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Elevated bone resorption and diminished bone formation have been recognized as the primary features of glucocorticoid-associated skeletal disorders. However, the direct effects of excess glucocorticoids on bone turnover remain unclear. Here, we explored the outcomes of exogenous glucocorticoid treatment on bone loss and delayed fracture healing in mice and found that reduced bone turnover was a dominant feature, resulting in a net loss of bone mass. The primary effect of glucocorticoids on osteogenic differentiation was not inhibitory; instead, they cooperated with macrophages to facilitate osteogenesis. Impaired local nutrient status--notably, obstructed fatty acid transportation--was a key factor contributing to glucocorticoid-induced impairment of bone turnover in vivo. Furthermore, fatty acid oxidation in macrophages fueled the ability of glucocorticoid-liganded receptors to enter the nucleus and then promoted the expression of BMP2, a key cytokine that facilitates osteogenesis. Metabolic reprogramming by localized fatty acid delivery partly rescued glucocorticoid-induced pathology by restoring a healthier immune-metabolic milieu. These data provide insights into the multifactorial metabolic mechanisms by which glucocorticoids generate skeletal disorders, thus suggesting possible therapeutic avenues., Introduction Synthetic glucocorticoids (GCs), acting as antiinflammatory and immunomodulatory agents, are the mainstays in the treatment of immune-related disorders (e.g., rheumatoid arthritis and Graves' disease). GCs are also prescribed to [...]

Nuclear factor [kappa]-B (NF[kappa]B) is activated in iPSC-cardiac myocytes from patients with arrhythmogenic cardiomyopathy (ACM) under basal conditions, and inhibition of NF[kappa]B signaling prevents disease in [Dsg2.sup.mut/mut] mice, a robust mouse model of ACM. Here, we used genetic approaches and single-cell RNA-Seq to define the contributions of immune signaling in cardiac myocytes and macrophages in the natural progression of ACM using [Dsg2.sup.mut/mut] mice. We found that NF[kappa]B signaling in cardiac myocytes drives myocardial injury, contractile dysfunction, and arrhythmias in [Dsg2.sup.mut/mut] mice. NF[kappa]B signaling in cardiac myocytes mobilizes macrophages expressing C-C motif chemokine receptor-2 ([CCR2.sup.+] cells) to affected areas within the heart, where they mediate myocardial injury and arrhythmias. Contractile dysfunction in [Dsg2.sup.mut/mut] mice is caused both by loss of heart muscle and negative inotropic effects of inflammation in viable muscle. Single nucleus RNA-Seq and cellular indexing of transcriptomes and epitomes (CITE-Seq) studies revealed marked proinflammatory changes in gene expression and the cellular landscape in hearts of [Dsg2.sup.mut/mut] mice involving cardiac myocytes, fibroblasts, and [CCR2.sup.+] macrophages. Changes in gene expression in cardiac myocytes and fibroblasts in [Dsg2.sup.mut/mut] mice were dependent on [CCR2.sup.+] macrophage recruitment to the heart. These results highlight complex mechanisms of immune injury and regulatory crosstalk between cardiac myocytes, inflammatory cells, and fibroblasts in the pathogenesis of ACM., Introduction Arrhythmogenic cardiomyopathy (ACM) is a familial heart muscle disease characterized by the early onset of arrhythmias and increased risk of sudden death followed by progressive myocardial injury ultimately leading [...]

Targeting tumor-associated macrophages (TAMs) is an emerging approach being tested in multiple clinical trials. TAMs, depending on their differentiation state, can exhibit pro- or antitumorigenic functions. For example, the M2-like phenotype represents a protumoral state that can stimulate tumor growth, angiogenesis, metastasis, therapy resistance, and immune evasion by expressing immune checkpoint proteins. In this issue of the JCI, Vaccaro and colleagues utilized an innovative drug screen approach to demonstrate that targeting driver oncogenic signaling pathways concurrently with anti-CD47 sensitizes tumor cells, causing them to undergo macrophage-induced phagocytosis. The combination treatment altered expression of molecules on the tumor cells that typically limit phagocytosis. It also reprogrammed macrophages to an M1-like antitumor state. Moreover, the approach was generalizable to tumor cells with different oncogenic pathways, opening the door to precision oncology-based rationale combination therapies that have the potential to improve outcomes for patients with oncogene-driven lung cancers and likely other cancer types., Macrophage checkpoint inhibitors expand the immunotherapy arsenal T cell checkpoint inhibitors (ICIs) have transformed the therapy landscape for a number of diseases; however, only a fraction of patients with cancer [...]

