Monocyte Function Research Articles

Pro-inflammatory Stimulation of Monocytes by ANCA Is Linked to Changes in Cellular Metabolism.

Clinical and experimental data suggest that pathogenesis in anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis is driven by ANCA-mediated activation of neutrophils and monocytes. While the role of neutrophils has been extensively investigated, the function of monocytes remains relatively understudied. We have previously demonstrated that stimulation of monocytes with anti-myeloperoxidase (MPO), but not anti-proteinase-3 (PR3), antibodies results in production of the pro-inflammatory cytokine IL-1β. Changes in cellular metabolism, particularly a switch to glycolysis, have recently been linked to activation of immune cells and production of IL-1β. Therefore, we investigated the metabolic profile of monocytes following ANCA stimulation. We found a significant increase in glucose uptake in anti-MPO stimulated monocytes. Interestingly, both anti-MPO and anti-PR3 stimulation resulted in an immediate increase in glycolysis, measured by Seahorse extracellular flux analysis. However, this increase in glycolysis was sustained (for up to 4 h) in anti-MPO- but not anti-PR3-treated cells. In addition, only anti-MPO-treated cells exhibited increased oxidative phosphorylation, a metabolic response that correlated with IL-1β production. These data indicate that monocyte metabolism is altered by ANCA, with divergent responses to anti-MPO and anti-PR3 antibodies. These metabolic changes may underlie pathologic immune activation in ANCA associated vasculitis, as well as potentially contributing to the differing clinical phenotype between PR3- and MPO-ANCA positive patients. These metabolic pathways may therefore be potential targets for therapeutic intervention.

Longitudinal immune profile reveals reduced function of pro-inflammatory monocytes with age following kidney transplantation.

Toxicity of immunosuppression, notably the risk of infection, increases with age. However, the dynamic changes in innate immune response following transplantation are unclear. Based on recent observations, we hypothesized that pro-inflammatory capacity would decrease with age. We analyzed approximately 300 PBMC samples collected longitudinally in 45 de novo, adult kidney recipients and performed detailed phenotypic and functional profiling of monocytes and T cell subsets. Inflammatory response to TLR4 stimulation and indirect allostimulation using mismatched HLA peptides were assessed. In patients aged ≥56years, TNF-α production by intermediate monocytes was similar to that in younger patients early posttransplant, but diminished substantially later. Adjusted analyses suggested that this was not attributable to confounding factors. In contrast, the alloimmune response to HLA peptides measured by IFN-γ in CD4+ T cells and TNF-α in monocytes was stable over time, but was low in older recipients. Measurement of CD80-86 surface expression revealed no signal for a lower costimulation capacity of APCs. These results suggest that older recipients have a reduced function of their innate pro-inflammatory immune cells posttransplant while maintaining a stable, low alloimmune response over time. The effect of reduced immunosuppressant doses on preventing this phenomenon needs to be clarified.

Non-cytotoxic doses of shikonin inhibit lipopolysaccharide-induced TNF-α expression via activation of the AMP-activated protein kinase signaling pathway.

Shikonin has been reported to exhibit a wide variety of medical functions. However, the strong non-selective cytotoxicity of shikonin can restrict its clinical application. The aim of the present study was to investigate the effects of shikonin at non-cytotoxic doses on the pro-inflammation functions of monocytes and macrophages. The present results suggested that the non-cytotoxic doses of shikonin effectively inhibited lipopolysaccharide (LPS)-induced reactive oxygen species production, NF-κB activation and TNF-α expression in RAW 264.7 mouse macrophages via AMP-activated protein kinase (AMPK) signaling pathway. In addition, the non-cytotoxic doses of shikonin downregulated LPS-induced TNF-α expression via AMPK signaling activation in primary murine bone marrow-derived macrophages, and also in monocytes cultured ex vivo from patients with chronic obstructive pulmonary disease (COPD). The present in vivo results indicated that the low-toxic dose of shikonin suppressed LPS-induced endotoxin shock and TNF-α expression in mice. Collectively, the present results may provide clinical and translational relevance for treating COPD and other TNF-α-related inflammatory disorders.

