Opsonophagocytosis of Chlamydia pneumoniae by Human Monocytes and Neutrophils

ABSTRACT The human respiratory tract pathogen Chlamydia pneumoniae, which causes mild to severe infections, has been associated with the development of chronic inflammatory diseases. To understand the biology of C. pneumoniae infections, several studies have investigated the interaction between C. pneumoniae and professional phagocytes. However, these studies have been conducted under nonopsonizing conditions, making the role of opsonization in C. pneumoniae infections elusive. Thus, we analyzed complement and antibody opsonization of C. pneumoniae and evaluated how opsonization affects chlamydial infectivity and phagocytosis in human monocytes and neutrophils. We demonstrated that IgG antibodies and activation products of complement C3 and C4 are deposited on the surface of C. pneumoniae elementary bodies when incubated in human serum. Complement activation limits C. pneumoniae infectivity in vitro and has the potential to induce bacterial lysis by the formation of the membrane attack complex. Coculture of C. pneumoniae and freshly isolated human leukocytes showed that complement opsonization is superior to IgG opsonization for efficient opsonophagocytosis of C. pneumoniae in monocytes and neutrophils. Neutrophil-mediated phagocytosis of C. pneumoniae was crucially dependent on opsonization, while monocytes retained minor phagocytic potential under nonopsonizing conditions. Complement opsonization significantly enhanced the intracellular neutralization of C. pneumoniae in peripheral blood mononuclear cells and neutrophils and almost abrogated the infectious potential of C. pneumoniae. In conclusion, we demonstrated that complements limit C. pneumoniae infection in vitro by interfering with C. pneumoniae entry into permissive cells by direct complement-induced lysis and by tagging bacteria for efficient phagocytosis in both monocytes and neutrophils.

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Evaluation of Immunostimulatory Activities of Synthetic Mannose-Containing Structures Mimicking the β-(1→2)-Linked Cell Wall Mannans of Candida albicans

Clinical and Vaccine Immunology ◽

10.1128/cvi.00298-12 ◽

2012 ◽

Vol 19 (11) ◽

pp. 1889-1893 ◽

Cited By ~ 14

Author(s):

Kaarina Ranta ◽

Kaisa Nieminen ◽

Filip S. Ekholm ◽

Moniká Poláková ◽

Mattias U. Roslund ◽

...

Keyword(s):

Candida Albicans ◽

Mononuclear Cells ◽

Human Peripheral Blood ◽

Peripheral Blood Mononuclear ◽

Content Type ◽

Mouse Macrophages ◽

Ifn Γ ◽

Low Levels ◽

Necrosis Factor

ABSTRACTImmunostimulatory properties of synthetic structures mimicking the β-(1→2)-linked mannans ofCandida albicanswere evaluatedin vitro. Contrary to earlier observations, tumor necrosis factor (TNF) production was not detected after stimulation with mannotetraose in mouse macrophages. Divalent disaccharide 1,4-bis(α-d-mannopyranosyloxy)butane induced TNF and some molecules induced low levels of gamma interferon (IFN-γ) in human peripheral blood mononuclear cells (PBMC).

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The Potential of IgG to Induce Murine and Human Thymic Maturation of IL-10+ B Cells (B10) Revealed in a Pilot Study

Cells ◽

10.3390/cells9102239 ◽

2020 ◽

Vol 9 (10) ◽

pp. 2239

Author(s):

Amanda Harumi Sabô Inoue ◽

Aline Aparecida de Lima Lira ◽

Marília Garcia de-Oliveira ◽

Thamires Rodrigues de Sousa ◽

Fábio da Ressureição Sgnotto ◽

...

