Central Macrophage Research Articles

Angelica sinensis polysaccharides ameliorate 5-FU-induced stress anemia via promoting extramedullary erythroblastic island central macrophage-mediated erythroid differentiation

BackgroundChronic anemia, especially chemotherapy-induced anemia, is a common and intractable symptom. Puzzlingly, the conventional anemic treatment may lead to various side effects, and the mechanism of stress anemia remains unclear. MethodsHere, peripheral blood, histopathological and transmission electron microscopical examination, colony forming test, flow cytometry, and qRT-PCR assay were used to investigate the effects of Angelia sinensis polysaccharide (ASP), one main active ingredient of Chinese herb medicine Angelica sinensis, on ameliorating 5-fluorouracil (5-FU)-induced stress anemia. ResultsWe found that intraperitoneal injection to a C57BL/6J mouse ASP 100 mg/kg per day for consecutive 10 days or 14 days, remarkably accelerated the recovery of RBC, hemoglobin, and hematocrit in blood. ASP alleviated 5-FU-caused impairment of bone marrow cell and BFU-E enumeration. Meanwhile, ASP antagonized 5-FU promoting extramedullary erythropoiesis in the spleen, inducing splenomegaly due to stress erythroblastic islands, and occurrence of megakaryocytes and hematopoietic precursors in splenic colonies. ASP increased splenic stress BFU-E enumeration, driving BFU-E differentiation towards Pro-E and end-stage erythroblasts. Furthermore, ASP increased the number of F4/80+VCAM-1+ splenic erythroblastic island central macrophages, upregulating genetic expression of EPOR, Emp, VCAM-1, Hmox-1, Trf, TfR1, Fpn1, Spi-C, DNase2a, Tim4, MertK, and Klf1 in splenocytes. ConclusionsOur findings indicate that the possible mechanism of chemotherapy-induced anemia is related to stress erythroid maturation arrest. Whereas, ASP may promote stress erythroid differentiation via elevated EPO sensitivity in extramedullary hematopoietic organs and enhanced macrophage-mediated adhesion, iron homeostasis and transfer, and nuclear engulfment, which may represent a promising therapeutic strategy.

Dichloroacetate Prevents Sepsis Associated Encephalopathy by Inhibiting Microglia Pyroptosis through PDK4/NLRP3.

Dichloroacetate (DCA), a pyruvate dehydrogenase kinase inhibitor, is often used to treat lactic acidosis and malignant tumors. Increasing studies have shown that DCA has neuroprotective effects. Here, we explored the role and mechanism of DCA in Sepsis associated encephalopathy (SAE). Single-cell analysis was used to determine the important role of PDK4 in SAE and identify the cell type. GO and GSEA analysis were used to determine the correlation between DCA and pyroptosis. Through LPS + ATP stimulation, a microglia pyroptosis model was established to observe the expression level of intracellular pyroptosis-related proteins under DCA intervention, and further detect the changes in intracellular ROS and JC-1. Additionally, a co-culture environment of microglia and neuron was simply constructed to evaluate the effect of DCA on activated microglia-mediated neuronal apoptosis. Finally, Novel object recognition test and the Morris water maze were used to explore the effect of DCA on cognitive function in mice from different groups after intervention. Based on the above experiments,this study concludes thatDCA can improve the ratio of peripheral and central M1 macrophages, inhibit NLRP3-mediated pyroptosis through ROS and mitochondrial membrane potential (MMP). DCA can reduce neuron death caused by SAE and improve cognitive function in LPS mice. In SAE, DCA may be a potential candidate drug for the treatment of microglia-mediated neuroinflammation.

First detection of PCV4 in swine in the United States: codetection with PCV2 and PCV3 and direct detection within tissues