Prediction Power on Cardiovascular Disease of Neuroimmune Guidance Cues Expression by Peripheral Blood Monocytes Determined by Machine-Learning Methods.

Atherosclerosis is the underlying pathology in a major part of cardiovascular disease, the leading cause of mortality in developed countries. The infiltration of monocytes into the vessel walls of large arteries is a key denominator of atherogenesis, making monocytes accountable for the development of atherosclerosis. With the development of high-throughput transcriptome profiling platforms and cytometric methods for circulating cells, it is now feasible to study in-depth the predicted functional change of circulating monocytes reflected by changes of gene expression in certain pathways and correlate the changes to disease outcome. Neuroimmune guidance cues comprise a group of circulating- and cell membrane-associated signaling proteins that are progressively involved in monocyte functions. Here, we employed the CIRCULATING CELLS study cohort to classify cardiovascular disease patients and healthy individuals in relation to their expression of neuroimmune guidance cues in circulating monocytes. To cope with the complexity of human datasets featured by noisy data, nonlinearity and multidimensionality, we assessed various machine-learning methods. Of these, the linear discriminant analysis, Naïve Bayesian model and stochastic gradient boost model yielded perfect or near-perfect sensibility and specificity and revealed that expression levels of the neuroimmune guidance cues SEMA6B, SEMA6D and EPHA2 in circulating monocytes were of predictive values for cardiovascular disease outcome.

Cancer-Specific Immune Prognostic Signature in Solid Tumors and Its Relation to Immune Checkpoint Therapies.

To elucidate the role of immune cell infiltration as a prognostic signature in solid tumors, we analyzed immune-function-related genes from four publicly available single-cell RNA-Seq data sets and twenty bulk tumor RNA-Seq data sets from The Cancer Genome Atlas (TCGA). Unsupervised clustering of pan-cancer transcriptomic signature showed two major immune function types: one related to NK-, T-, and B-cell functions and the other related to monocyte, macrophage, dendritic cell, and Toll-like receptor functions. Kaplan–Meier analysis showed differential prognosis of these two groups, dependent on the cancer type. Our analysis of TCGA solid tumors with an elastic net model identified 155 genes associated with disease-free survival in different tumor types with varied influence across different cancer types. With this gene set, we computed cancer-specific prognostic immune score models for individual cancer types that predicted disease-free and overall survival. Validation of our model on available published data of immune checkpoint blockade therapies on melanoma, kidney renal cell carcinoma, non-small cell lung cancer, gastric cancer and bladder cancer confirmed that cancer-specific higher immune scores are associated with response to immunotherapy. Our analysis provides a comprehensive map of cancer-specific immune-related prognostic gene sets that are associated with immunotherapy response.

Dietary 23-hydroxy ursolic acid protects against diet-induced weight gain and hyperglycemia by protecting monocytes and macrophages against nutrient stress-triggered reprogramming and dysfunction and preventing adipose tissue inflammation

The aim of this study was to determine whether the atheroprotective phytochemical 23-hydroxy ursolic acid protects against diet-induced obesity and hyperglycemia by preventing nutrient stress-induced monocyte reprogramming. After a two week run-in period on a defined, phytochemical-free low-fat maintenance diet, 12-week old female C57BL/6J mice were either kept on the maintenance diet for additional 13 weeks or switched to either a high-calorie diet, a high-calorie diet supplemented with either 0.05% 23-hydroxy ursolic acid or a high-calorie diet supplemented with 0.2% 23-hydroxy ursolic acid. Dietary supplementation with 23-hydroxy ursolic acid reduced weight gain and adipose tissue mass, prevented hyperglycemia, hyperleptinemia and adipose tissue inflammation, and preserved glucose tolerance. 23-Hydroxy ursolic acid also preserved blood monocyte mitogen-activated protein kinase phosphatase-1 activity, a biomarker of monocyte health, and reduced macrophage content in the adipose tissue. Targeted gene profiling by qRT-PCR using custom-designed TaqMan® Array Cards revealed that dietary 23-hydroxy ursolic acid converts macrophages into a transcriptionally hyperactive phenotype with enhanced antioxidant defenses and anti-inflammatory potential. In conclusion, our findings show that dietary 23-hydroxy ursolic acid exerts both anti-obesogenic effects through multiple mechanisms. These include improving glucose tolerance, preventing hyperleptinemia, maintaining blood monocyte function, reducing recruitment of monocyte-derived macrophages into adipose tissues during nutrient stress, and converting these macrophages into an anti-inflammatory, potentially inflammation-resolving phenotype, all contributing to reduced adipose tissue inflammation. Our data suggest that 23-hydroxy ursolic acid may serve as an oral therapeutic and dietary supplement suited for patients at risk for obesity, impaired glucose tolerance and cardiovascular disease.