Keyword(s):

B Cells ◽

Mononuclear Cells ◽

Inflammatory Diseases ◽

Allergen Challenge ◽

Serum Samples ◽

Peripheral Blood Mononuclear ◽

Maturation Stages ◽

B10 Cells

Regulatory B (B10) cells can control several inflammatory diseases, including allergies; however, the origin of peripheral B10 cells is not fully understood, and the involvement of primary lymphoid organs (PLOs) as a primary site of maturation is not known. Here, using a murine model of allergy inhibition mediated by maternal immunization with ovalbumin (OVA), we aimed to evaluate whether B10 cells can mature in the thymus and whether IgG can mediate this process. Female mice were immunized with OVA, and offspring thymus, bone marrow, spleen, lung, and serum samples were evaluated at different times and after passive transfer of purified IgG or thymocytes. A translational approach was implemented using human nonatopic thymus samples, nonatopic peripheral blood mononuclear cells (PBMCs), and IgG from atopic or nonatopic individuals. Based on the expression of CD1d on B cells during maturation stages, we suggest that B10 cells can also mature in the murine thymus. Murine thymic B10 cells can be induced in vitro and in vivo by IgG and be detected in the spleen and lungs in response to an allergen challenge. Like IgG from atopic individuals, human IgG from nonatopic individuals can induce B10 cells in the infant thymus and adult PBMCs. Our observations suggest that B10 cells may mature in the thymus and that this mechanism may be mediated by IgG in both humans and mice. These observations may support the future development of IgG-based immunoregulatory therapeutic strategies.

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2221. Chlamydia pneumoniae (Cpn) Induces IFN-γ Responses in Peripheral Blood Mononuclear Cells (PBMC) from Pediatric and Adult Asthma Patients: Effects of Age and Inhaled Corticosteroid Use

Open Forum Infectious Diseases ◽

10.1093/ofid/ofz360.1899 ◽

2019 ◽

Vol 6 (Supplement_2) ◽

pp. S758-S758

Author(s):

Aviva Szigeti ◽

Margaret Hammerschlag ◽

Diana Weaver ◽

Tamar Smith-Norowitz ◽

Stephan Kohlhoff

Keyword(s):

Peripheral Blood Mononuclear Cells ◽

Peripheral Blood ◽

Chlamydia Pneumoniae ◽

Mononuclear Cells ◽

Inhaled Corticosteroid ◽

Peripheral Blood Mononuclear ◽

Blood Mononuclear Cells ◽

Asthma Patients ◽

Ifn Γ

Abstract Background Chlamydia pneumoniae (Cpn) is unique in its ability to cause chronic infections, potentially triggering asthma exacerbations as well as subsequent asthma development. Th1-mediated immunity and IFN-γ are critical for clearing chlamydial infections. Persistent or recent Cpn infection may be identified in vitro by detecting T-helper cytokine IFN-γ produced by peripheral blood mononuclear cells (PBMC) stimulated by Cpn. Inhaled corticosteroids (ICS) may have an inhibitory effect on IFN-γ. Prior studies have shown increased Th2 responses upon in vitro Cpn stimulation with increased age. Our aim was to determine whether age and inhaled corticosteroid (ICS) use affect Cpn-induced PBMC produced IFN-γ levels. Methods Pediatric and adult subjects with (n = 23) and without (n = 10) asthma were enrolled. PBMC obtained from all subjects were stimulated with Cpn (MOI = 0.1 x48h) in vitro. IFN-γ levels in culture supernatants were determined by ELISA and reported as pg/mL. Nasopharyngeal (NP) swabs were tested for Cpn using Real-Time PCR. Statistical analysis for continuous variables was performed using the Mann–Whitney U test. Results None of the subjects were positive for Cpn by PCR on NP swab. Levels of IFN-γ produced by PBMC stimulated by Cpn were similar between asthmatic vs. control subjects (41.7 vs. 68.8, respectively; P = 0.72) and between pediatric and adult subjects with asthma (IFN-γ 54 vs. 20.1 respectively, P = 0.95). Pediatric subjects with asthma who received ICS had lower IFN-γ levels than those who did not (median IFN-γ 25.5 vs. 209; P = 0.003). Conclusion Our finding of lower IFN-γ levels among asthma patients on ICS compared with those not on ICS suggests that ICS use may dampen the systemic inflammatory response. While we did not find a statistically significant difference between pediatric and adult age groups in this pilot study, there was a trend to higher Cpn-induced IFN-γ levels among younger pediatric subjects. Future prospective studies should further define predictors of diminished IFN-γ responses in patients with asthma. Disclosures All authors: No reported disclosures.

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PRTX-100 Inhibits Platelet Phagocytosis In Vitro.