Since PCV4 was first described in 2019, the virus has been identified in several countries in Southeast Asia and Europe. Most studies have been limited to detecting PCV4 by PCR. Thus, PCV4 has an unclear association with clinical disease. This study utilized 512 porcine clinical lung, feces, spleen, serum, lymphoid tissue, and fetus samples submitted to the ISU-VDL from June–September 2023. PCV4 was detected in 8.6% of samples with an average Ct value of 33. While detection rates among sample types were variable, lymphoid tissue had the highest detection rate (18.7%). Two ORF2 sequences were obtained from lymphoid tissue samples and had 96.36–98.98% nucleotide identity with reference sequences. Direct detection of PCV4 by RNAscope revealed viral replication in B lymphocytes and macrophages in lymph node germinal centers and histiocytic and T lymphocyte infiltration in the lamina propria of the small intestine. PCV4 detection was most commonly observed in nursery to finishing aged pigs displaying respiratory and enteric disease. Coinfection with PCV2, PCV3, and other endemic pathogens was frequently observed, highlighting the complex interplay between different PCVs and their potential roles in disease pathogenesis. This study provides insights into the frequency of detection, tissue distribution, and genetic characteristics of PCV4 in the US.

Central nervous system-associated macrophages modulate the immune response following stroke in aged mice.

Age is a major nonmodifiable risk factor for ischemic stroke. Central nervous system-associated macrophages (CAMs) are resident immune cells located along the brain vasculature at the interface between the blood circulation and the parenchyma. By using a clinically relevant thromboembolic stroke model in young and aged male mice and corresponding human tissue samples, we show that during aging, CAMs acquire a central role in orchestrating immune cell trafficking after stroke through the specific modulation of adhesion molecules by endothelial cells. The absence of CAMs provokes increased leukocyte infiltration (neutrophils and CD4+ and CD8+ T lymphocytes) and neurological dysfunction after stroke exclusively in aged mice. Major histocompatibility complex class II, overexpressed by CAMs during aging, plays a significant role in the modulation of immune responses to stroke. We demonstrate that during aging, CAMs become central coordinators of the neuroimmune response that ensure a long-term fine-tuning of the immune responses triggered by stroke.

TFPI from erythroblasts drives heme production in central macrophages promoting erythropoiesis in polycythemia

Bleeding and thrombosis are known as common complications of polycythemia for a long time. However, the role of coagulation system in erythropoiesis is unclear. Here, we discover that an anticoagulant protein tissue factor pathway inhibitor (TFPI) plays an essential role in erythropoiesis via the control of heme biosynthesis in central macrophages. TFPI levels are elevated in erythroblasts of human erythroblastic islands with JAK2V617F mutation and hypoxia condition. Erythroid lineage-specific knockout TFPI results in impaired erythropoiesis through decreasing ferrochelatase expression and heme biosynthesis in central macrophages. Mechanistically, the TFPI interacts with thrombomodulin to promote the downstream ERK1/2-GATA1 signaling pathway to induce heme biosynthesis in central macrophages. Furthermore, TFPI blockade impairs human erythropoiesis in vitro, and normalizes the erythroid compartment in mice with polycythemia. These results show that erythroblast-derived TFPI plays an important role in the regulation of erythropoiesis and reveal an interplay between erythroblasts and central macrophages.

Significance of erythropoesis correction in preoperative preparation in the surgical treatment of patients with colorectal cancer

Background. Anemic syndrome, which accompanies the development of pathology in patients with colorectal cancer, can cause serious early postoperative complications. The question of how preoperative preparation activities influence the state of the intestinal structures and the clinical situation in general is relevant. Of particular importance in this situation is the correction of anemic syndrome before surgery, using hematopoietic stimulants. Objective. The purpose of the study is to analyze the dynamics of erythropoiesis and changes in tissue and cell ultrastructure in intraoperative colon biopsy specimens after stimulation of erythropoiesis in preoperative preparation in the surgical treatment of patients with colorectal cancer. Methods. The study of red bone marrow and the wall of the large intestine using transmission electron microscopy. Results. Most patients with colorectal cancer are characterized by the development of signs of ineffective erythropoiesis with a moderate slowdown in the differentiation of erythroblasts and a delay in the release of mature erythrocytes into the systemic circulation. Stimulation of erythropoiesis with the help of epoetin leads to an increase in the functional activity of the central macrophages of the erythropoietic islets of the red bone marrow and the enhancement of their intercellular association with maturing erythroid cells. This is accompanied by the formation of a large number of full-fledged erythrocytes, an increase in the numerical density of cells of the erythroid series and the normalization of their ultrastructure. Conclusion. Stimulation of erythropoiesis with the help of epoetin in the preoperative period causes the inhibition of endothelial dysfunction due to the restriction of dystrophic and destructive changes in endothelial cell organelles, stabilization of their membranes, prevention of arteriole spasm, erythrocyte stasis and erythrocyte phenomena. As a result, the phenomena of infiltration, perivascular and interstitial edema in the composition of the membranes of the intestinal wall are much less pronounced than in the group of patients without preoperative activation of erythropoiesis.

