Macrophage Receptor Research Articles

Neuropeptide Y regulates cholesterol uptake and efflux in macrophages and promotes foam cell formation.

The dysregulation of lipid metabolic pathways (cholesterol uptake and efflux) in macrophages results in the formation of lipid-dense macrophages, named foam cells, that participate in plaque formation. NPY binding to NPY receptors in macrophages can modulate cell functions and affect the process of atherosclerotic plaques. The present study aimed to determine whether NPY affects the formation of macrophage-derived foam cells and its underlying mechanisms in macrophages. THP-1-derived macrophages were incubated with oxidized low-density lipoprotein (ox-LDL) and treated with different concentrations of NPY. We analysed the relative levels of proteins related to cholesterol uptake and efflux. We found that NPY effectively increased cholesterol uptake and intracellular cholesterol content via the Y1 and Y5 receptors, and this effect was blocked by Y1 and Y5 antagonists. Mechanistically, NPY enhanced the expression of SRA and CD36 via the PKC/PPARγ pathways, promoting macrophage cholesterol uptake. Moreover, NPY significantly decreased cholesterol efflux to the extracellular cholesterol acceptors ApoA1 and HDL in macrophages. NPY mediated decreases in ABCA1, ABCG1 and SR-BI expression through the inhibition of the JAK/STAT3 pathways. Our results suggest that NPY binding to the Y1 and Y5 receptors enhances foam cell formation by regulating cholesterol uptake and efflux in macrophages.

Emerging Nuclear Medicine Imaging of Atherosclerotic Plaque Formation.

Atherosclerosis is a chronic widespread cardiovascular disease and a major predisposing factor for cardiovascular events, among which there are myocardial infarction and ischemic stroke. Atherosclerotic plaque formation is a process that involves different mechanisms, of which inflammation is the most common. Plenty of radiopharmaceuticals were developed to elucidate the process of plaque formation at different stages, some of which were highly specific for atherosclerotic plaque. This review summarizes the current nuclear medicine imaging landscape of preclinical and small-scale clinical studies of these specific RPs, which are not as widespread as labeled FDG, sodium fluoride, and choline. These include oxidation-specific epitope imaging, macrophage, and other cell receptors visualization, neoangiogenesis, and macrophage death imaging. It is shown that specific radiopharmaceuticals have strength in pathophysiologically sound imaging of the atherosclerotic plaques at different stages, but this also may induce problems with the signal registration for low-volume plaques in the vascular wall.

Teleost fish IgM+ plasma-like cells possess IgM-secreting, phagocytic, and antigen-presenting capacities.

Plasma cells are terminally differentiated antibody-secreting B lymphocytes that contribute to humoral immunity by producing large numbers of antibodies. Increasing evidence suggests that teleost fish B cells share certain characteristics with mammalian B1 B cells, including antibody-secreting, phagocytic, and antigen-presenting capacities. However, the difference between mature B cells and plasma cells remains unclear. In this study, we found that, based on their light-scattering characteristics, tilapia anterior kidney (AK) leukocytes can be categorized into two IgM+ B-cell subsets: the lymphoid (L) gate and granulocyte–monocyte/macrophage (G-M) subsets. G-M gate cells are more numerous than L-gate cells and have higher mean fluorescence, but lower forward scatter and side scatter. We analyzed the morphological and ultrastructural features of sorted IgM+ cells and found that L-gate IgM+ cells have a high nucleus–cytoplasm ratio and lymphocyte-like morphology, whereas G-M gate IgM+ cells have a small nucleus, more abundant endoplasmic reticulum, and a larger number of mitochondria, and have a plasma cell-like or macrophage-like morphology. To further characterize the cell types, we examined the specific patterns of expression of B-cell- and T-cell-related genes. We found that B-cell-specific genes were expressed by both L-gate and G-M gate IgM+ cells, and that G-M gate IgM+ cells secreted extremely high levels of IgM. However, T-cell-related genes were highly expressed only in L-gate IgM– cells. These results suggest that G-M gate IgM+ cells are similar to plasma-like cells, with high antibody-secreting capacity. Given that G-M gate cells include the granulocyte, monocyte, and macrophage cell types, but not B cells, monocyte/macrophage markers were used to investigate the cell types further. A macrophage receptor with a collagenous structure was frequently observed, and macrophage-expressed gene-1 was highly expressed, in the G-M gate IgM+ cells. Phagocytic capacity, as determined by ingestion of beads or bacteria, was significantly higher in G-M gate IgM+ cells than in L-gate IgM+ cells, as was antigen-processing capacity. Our findings show that tilapia AK leukocytes can be divided into two IgM+ B-cell subsets and that G-M gate IgM+ cells resemble plasma-like cells, having high antibody-secreting, phagocytic, and antigen-presenting capacities. Thus, this study increases our understanding of the functions of teleost fish plasma-like cells.