Blood ◽

10.1182/blood.v108.11.1081.1081 ◽

2006 ◽

Vol 108 (11) ◽

pp. 1081-1081 ◽

Cited By ~ 1

Author(s):

Chris Yatko ◽

Christopher Herrem ◽

Samia Siddiqui ◽

Victor S. Sloan

Keyword(s):

Mononuclear Cells ◽

Therapeutic Option ◽

Protein A ◽

Flow Cytometric Analysis ◽

Human Platelets ◽

Staphylococcal Protein A ◽

Human Monocytes ◽

Peripheral Blood Mononuclear ◽

Vitro Assay

Abstract Background: In idiopathic thrombocytopenic purpura (ITP), autoantibodies bind to platelets which are then phagocytosed by monocytes/macrophages and removed by the reticuloendothelial system. PRTX-100 (Staphylococcal protein A) is being investigated for the treatment of ITP. Objective: To assess the effect of PRTX-100 on phagocytosis of platelets in an in vitro assay. Methods: Human monocytes were isolated from whole blood peripheral blood mononuclear cells (PBMCs) by adherence and cultured for 6 days in RPMI + 5% human serum. 48 hours prior to phagocytosis assay, PRTX-100 was added at 250, 25, and 2.5ng/ml. Human platelets were labeled with a fluorescent (PerCP) lipophilic dye and opsonized with an antibody to MHC Class I (W632). 2×10−5 monocytes were co-cultured with 2×10−7 labeled platelets for 1 hour at 37 ° C. All conditions were performed in triplicate. After an hour, phycoerythrin (PE) labeled anti-CD61 antibody was added to assess surface bound platelets versus ingested platelets. Phagocytosis was determined by flow cytometric analysis. The monocyte population was gated upon by forward and side scatter properties, then verified by staining with CD14-FITC. Percent phagocytosis was calculated as the fraction of ingested platelets (PerCP +/CD61−) to the total PerCP population (PerCP +/CD61−) + ( PerCP+/CD61+) within the gated monocyte population. Results: PRTX-100 inhibits the phagocytosis of W632 opsonized platelets by human monocytes. Phagocytosis of W632 opsonized platelets was 40%, while phagocytosis in the presence of PRTX-100 at concentrations of 250, 25, and 2.5ng/ml was 18.3%, 23%, and 24.3%, respectively. Phagocytosis at 250ng/ml and 25ng/ml was significantly different from control phagocytosis with p values of 0.014 and 0.001 respectively by Student’s t test. Conclusions: PRTX-100 inhibits the phagocytosis of platelets by monocytes, the effector limb of ITP. Prevention of platelet phagocytosis is an important treatment goal in ITP. PRTX-100 has been shown to be generally safe and well-tolerated in a phase I study in healthy volunteers (J Clin Pharmacol, in press). PRTX -100 is a promising therapeutic option for ITP and deserves further study. Effect of PRTX-100 on In Vitro Phagocytosis of Opsonized Human Platelets Effect of PRTX-100 on In Vitro Phagocytosis of Opsonized Human Platelets

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Unravelling Atrioventricular Block Risk in Inflammatory Diseases: Systemic Inflammation Acutely Delays Atrioventricular Conduction via a Cytokine‐Mediated Inhibition of Connexin43 Expression

Journal of the American Heart Association ◽

10.1161/jaha.121.022095 ◽

2021 ◽

Author(s):

Pietro Enea Lazzerini ◽

Maurizio Acampa ◽

Michael Cupelli ◽

Alessandra Gamberucci ◽

Ujala Srivastava ◽

...

Keyword(s):