Beyond antiviral: role of IFN-I in brain development

Interferons and central nervous system resident macrophages, microglia, are well-known for their respective roles in antiviral defense and phagocytosis. Using a classic experimental paradigm for examining activity-dependent neural plasticity, Escoubas, Dorman, et al. recently identified a role for microglial type I interferon signaling in the clearance of unwanted neurons during mouse brain development.

Deciphering the regulatory landscape of murine splenic response to anemic stress at single-cell resolution

AbstractStress erythropoiesis can be influenced by multiple mediators through both intrinsic and extrinsic mechanisms in early erythroid precursors. Single-cell RNA sequencing was conducted on spleen tissue isolated from mice subjected to phenylhydrazine and serial bleeding to explore novel molecular mechanisms of stress erythropoiesis. Our results showed prominent emergence of early erythroblast populations under both modes of anemic stress. Analysis of gene expression revealed distinct phases during the development of emerging erythroid cells. Interestingly, we observed the presence of a "hiatus" subpopulation characterized by relatively low level of transcriptional activities that transitions between early stages of emerging erythroid cells, with moderate protein synthesis activities. Moreover, single-cell analysis conducted on macrophage populations revealed distinct transcriptional programs in Vcam1+ macrophages under stress. Notably, a novel marker, CD81, was identified for labeling central macrophages in erythroblastic islands (EBIs), which is functionally required for EBIs to combat anemic stress. These findings offer fresh insights into the intrinsic and extrinsic pathways of early erythroblasts' response to stress, potentially informing the development of innovative therapeutic approaches for addressing anemic-related conditions.

Mesenchymal stromal cells suppress microglial activation and tumor necrosis factor production

Background aimsWhite matter diseases are commonly associated with microglial activation and neuroinflammation. Mesenchymal stromal cells (MSCs) have immunomodulatory properties and thus have the potential to be developed as cell therapy for white matter disease. MSCs interact with resident macrophages to alter the trajectory of inflammation; however, the impact MSCs have on central nervous system macrophages and the effect this has on the progression of white matter disease are unclear. MethodsIn this study, we utilized numerous assays of varying complexity to model different aspects of white matter disease. These assays ranged from an in vivo spinal cord acute demyelination model to a simple microglial cell line activation assay. Our goal was to investigate the influence of human umbilical cord tissue MSCs on the activation of microglia. ResultsMSCs reduced the production of tumor necrosis factor (TNF) by microglia and decreased demyelinated lesions in the spinal cord after acute focal injury. To determine if MSCs could directly suppress the activation of microglia and to develop an efficient potency assay, we utilized isolated primary microglia from mouse brains and the Immortalized MicroGlial Cell Line (IMG). MSCs suppressed the activation of microglia and the release of TNF after stimulation with lipopolysaccharide, a toll-like receptor agonist. ConclusionsIn this study, we demonstrated that MSCs altered the immune response after acute injury in the spinal cord. In numerous assays, MSCs suppressed activation of microglia and release of the pro-inflammatory cytokine TNF. Of these assays, IMG could be standardized and used as an effective potency assay to determine the efficacy of MSCs for treating white matter disease or other neuroinflammatory conditions associated with microglial activation.

Irf5 Expression in Macrophages Contributes to Iron Regulation within Erythromyeloblastic Islands

