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Introduction: Innate immunity is crucial to reducing parasite burden and contributing to survival in severe malaria. Monocytes are key actors in the innate response and, like macrophages, are plastic cells whose function and phenotype are regulated by the signals from the microenvironment. In the context of cerebral malaria (CM), monocyte response constitutes an important issue to understand. We previously demonstrated that decreased percentages of nonclassical monocytes were associated with death outcomes in CM children. In the current study, we postulated that monocyte phagocytosis function is impacted by the severity of malaria infection. Methods: To study this hypothesis, we compared the opsonic and nonopsonic phagocytosis capacity of circulant monocytes from Beninese children with uncomplicated malaria (UM) and CM. For the CM group, samples were obtained at inclusion (D0) and 3 and 30 days after treatment (D3, D30). The phagocytosis capacity of monocytes and their subsets was characterized by flow cytometry and transcriptional profiling by studying genes known for their functional implication in infected-red blood cell (iRBC) elimination or immune escape. Results: Our results confirm our hypothesis and highlight the higher capacity of nonclassical monocytes to phagocyte iRBC. We also confirm that a low number of nonclassical monocytes is associated with CM outcome when compared to UM, suggesting a mobilization of this subpopulation to the cerebral inflammatory site. Finally, our results suggest the implication of the inhibitory receptors LILRB1, LILRB2, and Tim3 in phagocytosis control. Discussion: Taken together, these data provide a better understanding of the interplay between monocytes and malaria infection in the pathogenicity of CM.

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Malaria Cerebral , Monocitos , Fagocitosis , Humanos , Malaria Cerebral/inmunología , Malaria Cerebral/parasitología , Monocitos/inmunología , Masculino , Preescolar , Femenino , Niño , Lactante , Plasmodium falciparum/inmunología , Proteínas Opsoninas/metabolismo , Proteínas Opsoninas/inmunología , Eritrocitos/parasitología , Eritrocitos/inmunología , Inmunidad Innata

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Ly6Clo monocytes are a myeloid subset that specializes in the surveillance of vascular endothelium. Ly6Clo monocytes have been shown to derive from Ly6Chi monocytes. NOTCH2 signaling has been implicated as a trigger for Ly6Clo monocyte development, but the basis for this effect is unclear. Here, we examined the impact of NOTCH2 signaling of myeloid progenitors on the development of Ly6Clo monocytes in vitro. NOTCH2 signaling induced by delta-like ligand 1 (DLL1) efficiently induced the transition of Ly6Chi TREML4- monocytes into Ly6Clo TREML4+ monocytes. We further identified two additional transcriptional requirements for development of Ly6Clo monocytes. Deletion of BCL6 from myeloid progenitors abrogated development of Ly6Clo monocytes. IRF2 was also required for Ly6Clo monocyte development in a cell-intrinsic manner. DLL1-induced in vitro transition into Ly6Clo TREML4+ monocytes required IRF2 but unexpectedly could occur in the absence of NUR77 or BCL6. These results imply a transcriptional hierarchy for these factors in controlling Ly6Clo monocyte development.

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Endotelio Vascular , Monocitos , Hematopoyesis , Transducción de Señal

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Background: Fully mature monocytes that express CD14, but not CD16, undergo phagocytosis within tissues, whereas non-classical monocytes, CD14-low CD16+, represent <11% of peripheral monocytes and have primary pro-inflammatory functions. Inflammation plays a major role in Covid-19 disease and adds to the inflammation caused by chronic hemodialysis. The aim of our study was to monitor monocyte subsets in five patients with end-stage kidney disease (ESKD) over a 1-year period after a mild Covid-19 infection. Five ESKD patients with a mild Covid-19 infection were monitored using CD14, CD16, CD300e, HLA-DR, CD64, and CD45 panels using a BD FACS Canto flow cytometer. Results: CD14-low CD16+ was dramatically (p=0,001) decreased in patients during Covid-19 infection, as previously described for patients without chronic renal failure. In addition, CD14-low CD16+ monocytes remained decreased for 10 months after recovery from Covid. Intermediate monocytes increased during Covid-19 infection and decreased 10 months after infection but this subtype of monocytes retained their inflammatory activity with a significant increase in HLA-DR expression after recovery from Covid infection. Conclusion: Our study shows that ESKD patients had a pro-inflammatory profile induced by Covid 19, but this status was prolonged significantly over a 10-month period. Thus, advanced renal failure treated by hemodialysis did not dramatically change the inflammatory response against to SARS Covid 2. It seems that monocytes retain their inflammatory status for many months in ESKD patients after a Covid-19 infection.