Systemic Inflammation ◽

Inflammatory Markers ◽

Cardiac Tissue ◽

Mononuclear Cells ◽

Inflammatory Diseases ◽

Atrioventricular Conduction ◽

Peripheral Blood Mononuclear ◽

In Vivo Animal Model

Background Recent data suggest that systemic inflammation can negatively affect atrioventricular conduction, regardless of acute cardiac injury. Indeed, gap‐junctions containing connexin43 coupling cardiomyocytes and inflammation‐related cells (macrophages) are increasingly recognized as important factors regulating the conduction in the atrioventricular node. The aim of this study was to evaluate the acute impact of systemic inflammatory activation on atrioventricular conduction, and elucidate underlying mechanisms. Methods and Results We analyzed: (1) the PR‐interval in patients with inflammatory diseases of different origins during active phase and recovery, and its association with inflammatory markers; (2) the existing correlation between connexin43 expression in the cardiac tissue and peripheral blood mononuclear cells (PBMC), and the changes occurring in patients with inflammatory diseases over time; (3) the acute effects of interleukin(IL)‐6 on atrioventricular conduction in an in vivo animal model, and on connexin43 expression in vitro. In patients with elevated C‐reactive protein levels, atrioventricular conduction indices are increased, but promptly normalized in association with inflammatory markers reduction, particularly IL‐6. In these subjects, connexin43 expression in PBMC, which is correlative of that measured in the cardiac tissue, inversely associated with IL‐6 changes. Moreover, direct IL‐6 administration increased atrioventricular conduction indices in vivo in a guinea pig model, and IL‐6 incubation in both cardiomyocytes and macrophages in culture, significantly reduced connexin43 proteins expression. Conclusions The data evidence that systemic inflammation can acutely worsen atrioventricular conduction, and that IL‐6‐induced down‐regulation of cardiac connexin43 is a mechanistic pathway putatively involved in the process. Though reversible, these alterations could significantly increase the risk of severe atrioventricular blocks during active inflammatory processes.

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In Vitro and In Vivo Intracellular Killing Effects of Tigecycline against Clinical Nontyphoid Salmonella Isolates Using Ceftriaxone as a Comparator

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01807-10 ◽

2011 ◽

Vol 55 (6) ◽

pp. 2755-2759 ◽

Cited By ~ 7

Author(s):

Hung-Jen Tang ◽

Wen-Chien Ko ◽

Chi-Chung Chen ◽

Po-Lin Chen ◽

Han Siong Toh ◽

...

Keyword(s):

Survival Rate ◽

Mononuclear Cells ◽

Human Peripheral Blood ◽

Pathogen Resistance ◽

Peripheral Blood Mononuclear ◽

Content Type ◽

Time Kill ◽

Peritonitis Model

ABSTRACTSalmonellais an important, worldwide food-borne pathogen. Resistance to fluoroquinolones and cephalosporins has been increasingly reported, and new therapeutic agents are desperately needed. In this study, we evaluated thein vitroantimicrobial susceptibility of clinical nontyphoidalSalmonellaisolates to tigecycline. Antibacterial activity of tigecycline, ceftriaxone, and ciprofloxacin were investigated by time-kill studies and the murine peritonitis model. The MIC50/MIC90values of tigecycline, ceftriaxone, and ciprofloxacin against 76Salmonellaisolates were 0.25/0.5, 1/8, and 0.125/0.5 μg/ml, respectively. The intracellular inhibitory activity of tigecycline at 0.5 μg/ml (1× MIC) againstSalmonellaisolates in human peripheral blood mononuclear cells was sustained for 24 h. In a mouse peritonitis model, tigecycline reduced the extracellular and intracellular bacterial counts from 107CFU/ml and 105CFU/ml, respectively, to an undetectable level within 96 h. The results were similar to those obtained with ceftriaxone. The survival rate of mice exposed to tigecycline after being infected by an inoculum of 1 × 105CFU was 80%, and that of mice exposed to ceftriaxone was 100%. When the inoculum was increased to 1.3 × 106CFU, the survival rate of mice treated by tigecycline was 20%, and that of mice exposed to ceftriaxone was 0% (P= 0.2). When a ceftriaxone- and ciprofloxacin-resistant but tigecycline-susceptible isolate was tested, mice treated by tigecycline had a higher survival rate than those treated by ceftriaxone (15/20 [75%] versus 6/20 [30%];P= 0.011). Our results suggest that tigecycline is at least as effective as ceftriaxone for murineSalmonellainfections and warrants further clinical investigations to delineate its potential against humanSalmonellainfections.