Erythroblastic islands (EBIs) classically consist of a central macrophage supporting surrounding erythroid precursors as they develop from CFU-Es to reticulocytes. More recently, it has been shown that, in the bone marrow, these structures also support granulopoiesis, expanding our understanding of the niche, and leading to the concept of erythromyeloblastic islands (EMBI). Under certain inflammatory conditions, there is an imbalance between erythropoiesis and granulopoiesis, eventually leading to anemia and increased neutrophil counts. The central macrophage has been proposed to play a role in regulating this balanced blood production; however, the molecular mechanisms remain unknown. Recently, we demonstrated that mice lacking the transcription factor interferon regulatory factor 5 ( Irf5 -/-) present iron deficiency anemia as they age, along with altered structures in the bone marrow EMBIs. Further, our CITEseq studies showed that Irf5 is specifically expressed by macrophages in the EMBIs. Therefore, we concluded that Irf5 is a regulator of the balance between erythropoiesis and granulopoiesis in the bone marrow erythromyeloblastic niche. To understand the mechanisms leading to iron deficiency anemia, as well as changes in the island, we first undertook a comprehensive analysis of hematopoietic progenitors. We measured the relative amounts of megakaryocytic-erythroid progenitors (MEP), granulocyte-monocyte progenitors (GMP) and common myeloid progenitors (CMP) in the bone marrow and observed decreased MEP (WT: 82.81% vs KO: 76.28%, p-value 0.011), increased GMP (WT: 14.94% vs KO: 22.16%, p-value 0.002) and unchanged CMP compared to wild-type (WT) as the mice aged. Given that Irf5 is expressed in both hematopoietic and non-hematopoietic cells, we generated Irf5fl/fl-Vav-iCre+ mice to evaluate the contribution of non-hematopoietic Irf5 to EMBI formation and composition. We observed a significant reduction in the number of EMBI, with overrepresentation of myeloid cells, suggesting that the phenotype observed in Irf5 -/- is mostly due to hematopoietic Irf5. However, when we measured the relative amounts of MEP and GMP in the Irf5fl/fl-Vav-iCre+ mice, these remained unchanged, suggesting that cell non-autonomous functions contribute to EMBI composition. scRNAseq analyses of EMBIs from 3 and 9 months-old littermate-matched WT and Irf5 -/-mice revealed that heme synthesis was the only downregulated pathway. In addition, central macrophages from Irf5 -/- EMBIs presented decreased expression of major iron regulatory proteins, including heme transporter Slc48a1 and Heme-Oxygenase-1 (HMOX1). Functional analyses using Calcein-AM staining and flow cytometry of freshly isolated central macrophages from enriched EMBIs of WT and Irf5 -/-mice revealed that the labile iron pool (LIP) of the central macrophage of Irf5 -/- EMBIs was significantly reduced (mean 14.0) compared to WT (mean 49.0, p-value 0.004). This apparent decrease in intracellular iron was concomitantly supported by significant decreases in serum iron (WT: 67.27 uM/L vs KO: 39.67 uM/L, p-value: 0.0059), increased serum EPO (WT: 108 pg/dL vs KO: 163.8 pg/dL, p-value: .0075), and Prussian blue staining of Irf5 -/-spleen indicating iron deposition in aging Irf5 -/- mice. Finally, these findings were conserved in Irf5fl/fl-Vav-iCre+ mice, highlighting a cell autonomous function for Irf5 in iron regulation within the island. Together, these data suggest that Irf5 may be a new regulator of iron scavenging, trafficking, and/or utilization within the central macrophage, potentially playing a major role in the red-cell production capacity of the EMBI.

Leveraging Transcriptomic Data to Study Erythromyeloblastic Island (EMBI) Macrophages: Using Spic-GFP As a Marker for Embi Macrophages