Modulation of Monocyte Activation and Function during Direct Antiviral Agent Treatment in Patients Coinfected with HIV and Hepatitis C Virus.

The activation phenotypes and functional changes in monocyte subsets during hepatitis C virus (HCV) elimination in HIV/HCV-coinfected patients were evaluated. Twenty-two HIV/HCV-coinfected patients on suppressive combination antiretroviral treatment (cART) achieving HCV elimination after direct-acting antiviral (DAA) therapy and 10 HIV-monoinfected patients were included. The activation phenotype (10 markers) and polyfunctionality (intracellular interleukin-1α [IL-1α], IL-1β, IL-6, IL-8, tumor necrosis factor alpha [TNF-α], and IL-10 production) in three monocyte subsets (classical, intermediate, and nonclassical) were evaluated by flow cytometry before and at the end of treatment. Cell-associated HIV DNA levels were assayed by droplet digital PCR. After HCV clearance, there was a significant increase in classical monocyte and decreases in intermediate and nonclassical monocyte levels. The levels of the activation markers CD49d, CD40, and CX3CR1 were decreased after treatment in the monocyte subsets, reaching the levels in HIV-monoinfected patients. After lipopolysaccharide (LPS) stimulation, although polyfunctionality significantly decreased in intermediate and nonclassical monocytes, some combinations, such as the IL-1α- (IL-1α-negative) IL-1β- IL-6+ (IL-6-producing) IL-8- TNF-α- IL-10- combination, were remarkably increased at the end of treatment compared to the control group. Cell-associated HIV DNA levels correlated with activation markers before but not after treatment. HCV clearance after DAA treatment in patients on cART exerts an anti-inflammatory profile on monocyte subsets, activation phenotypes, and polyfunctionality. However, there is not a complete normalization compared with HIV-monoinfected patients.

Abstract 6497: Early assessment of liquid biomarkers to predict pneumonitis after chemoradiation in patients with locally advanced non-small cell lung cancer (LA-NSCLC)