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Soluble ST2 Does Not Regulate TNF-α and IL-6 Production in Dengue Virus-Infected Human Monocytes

ISRN Tropical Medicine ◽

10.1155/2013/230273 ◽

2013 ◽

Vol 2013 ◽

pp. 1-7 ◽

Cited By ~ 1

Author(s):

Marisol Pérez-Acosta ◽

Félix Giovanni Delgado ◽

Jaime E. Castellanos

Keyword(s):

Dengue Virus ◽

Viral Antigen ◽

Mononuclear Cells ◽

Acute Infection ◽

Denv Infection ◽

Human Monocytes ◽

Peripheral Blood Mononuclear ◽

Soluble St2 ◽

Tnf Α

Dengue virus (DENV) produces an acute infection that results in the overproduction of proinflammatory cytokines. Although increased levels of the immunoregulator soluble ST2 (sST2) protein have been reported in the serum of patients with dengue, its importance during DENV infection remains unclear. The purpose of this study was to evaluate the effect of a recombinant human sST2 protein on the production of TNF-α and IL-6 in an in vitro model of DENV infection. Peripheral blood mononuclear cells (PBMCs) were permissive to in vitro DENV infection since viral antigen was detected in CD14+ monocytes by flow cytometry (median, 1%; range, 0–2.2), and in their supernatants TNF-α and IL-6 were detected. However, sST2 protein was not detected. Using multiple staining on infected PBMC we found that only CD14+ cells produced TNF-α and IL-6. Treatment with human recombinant sST2 protein decreased lipopolysaccharide-induced monocyte TNF-α and IL-6 production. However, this effect was not observed when the monocytes were pretreated with sST2 and later infected with DENV-2. These results suggest that sST2 has different roles in the regulation of TNF-α and IL-6 expression in human monocytes stimulated with LPS and DENV-2.

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Immunization with Brucella VirB Proteins Reduces Organ Colonization in Mice through a Th1-Type Immune Response and Elicits a Similar Immune Response in Dogs

Clinical and Vaccine Immunology ◽

10.1128/cvi.00653-14 ◽

2014 ◽

Vol 22 (3) ◽

pp. 274-281 ◽

Cited By ~ 10

Author(s):

Cora N. Pollak ◽

María Magdalena Wanke ◽

Silvia M. Estein ◽

M. Victoria Delpino ◽

Norma E. Monachesi ◽

...

Keyword(s):

Immune Response ◽

Mononuclear Cells ◽

Interleukin 4 ◽

Delayed Type Hypersensitivity ◽

Peripheral Blood Mononuclear ◽

Content Type ◽

Type Iv ◽

Ifn Γ ◽

Organ Colonization

ABSTRACTVirB proteins fromBrucellaspp. constitute the type IV secretion system, a key virulence factor mediating the intracellular survival of these bacteria. Here, we assessed whether a Th1-type immune response against VirB proteins may protect mice fromBrucellainfection and whether this response can be induced in the dog, a natural host forBrucella. Splenocytes from mice immunized with VirB7 or VirB9 responded to their respective antigens with significant and specific production of gamma interferon (IFN-γ), whereas interleukin-4 (IL-4) was not detected. Thirty days after an intraperitoneal challenge with liveBrucella abortus, the spleen load of bacteria was almost 1 log lower in mice immunized with VirB proteins than in unvaccinated animals. As colonization reduction seemed to correlate with a Th1-type immune response against VirB proteins, we decided to assess whether such a response could be elicited in the dog. Peripheral blood mononuclear cells (PBMCs) from dogs immunized with VirB proteins (three subcutaneous doses in QuilA adjuvant) produced significantly higher levels of IFN-γ than cells from control animals uponin vitrostimulation with VirB proteins. A skin test to assess specific delayed-type hypersensitivity was positive in 4 out of 5 dogs immunized with either VirB7 or VirB9. As both proteins are predicted to locate in the outer membrane ofBrucellaorganisms, the ability of anti-VirB antibodies to mediate complement-dependent bacteriolysis ofB. caniswas assessedin vitro. Sera from dogs immunized with either VirB7 or VirB9, but not from those receiving phosphate-buffered saline (PBS), produced significant bacteriolysis. These results suggest that VirB-specific responses that reduce organ colonization byBrucellain mice can be also elicited in dogs.