We recently demonstrated that the majority of the erythroblastic islands in the mouse bone marrow are supporting not only terminal erythropoiesis but also granulopoiesis, being common niches for terminal hematopoiesis (Romano, Seu, et al. Blood 2022). Using single cell RNA-sequencing (scRNA-seq) and Cellular Indexing of Transcriptome and Epitopes by Sequencing (CITE-seq) we demonstrated several macrophage populations captured from EMBI-enriched bone marrow. Notably, the population of the classically defined EBI macrophages characterized by the genes Mertk, Dnase2a, and Hmox1, also showed sensitive and specific expression of the heme-induced Spic transcription factor (Figure 1). In agreement with the CITE-seq data, imaging flow cytometry (IFC) of EMBIs from SpiC igfp/igfp mouse bone marrow demonstrated that at least 80% of the central macrophages were GFP +, confirming their expression of Spic (Figure 2). After stimulation with erythropoietin (Epo), which leads to an increase of both the number and size of EMBIs, both Spic-GFP-positive and negative central macrophages doubled in number, as determined by IFC studies. Moreover, Spic-GFP + EMBI macrophages isolated from mouse bone marrow reconstituted efficiently into EMBIs after incubation with CD71 + and CD11b + cells isolated from another ( Spic-GFP neg) bone marrow. Characterizing EMBIs by IFC allows for identification of phenotypic characteristics of macrophages participating in the EMBI structure. The strong and intimate interactions between the erythroblasts and granulocytic precursors to the central macrophage facilitates the preservation of EMBIs and their analysis by IFC; however, these same strong interactions add a layer of complexity to methods that require single cells, such as scRNA-seq. Previous attempts to capture EMBI macrophages have had lower than expected yields. Utilizing the Spic-GFP reporter mouse model permitted the use of harsher dissociation by trypsinization and sorting based on GFP positivity, optimizing EMBI macrophage recovery. The resulting captures yielded thousands of EMBI macrophages based on cells expressing Adgre1, Vcam1, Hmox1, DNase2a, and Mertk that show a marked degree of transcriptional heterogeneity in other macrophage surface markers. Comparison of the captures from control (saline-) and Epo-stimulated EMBI macrophages revealed an emergent macrophage population, increasing up to 25-fold after Epo in comparison to control. This Epo-induced macrophage population has approximately 900 globally distinguishing genes including markers characteristic of monocytes such as Ace (encoding the angiotensin-converting enzyme), implicating monocyte recruitment to serve as stress erythropoiesis-supporting macrophages within the murine bone marrow. Additional clusters of macrophages with the characteristics of classically defined EMBI macrophages are also increasing with Epo- stimulation, indicating heterogeneity within the Spic-GFP + population. To verify these transcriptomic results on the protein level, we performed IFC with CD169, Mertk, and other EMBI macrophage markers providing further insights to the heterogeneity of their expression on the central macrophages within intact EMBIs. We anticipate that the Spic-GFP + reporter mouse will be a useful tool in EMBI studies, providing a specific and sensitive marker to trace and isolate the majority of EMBI macrophages.

Ineffective Erythropoiesis Negatively Impacts the Erythroblastic Island in Sickle Cell Disease

Sickle cell disease (SCD) is a genetic recessive disorder, characterized by painful episodes of vaso-occlusion, chronic hemolytic anemia, and progressive organ failure. It is one of the most common and severe monogenic diseases worldwide. SCD is caused by a point mutation in the β-globin gene, leading to the expression of an abnormal hemoglobin (HbS) that polymerizes under hypoxic conditions, driving red blood cells (RBC) sickling. Peripheral RBCs are extensively studied in SCD, as hemolysis is the major contributor to anemia, but little is known about erythroid differentiation in this pathology. We recently demonstrated the occurrence of ineffective erythropoiesis in the bone marrow (BM) of SCD patients, characterized by the death of a significant proportion of erythroblasts between the polychromatic and the orthochromatic stages. We hypothesize that this increased cell death could negatively impact the central macrophages of the erythroblastic islands (EBIs) as they would engulf high numbers of hemoglobin-rich apoptotic erythroblasts and undergo ferroptosis because of excess iron. To investigate our hypothesis, we used an in vitro system whereby CD14 + monocytes were isolated from peripheral blood and cultured in media supplemented with M-CSF and dexamethasone. This led them to differentiate towards anti-inflammatory tissue macrophages, with an “EBI-like” phenotype, expressing markers such as CD16, CD163, CD169, CD206. Phagocytosis of high numbers of RBCs by “EBI-like” macrophages led to increased cell death, reactive oxygen species, and lipid peroxidation, and to the upregulation of genes involved in ferroptosis pathways such as HO-1 and SLC7A11. This was circumvented by the ferroptosis inhibitors ferrostatin-1 and liproxtatin-1. Furthermore, phagocytosis of apoptotic in vitro-generated SCD erythroblasts also induced ferroptosis in EBI-like macrophages, with increased cell death and lipid peroxidation in a time-dependent manner. Interestingly, while this was the case with hemoglobin-rich, terminally differentiating erythroblasts, phagocytosis of cells without hemoglobin, such as K562 or GPA - progenitor cells did not affect cell viability, indicating that the observed ferroptosis was due to the iron content of the internalized cells. We investigated the effect of increased RBC phagocytosis on the phenotype of surviving macrophages and found an M2 to M1 shift indicating a switch to a pro-inflammatory phenotype under the condition of elevated levels of intracellular iron and oxidative stress. We challenged these findings in vivo using the Townes humanized SCD mouse model. Data on 4-month-old mice showed the expected switch in erythroid production from steady-state erythropoiesis in the BM to a stress erythropoiesis in the spleen. This was associated with a decrease in the number of EBI macrophages (F4/80 HighVCAM+CD169+Ly6G-) in the BM of SS mice when compared to AS and AA mice, and an expansion of red pulp macrophages (RPMs) F4/80 HighCD169+VCAM+. Using a flow cytometry-based assay we found high lipid peroxidation levels in BM EBI macrophages of SS mice as compared to AA, suggesting occurrence of ferroptosis in these cells. Our study suggests that IE in SCD may contribute to anemia, directly by cell death of differentiating erythroblasts, and indirectly by disrupting the erythroblastic island or by inducing its collapse following ferroptosis of the central macrophage. Our work with the Townes mouse model and BM samples from SCD patients will help to elucidate this hypothesis.