Abstract PURPOSE: Patients undergoing chemoradiation (CRT) for LA-NSCLC are at significant risk of developing radiation pneumonitis. As consolidation checkpoint inhibitor immunotherapy (IO) becomes standard of care after chemoradiation, pneumonitis (either radiation or immune-mediated) is also the most common rationale for cessation of IO and management with steroids. Our goal is to develop an early predictive biomarker for pneumonitis post-CRT, to risk stratify patients for treatment toxicity. We analyzed relationships between plasma cytokine levels, single cell functional assays, and pneumonitis prevalence within a prospective phase II clinical trial of chemoradiation for LA-NSCLC. METHODS: 35 Patients with AJCC v7 stage IIB-IIIB NSCLC were prospectively enrolled on the FLARE-RT trial (NCT02773238) from 2016-2019. All patients underwent chemoradiation; 18 patients also received consolidation IO (durvalumab) after it became standard of care in late 2017. 24 patients consented to peripheral blood collection at baseline and 21 patients during week 3 of CRT. Plasma concentrations of 43 common inflammatory cytokines were measured at both time points. Pneumonitis was defined as CTCAE v4 grade 2 or higher, irrespective of radiation-induced or immune-mediated causality. Bootstrapping over 100 iterations of LASSO feature selection was performed to reduce dimensionality and guard against false discoveries. Univariate Cox regression of selected cytokine concentrations were evaluated for associations to time-dependent pneumonitis incidence. RESULTS: Grade 2+ pneumonitis occurred in 13/35 patients, 7 of whom received durvalumab. Four plasma cytokines were most consistently associated with pneumonitis: GMCSF, E selectin, sIL6R (soluble Interleukin-6 Receptor), and SCF (stem cell factor). Higher baseline GMCSF and sIL6R (HR 2.1-4.2, p=0.01-0.03) were correlated with pneumonitis. Higher mid-CRT GMCSF, sIL6R and E selectin (HR 1.6-3.2, p=0.02-0.05), along with decline in mid-CRT SCF (HR 0.2, p=0.02), were associated with pneumonitis. The 4-cytokine signature at mid-CRT predicted for subsequent pneumonitis incidence (cross-validated AUC=0.84). Since GMCSF and SCF are linked to monocyte development, and sIL6R and E selectin are both associated with monocyte function, we performed single cell functional assay (Isocode chips) of mid-CRT peripheral monocytes from two patients with different pneumonitis status. A patient who subsequently developed grade 3 pneumonitis showed higher peripheral monocyte function (5.7% producing MIP1b) relative to a patient who did not develop pneumonitis (0% producing MIP1b). CONCLUSIONS: Our results show it may be possible to predict development of pneumonitis after chemoradiation in patients with LA-NSCLC, with mid-treatment assessment of 4 plasma cytokines: GMCSF, E selectin, sIL6R, and SCF. Changes in these cytokines may be related to monocyte function. These biomarkers need to be validated in other independent patient populations. Citation Format: Jing Zeng, Ramesh Rengan, Rafael Santana-Davila, Daniel S. Hippe, Ashley M. Houghton, Paul E. Kinahan, Hubert J. Vesselle, Paul Lampe, Stephen R. Bowen. Early assessment of liquid biomarkers to predict pneumonitis after chemoradiation in patients with locally advanced non-small cell lung cancer (LA-NSCLC) [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6497.

Innate Rhythms: Clocks at the Center of Monocyte and Macrophage Function.

The circadian cycle allows organisms to track external time of day and predict/respond to changes in the external environment. In higher order organisms, circadian rhythmicity is a central feature of innate and adaptive immunity. We focus on the role of the molecular clock and circadian rhythmicity specifically in monocytes and macrophages of the innate immune system. These cells display rhythmicity in their internal functions, such as metabolism and inflammatory mediator production as well as their external functions in pathogen sensing, phagocytosis, and migration. These inflammatory mediators are of clinical interest as many are therapeutic targets in inflammatory disease such as cardiovascular disease, diabetes, and rheumatoid arthritis. Moreover, circadian rhythm disruption is closely linked with increased prevalence of these conditions. Therefore, understanding the mechanisms by which circadian disruption affects monocyte/macrophage function will provide insights into novel therapeutic opportunities for these chronic inflammatory diseases.

Characterization of ovine monocyte activity when cultured with Haemonchus contortus larvae in vitro.

The objective of this study was to identify and characterize cell populations within ovine peripheral blood mononuclear cells (PBMCs) associated with Haemonchus contortus (Hc) larval morbidity and impairment in vitro. Monocytes and lymphocytes were separated from PBMC from parasite-resistant St. Croix (STC) sheep and parasite-susceptible Suffolk (SUF) sheep. Cells were cultured with Hc third stage larvae (L3) for 9h. Larval morbidity was assessed using ATP concentration. Activation status was determined through gene expression analysis and enzyme inhibition. Enzymes arginase-1 (Arg1) and inducible nitric oxide synthase (iNOS) were inhibited using BEC (S-(2-boronoethyl)-I-cysteine) and 1400W (N-(3-(aminomethyl)benzyl)acetamidine), respectively. Larval ATP was lower when cultured with STC-derived monocytes (0.015μmol/L ATP) compared to SUF-derived monocytes (0.067μmol/L ATP) (P<.001), or lymphocytes from either breed (STC: 0.085μmol/L, SUF: 0.112μmol/L ATP) (P<.001). SUF-derived monocytes displayed higher expression of M1 genes, whereas STC-derived monocytes displayed M2 genes continuously. Inhibition of Arg1 decreased monocyte function in both breeds, whereas iNOS inhibition restored SUF-derived monocyte function. Together, these data indicate STC-derived monocytes favour M2 phenotype when exposed to L3, where SUF-derived monocyte function resembled M1 phenotype and described potential for improving Suffolk sheep through modulating inflammatory responses.