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COVID-19 , Fallo Renal Crónico , Humanos , Monocitos , SARS-CoV-2 , Diálisis Renal , Fallo Renal Crónico/terapia , Inflamación

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BACKGROUND: Emerging evidence from mouse models is beginning to elucidate the brain's immune response to tau pathology, but little is known about the nature of this response in humans. In addition, it remains unclear to what extent tau pathology and the local inflammatory response within the brain influence the broader immune system. METHODS: To address these questions, we performed single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) from carriers of pathogenic variants in MAPT, the gene encoding tau (n = 8), and healthy non-carrier controls (n = 8). Primary findings from our scRNA-seq analyses were confirmed and extended via flow cytometry, droplet digital (dd)PCR, and secondary analyses of publicly available transcriptomics datasets. RESULTS: Analysis of ~ 181,000 individual PBMC transcriptomes demonstrated striking differential expression in monocytes and natural killer (NK) cells in MAPT pathogenic variant carriers. In particular, we observed a marked reduction in the expression of CX3CR1-the gene encoding the fractalkine receptor that is known to modulate tau pathology in mouse models-in monocytes and NK cells. We also observed a significant reduction in the abundance of nonclassical monocytes and dysregulated expression of nonclassical monocyte marker genes, including FCGR3A. Finally, we identified reductions in TMEM176A and TMEM176B, genes thought to be involved in the inflammatory response in human microglia but with unclear function in peripheral monocytes. We confirmed the reduction in nonclassical monocytes by flow cytometry and the differential expression of select biologically relevant genes dysregulated in our scRNA-seq data using ddPCR. CONCLUSIONS: Our results suggest that human peripheral immune cell expression and abundance are modulated by tau-associated pathophysiologic changes. CX3CR1 and nonclassical monocytes in particular will be a focus of future work exploring the role of these peripheral signals in additional tau-associated neurodegenerative diseases.

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Monocitos , Tauopatías , Ratones , Animales , Humanos , Monocitos/metabolismo , Leucocitos Mononucleares , Análisis de Expresión Génica de una Sola Célula , Tauopatías/genética , Tauopatías/metabolismo , Tauopatías/patología , Microglía/metabolismo , Análisis de la Célula Individual , Receptor 1 de Quimiocinas CX3C/genética , Receptor 1 de Quimiocinas CX3C/metabolismo , Proteínas de la Membrana/metabolismo

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CD8+ T cells drive anti-cancer immunity in response to antigen-presenting cells such as dendritic cells and subpopulations of monocytes and macrophages. While CD14+ classical monocytes modulate CD8+ T cell responses, the contributions of CD16+ nonclassical monocytes to this process remain unclear. Herein we explored the role of nonclassical monocytes in CD8+ T cell activation by utilizing E2-deficient (E2-/-) mice that lack nonclassical monocytes. During early metastatic seeding, modeled by B16F10-OVA cancer cells injected into E2-/- mice, we noted lower CD8+ effector memory and effector T cell frequencies within the lungs as well as in lung-draining mediastinal lymph nodes in the E2-/- mice. Analysis of the myeloid compartment revealed that these changes were associated with depletion of MHC-IIloLy6Clo nonclassical monocytes within these tissues, with little change in other monocyte or macrophage populations. Additionally, nonclassical monocytes preferentially trafficked to primary tumor sites in the lungs, rather than to the lung-draining lymph nodes, and did not cross-present antigen to CD8+ T cells. Examination of the lung microenvironment in E2-/- mice revealed reduced CCL21 expression in endothelial cells, which is chemokine involved in T cell trafficking. Our results highlight the previously unappreciated importance of nonclassical monocytes in shaping the tumor microenvironment via CCL21 production and CD8+ T cell recruitment.