Spatial Mapping Reveals Distinct Erythroid Niches in Mice and Humans during Development and Stress

Like hematopoiesis, erythropoiesis requires erythroid niche nurturing and support. While extensive studies have revealed various components and functions of the HSC niches, the erythroid niche is less characterized. The most studied erythroid niche is the bone marrow erythroblastic island (EBI), which comprises a central macrophage surrounded by developing erythroblasts. However, most of the reported studies of EBI involve in vitro reconstitution of mixed cell populations that do not recapitulate in vivo niches. Here, we utilized multiple spatial mapping technologies, including spatial transcriptomics and whole-mount imaging studies, to characterize erythropoiesis in mouse and human hematopoietic tissues during development and under stress. We first utilized the Visium platform from 10x Genomics to study the spatial transcriptomic profiles of the E14.5 mouse fetal liver, which is the major organ for murine fetal definitive erythropoiesis. The Visium platform contains barcoded spots with a diameter of 55 µm, which is suboptimal for single-cell analyses but ideal for the investigation of cell-cell interactions within their microenvironment. We found that each hepatic lobe exhibited different cluster distributions, indicating distinct cell compositions among different anatomic loci. Further deconvolution of the clusters and co-analysis with corresponding single-cell RNA sequencing data revealed that there was a higher positive correlation between erythroid cells and C1q+ macrophages compared to other macrophages, suggesting that C1q+ macrophages are more likely to be EBI macrophages in mouse. C1q+ macrophages exhibited high expression of known genes involved in EBI macrophage function and erythropoiesis. Cyclic immunofluorescence (CyCIF) and whole-mount immunofluorescence analyses further demonstrated the co-localization of C1q+ macrophages with erythroid cells in the E14.5 fetal liver. This strong positive correlation between C1q+ macrophages and erythroid cells was also observed across different stages of fetal liver development, newborn bone marrow, and adult spleen under physiologic and stress conditions. These findings demonstrate that C1q+ macrophages as EBI macrophages are conserved across different hematopoietic organs in mice. We next applied the same technologies to human hematopoietic tissues during development and stress. In contrast to the mouse, we did not observe a strong positive correlation between erythroid cells and C1q+ macrophages or other macrophages in the human fetal liver. Instead, there is a strong association between erythroid progenitor and maturing erythroid cells. This strong association is also found in different developmental stages of the human fetal liver as well as the bone marrow, which becomes even stronger in regenerating human bone marrow after chemotherapy. We also recapitulated these findings in a human bone marrow organoid model. These results demonstrate the erythroid cell self-sustaining capacities in human erythropoiesis. Taken together, our findings reveal the distinct erythroid niches in mice and humans during development and stress, which provides a comprehensive understanding of erythropoiesis during evolution.

Characterization of the responses of brain macrophages to focused ultrasound-mediated blood-brain barrier opening.