Endocannabinoids Modulate Production of Osteoclastogenic Factors by Stem Cells of the Apical Papilla In Vitro

IntroductionMany mediators are produced during pulp inflammation and necrosis, including endocannabinoids (ECbs), which might affect the function of stem cells of the apical papilla (SCAP), cells of paramount importance for root formation, and regenerative endodontic treatment. The aim of this study was to evaluate the production of osteoclastogenesis-related mediators by SCAP modulated by ECbs and lipopolysaccharide (LPS) in vitro. MethodsSCAP were cultured and treated with ECb anandamide (AEA), 2-arachidonoylglycerol, or N-arachidonoylaminophenol. All groups were incubated in the presence of a vehicle or LPS and the antagonist of transient receptor potential cation channel subfamily V member 1, capsazepine. After 24 hours, the culture medium supernatants were collected for further quantification of tumor necrosis factor alpha, CCL2, macrophage colony-stimulating factor, osteoprotegerin, and receptor activator of nuclear factor kappa B ligand. ResultsSmall amounts of tumor necrosis factor alpha and receptor activator of nuclear factor kappa B ligand were detected in SCAP supernatants, and none of the experimental conditions altered their production. A down-regulation in constitutive CCL2 production was observed in the AEA group compared with that in the LPS group. The production of macrophage colony-stimulating factor was significantly increased in all groups treated with AEA compared with the control and LPS-treated groups. Osteoprotegerin was significantly increased by AEA alone and by 2-arachidonoylglycerol and N-arachidonoylaminophenol in the presence of LPS and capsazepine. ConclusionsAEA modulates some of the osteoclastogenic factors produced by SCAP in a bone resorption protective fashion.

Investigating links between air pollution, COVID-19 and lower respiratory infectious diseases – A review

Background/aim Hundreds of air pollution and COVID-19 papers have been published in a short time varying widely in quality. The Greater London Authority commissioned a comprehensive overview of the most credible evidence for the links between air pollution and COVID-19 and other lung infections. Methods Building on the Brunekreef et al (2021) review, a literature search was performed from November 2020 – May 2021, classifying the evidence by exposure, the potential mechanism and the health outcome. Studies on hospital admissions for lower respiratory infections were reviewed from 2011 Results Long-term exposure increased the risk of hospitalization in people already infected with COVID-19 in two good-quality cohort studies with individual data, perhaps due to respiratory and cardiovascular disease from air pollution increasing vulnerability to COVID-19 adverse outcomes. Mortality results were conflicting. Inconsistent results were found for long-term exposure and COVID-19 cases in ecological studies. Studies on short-term exposure were unclear and hard to evaluate. Several studies demonstrated that inhaled pollutant increased expression of the ACE2 receptor and other host susceptibility genes in both alveolar epithelial cells and macrophages. Susceptibility to infection by SARS-CoV-2 can be inferred from these results but only one study used SARS-CoV-2 directly. Several studies from 2011-2021 showed a link between air pollution and hospital admissions for lung infections although the studies were spread across different age groups and disease definitions. Particulate matter does not appear to play any important part in transporting COVID-19 in the environment, as had been suggested by some earlier studies. Conclusions The publication of cohort studies with individual data in this time period is an important step forward. Results are suggestive for long-term exposure and hospital admissions for COVID-19 and some toxicological effects are plausible but further research will be necessary to strengthen these emerging findings. Keywords Air pollution, COVID-19, lower respiratory infections.

P53 Promotes Ferroptosis in Macrophages Treated with Fe3O4 Nanoparticles.