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Monocitos , Neoplasias , Ratones , Animales , Linfocitos T CD8-positivos , Células Endoteliales , Pulmón , Neoplasias/metabolismo , Microambiente Tumoral

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Social determinants of health (SDoH) include socioeconomic, environmental, and psychological factors that impact health. Neighborhood socioeconomic deprivation (NSD) and low individual-level socioeconomic status (SES) are SDoH that associate with incident heart failure, stroke, and cardiovascular mortality, but the underlying biological mechanisms are not well understood. Previous research has demonstrated an association between NSD, in particular, and key components of the neural-hematopoietic-axis including amygdala activity as a marker of chronic stress, bone marrow activity, and arterial inflammation. Our study further characterizes the role of NSD and SES as potential sources of chronic stress related to downstream immunological factors in this stress-associated biologic pathway. We investigated how NSD, SES, and catecholamine levels (as proxy for sympathetic nervous system activation) may influence monocytes which are known to play a significant role in atherogenesis. First, in an ex vivo approach, we treated healthy donor monocytes with biobanked serum from a community cohort of African Americans at risk for CVD. Subsequently, the treated monocytes were subjected to flow cytometry for characterization of monocyte subsets and receptor expression. We determined that NSD and serum catecholamines (namely dopamine [DA] and norepinephrine [NE]) associated with monocyte C-C chemokine receptor type 2 (CCR2) expression (p < 0.05), a receptor known to facilitate recruitment of monocytes towards arterial plaques. Additionally, NSD associated with catecholamine levels, especially DA in individuals of low SES. To further explore the potential role of NSD and the effects of catecholamines on monocytes, monocytes were treated in vitro with epinephrine [EPI], NE, or DA. Only DA increased CCR2 expression in a dose-dependent manner (p < 0.01), especially on non-classical monocytes (NCM). Furthermore, linear regression analysis between D2-like receptor surface expression and surface CCR2 expression suggested D2-like receptor signaling in NCM. Indicative of D2-signaling, cAMP levels were found to be lower in DA-treated monocytes compared to untreated controls (control 29.78 pmol/ml vs DA 22.97 pmol/ml; p = 0.038) and the impact of DA on NCM CCR2 expression was abrogated by co-treatment with 8-CPT, a cAMP analog. Furthermore, Filamin A (FLNA), a prominent actin-crosslinking protein, that is known to regulate CCR2 recycling, significantly decreased in DA-treated NCM (p < 0.05), indicating a reduction of CCR2 recycling. Overall, we provide a novel immunological mechanism, driven by DA signaling and CCR2, for how NSD may contribute to atherogenesis. Future studies should investigate the importance of DA in CVD development and progression in populations disproportionately experiencing chronic stress due to SDoH.

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[This corrects the article DOI: 10.3389/fimmu.2022.889175.].