The opening of the blood-brain barrier (BBB) by focused ultrasound (FUS) coupled with intravenously injected microbubbles can be leveraged as a form of immunotherapy for the treatment of neurodegenerative disorders. However, how FUS BBB opening affects brain macrophages is not well understood. Here by using single-cell sequencing to characterize the distinct responses of microglia and central nervous system-associated macrophages (CAMs) to FUS-mediated BBB opening in mice, we show that the treatment remodels the immune landscape via the recruitment of CAMs and the proliferation of microglia and via population size increases in disease-associated microglia. Both microglia and CAMs showed early and late increases in population sizes, yet only the proliferation of microglia increased at both timepoints. The population of disease-associated microglia also increased, accompanied by the upregulation of genes associated with gliogenesis and phagocytosis, with the depletion of brain macrophages significantly decreasing the duration of BBB opening.

Cell-autonomous effects of APOE4 in restricting microglial response in brain homeostasis and Alzheimer's disease.

Microglial involvement in Alzheimer's disease (AD) pathology has emerged as a risk-determining pathogenic event. While apolipoprotein E (APOE) is known to modify AD risk, it remains unclear how microglial apoE impacts brain cognition and AD pathology. Here, using conditional mouse models expressing apoE isoforms in microglia and central nervous system-associated macrophages (CAMs), we demonstrate a cell-autonomous effect of apoE3-mediated microglial activation and function, which are negated by apoE4. Expression of apoE3 in microglia/CAMs improves cognitive function, increases microglia surrounding amyloid plaque and reduces amyloid pathology and associated toxicity, whereas apoE4 expression either compromises or has no effects on these outcomes by impairing lipid metabolism. Single-cell transcriptomic profiling reveals increased antigen presentation and interferon pathways upon apoE3 expression. In contrast, apoE4 expression downregulates complement and lysosomal pathways, and promotes stress-related responses. Moreover, in the presence of mouse endogenous apoE, microglial apoE4 exacerbates amyloid pathology. Finally, we observed a reduction in Lgals3-positive responsive microglia surrounding amyloid plaque and an increased accumulation of lipid droplets in APOE4 human brains and induced pluripotent stem cell-derived microglia. Our findings establish critical isoform-dependent effects of microglia/CAM-expressed apoE in brain function and the development of amyloid pathology, providing new insight into how apoE4 vastly increases AD risk.

Neutrophil biology in injuries and diseases of the central and peripheral nervous systems.

The role of inflammation in nervous system injury and disease is attracting increased attention. Much of that research has focused on microglia in the central nervous system (CNS) and macrophages in the peripheral nervous system (PNS). Much less attention has been paid to the roles played by neutrophils. Neutrophils are part of the granulocyte subtype of myeloid cells. These cells, like macrophages, originate and differentiate in the bone marrow from which they enter the circulation. After tissue damage or infection, neutrophils are the first immune cells to infiltrate into tissues and are directed there by specific chemokines, which act on chemokine receptors on neutrophils. We have reviewed here the basic biology of these cells, including their differentiation, the types of granules they contain, the chemokines that act on them, the subpopulations of neutrophils that exist, and their functions. We also discuss tools available for identification and further study of neutrophils. We then turn to a review of what is known about the role of neutrophils in CNS and PNS diseases and injury, including stroke, Alzheimer's disease, multiple sclerosis, amyotrophic lateral sclerosis, spinal cord and traumatic brain injuries, CNS and PNS axon regeneration, and neuropathic pain. While in the past studies have focused on neutrophils deleterious effects, we will highlight new findings about their benefits. Studies on their actions should lead to identification of ways to modify neutrophil effects to improve health.

Peripheral and central macrophages in obesity.

Obesity is associated with chronic, low-grade inflammation. Excessive nutrient intake causes adipose tissue expansion, which may in turn cause cellular stress that triggers infiltration of pro-inflammatory immune cells from the circulation as well as activation of cells that are residing in the adipose tissue. In particular, the adipose tissue macrophages (ATMs) are important in the pathogenesis of obesity. A pro-inflammatory activation is also found in other organs which are important for energy metabolism, such as the liver, muscle and the pancreas, which may stimulate the development of obesity-related co-morbidities, including insulin resistance, type 2 diabetes (T2D), cardiovascular disease (CVD) and non-alcoholic fatty liver disease (NAFLD). Interestingly, it is now clear that obesity-induced pro-inflammatory signaling also occurs in the central nervous system (CNS), and that pro-inflammatory activation of immune cells in the brain may be involved in appetite dysregulation and metabolic disturbances in obesity. More recently, it has become evident that microglia, the resident macrophages of the CNS that drive neuroinflammation, may also be activated in obesity and can be relevant for regulation of hypothalamic feeding circuits. In this review, we focus on the action of peripheral and central macrophages and their potential roles in metabolic disease, and how macrophages interact with other immune cells to promote inflammation during obesity.