Fe3O4 nanoparticles are the most widely used magnetic nanoparticles in the biomedicine field. The biodistribution of most nanoparticles in vivo is determined by the capture of macrophages; however, the effects of nanoparticles on macrophages remain poorly understood. Here, we demonstrated that Fe3O4 nanoparticles could reduce macrophage viability after 48 h of treatment and induce a shift in macrophage polarization toward the M1 phenotype; RNA sequencing revealed the activation of the ferroptosis pathway and p53 upregulation compared to the control group. The expression in p53, xCT, glutathione peroxidase 4 (GPX4), and transferrin receptor (TFR) in macrophages was similar to that in erastin-induced ferroptosis in macrophages, and the ultrastructural morphology of mitochondria was consistent with that of erastin-treated cells. We used DCFH-DA to estimate the intracellular reactive oxygen species content in Fe3O4 nanoparticles treated with Ana-1 and JC-1 fluorescent probes to detect the mitochondrial membrane potential change; both showed to be time-dependent. Fer-1 inhibited the reduction of the glutathione/oxidized glutathione (GSH/GSSG) ratio and inhibited intracellular oxidative stress states; therefore, Fe3O4 nanoparticles induced ferroptosis in macrophages. Finally, we used pifithrin-α hydrobromide (PFT) as a p53 inhibitor to verify whether the high expression of p53 is involved in mediating this process. After PFT treatment, the live/dead cell rate, TFR, p53 expression, and GPX4 consumption were inhibited and mitigated the GSH/GSSG ratio reduction as well. This indicates that p53 may contribute to Fe3O4 nanoparticle-induced ferroptosis of macrophages. We provide a theoretical basis for the molecular mechanisms of ferroptosis in macrophages and the biotoxicity in vivo induced by Fe3O4 nanoparticles.

The P2Y11 receptor of human M2 macrophages activates canonical and IL-1 receptor signaling to translate the extracellular danger signal ATP into anti-inflammatory and pro-angiogenic responses

The cytoprotective ATP receptor P2Y11 is upregulated during M2 macrophage differentiation and contributes to the anti-inflammatory properties of this macrophage subset. Here, we studied P2Y11-induced reprogramming of human M2 macrophages at the level of mRNA and protein expression. Upregulation of IL-1 receptor (IL-1R) and its known downstream effectors VEGF, CCL20 and SOCS3 as well as downregulation of the ATP-degrading ecto-ATPase CD39 emerged as hallmarks of P2Y11 activation. The anti-inflammatory signature of the P2Y11 transcriptome was further characterized by the downregulation of P2RX7, toll-like receptors and inflammasome components. P2Y11-induced IL-1R upregulation formed the basis for reinforced IL-1 responsiveness of activated M2 macrophages, as IL-1α and IL-1ß each enhanced P2Y11-induced secretion of VEGF and CCL20 as well as the previously reported shedding of soluble tumor necrosis factor receptor 2 (sTNFR2). Raising intracellular cyclic AMP (cAMP) in M2 macrophages through phosphodiesterase 4 inhibition enhanced P2Y11-driven responses. The cAMP-binding effector, exchange protein activated by cAMP 1 (Epac1), which is known to induce SOCS3, differentially regulated the P2Y11/IL-1R response because pharmacological Epac1 inhibition enhanced sTNFR2 and CCL20 release, but had no effect on VEGF secretion. In addition to cAMP, calcium and protein kinase C participated in P2Y11 signaling. Our study reveals how P2Y11 harnesses canonical and IL-1R signaling to promote an anti-inflammatory and pro-angiogenic switch of human M2 macrophages, which may be controlled in part by an Epac1-SOCS3 axis.

Dysregulation of miR-103a Mediates Cigarette Smoking-induced Lipid-laden Macrophage Formation.

Cigarette smoke (CS) is considered a major risk factor for chronic obstructive pulmonary disease (COPD) that is currently the third leading cause of death in the United States. Studies have indicated that patients with COPD have elevated blood low-density lipoprotein levels, which may contribute to the dysregulation of lipid metabolism. Accumulating data show that microRNAs (miRNAs) are involved in various human diseases. However, the role of microRNAs in the pathogenesis of COPD remains poorly defined. In this study, we found that miR-103a expression was significantly reduced in alveolar macrophages from smokers and patients with COPD versus that in alveolar macrophages from nonsmokers. Our data indicated that reactive oxygen species negatively regulate miR-103a in macrophages. Functionally, miR-103a modulates the expressions of genes involved in lipid metabolism and directly targets low-density lipoprotein receptors in macrophages. Furthermore, overexpression of miR-103a suppressed the accumulation of lipid droplets and reduced the reactive oxygen species, both invitro and invivo. Taken together, our findings indicate that downregulation of miR-103a contributes to cigarette smoke-induced lipid-laden macrophage formation and plays a critical role in lipid homeostasis in lung macrophages in the pathogenesis of COPD.