The role and function of I\u03baK\u03b1/\u03b2 in monocyte impairment

Following major trauma, sepsis or surgery, some patients exhibit an impaired monocyte inflammatory response that is characterized by a decreased response to a subsequent bacterial challenge. To investigate this poorly understood phenomenon, we adopted an in-vitro model of endotoxin tolerance utilising primary human CD14 + monocytes to focus on the effect of impairment on IκKα/β, a critical part of the NFκB pathway. Impaired monocytes had decreased IκKα mRNA and protein expression and decreased phosphorylation of the IκKα/β complex. The impaired monocyte secretome demonstrated a distinct cytokine/chemokine footprint from the naïve monocyte, and that TNF-α was the most sensitive cytokine or chemokine in this setting of impairment. Inhibition of IκKα/β with a novel selective inhibitor reproduced the impaired monocyte phenotype with decreased production of TNF-α, IL-6, IL-12p70, IL-10, GM-CSF, VEGF, MIP-1β, TNF-β, IFN-α2 and IL-7 in response to an LPS challenge. Surgical patients with infection also exhibited an impaired monocyte phenotype and had decreased SITPEC, TAK1 and MEKK gene expression, which are important for IκKα/β activation. Our results emphasize that impaired monocyte function is, at least in part, related to dysregulated IκKα/β activation, and that IκKα/β is likely involved in mounting a sufficient monocyte inflammatory response. Future studies may wish to focus on adjuvant therapies that augment IκKα/β function to restore monocyte function in this clinically important problem.

In vitro effects of conjugated linoleic acid (CLA) on inflammatory functions of bovine monocytes

The conjugated linoleic acid (CLA) isomers, a group of naturally occurring isomers of the essential fatty acid (FA) linoleic acid, have received special attention in animal and human nutrition. Although they have long been used as dietary integrators in dairy cows, the effects of CLA isomers on bovine immune cells remain mostly undisclosed. The present study aimed to cover this gap and investigate the in vitro effects of CLA on inflammatory functions, including chemotaxis, phagocytosis, killing capability, and extracellular respiratory burst of purified bovine monocytes (CD14+). The apoptosis rate of monocytes was addressed as well. Once assessed, the effects of different concentrations (10, 50, 100, and 500 μM) of the 2 main CLA isomers, namely cis-9,trans-11 and trans-10,cis-12, the experiments were carried out using a concentration of 50 μM of the CLA isomers, both individually and in a mixture (50:50). The immunomodulatory activities of linoleic acid, an essential FA, and stearic acid, a saturated FA, were also investigated. Only the 50:50 CLA mixture was able to reduce monocyte apoptosis and to increase the extracellular respiratory burst during experimental proinflammatory conditions, as assessed by measuring production of reactive oxygen species. Linoleic acid and CLA had no effects on chemotaxis, phagocytosis, or killing capability. Remarkably, treatment of monocytes with stearic acid significantly reduced their chemotactic capability. The present results demonstrated that CLA isomers do have immunomodulatory effects on some functions of bovine monocytes, and that the mixture of the 2 CLA isomers is more effective than the CLA isomers individually.

The Diverse Roles of Monocytes in Cryptococcosis.