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INTRODUCTION: Neutrophils are crucial to antimicrobial defense, but excessive neutrophilic inflammation elicits immune pathology. Currently, no effective treatment exists to curb neutrophil activation. However, neutrophils express a variety of inhibitory receptors which may represent potential therapeutic targets to limit neutrophilic inflammation. Indeed, we previously showed that the inhibitory collagen receptor leukocyte-associated immunoglobulin-like receptor 1 (LAIR-1) regulates neutrophilic airway inflammation and inhibits neutrophil extracellular trap formation. The inhibitory receptor Allergin-1 is expressed by myeloid cells and B cells. Allergin-1 suppresses mast cell and basophil activation, but a potential regulatory role on neutrophils remains unexplored. We aimed to demonstrate the regulation of neutrophils by Allergin-1. METHODS: We examine Allergin-1 isoform expression on human neutrophils during homeostatic (healthy donors) and chronic inflammatory (systemic lupus erythematosus patients) conditions in comparison to other circulating leukocytes by flow cytometry. To reveal a potential role for Allergin-1 in regulating neutrophilic inflammation, we experimentally infect wild-type (WT) and Allergin-1-deficient mice with a respiratory syncytial virus (RSV) and monitor disease severity and examine cellular airway infiltrate. Flow cytometry was used to confirm Allergin-1 expression by airway-infiltrated neutrophils in RSV infection-induced bronchiolitis patients. RESULTS: Only the short 1 (S1) isoform, but not the long (L) or S2 isoform could be detected on blood leukocytes, with the exception of nonclassical monocytes, which exclusively express the S2 isoform. Allergin-1 expression levels did not vary significantly between healthy individuals and patients with the systemic inflammatory disease on any interrogated cell type. Airway-infiltrated neutrophils of pediatric RSV bronchiolitis patients were found to express Allergin-1S1. However, Allergin-1-deficient mice experimentally infected with RSV did not show exacerbated disease or increased neutrophil airway infiltration compared to WT littermates. CONCLUSION: Allergin-1 isoform expression is unaffected by chronic inflammatory conditions. In stark contrast to fellow inhibitory receptor LAIR-1, Allergin-1 does not regulate neutrophilic inflammation in a mouse model of RSV bronchiolitis.

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Bronquiolitis , Inflamación , Receptores Inmunológicos , Infecciones por Virus Sincitial Respiratorio , Animales , Niño , Humanos , Ratones , Inflamación/genética , Inflamación/metabolismo , Neutrófilos , Isoformas de Proteínas/genética , Receptores Inmunológicos/genética , Receptores Inmunológicos/metabolismo , Infecciones por Virus Sincitial Respiratorio/genética , Infecciones por Virus Sincitial Respiratorio/metabolismo , Virus Sincitiales Respiratorios

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The differential contribution of monocyte subsets expressing the C-C chemokine receptor 2 (CCR2) to subclinical atherosclerosis in girls and boys is unclear. In this pilot study, we compared classical, intermediate, and nonclassical monocyte subsets expressing CCR2 in 33 obese children of both sexes aged 8 to 16 divided by carotid intima-media thickness (IMT), considering values above the 75th percentile (p75) as abnormally high IMT. Obesity was defined as body mass index above the 95th percentile according to age and sex. Flow cytometry analyses revealed that boys but not girls with IMT ≥ p75 displayed increased CCR2+ cell percentage and CCR2 expression in the three monocyte subsets, compared to boys with IMT < p75. The CCR2+ cell percentage and CCR2 expression in the three monocyte subsets significantly correlated with increased IMT and insulin resistance in boys but not girls, where the CCR2+ nonclassical monocyte percentage had the strongest associations (r = 0.73 and r = 0.72, respectively). The role of CCR2+ monocyte subpopulations in identifying an abnormally high IMT shows a marked sexual dimorphism, where boys seem to be at higher subclinical atherosclerosis risk than girls.

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Similar to human monocytes, bovine monocytes can be split into CD14highCD16- classical, CD14highCD16high intermediate and CD14-/dimCD16high nonclassical monocytes (cM, intM, and ncM, respectively). Here, we present an in-depth analysis of their steady-state bulk- and single-cell transcriptomes, highlighting both pronounced functional specializations and transcriptomic relatedness. Bulk gene transcription indicates pro-inflammatory and antibacterial roles of cM, while ncM and intM appear to be specialized in regulatory/anti-inflammatory functions and tissue repair, as well as antiviral responses and T-cell immunomodulation. Notably, intM stood out by high expression of several genes associated with antigen presentation. Anti-inflammatory and antiviral functions of ncM are further supported by dominant oxidative phosphorylation and selective strong responses to TLR7/8 ligands, respectively. Moreover, single-cell RNA-seq revealed previously unappreciated heterogeneity within cM and proposes intM as a transient differentiation intermediate between cM and ncM.