Positive Association between Autoantibodies Against Oxidized LDL and HDL-C: A Novel Mechanism for HDL Cardioprotection?

In the atherosclerotic plaque microenvironment, oxidized phospholipids expressed in the oxidized low-density lipoprotein (oxLDL) surface bind to scavenger receptors of macrophages eliciting foam cell formation and plaque progression. Auto-antibodies against oxLDL (oxLDL-Ab) interact with oxidative epitopes leading to the formation of immune complexes that are unable to interact with macrophage receptors, thus abrogating atherogenesis. Release of oxLDL-Ab by B cells involves interleukin 5 and Th2 response, which in turn are potentiated by HDL. Thereby, we hypothesized that individuals with higher levels of HDL-C may plausibly display elevated titers of oxLDL-Ab. To evaluate the relationship between HDL-C and oxLDL-Ab levels. Asymptomatic individuals (n = 193) were grouped according to their HDL-C concentration to one of three categories: low (< 68 mg/dL), intermediate (68 to 80 mg/dL) or high (> 80 mg/dL). P values < 0.05 were considered statistically significant. Our analysis included 193 individuals (mean age: 47 years; male: 26.3%). Compared to individuals in the lowest HDL-C tertile, those in the highest tertile were older (36 versus 53 years; p = 0.001) and less frequently male (42.6% versus 20.9%; p = 0.001). Mean values of oxLDL-Ab increased as the HDL-C group escalated (0.31, 0.33 and 0.43 units, respectively; p = 0.001 for trend). Simple linear regression found a significant, positive relationship between the independent variable, HDL-C, and the dependent variable, oxLDL-Ab (R = 0.293; p = 0.009). This relation remained significant (R = 0.30; p = 0.044), after adjustment by covariates. Apolipoprotein AI levels were also related to oxLDL-Ab in both simple and adjusted linear regression models. HDL-C and oxLDL-Ab are independently related.

Erythropoietin mediates re-programming of endotoxin-tolerant macrophages through PI3K/AKT signaling and protects mice against secondary infection.

Initial lipopolysaccharide (LPS) exposure leads to a hypo-responsive state by macrophages to a secondary stimulation of LPS, known as endotoxin tolerance. However, recent findings show that functions of endotoxin-tolerant macrophages are not completely suppressed, whereas they undergo a functional re-programming process with upregulation of a panel of molecules leading to enhanced protective functions including antimicrobial and tissue-remodeling activities. However, the underlying molecular mechanisms are still elusive. Erythropoietin (EPO), a glycoprotein regulated by hypoxia-inducible factor 1α (HIF-1α), exerts anti-inflammatory and tissue-protective activities. Nevertheless, the potential effects of EPO on functional re-programming of endotoxin-tolerant macrophages have not been investigated yet. Here, we found that initial LPS exposure led to upregulation of HIF-1α/EPO in macrophages and that EPO enhanced tolerance in tolerized macrophages and mice as demonstrated by suppressed proinflammatory genes such as Il1b, Il6, and Tnfa after secondary LPS stimulation. Moreover, we showed that EPO improved host protective genes in endotoxin-tolerant macrophages and mice, such as the anti-bacterial genes coding for cathelicidin-related antimicrobial peptide (Cnlp) and macrophage receptor with collagenous structure (Marco), and the tissue-repairing gene vascular endothelial growth factor C (Vegfc). Therefore, our findings indicate that EPO mediates the functional re-programming of endotoxin-tolerant macrophages. Mechanistically, we found that PI3K/AKT signaling contributed to EPO-mediated re-programming through upregulation of Irak3 and Wdr5 expression. Specifically, IL-1 receptor-associated kinase 3 (IRAK3) was responsible for inhibiting proinflammatory genes Il1b, Il6, and Tnfa in tolerized macrophages after LPS rechallenge, whereas WDR5 contributed to the upregulation of host beneficial genes including Cnlp, Marco, and Vegfc. In a septic model of mice, EPO pretreatment significantly promoted endotoxin-tolerant re-programming, alleviated lung injury, enhanced bacterial clearance, and decreased mortality in LPS-tolerized mice after secondary infection of Escherichia coli. Collectively, our results reveal a novel role for EPO in mediating functional re-programming of endotoxin-tolerant macrophages; thus, targeting EPO appears to be a new therapeutic option in sepsis and other inflammatory disorders.

