Together, these results support a role of PrP^C in the modulation of the shift of microglia from a quiescent state to an activated phenotype and in the regulation of the microglial response during classical and alternative activation.     

Microglial replacement in the aged brain restricts neuroinflammation following intracerebral hemorrhage

Aged microglia display augmented inflammatory activity after neural injury. Although aging is a risk factor for poor outcome after brain insults, the precise impact of aging-related alterations in microglia on neural injury remains poorly understood. Microglia can be eliminated via pharmacological inhibition of the colony–stimulating factor 1 receptor (CSF1R). Upon withdrawal of CSF1R inhibitors, microglia rapidly repopulate the entire brain, leading to replacement of the microglial compartment. In this study, we investigated the impact of microglial replacement in the aged brain on neural injury using a mouse model of intracerebral hemorrhage (ICH) induced by collagenase injection. We found that replacement of microglia in the aged brain reduced neurological deficits and brain edema after ICH. Microglial replacement-induced attenuation of ICH injury was accompanied with alleviated blood-brain barrier disruption and leukocyte infiltration. Notably, newly repopulated microglia had reduced expression of IL-1β, TNF-α and CD86, and upregulation of CD206 in response to ICH. Our findings suggest that replacement of microglia in the aged brain restricts neuroinflammation and brain injury following ICH.Subject terms: Neuroimmunology, Cognitive ageing     

Development of peritoneal macrophage along a dendritic cell lineage in response to uptake of oligomannose-coated liposomes

In this study, we investigate the potential of peritoneal macrophages to differentiate into dendritic cell (DCs) in response to preferential uptake of oligomannose-coated liposomes (OMLs). About 30% of peritoneal cells (PECs) preferentially took up OMLs that were administered into the peritoneal cavity. The OML-ingesting cells expressed CD11b and F4/80, but lacked CD11c expression, indicating that the OML-ingesting PECs with a CD11b^highCD11c⁻ phenotype are resident peritoneal macrophages. During in vitro cultivation, CD11c⁺ cells arose among the PECs with ingested OMLs. CD11c⁺ cells also developed among enriched peritoneal CD11b^highCD11⁻ cells from OML-treated mice, and the resulting CD11c⁺ cells expressed co-stimulatory molecules and MHC class II. In addition, OML-ingesting CD11b^highCD11c⁺ cells were found in spleen after the enriched peritoneal macrophages with ingested OMLs were transplanted in the peritoneal cavity of mice. These results show that a fraction of peritoneal macrophages can differentiate into mature DCs following uptake of OMLs.     

MBP-Positive and CD11c-Positive Cells Are Associated with Different Phenotypes of Korean Patients with Non-Asthmatic Chronic Rhinosinusitis

Background

Asthmatic nasal polyps primarily exhibit eosinophilic infiltration. However, the identities of the immune cells that infiltrate non-asthmatic nasal polyps remain unclear. Thus, we thought to investigate the distribution of innate immune cells and its clinical relevance in non-asthmatic chronic rhinosinusitis (CRS) in Korea.

Methods

Tissues from uncinate process (UP) were obtained from controls (n = 18) and CRS without nasal polyps (CRSsNP, n = 45). Nasal polyps (NP) and UP were obtained from CRS with nasal polyps (CRSwNP, n = 56). The innate immune cells was evaluated by immunohistochemistry such as, eosinophil major basic protein (MBP), tryptase, CD68, CD163, CD11c, 2D7, human neutrophil elastase (HNE) and its distribution was analyzed according to clinical parameters.

Results

In comparisons between UP from each group, CRSwNP had a higher number of MPB⁺, CD68⁺, and CD11c⁺ cells relative to CRSsNP. Comparisons between UP and NP from CRSwNP indicated that NP have a higher infiltrate of MBP⁺, CD163⁺, CD11c⁺, 2D7⁺ and HNE⁺ cells, whereas fewer CD68⁺ cells were found in NP. In addition, MBP⁺ and CD11c⁺ cells were increased from UP of CRSsNP, to UP of CRSwNP, and to NP of CRSwNP. Moreover, in UP from CRSwNP, the number of MBP⁺ and CD11c⁺ cells positively correlated with CT scores. In the analysis of CRSwNP phenotype, allergic eosinophilic polyps had a higher number of MBP⁺, tryptase⁺, CD11c⁺, 2D7⁺ cells than others, whereas allergic non-eosinophilic polyps showed mainly infiltration of HNE⁺ and 2D7⁺ cells.

Conclusions

The infiltration of MBP⁺ and CD11c⁺ innate immune cells show a significant association with phenotype and disease extent of CRS and allergic status also may influences cellular phenotype in non-asthmatic CRSwNP in Korea.     

Young microglia restore amyloid plaque clearance of aged microglia

Alzheimer′s disease (AD) is characterized by deposition of amyloid plaques, neurofibrillary tangles, and neuroinflammation. In order to study microglial contribution to amyloid plaque phagocytosis, we developed a novel ex vivo model by co‐culturing organotypic brain slices from up to 20‐month‐old, amyloid‐bearing AD mouse model (APPPS1) and young, neonatal wild‐type (WT) mice. Surprisingly, co‐culturing resulted in proliferation, recruitment, and clustering of old microglial cells around amyloid plaques and clearance of the plaque halo. Depletion of either old or young microglial cells prevented amyloid plaque clearance, indicating a synergistic effect of both populations. Exposing old microglial cells to conditioned media of young microglia or addition of granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) was sufficient to induce microglial proliferation and reduce amyloid plaque size. Our data suggest that microglial dysfunction in AD may be reversible and their phagocytic ability can be modulated to limit amyloid accumulation. This novel ex vivo model provides a valuable system for identification, screening, and testing of compounds aimed to therapeutically reinforce microglial phagocytosis.