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Presentación de Antígeno , Monocitos , Animales , Antivirales/metabolismo , Biología , Bovinos , Diferenciación Celular , Humanos

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Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can result in severe immune dysfunction, hospitalization, and death. Many patients also develop long-COVID-19, experiencing symptoms months after infection. Although significant progress has been made in understanding the immune response to acute SARS-CoV-2 infection, gaps remain in our knowledge of how innate immunity influences disease kinetics and severity. We hypothesized that cytometry by time-of-flight analysis of PBMCs from healthy and infected subjects would identify novel cell surface markers and innate immune cell subsets associated with COVID-19 severity. In this pursuit, we identified monocyte and dendritic cell subsets that changed in frequency during acute SARS-CoV-2 infection and correlated with clinical parameters of disease severity. Subsets of nonclassical monocytes decreased in frequency in hospitalized subjects, yet increased in the most severe patients and positively correlated with clinical values associated with worse disease severity. CD9, CD163, PDL1, and PDL2 expression significantly increased in hospitalized subjects, and CD9 and 6-Sulfo LacNac emerged as the markers that best distinguished monocyte subsets amongst all subjects. CD9+ monocytes remained elevated, whereas nonclassical monocytes remained decreased, in the blood of hospitalized subjects at 3-4 months postinfection. Finally, we found that CD9+ monocytes functionally released more IL-8 and MCP-1 after LPS stimulation. This study identifies new monocyte subsets present in the blood of COVID-19 patients that correlate with disease severity, and links CD9+ monocytes to COVID-19 progression.

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COVID-19 , Humanos , Monocitos , SARS-CoV-2 , Interleucina-8/metabolismo , Lipopolisacáridos/metabolismo , Células Mieloides , Hospitalización , Tetraspanina 29/metabolismo , Síndrome Post Agudo de COVID-19

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Nonclassical monocytes maintain vascular homeostasis by patrolling the vascular endothelium, responding to inflammatory signals, and scavenging cellular debris. Nonclassical monocytes also prevent metastatic tumor cells from seeding new tissues, but whether the patrolling function of nonclassical monocytes is required for this process is unknown. To answer this question, we utilized an inducible-knockout mouse that exhibits loss of the integrin-adaptor protein Kindlin-3 specifically in nonclassical monocytes. We show that Kindlin-3-deficient nonclassical monocytes are unable to patrol the vascular endothelium in either the lungs or periphery. We also find that Kindlin-3-deficient nonclassical monocytes cannot firmly adhere to, and instead "slip" along, the vascular endothelium. Loss of patrolling activity by nonclassical monocytes was phenocopied by ablation of LFA-1, an integrin-binding partner of Kindlin-3. When B16F10 murine melanoma tumor cells were introduced into Kindlin-3-deficient mice, nonclassical monocytes showed defective patrolling towards tumor cells and failure to ingest tumor particles in vivo. Consequently, we observed a significant, 4-fold increase in lung tumor metastases in mice possessing Kindlin-3-deficient nonclassical monocytes. Thus, we conclude that the patrolling function of nonclassical monocytes is mediated by Kindlin-3 and essential for these cells to maintain vascular endothelial homeostasis and prevent tumor metastasis to the lung.

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Proteínas del Citoesqueleto/genética , Regulación Neoplásica de la Expresión Génica , Antígeno-1 Asociado a Función de Linfocito/genética , Melanoma Experimental/genética , Monocitos/inmunología , Fagocitosis , Neoplasias Cutáneas/genética , Animales , Médula Ósea/inmunología , Trasplante de Médula Ósea , Adhesión Celular , Comunicación Celular/inmunología , Proteínas del Citoesqueleto/deficiencia , Proteínas del Citoesqueleto/inmunología , Endotelio Vascular/inmunología , Endotelio Vascular/patología , Femenino , Humanos , Inyecciones Intravenosas , Pulmón/irrigación sanguínea , Pulmón/inmunología , Pulmón/patología , Antígeno-1 Asociado a Función de Linfocito/inmunología , Melanoma Experimental/inmunología , Melanoma Experimental/secundario , Ratones , Ratones Noqueados , Monocitos/patología , Células Neoplásicas Circulantes/inmunología , Células Neoplásicas Circulantes/patología , Cultivo Primario de Células , Transducción de Señal , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/patología , Irradiación Corporal Total