Osteostatin Inhibits M-CSF+RANKL-Induced Human Osteoclast Differentiation by Modulating NFATc1.

Parathyroid hormone-related protein (PTHrP) C-terminal peptides regulate the metabolism of bone cells. PHTrP [107–111] (osteostatin) promotes bone repair in animal models of bone defects and prevents bone erosion in inflammatory arthritis. In addition to its positive effects on osteoblasts, osteostatin may inhibit bone resorption. The aim of this study was to determine the effects of osteostatin on human osteoclast differentiation and function. We used macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor κB ligand (RANKL) to induce the osteoclast differentiation of adherent human peripheral blood mononuclear cells. Tartrate-resistant acid phosphatase (TRAP) staining was performed for the detection of the osteoclasts. The function of mature osteoclasts was assessed with a pit resorption assay. Gene expression was evaluated with qRT-PCR, and nuclear factor of activated T cells, cytoplasmic 1 (NFATc1) nuclear translocation was studied by immunofluorescence. We observed that osteostatin (100, 250 and 500 nM) decreased the differentiation of osteoclasts in a concentration-dependent manner, but it did not modify the resorptive ability of mature osteoclasts. In addition, osteostatin decreased the mRNA levels of cathepsin K, osteoclast associated Ig-like receptor (OSCAR) and NFATc1. The nuclear translocation of the master transcription factor in osteoclast differentiation NFATc1 was reduced by osteostatin. Our results suggest that the anti-resorptive effects of osteostatin may be dependent on the inhibition of osteoclastogenesis. This study has shown that osteostatin controls human osteoclast differentiation in vitro through the downregulation of NFATc1.

BAP1 Loss Promotes Suppressive Tumor Immune Microenvironment via Upregulation of PROS1 in Class 2 Uveal Melanomas.

Simple SummaryUveal melanoma is a highly metastatic cancer of the eye which is notoriously resistant to therapy. Elucidating the mechanisms of metastasis in order to devise effective therapies has been a major challenge. The strongest genetic risk factor for metastasis in uveal melanoma is the mutational inactivation of the BAP1 tumor-suppressor gene. However, it remains unknown how BAP1 loss promotes tumor progression. Here, we show that BAP1 loss leads to increased expression of PROS1 in uveal melanocytes and melanoma cells, which in turn leads to phosphorylation and activation of the receptor tyrosine kinase MERTK on adjacent macrophages, driving them into a suppressive M2-polarized state. This mechanism could help explain the suppressive tumor immune microenvironment that is characteristic of BAP1-mutant uveal melanomas, and it suggests that BAP1 loss may lead to metastasis at least in part by facilitating immune escape. These findings provide new insights into the role of BAP1 in uveal melanoma, and they nominate new strategies for increasing the efficacy of immunotherapy in this cancer.Uveal melanoma (UM) is the most common primary cancer of the eye and is associated with a high rate of metastatic death. UM can be stratified into two main classes based on metastatic risk, with class 1 UM having a low metastatic risk and class 2 UM having a high metastatic risk. Class 2 UM have a distinctive genomic, transcriptomic, histopathologic, and clinical phenotype characterized by biallelic inactivation of the BAP1 tumor-suppressor gene, an immune-suppressive microenvironment enriched for M2-polarized macrophages, and poor response to checkpoint-inhibitor immunotherapy. To identify potential mechanistic links between BAP1 loss and immune suppression in class 2 UM, we performed an integrated analysis of UM samples, as well as genetically engineered UM cell lines and uveal melanocytes (UMC). Using RNA sequencing (RNA-seq), we found that the most highly upregulated gene associated with BAP1 loss across these datasets was PROS1, which encodes a ligand that triggers phosphorylation and activation of the immunosuppressive macrophage receptor MERTK. The inverse association between BAP1 and PROS1 in class 2 UM was confirmed by single-cell RNA-seq, which also revealed that MERTK was upregulated in CD163+ macrophages in class 2 UM. Using ChIP-seq, BAP1 knockdown in UM cells resulted in an accumulation of H3K27ac at the PROS1 locus, suggesting epigenetic regulation of PROS1 by BAP1. Phosphorylation of MERTK in RAW 264.7 monocyte–macrophage cells was increased upon coculture with BAP1−/− UMCs, and this phosphorylation was blocked by depletion of PROS1 in the UMCs. These findings were corroborated by multicolor immunohistochemistry, where class 2/BAP1-mutant UMs demonstrated increased PROS1 expression in tumor cells and increased MERTK phosphorylation in CD163+ macrophages compared with class 1/BAP1-wildtype UMs. Taken together, these findings provide a mechanistic link between BAP1 loss and the suppression of the tumor immune microenvironment in class 2 UMs, and they implicate the PROS1–MERTK pathway as a potential target for immunotherapy in UM.