Macrophage immune checkpoint inhibitors, such as anti-CD47 antibodies, show promise in clinical trials for solid and hematologic malignancies. However, the best strategies to use these therapies remain unknown, and ongoing studies suggest they may be most effective when used in combination with other anticancer agents. Here, we developed an unbiased, high-throughput screening platform to identify drugs that render lung cancer cells more vulnerable to macrophage attack, and we found that therapeutic synergy exists between genotype-directed therapies and anti-CD47 antibodies. In validation studies, we found that the combination of genotype- directed therapies and CD47 blockade elicited robust phagocytosis and eliminated persister cells in vitro and maximized antitumor responses in vivo. Importantly, these findings broadly applied to lung cancers with various RTK/MAPK pathway alterations--including EGFR mutations, ALK fusions, or [KRAS.sup.G12C] mutations. We observed downregulation of [beta]2-microglobulin and CD73 as molecular mechanisms contributing to enhanced sensitivity to macrophage attack. Our findings demonstrate that dual inhibition of the RTK/MAPK pathway and the CD47/SIRPa axis is a promising immunotherapeutic strategy. Our study provides strong rationale for testing this therapeutic combination in patients with lung cancers bearing driver mutations., Introduction Many cancers arise from a single mutation in a proto-oncogene that drives tumor growth. These cancers are often treated with genotype-directed 'targeted therapies,' small-molecule inhibitors that specifically disable growth [...]

Neuropathic pain remains poorly managed by current therapies, highlighting the need to improve our knowledge of chronic pain mechanisms. In neuropathic pain models, dorsal root ganglia (DRG) nociceptive neurons transfer miR-21 packaged in extracellular vesicles to macrophages that promote a proinflammatory phenotype and contribute to allodynia. Here we show that miR-21 conditional deletion in DRG neurons was coupled with lack of upregulation of chemokine CCL2 after nerve injury and reduced accumulation of CCR2-expressing macrophages, which showed TGF-[beta]-related pathway activation and acquired an M2-like antinociceptive phenotype. Indeed, neuropathic allodynia was attenuated after conditional knockout of miR-21 and restored by TGF-[beta]R inhibitor (SB431542) administration. Since TGF-[beta]R2 and TGF-[beta]1 are known miR- 21 targets, we suggest that miR-21 transfer from injured neurons to macrophages maintains a proinflammatory phenotype via suppression of such an antiinflammatory pathway. These data support miR-21 inhibition as a possible approach to maintain polarization of DRG macrophages at an M2-like state and attenuate neuropathic pain., Introduction Peripheral neuropathy is a debilitating condition that often leads to severe and chronic neuropathic pain, for which novel treatment strategies are needed because current drugs have limited efficacy and [...]

Induction of lipid-laden foamy macrophages is a cellular hallmark of tuberculosis (TB) disease, which involves the transformation of infected phagolysosomes from a site of killing into a nutrient-rich replicative niche. Here, we show that a terpenyl nucleoside shed from Mycobacterium tuberculosis, 1-tuberculosinyladenosine (1-TbAd), caused lysosomal maturation arrest and autophagy blockade, leading to lipid storage in M1 macrophages. Pure 1-TbAd, or infection with terpenyl nucleoside-producing M. tuberculosis, caused intralysosomal and peribacillary lipid storage patterns that matched both the molecules and subcellular locations known in foamy macrophages. Lipidomics showed that 1-TbAd induced storage of triacylglycerides and cholesterylesters and that 1-TbAd increased M. tuberculosis growth under conditions of restricted lipid access in macrophages. Furthermore, lipidomics identified 1-TbAd-induced lipid substrates that define Gaucher's disease, Wolman's disease, and other inborn lysosomal storage diseases. These data identify genetic and molecular causes of M. tuberculosis-induced lysosomal failure, leading to successful testing of an agonist of TRPML1 calcium channels that reverses lipid storage in cells. These data establish the host-directed cellular functions of an orphan effector molecule that promotes survival in macrophages, providing both an upstream cause and detailed picture of lysosome failure in foamy macrophages., Introduction To benefit from immune cloaking and gain access to nutrients within host cells, intracellular bacteria must evade, withstand, or defeat host killing. Bacteria enter macrophages through phagocytosis, where infected [...]