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Pharmacokinetics of Miltefosine in Children and Adults with Cutaneous Leishmaniasis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02198-16 ◽

2016 ◽

Vol 61 (3) ◽

Cited By ~ 18

Author(s):

María del Mar Castro ◽

Maria Adelaida Gomez ◽

Anke E. Kip ◽

Alexandra Cossio ◽

Eduardo Ortiz ◽

...

Keyword(s):

Cutaneous Leishmaniasis ◽

Drug Exposure ◽

Mononuclear Cells ◽

Open Label ◽

Time Curve ◽

Peripheral Blood Mononuclear ◽

Content Type ◽

Liquid Chromatography Tandem Mass ◽

Intracellular Compartments

ABSTRACT An open-label pharmacokinetics (PK) clinical trial was conducted to comparatively assess the PK and explore the pharmacodynamics (PD) of miltefosine in children and adults with cutaneous leishmaniasis (CL) in Colombia. Sixty patients, 30 children aged 2 to 12 years and 30 adults aged 18 to 60 years, were enrolled. Participants received miltefosine (Impavido) at a nominal dose of 2.5 mg/kg/day for 28 days. Miltefosine concentrations were measured in plasma and peripheral blood mononuclear cells by liquid chromatography-tandem mass spectrometry of samples obtained during treatment and up to 6 months following completion of treatment, when therapeutic outcome was determined. Fifty-two patients were cured, 5 pediatric patients failed treatment, and 3 participants were lost to follow-up. Leishmania (Viannia) panamensis predominated among the strains isolated (42/46; 91%). Noncompartmental analysis demonstrated that plasma and intracellular miltefosine concentrations were, overall, lower in children than in adults. Exposure to miltefosine, estimated by area under the concentration-time curve and maximum concentration, was significantly lower in children in both the central and intracellular compartments (P < 0.01). Leishmania persistence was detected in 43% of study participants at the end of treatment and in 27% at 90 days after initiation of treatment. Clinical response was not dependent on parasite elimination. In vitro miltefosine susceptibility was similar for Leishmania strains from adults and children. Our results document PK differences for miltefosine in children and adults with cutaneous leishmaniasis that affect drug exposure and could influence the outcome of treatment, and they provide bases for optimizing therapeutic regimens for CL in pediatric populations. (This study has been registered at ClinicalTrials.gov under identifier NCT01462500.)

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Hypoxia directed migration of human naïve monocytes is associated with an attenuation of cytokine release: indications for a key role of CCL26

Journal of Translational Medicine ◽

10.1186/s12967-020-02567-7 ◽

2020 ◽

Vol 18 (1) ◽

Author(s):

Lars Hummitzsch ◽

Rouven Berndt ◽

Matthias Kott ◽

Rene Rusch ◽

Fred Faendrich ◽

...

Keyword(s):

Cell Migration ◽

Mononuclear Cells ◽

Human Monocytes ◽

Peripheral Blood Mononuclear ◽

Paracrine Factors ◽

Directed Migration ◽

Hypoxic Conditions ◽

Monocyte Migration

Abstract Background Numerous tissue-derived factors have been postulated to be involved in tissue migration of circulating monocytes. The aim of this study was to evaluate whether a defined hypoxic gradient can induce directed migration of naïve human monocytes and to identify responsible autocrine/paracrine factors. Methods Monocytes were isolated from peripheral blood mononuclear cells, transferred into chemotaxis chambers and subjected to a defined oxygen gradient with or without the addition of CCL26. Cell migration was recorded and secretome analyses were performed. Results Cell migration recordings revealed directed migration of monocytes towards the source of hypoxia. Analysis of the monocyte secretome demonstrated a reduced secretion of 70% (19/27) of the analyzed cytokines under hypoxic conditions. The most down-regulated factors were CCL26 (− 99%), CCL1 (− 95%), CX3CL1 (− 95%), CCL17 (− 85%) and XCL1 (− 83%). Administration of recombinant CCL26 abolished the hypoxia-induced directed migration of human monocytes, while the addition of CCL26 under normoxic conditions resulted in a repulsion of monocytes from the source of CCL26. Conclusions Hypoxia induces directed migration of human monocytes in-vitro. Autocrine/paracrine released CCL26 is involved in the hypoxia-mediated monocyte migration and may represent a target molecule for the modulation of monocyte migration in-vivo.

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