Dry Powder Comprised of Isoniazid-Loaded Nanoparticles of Hyaluronic Acid in Conjugation with Mannose-Anchored Chitosan for Macrophage-Targeted Pulmonary Administration in Tuberculosis.

Marketed dosage forms fail to deliver anti-tubercular drugs directly to the lungs in pulmonary Tuberculosis (TB). Therefore, nanomediated isoniazid (INH)-loaded dry powder for inhalation (Nano-DPI) was developed for macrophage-targeted delivery in TB. Mannosylated chitosan (MC) and hyaluronic acid (HA) with an affinity for the surface mannose and CD44 receptors of macrophages were used in conjugation to prepare hybrid nanosuspension by ionic gelation method using cross-linker, sodium tri-polyphosphate (TPP) followed by freeze-drying to obtain a dry powder composed of nanoparticles (INH-MC/HA NPs). Nanoformulations were evaluated for aerodynamic characteristics, cytotoxicity, hemocompatibility, macrophage phenotype analysis, and immune regulation. Cellular uptake imaging was also conducted to evaluate the uptake of NPs. The nanopowders did not pose any significant toxicity to the cells, along with good compatibility with red blood cells (RBCs). The pro-inflammatory costimulatory markers were upregulated, demonstrating the activation of T-cell response. Moreover, the NPs did not show any tolerogenic effect on the macrophages. Furthermore, confocal imaging exhibited the translocation of NPs in the cells. Altogether, the findings present that nano-DPI was found to be a promising vehicle for targeting macrophages.

Proteomic Changes of Osteoclast Differentiation in Rheumatoid and Psoriatic Arthritis Reveal Functional Differences.

BackgroundOsteoclasts play a crucial role in the maintenance, repair, and remodeling of bones of the adult vertebral skeleton due to their bone resorption capability. Rheumatoid arthritis (RA) and psoriatic arthritis (PsA) are associated with increased activity of osteoclasts.ObjectivesOur study aimed to investigate the dynamic proteomic changes during osteoclast differentiation in healthy donors, in RA, and PsA.MethodsBlood samples of healthy donors, RA, and PsA patients were collected, and monocytes were isolated and differentiated into osteoclasts in vitro using macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor κB ligand (RANK-L). Mass spectrometry-based proteomics was used to analyze proteins from cell lysates. The expression changes were analyzed with Gene Set Enrichment Analysis (GSEA).ResultsThe analysis of the proteomic changes revealed that during the differentiation of the human osteoclasts, expression of the proteins involved in metabolic activity, secretory function, and cell polarity is increased; by contrast, signaling pathways involved in the immune functions are downregulated. Interestingly, the differences between cells of healthy donors and RA/PsA patients are most pronounced after the final steps of differentiation to osteoclasts. In addition, both in RA and PsA the differentiation is characterized by decreased metabolic activity, associated with various immune pathway activities; furthermore by accelerated cytokine production in RA.ConclusionsOur results shed light on the characteristic proteomic changes during human osteoclast differentiation and expression differences in RA and PsA, which reveal important pathophysiological insights in both diseases.