Uncontrolled inflammation occurred in sepsis results in multiple organ injuries and shock, which contributes to the death of patients with sepsis. However, the regulatory mechanisms that restrict excessive inflammation are still elusive. Here, we identified an Ig-like receptor called signaling lymphocyte activation molecular family 7 (SLAMF7) as a key suppressor of inflammation during sepsis. We found that the expression of SLAMF7 on monocytes/macrophages was significantly elevated in patients with sepsis and in septic mice. SLAMF7 attenuated TLR-dependent MAPK and NF-[kappa]B signaling activation in macrophages by cooperating with Src homology 2-containing inositol-5'-phosphatase 1 (SHIP1). Furthermore, SLAMF7 interacted with SHIP1 and TNF receptor-associated factor 6 (TRAF6) to inhibit K63 ubiquitination of TRAF6. In addition, we found that tyrosine phosphorylation sites within the intracellular domain of SLAMF7 and the phosphatase domain of SHIP1 were indispensable for the interaction between SLAMF7, SHIP1, and TRAF6 and SLAMF7-mediated modulation of cytokine production. Finally, we demonstrated that SLAMF7 protected against lethal sepsis and endotoxemia by downregulating macrophage proinflammatory cytokines and suppressing inflammation-induced organ damage. Taken together, our findings reveal a negative regulatory role of SLAMF7 in polymicrobial sepsis, thus providing sights into the treatment of sepsis., Introduction Sepsis is one of the leading causes of in-hospital death, but the pathological mechanisms underlying this disease have yet to be uncovered (1). Defining the clinical criteria for the [...]

* Context.--Women with diabetes have increased stillbirth risk. Although the underlying pathophysiological processes are poorly understood, stillbirth is frequently related to abnormal placental structure and function. Objective.--To investigate placental morphology and cellular characteristics in the placentas of women with diabetes who had stillbirths and stillbirths of unexplained cause. Design.--Placentas from women with uncomplicated live births, live births in women with diabetes, unexplained stillbirths, and stillbirths related to diabetes (n = 10/group) underwent clinical histopathologic assessment and were also investigated using immunohistochemical staining to quantify syncytial nuclear aggregates, proliferation, trophoblast area, vascularization, T cells, placental macrophages (Hofbauer cells), and the receptor for advanced glycation end products. Results.--[Ki67.sup.+] cells were decreased in unexplained stillbirths compared with live births in women with diabetes. Both stillbirth groups had increased cytokeratin 7+/nuclear area compared with controls. Blood vessels/villi were decreased in unexplained stillbirth compared with live births from women with diabetes. Compared with uncomplicated controls, [CD163.sup.+] macrophages were increased in live births in women with diabetes and unexplained stillbirths, and further increased in stillbirths related to diabetes. There was no change in [CD3.sup.+] T cells or syncytial nuclear aggregates. Receptor for advanced glycation end products-positive cells were decreased in both stillbirth groups compared with diabetes-related live births. Co-localization of receptor for advanced glycation end products in macrophages was increased in both stillbirth groups compared with live birth groups. Conclusions.--Stillbirths related to diabetes exhibit placental phenotypic differences compared with live births. Further investigation of these parameters may provide understanding of the pathologic mechanisms of stillbirth and aid the development of stillbirth prevention strategies. (Arch Pathol Lab Med. 2021;145:82-89; doi: 10.5858/arpa.2019-0524-OA), Stillbirth is defined in the United Kingdom as intrauterine fetal death after 24-weeks gestation and in 2017 the stillbirth rate was 4.2 per 1000 total births. (1) The stillbirth rate [...]

The molecular mechanisms underlying obesity-induced increases in [beta] cell mass and the resulting [beta] cell dysfunction need to be elucidated further. Our study revealed that GPR92, expressed in islet macrophages, is modulated by dietary interventions in metabolic tissues. Therefore, we aimed to define the role of GPR92 in islet inflammation by using a high-fat diet- induced (HFD-induced) obese mouse model. GPR92-KO mice exhibited glucose intolerance and reduced insulin levels--despite the enlarged pancreatic islets--as well as increased islet macrophage content and inflammation level compared with WT mice. These results indicate that the lack of GPR92 in islet macrophages can cause [beta] cell dysfunction, leading to disrupted glucose homeostasis. Alternatively, stimulation with the GPR92 agonist farnesyl pyrophosphate results in the inhibition of HFD- induced islet inflammation and increased insulin secretion in WT mice, but not in GPR92-KO mice. Thus, our study suggests that GPR92 can be a potential target to alleviate [beta] cell dysfunction via the inhibition of islet inflammation associated with the progression of diabetes., Introduction Obesity-induced low-grade tissue inflammation manifests in multiple tissues, including pancreatic islets, and plays a critical role in mediating insulin resistance and [beta] cell dysfunction (1-3). Obesity-associated insulin resistance stimulates [...]

