RESUMO
Early atherosclerosis depends upon responses by immune cells resident in the intimal aortic wall. Specifically, the healthy intima is thought to be populated by vascular dendritic cells (DCs) that, during hypercholesterolemia, initiate atherosclerosis by being the first to accumulate cholesterol. Whether these cells remain key players in later stages of disease is unknown. Using murine lineage-tracing models and gene expression profiling, we reveal that myeloid cells present in the intima of the aortic arch are not DCs but instead specialized aortic intima resident macrophages (MacAIR) that depend upon colony-stimulating factor 1 and are sustained by local proliferation. Although MacAIR comprise the earliest foam cells in plaques, their proliferation during plaque progression is limited. After months of hypercholesterolemia, their presence in plaques is overtaken by recruited monocytes, which induce MacAIR-defining genes. These data redefine the lineage of intimal phagocytes and suggest that proliferation is insufficient to sustain generations of macrophages during plaque progression.
Assuntos
Aorta/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Placa Aterosclerótica/imunologia , Túnica Íntima/imunologia , Animais , Diferenciação Celular , Linhagem da Célula , Movimento Celular , Proliferação de Células , Células Cultivadas , Colesterol/metabolismo , Progressão da Doença , Humanos , Fator Estimulador de Colônias de Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Parabiose , FagocitoseRESUMO
BACKGROUND: Trem2 (triggering receptor on myeloid cells 2), a surface lipid receptor, is expressed on foamy macrophages within atherosclerotic lesions and regulates cell survival, proliferation, and anti-inflammatory responses. Studies examining the role of Trem2 in atherosclerosis have shown that deletion of Trem2 leads to impaired foamy macrophage lipid uptake, proliferation, survival, and cholesterol efflux. Thus, we tested the hypothesis that administration of a Trem2 agonist antibody (AL002a) to atherogenic mice would enhance macrophage survival and decrease necrotic core formation to improve plaque stability. METHODS: To model a therapeutic intervention approach, atherosclerosis-prone mice (Ldlr [low-density lipoprotein receptor]-/-) were fed a high-fat diet for 8 weeks, then transitioned to treatment with AL002a or isotype control for an additional 8 weeks while continuing on a high-fat diet. RESULTS: AL002a-treated mice had increased lesion size in both the aortic root and whole mount aorta, which correlated with an expansion of plaque macrophage area. This expansion was due to increased macrophage survival and proliferation in plaques. Importantly, plaques from AL002a-treated mice showed improved features of plaque stability, including smaller necrotic cores, increased fibrous caps, and greater collagen deposition. Single-cell RNA sequencing of whole aorta suspensions from isotype- and AL002a-treated atherosclerotic mice revealed that Trem2 agonism dramatically altered foamy macrophage transcriptome. This included upregulation of oxidative phosphorylation and increased expression of collagen genes. In vitro studies validated that Trem2 agonism with AL002a promoted foamy macrophage oxidized low-density lipoprotein uptake, survival, and cholesterol efflux. CONCLUSIONS: Trem2 agonism expands atherosclerotic plaque macrophages by promoting cell survival and proliferation but improves features of plaque stability by rewiring foamy macrophage function to enhance cholesterol efflux and collagen deposition.
Assuntos
Aterosclerose , Modelos Animais de Doenças , Células Espumosas , Glicoproteínas de Membrana , Camundongos Endogâmicos C57BL , Camundongos Knockout , Placa Aterosclerótica , Receptores Imunológicos , Animais , Receptores Imunológicos/agonistas , Receptores Imunológicos/metabolismo , Receptores Imunológicos/genética , Glicoproteínas de Membrana/agonistas , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/genética , Camundongos , Aterosclerose/patologia , Aterosclerose/metabolismo , Aterosclerose/genética , Aterosclerose/tratamento farmacológico , Aterosclerose/prevenção & controle , Células Espumosas/metabolismo , Células Espumosas/patologia , Células Espumosas/efeitos dos fármacos , Masculino , Receptores de LDL/genética , Receptores de LDL/metabolismo , Receptores de LDL/deficiência , Proliferação de Células/efeitos dos fármacos , Dieta Hiperlipídica , Sobrevivência Celular/efeitos dos fármacos , Necrose , Doenças da Aorta/patologia , Doenças da Aorta/genética , Doenças da Aorta/metabolismo , Doenças da Aorta/prevenção & controleRESUMO
Glucocorticoid synthesis by adrenal glands (AGs) is regulated by the hypothalamic-pituitary-adrenal axis to facilitate stress responses when the host is exposed to stimuli. Recent studies implicate macrophages as potential steroidogenic regulators, but the molecular mechanisms by which AG macrophages exert such influence remain unclear. In this study, we investigated the role of AG macrophages in response to cold challenge or atherosclerotic inflammation as physiologic models of acute or chronic stress. Using single-cell RNA sequencing, we observed dynamic AG macrophage polarization toward classical activation and lipid-associated phenotypes following acute or chronic stimulation. Among transcriptional alterations induced in macrophages, triggering receptor expressed on myeloid cells 2 (Trem2) was highlighted because of its upregulation following stress. Conditional deletion of macrophage Trem2 revealed a protective role in stress responses. Mechanistically, Trem2 deletion led to increased AG macrophage death, abolished the TGF-ß-producing capacity of AG macrophages, and resulted in enhanced glucocorticoid production. In addition, enhanced glucocorticoid production was replicated by blockade of TGF-ß signaling. Together, these observations suggest that AG macrophages restrict steroidogenesis through Trem2 and TGF-ß, which opens potential avenues for immunotherapeutic interventions to resolve stress-related disorders.