Abstract 3457: Rebastinib, a TIE2 antagonist improves chemotherapy response in homologous recombination proficient epithelial ovarian cancer murine models

Abstract Objectives: The Angiopoietin (ANGPT)/TIE2 kinase axis plays a critical role in cancer angiogenesis. Rebastinib, an investigational TIE2 inhibitor inhibits Tie2 receptor in endothelial cells and macrophages. M2-like Tie2+ macrophages, being pro-tumorigenic, are an attractive target for cancer therapy. Current clinical trials are testing rebastinib with chemotherapy in platinum-resistant ovarian cancer. However, the mechanism is not fully understood. The objective of this study was to determine the effects of rebastinib alone and combined with chemotherapy in preclinical models of ovarian cancer. Methods: RNA-Seq was carried out to determine rebastinib-induced gene expression in ID8 cells and PMJ2R murine peritoneal macrophages. To determine the pharmacodynamics of immune cells in ascites, syngeneic ID8 mice were pre-treated with 10 mg/kg/day rebastinib/control for a week, followed by rebastinib with or without 20 mg/kg carboplatin + 12 mg/kg paclitaxel (chemo) for two weeks. Mice were sacrificed 24h after the last treatment to harvest ascites for flow cytometry. Cohorts of mice from syngeneic ID8 murine model and from a patient derived xenograft (PDX) model of ovarian cancer were used for survival analysis. Results: RNA-Seq showed that in ID8 cells, 1528 genes were upregulated and 3115 genes were downregulated by rebastinib. In macrophages, 2302 genes were upregulated and 2970 genes were downregulated. Several ANGPT-like genes (ANGPTL2, ANGPTL4, ANGPTL6) involved in tumorigenesis, angiogenesis, proliferation were downregulated 2X-10X (p&amp;lt;0.001) in ID8 cells and macrophages. ANGPTL1, an anti-angiogenic and anti-apoptotic gene, was increased 10X in ID8 cells (p&amp;lt;0.001) but not altered in macrophages. ANGPT2, a context-dependent agonist/antagonist of TIE2 pathway, was increased 1.42X in ID8 (P&amp;lt;0.05) cells and 4.6X in macrophages (p&amp;lt;0.001). Flow studies showed that rebastinib significantly increased CD45+ macrophage (p&amp;lt;0.03 vs. chemo; p&amp;lt;0.02 vs. rebastinib+chemo), maintained Th leukocytes (p&amp;lt;0.03, vs. chemo) and significantly increased cytotoxic T cells (p&amp;lt;0.02 vs. control; p&amp;lt;0.0004, p&amp;lt;0.0001 vs. chemo and rebastinib+chemo, respectively). Rebastinib had no significant effect on regulatory T cells, Tie2+ macrophages or Tie2+ M2 macrophages. Rebastinib combined with chemotherapy improved median survival significantly in murine models of ovarian cancer - ID8 (p&amp;lt;0.01) and PDX (p&amp;lt;0.0001). Conclusions: Rebastinib exerts differential effects on tumor cells and macrophages that could contribute to its mechanism of action. Rebastinib alters immune cells and increases cytotoxic T cells in ascites. Rebastinib combined with chemotherapy extends survival in PDX and syngeneic ID8 murine models of ovarian cancer. Further investigation using rational combinations of rebastinib and chemotherapy are underway. Citation Format: Vijayalaxmi G. Gupta, Katherine F. Roby, Harsh B. Pathak, Andrew K. Godwin, Sumedha Gunewardena, Dineo Khabele. Rebastinib, a TIE2 antagonist improves chemotherapy response in homologous recombination proficient epithelial ovarian cancer murine models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3457.

A vitronectin-derived dimeric peptide suppresses osteoclastogenesis by binding to c-Fms and inhibiting M-CSF signaling

Vitronectin is an abundant multifunctional glycoprotein found in serum, the extracellular matrix, and bone, and is involved in diverse physiological processes. Here, we developed a new bioactive dimeric peptide (VnP-8-DN1 dimer) from a human vitronectin-derived motif (IDAAFTRINCQG; residues 206–217; VnP-8) via removal of an isoleucine residue at the N-terminus of VnP-8 and spontaneous air oxidation. The VnP-8-DN1 dimer potently enhanced cell attachment activity, and this activity was mediated by binding to cellular heparan sulfate proteoglycan receptors. Moreover, the VnP-8-DN1 dimer suppressed osteoclast differentiation by blocking the early stage of osteoclastogenesis induced by macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). Furthermore, the VnP-8-DN1 dimer decreased the bone-resorbing activity of osteoclasts and increased the survival of osteoclast precursor cells by decreasing the cellular level of c-Fms and reducing RANK expression. Taken together, these results demonstrate that the VnP-8-DN1 dimer inhibits the early stages of M–CSF– and RANK-induced osteoclast differentiation by binding to c-Fms and inhibiting M-CSF signaling.