Monocytes are considered to play a central role in the pathogenesis of Cryptococcus neoformans infection. Monocytes and monocyte-derived macrophages and dendritic cells are key components for the control of infection, but paradoxically they can also contribute to detrimental host responses and may even support fungal proliferation and dissemination. Simultaneously, the C. neoformans polysaccharide capsule can impair the functions of monocytes. Although monocytes are often seen as simple precursor cells, they also function as independent immune effector cells. In this review, we summarize these monocyte-specific functions during cryptococcal infection and the influence of C. neoformans on monocyte responses. We also cover the most recent findings on the functional and phenotypic heterogeneity of monocytes and discuss how new advanced technologies provide a platform to address outstanding questions in the field.

Platelet-Monocyte Aggregates: Understanding Mechanisms and Functions in Sepsis.

Platelets have been shown to play an important immunomodulatory role in the pathogenesis of various diseases through their interactions with other immune and nonimmune cells. Sepsis is a major cause of death in the United States, and many of the mechanisms driving sepsis pathology are still unresolved. Monocytes have recently received increasing attention in sepsis pathogenesis, and multiple studies have associated increased levels of platelet-monocyte aggregates observed early in sepsis with clinical outcomes in sepsis patients. These findings suggest platelet-monocyte aggregates may be an important prognostic indicator. However, the mechanisms leading to platelet interaction and aggregation with monocytes, and the effects of aggregation during sepsis are still poorly defined. There are few studies that have really investigated functions of platelets and monocytes together, despite a large body of research showing separate functions of platelets and monocytes in inflammation and immune responses during sepsis. The goal of this review is to provide insights into what we do know about mechanisms and biological meanings of platelet-monocyte interactions, as well as some of the technical challenges and limitations involved in studying this important potential mechanism in sepsis pathogenesis. Improving our understanding of platelet and monocyte biology in sepsis may result in identification of novel targets that can be used to positively affect outcomes in sepsis.

Future of Statins in Sepsis: A Review on Its Safety and Efficacy

Sepsis is a life-threatening organ dysfunction caused by a dysregulated host response to infection. Statins [Hydroxymethylglutaryl-CoA reductase inhibitors] not only lower cholesterol levels but also have been proposed as adjunctive therapy in sepsis due to their pleiotropic effects. They act on several stages in sepsis: the generation of proinflammatory cytokines, modulation of leukocyte and monocyte functions, and reduction of oxidative stress as well as improvement in endothelial function and platelet activity. However, it has been argued if the observed beneficial effect of statins in sepsis is related to preadmission or post-admission use of statins. Also, the positive impact of statins on the clinical outcome of patients with sepsis has shown conflicting results. Accordingly, this review will discuss recent evidence regarding the use of statins in sepsis. Also, adequate use of statins based on the right drug, at the right time, at the right dose and in the right population will be discussed. The information in this review shows that the effect of statins is a drug, not a class effect, with the most effective drug in sepsis being simvastatin. Besides, it highlights the importance of proper timing and dosing of statins to manifest their antibacterial and pleiotropic effects. Finally, the effect of statins in sepsis is restricted to early phases of sepsis or sepsis prevention, not sepsis complicated with organ dysfunction or septic shock. However, more in vivo and clinical trials are required to determine the final decision about statin use in sepsis.

Transcatheter Aortic Valve Implantation Represents an Anti-Inflammatory Therapy Via Reduction of Shear Stress-Induced, Piezo-1-Mediated Monocyte Activation.