Assuntos
Glândulas Suprarrenais , Glucocorticoides , Macrófagos , Glicoproteínas de Membrana , Receptores Imunológicos , Fator de Crescimento Transformador beta , Animais , Macrófagos/metabolismo , Macrófagos/imunologia , Camundongos , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/genética , Glucocorticoides/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Receptores Imunológicos/metabolismo , Receptores Imunológicos/genética , Glândulas Suprarrenais/metabolismo , Masculino , Camundongos Knockout , Camundongos Endogâmicos C57BL , Transdução de SinaisRESUMO
Pancreatic ductal adenocarcinoma (PDA) orchestrates a suppressive tumor microenvironment that fosters immunotherapy resistance. Tumor-associated macrophages (TAMs) are the principal immune cell infiltrating PDA and are heterogeneous. Here, by employing macrophage fate-mapping approaches and single-cell RNA sequencing, we show that monocytes give rise to most macrophage subsets in PDA. Tumor-specific CD4, but not CD8, T cells promote monocyte differentiation into MHCIIhi anti-tumor macrophages. By conditional major histocompatibility complex (MHC) class II deletion on monocyte-derived macrophages, we show that tumor antigen presentation is required for instructing monocyte differentiation into anti-tumor macrophages, promoting Th1 cells, abrogating Treg cells, and mitigating CD8 T cell exhaustion. Non-redundant IFNγ and CD40 promote MHCIIhi anti-tumor macrophages. Intratumoral monocytes adopt a pro-tumor fate indistinguishable from that of tissue-resident macrophages following loss of macrophage MHC class II or tumor-specific CD4 T cells. Thus, tumor antigen presentation by macrophages to CD4 T cells dictates TAM fate and is a major determinant of macrophage heterogeneity in cancer.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Monócitos , Linfócitos T CD4-Positivos , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/genética , Antígenos de Neoplasias , Antígenos de Histocompatibilidade Classe II , Microambiente Tumoral , Neoplasias PancreáticasRESUMO
Atherosclerosis is driven by the expansion of cholesterol-loaded 'foamy' macrophages in the arterial intima. Factors regulating foamy macrophage differentiation and survival in plaque remain poorly understood. Here we show, using trajectory analysis of integrated single-cell RNA sequencing data and a genome-wide CRISPR screen, that triggering receptor expressed on myeloid cells 2 (Trem2) is associated with foamy macrophage specification. Loss of Trem2 led to a reduced ability of foamy macrophages to take up oxidized low-density lipoprotein (oxLDL). Myeloid-specific deletion of Trem2 showed an attenuation of plaque progression, even when targeted in established atherosclerotic lesions, and was independent of changes in circulating cytokines, monocyte recruitment or cholesterol levels. Mechanistically, we link Trem2-deficient macrophages with a failure to upregulate cholesterol efflux molecules, resulting in impaired proliferation and survival. Overall, we identify Trem2 as a regulator of foamy macrophage differentiation and atherosclerotic plaque growth and as a putative therapeutic target for atherosclerosis.
RESUMO
Tissue-resident macrophages are present in all tissues where they perform homeostatic and immune surveillance functions. In many tissues, resident macrophages develop from embryonic progenitors, which mature into a self-maintaining population through local proliferation. However, tissue-resident macrophages can be supported by recruited monocyte-derived macrophages during scenarios such as tissue growth, infection, or sterile inflammation. Circulating blood monocytes arise from hematopoietic stem cell progenitors and possess unique gene profiles that support additional functions within the tissue. Determining cell origins (ontogeny) and cellular turnover within tissues has become important to understanding monocyte and macrophage contributions to tissue homeostasis and disease. Fate mapping, or lineage tracing, is a promising approach to tracking cells based on unique gene expression driving reporter systems, often downstream of a Cre-recombinase-mediated excision event, to express a fluorescent protein. This approach is typically deployed temporally with developmental stage, disease onset, or in association with key stages of inflammation resolution. Importantly, myeloid fate mapping can be combined with many emerging technologies, including single-cell RNA-sequencing and spatial imaging. The application of myeloid cell fate mapping approaches has allowed for impactful discoveries regarding myeloid ontogeny, tissue residency, and monocyte fate within disease models. This protocol outline will discuss a variety of myeloid fate mapping approaches, including constitutive and inducible labeling approaches in adult and embryo tissues. This article outlines basic approaches and models used in mice for fate mapping macrophages. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Adult Fate Mapping Basic Protocol 2: Embryonic Fate Mapping.
Assuntos
Macrófagos , Monócitos , Animais , Diferenciação Celular/fisiologia , Células-Tronco Hematopoéticas , Inflamação/metabolismo , Camundongos , Monócitos/metabolismoRESUMO
Despite the ubiquitous function of macrophages across the body, the diversity, origin, and function of adrenal gland macrophages remain largely unknown. We define the heterogeneity of adrenal gland immune cells using single-cell RNA sequencing and use genetic models to explore the developmental mechanisms yielding macrophage diversity. We define populations of monocyte-derived and embryonically seeded adrenal gland macrophages and identify a female-specific subset with low major histocompatibility complex (MHC) class II expression. In adulthood, monocyte recruitment dominates adrenal gland macrophage maintenance in female mice. Adrenal gland macrophage sub-tissular distribution follows a sex-dimorphic pattern, with MHC class IIlow macrophages located at the cortico-medullary junction. Macrophage sex dimorphism depends on the presence of the cortical X-zone. Adrenal gland macrophage depletion results in altered tissue homeostasis, modulated lipid metabolism, and decreased local aldosterone production during stress exposure. Overall, these data reveal the heterogeneity of adrenal gland macrophages and point toward sex-restricted distribution and functions of these cells.