THE NEURONAL ANTIINFLAMMATORY EFFECTS OF APELIN-13

Objective: Apelin, a peptide with growth factor characteristic, is expressed in many cells and acts by binding to the APJ receptor. Apelin mRNA was detected in the central nervous system (CNS), macrophages and many peripheral tissues. According to recent studies, apelin protects neurons from hypoxia and glutamate-mediated excitotoxicity and can protect cells from oxidative stres (stress) (OS) damage by stimulating catalase activity. Pituitary and neurodegenerative diseases are associated with progressive neuronal loss in the CNS. Neuroinflammation and OS-induced cell damage, caused by microglia and astrocytes activation, have an important role in the pathogenesis and prognosis of these diseases. The aim of this study is to investigate the effects of apelin-13 molecule on the development of inflammation and OS caused by bacterial endotoxin in an invitro astrocyte cell culture medium. Methods: The effect of apelin -13 on cell viability changes in lipopolysaccharide (LPS) induced astrocyte inflammation was analyzed by enzymatic MTT test. The anti-inflammatory effects were determined by qRT-PCR and ELISA analysis at the level of TNF-α, IL-1β, IL-6 and mRNA transcription and protein. The effects of apelin on endotoxin mediated OS and possible DNA damage, lipid peroxidation, and nitric oxide (NO) production were evaluated by ELISA method. Results: In our study, LPS application to astrocytes caused inflammation and a significant decrease in inflammation was found with apelin treatment.
 Conclusion: The results obtained from this study reveal the antiinflammatory, antiapoptotic and antioxidant effects of apelin, that have widely expressed receptors and role in the development of immune response on the development of neuroinflammation and oxidative cell damage.

Human TLR8 induces inflammatory bone marrow erythromyeloblastic islands and anemia in SLE-prone mice.

Anemia commonly occurs in systemic lupus erythematosus, a disease characterized by innate immune activation by nucleic acids. Overactivation of cytoplasmic sensors by self-DNA or RNA can cause erythroid cell death, while sparing other hematopoietic cell lineages. Whereas chronic inflammation is involved in this mechanism, less is known about the impact of systemic lupus erythematosus on the BM erythropoietic niche. We discovered that expression of the endosomal ssRNA sensor human TLR8 induces fatal anemia in Sle1.Yaa lupus mice. We observed that anemia was associated with a decrease in erythromyeloblastic islands and a block in differentiation at the CFU-E to proerythroblast transition in the BM. Single-cell RNAseq analyses of isolated BM erythromyeloblastic islands from human TLR8-expressing mice revealed that genes associated with essential central macrophage functions including adhesion and provision of nutrients were down-regulated. Although compensatory stress erythropoiesis occurred in the spleen, red blood cell half-life decreased because of hemophagocytosis. These data implicate the endosomal RNA sensor TLR8 as an additional innate receptor whose overactivation causes acquired failure of erythropoiesis via myeloid cell dysregulation.

Lineage tracing identifies in vitro microglia-to-neuron conversion by NeuroD1 expression.

Neuronal regeneration to replenish lost neurons after injury is critical for brain repair. Microglia, brain-resident macrophages that have the propensity to accumulate at the site of injury, can be a potential source for replenishing lost neurons through fate conversion into neurons, induced by forced expression of neuronal lineage-specific transcription factors. However, it has not been strictly demonstrated that microglia, rather than central nervous system-associated macrophages, such as meningeal macrophages, convert into neurons. Here, we show that NeuroD1-transduced microglia can be successfully converted into neurons in vitro using lineage-mapping strategies. We also found that a chemical cocktail treatment further promoted NeuroD1-induced microglia-to-neuron conversion. NeuroD1 with loss-of-function mutation, on the other hand, failed to induce the neuronal conversion. Our results indicate that microglia are indeed reprogrammed into neurons by NeuroD1 with neurogenic transcriptional activity.