Interstitial cells of Cajal (ICCs) are pacemaker cells in the intestine, and their function can be compromised by loss of C-KIT expression. Macrophage activation has been identified in intestine affected by Hirschsprung disease-associated enterocolitis (HAEC). In this study, we examined proinflammatory macrophage activation and explored the mechanisms by which it downregulates C-KIT expression in ICCs in colon affected by HAEC. We found that macrophage activation and TNF-[alpha] production were dramatically increased in the proximal dilated colon of HAEC patients and 3-week-old [Ednrb.sup.-/-] mice. Moreover, ICCs lost their [C-KIT.sup.+] phenotype in the dilated colon, resulting in damaged pacemaker function and intestinal dysmotility. However, macrophage depletion or TNF-[alpha] neutralization led to recovery of ICC phenotype and restored their pacemaker function. In isolated ICCs, TNF-[alpha]-mediated phosphorylation of p65 induced overexpression of microRNA-221 (miR-221), resulting in suppression of C-KIT expression and pacemaker currents. We also identified a TNF-[alpha]/NF-[kappa]B/miR-221 pathway that downregulated C-KIT expression in ICCs in the colon affected by HAEC. These findings suggest the important roles of proinflammatory macrophage activation in a phenotypic switch of ICCs, representing a promising therapeutic target for HAEC., Introduction Hirschsprung disease (HSCR) is a birth defect characterized by the absence of ganglion cells in the distal bowel (1). HSCR-associated enterocolitis (HAEC), the most common and serious complication of [...]

Elevated hematocrit is associated with cardiovascular risk; however, the causality and mechanisms are unclear. The [JAK2.sup.V617F] ([JAK2.sup.VF]) mutation increases cardiovascular risk in myeloproliferative disorders and in clonal hematopoiesis. [JAK2.sup.VF] mice with elevated WBCs, platelets, and RBCs display accelerated atherosclerosis and macrophage erythrophagocytosis. To investigate whether selective erythroid [JAK2.sup.VF] expression promotes atherosclerosis, we developed hyperlipidemic erythropoietin receptor Cre mice that express [JAK2.sup.VF] in the erythroid lineage (VFEpoR mice). VFEpoR mice without elevated blood cell counts showed increased atherosclerotic plaque necrosis, erythrophagocytosis, and ferroptosis. Selective induction of erythrocytosis with low-dose erythropoietin further exacerbated atherosclerosis with prominent ferroptosis, lipid peroxidation, and endothelial damage. VFEpoR RBCs had reduced antioxidant defenses and increased lipid hydroperoxides. Phagocytosis of human or murine WT or [JAK2.sup.VF] RBCs by WT macrophages induced ferroptosis, which was prevented by the ferroptosis inhibitor liproxstatin-1. Liproxstatin-1 reversed increased atherosclerosis, lipid peroxidation, ferroptosis, and endothelial damage in VFEpoR mice and in [JAK2.sup.VF] chimeric mice simulating clonal hematopoiesis, but had no impact in controls. Erythroid lineage [JAK2.sup.VF] expression led to qualitative and quantitative defects in RBCs that exacerbated atherosclerosis. Phagocytosis of RBCs by plaque macrophages promoted ferroptosis, suggesting a therapeutic target for reducing RBC-mediated cardiovascular risk., Introduction Elevated hematocrit is associated with increased atherothrombotic cardiovascular disease (ACD), the number one cause of death and disability in Western societies (1, 2). Observational studies have shown that human [...]

Immune-regulatory mechanisms of drug-free remission in rheumatoid arthritis (RA) are unknown. We hypothesized that synovial tissue macrophages (STM), which persist in remission, contribute to joint homeostasis. We used single-cell transcriptomics to profile 32,000 STMs and identified phenotypic changes in patients with early/active RA, treatment-refractory/active RA and RA in sustained remission. Each clinical state was characterized by different frequencies of nine discrete phenotypic clusters within four distinct STM subpopulations with diverse homeostatic, regulatory and inflammatory functions. This cellular atlas, combined with deep-phenotypic, spatial and functional analyses of synovial biopsy fluorescent activated cell sorted STMs, revealed two STM subpopulations (MerTK.sup.posTREM2.sup.high and MerTK.sup.posLYVE1.sup.pos) with unique remission transcriptomic signatures enriched in negative regulators of inflammation. These STMs were potent producers of inflammation-resolving lipid mediators and induced the repair response of synovial fibroblasts in vitro. A low proportion of MerTK.sup.pos STMs in remission was associated with increased risk of disease flare after treatment cessation. Therapeutic modulation of MerTK.sup.pos STM subpopulations could therefore be a potential treatment strategy for RA. Multiple subpopulations of synovial tissue macrophages with varied transcriptional, phenotypic and functional features may contribute to disease flare and tissue repair in patients with active rheumatoid arthritis and patients in clinical remission., Author(s): Stefano Alivernini [sup.1] [sup.2] [sup.3] [sup.4] , Lucy MacDonald [sup.1] [sup.4] , Aziza Elmesmari [sup.1] [sup.4] , Samuel Finlay [sup.1] [sup.4] , Barbara Tolusso [sup.2] , Maria Rita Gigante [...]