Aortic valve stenosis is an increasingly prevalent degenerative and inflammatory disease. Transcatheter aortic valve implantation (TAVI) has revolutionized its treatment, thereby avoiding its life-threatening/disabling consequences. Whether aortic valve stenosis is accelerated by inflammation and whether it is itself a cause of inflammation are unclear. We hypothesized that the large shear forces exerted on circulating cells, particularly on the largest circulating cells, monocytes, while passing through stenotic aortic valves result in proinflammatory effects that are resolved with TAVI. TAVI provides a unique opportunity to compare the activation status of monocytes under high shear stress (before TAVI) and under low shear stress (after TAVI). The activation status of monocytes was determined with a single-chain antibody, MAN-1, which is specific for the activated β2-integrin Mac-1. Monocyte function was further characterized by the adhesion of myocytes to stimulated endothelial cells, phagocytic activity, uptake of oxidized low-density lipoprotein, and cytokine expression. In addition, we designed a microfluidic system to recapitulate the shear rate conditions before and after TAVI. We used this tool in combination with functional assays, Ca2+ imaging, siRNA gene silencing, and pharmacological agonists and antagonists to identify the key mechanoreceptor mediating the shear stress sensitivity of monocytes. Last, we stained for monocytes in explanted stenotic aortic human valves. The resolution of high shear stress through TAVI reduces Mac-1 activation, cellular adhesion, phagocytosis, oxidized low-density lipoprotein uptake, and expression of inflammatory markers in monocytes and plasma. Using microfluidics and pharmacological and genetic studies, we could recapitulate high shear stress effects on isolated human monocytes under highly controlled conditions, showing that shear stress-dependent calcium influx and monocyte adhesion are mediated by the mechanosensitive ion channel Piezo-1. We also demonstrate that the expression of this receptor is shear stress dependent and downregulated in patients receiving TAVI. Last, we show monocyte accumulation at the aortic side of leaflets of explanted aortic valves. We demonstrate that high shear stress, as present in patients with aortic valve stenosis, activates multiple monocyte functions, and we identify Piezo-1 as the mainly responsible mechanoreceptor, representing a potentially druggable target. We demonstrate an anti-inflammatory effect and therefore a novel therapeutic benefit of TAVI.

Methods in isolation and characterization of bovine monocytes and macrophages.

Monocytes and macrophages belong to the mononuclear phagocyte system and play important roles in both physiological and pathological processes. The cells belonging to the monocyte/macrophage system are structurally and functionally heterogeneous. Several subsets of monocytes have been previously identified in mammalian blood, generating different subpopulations of macrophages in tissues. Although their distribution and phenotype are similar to their human counterpart, bovine monocytes and macrophages feature differences in both functions and purification procedures. The specific roles that monocytes and macrophages fulfil in several important diseases of bovine species, including among the others tuberculosis and paratuberculosis, brucellosis or the disease related to peripartum, remain still partially elusive. The purpose of this review is to discuss the current knowledge of bovine monocytes and macrophages. We will describe methods for their purification and characterization of their major functions, including chemotaxis, phagocytosis and killing, oxidative burst, apoptosis and necrosis. An overview of the flow cytometry and morphological procedures, including cytology, histology and immunohistochemistry, that are currently utilized to describe monocyte and macrophage main populations and functions is presented as well.

Human mesenchymal stem cells inhibit the differentiation and effector functions of monocytes.

Although monocytes represent an essential part of the host defence system, their accumulation and prolonged stimulation could be detrimental and may aggravate chronic inflammatory diseases. The present study has explored the less-understood immunomodulatory effects of mesenchymal stem cells on monocyte functions. Isolated purified human monocytes were co-cultured with human umbilical cord-derived mesenchymal stem cells under appropriate culture conditions to assess monocytes’ vital functions. Based on the surface marker analysis, mesenchymal stem cells halted monocyte differentiation into dendritic cells and macrophages and reduced their phagocytosis functions, which rendered an inability to stimulate T-cell proliferation. The present study confers that mesenchymal stem cells exerted potent immunosuppressive activity on monocyte functions such as differentiation, phagocytosis and Ag presentation; hence, they promise a potential therapeutic role in down-regulating the unwanted monocyte-mediated immune responses in the context of chronic inflammatory diseases.