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Neutrophils and monocytes encompassing the classical, intermediate, and nonclassical population constitute the majority of circulating myeloid cells in humans and represent the first line of innate immune defense. As such, changes in their relative and absolute amounts serve as sensitive markers of diverse inflammatory conditions. Multiple sclerosis (MS) is a chronic autoimmune disease of the central nervous system, causing demyelination and axonal loss, affecting various neuron functions and often causing irreversible neurological disability. MS disease course is individually highly heterogeneous but can be classified as progressive (PMS) or relapsing-remitting (RRMS). Each MS course type may be further characterized as active or inactive, depending on the recent disability progression and/or current relapses. Data on specific alterations of the myeloid compartment in association with MS disease course are scarce and conflicting. In the current study, we systematically immunophenotyped blood myeloid leukocytes by flow cytometry in 15 healthy and 65 MS subjects. We found a highly significant expansion of granulocytes, CD15+ neutrophils, and classical and nonclassical monocytes in inactive RRMS (RRMSi) with concomitant shrinkage of the lymphocyte compartment, which did not correlate with biochemical readouts of systemic inflammation. Each of these leukocyte populations and the combined myeloid signature accurately differentiated RRMSi from other MS forms. Additionally, nonclassical monocyte proportions were particularly elevated in RRMSi individuals receiving disease-modifying therapy (DMT), such as natalizumab. Our results suggest that flow cytometry-based myeloid cell immunophenotyping in MS may help to identify RRMSi earlier and facilitate monitoring of DMT response.

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Biomarcadores/sangre , Monocitos/inmunología , Esclerosis Múltiple Recurrente-Remitente/diagnóstico , Esclerosis Múltiple Recurrente-Remitente/inmunología , Neutrófilos/inmunología , Adulto , Anciano , Femenino , Citometría de Flujo/métodos , Humanos , Factores Inmunológicos/uso terapéutico , Inmunofenotipificación/métodos , Masculino , Persona de Mediana Edad , Monocitos/efectos de los fármacos , Esclerosis Múltiple Recurrente-Remitente/tratamiento farmacológico , Natalizumab/uso terapéutico , Neutrófilos/efectos de los fármacos

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AIM: To establish the effect of poly(acrylic acid)-coated iron oxide nanoparticles (PAC-IONs) and later exposure to a magnetic field on the differentiation of mononuclear phagocytes into macrophages. METHODS: By flow cytometry, cell death was evaluated with DIOC6 and PI, Poly (ADP-ribose) Polymerases (PARP) fragmentation, H2AX phosphorylation and TUNEL assay. Cytokines by Cytokine bead array and the intracellular amount of iron by atomic absorption spectrometry. RESULTS: PAC-IONs did not induce apoptosis, modify the cell membrane integrity or alter the mitochondrial membrane potential. They did not affect the cell morphology, the pattern of cytokine accumulation or the activating role of differentiation of mononuclear phagocytes into macrophages on the proliferation of autologous T cells. CONCLUSION: This evidence indicates that the PAC-IONs are safe and biocompatible. Moreover, the selectivity of the PAC-IONs for mononuclear phagocytes, as well as their increased uptake by non-classical monocytes, warrant future research with a view to their use as a contrast agent, a useful tool for in vivo tracking of tissue-infiltrating mononuclear phagocytes.

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Síndrome Coronario Agudo , Monocitos , Receptor 1 de Quimiocinas CX3C , Humanos , Receptores de Quimiocina