Extracellular Hsp90α stimulates a unique innate gene profile in microglial cells with simultaneous activation of Nrf2 and protection from oxidative stress.

Delivery of exogenous heat shock protein 90α (Hsp90α) and/or its induced expression in neural tissues has been suggested as a potential strategy to combat neurodegenerative disease. However, within a neurodegenerative context, a pro-inflammatory response to extracellular Hsp90α (eHsp90α) could undermine strategies to use it for therapeutic intervention. The aim of this study was to investigate the biological effects of eHsp90α on microglial cells, the primary mediators of inflammatory responses in the brain. Transcriptomic profiling by RNA-seq of primary microglia and the cultured EOC2 microglial cell line treated with eHsp90α showed the chaperone to stimulate activation of innate immune responses in microglia that were characterized by an increase in NF-kB-regulated genes. Further characterization showed this response to be substantially lower in amplitude than the effects of other inflammatory stimuli such as fibrillar amyloid-β (fAβ) or lipopolysaccharide (LPS). Additionally, the toxicity of conditioned media obtained from microglia treated with fAβ was attenuated by addition of eHsp90α. Using a co-culture system of microglia and hippocampal neuronal cell line HT22 cells separated by a chamber insert, the neurotoxicity of medium conditioned by microglia treated with fAβ was reduced when eHsp90α was also added. Mechanistically, eHsp90α was shown to activate Nrf2, a response which attenuated fAβ-induced nitric oxide production. The data thus suggested that eHsp90α protects against fAβ-induced oxidative stress. We also report eHsp90α to induce expression of macrophage receptor with collagenous structure (Marco), which would permit receptor-mediated endocytosis of fAβ.

1,25-Dihydroxyvitamin D Inhibits Osteoarthritis by Modulating Interaction Between Vitamin D Receptor and NLRP3 in Macrophages [Retraction

[This retracts the article DOI: 10.2147/JIR.S339670.].

Phosphatidylcholine Liposomes Down-Modulate CD4 Expression Reducing HIV Entry in Human Type-1 Macrophages.

A strategy adopted to combat human immunodeficiency virus type-1 (HIV-1) infection is based on interfering with virus entry into target cells. In this study, we found that phosphatidylcholine (PC) liposomes reduced the expression of the CD4 receptor in human primary type-1 macrophages but not in CD4+ T cells. The down-regulation was specific to CD4, as any effect was not observed in CCR5 membrane expression. Moreover, the reduction of membrane CD4 expression required the Ca2+-independent protein kinase C (PKC), which in turn mediated serine phosphorylation in the intracytoplasmic tail of the CD4 receptor. Serine phosphorylation of CD4 was also associated with its internalization and degradation in acidic compartments. Finally, the observed CD4 downregulation induced by PC liposomes in human primary macrophages reduced the entry of both single-cycle replication and replication competent R5 tropic HIV-1. Altogether, these results show that PC liposomes reduce HIV entry in human macrophages and may impact HIV pathogenesis by lowering the viral reservoir.

Estradiol suppresses psoriatic inflammation in mice by regulating neutrophil and macrophage functions

Psoriasis is a common inflammatory skin disease resulting from dysregulation of the IL-23/TH17 immune axis. The prevalence and severity of psoriasis is higher in men than in women, although the underlying reasons for this are unclear. We studied whether estradiol, a female hormone, plays protective roles in imiquimod-induced psoriatic inflammation in mice by regulating neutrophil and macrophage functions. Wild-type mice and conditional knockout mice were ovariectomized, supplemented with placebo or estradiol pellets, and an imiquimod-containing cream applied. Mice without endogenous ovarian hormones exhibited exacerbated psoriatic inflammation including increased production of IL-17A and IL-1β, which was reversed by exogenously added estradiol. The suppressive effect of estradiol on the production of IL-1β and IL-17A was abolished in mice lacking estrogen receptors in neutrophils and macrophages (Esr1f/fEsr2f/fLysM-Cre+ mice). IL-1β, which is required for production of IL-17A in the psoriasis model, was mainly produced by neutrophils and inflammatory macrophages. Estradiol suppressed IL-1β production from neutrophils and macrophages in mice both invivo and invitro and from human neutrophils invitro. Our results suggest a novel mechanism for sex-dependent differences in psoriasis clinical phenotypes that may shed new light on the pathology of psoriasis.