MONOCYTE FUNCTION FOLLOWING ACUTE EXERCISE IN BREAST CANCER SURVIVORS BEFORE AND AFTER EXERCISE TRAINING

Breast cancer therapy impairs immune function that may be attenuated with exercise, though the specific changes that occur remain unclear. PURPOSE: 1) To examine monocyte function in breast cancer survivors (BCS) following acute exercise and 2) to determine if this response changes with exercise training. METHODS: 9 BCS [Age: 58±8, BMI: 27.9±6.7] completed a cardiopulmonary exercise test (CPET). In a subsequent trial, 45 min of intermittent cycling at 60% of CPET peak wattage was performed. Blood was taken at rest, immediately (0h) and 1h after exercise. Monocyte phagocytosis and oxidative burst were assessed following E.coli exposure. Toll-like receptor 2 (TLR2) and 4 (TLR4) expression was determined on CD14+CD16- and CD14+CD16+ monocytes. All assays were performed before (pre) and after (post) 16 wk of combined aerobic and resistance training. Data are presented as mean fluorescence intensity ± SD. RESULTS: Phagocytosis increased 1h after acute exercise before (rest: 3396±941 0h: 3257±772, 1h: 3692±824, p=0.035) but not after training. There was a trend for greater phagocytosis with training (pre: 3533±815 post: 5027±2039, p=0.078). Oxidative burst was unchanged with acute exercise but improved following training (pre: 4264±1061 post: 5446±1287, p=0.026). CD16- TLR2 expression decreased acutely at 1h compared to rest and 0h both before and after training (rest: 350±70, 0h: 328±73, 1h: 287±41, both p<0.05) while CD16+ decreased acutely before training only (pre rest: 355±115 0h: 339±98 1h: 291±87. CD16- TLR4 expression tended to decrease with acute exercise (p=0.067) whereas CD16+ TLR4 expression decreased across all time points (rest: 140±15, 0h: 135±15, 1h: 123±18, all p<0.05) with neither population affected by training. CONCLUSIONS: In BCS, monocyte phagocytic capacity of bacteria increased following acute exercise, while training increased both phagocytosis and oxidative burst. Training appeared to mitigate the acute response, possibly due to higher resting function. Expression of TLR2 and TLR4 were progressively reduced with acute exercise that was mostly independent of training. The reduction of monocyte TLR2 and TLR4 may represent an anti-inflammatory response to acute exercise that promotes enhanced elimination of bacteria. Supported by Breast Cancer Research Foundation (New York, NY).

Paracoxib Alleviates Ventilator-Induced Lung Injury Through Functional Modulation of Lung-Recruited CD11bloLy6Chi Monocytes.

Lung-recruited Ly6Chi monocytes had been shown to be involved in ventilator-induced lung injury (VILI). Our present study aimed to investigate whether the cyclooxygenase-2 (COX-2) inhibition modulates the function of lung-recruited Ly6Chi monocytes in a mouse model of VILI. Mice were exposed to lipopolysaccharide (LPS; 20 ng) intraperitoneally prior to injurious mechanical ventilation (Vt = 30 mL/kg, PEEP = 0 cmH2O). A subgroup of mice was treated with intravenous parecoxib (30 mg/kg), a COX-2 inhibitor, 1 h prior to ventilation. Control mice received saline and were not ventilated. At the end of the experiment, blood gas analysis was performed and lung tissue was collected for histological assessment. Flow cytometry was employed to quantify the different populations of lung monocytes/macrophages and their function. Isolated Ly6Chi cells were used to measure the intracellular concentrations of reactive oxygen species (ROS) and nitric oxide (NO) by fluorescent probes, and cytokine production by cytometric bead array. Exposure to LPS and injurious ventilation was associated with severe lung histological damage, oxygenation impairment, and pulmonary edema; all of which were largely attenuated following the treatment of parecoxib. Furthermore, flow cytometry analysis revealed that parecoxib caused a reduction in the number of the lung-recruited CD11bloLy6Chi monocytes while there was no effect on tissue-resident CD64+ alveolar macrophages. In addition, the production of oxidative stress products (ROS, NO), MHC-II expression, and inflammatory cytokines in response to LPS and VILI in CD11bloLy6Chi monocytes was ameliorated by parecoxib. Parecoxib-induced alleviation of oxidative stress and inflammation in lung-recruited Ly6Chi monocytes may partly explain the beneficial action of COX-2 inhibition in VILI.