Monocytes arriving at the site of infection differentiate into functional effector macrophages to replenish the resident sentinel cells. Bordetella pertussis, the pertussis agent, secretes an adenylate cyclase toxin-hemolysin (CyaA) that binds myeloid phagocytes through complement receptor 3 (CD11b/CD18) and swiftly delivers its adenylyl cyclase enzyme domain into phagocytes. This ablates the bactericidal capacities of phagocytes through massive and unregulated conversion of cytosolic ATP into the key signaling molecule cAMP. We show that exposure of primary human monocytes to as low a concentration as 22.5 pM CyaA, or a low (2:1) multiplicity of infection by CyaA-producing B. pertussis bacteria, blocks macrophage colony-stimulating factor (M-CSF)-driven differentiation of monocytes. CyaA-induced cAMP signaling mediated through the activity of protein kinase A (PKA) efficiently blocked expression of macrophage markers, and the monocytes exposed to 22.5 pM CyaA failed to acquire the characteristic intracellular complexity of mature macrophage cells. Neither M-CSF-induced endoplasmic reticulum (ER) expansion nor accumulation of Golgi bodies, mitochondria, or lysosomes was observed in toxin-exposed monocytes, which remained small and poorly phagocytic and lacked pseudopodia. Exposure to 22.5 pM CyaA toxin provoked loss of macrophage marker expression on in vitro differentiated macrophages, as well as on primary human alveolar macrophages, which appeared to dedifferentiate into monocyte-like cells with upregulated CD14 levels. This is the first report that terminally differentiated tissue-resident macrophage cells can be dedifferentiated in vitro The results suggest that blocking of monocyte-to-macrophage transition and/or dedifferentiation of the sentinel cells of innate immunity through cAMP-elevating toxin action may represent a novel immune evasion strategy of bacterial pathogens.IMPORTANCE Macrophages are key sentinel cells of the immune system, and, as such, they are targeted by the toxins produced by the pertussis agent Bordetella pertussis The adenylate cyclase toxin (CyaA) mediates immune evasion of B. pertussis by suspending the bactericidal activities of myeloid phagocytes. We reveal a novel mechanism of potential subversion of host immunity, where CyaA at very low (22 pM) concentrations could inhibit maturation of human monocyte precursors into the more phagocytic macrophage cells. Furthermore, exposure to low CyaA amounts has been shown to trigger dedifferentiation of mature primary human alveolar macrophages back into monocyte-like cells. This unprecedented capacity is likely to promote survival of the pathogen in the airways, both by preventing maturation of monocytes attracted to the site of infection into phagocytic macrophages and by dedifferentiation of the already airway-resident sentinel cells.

• Klíčová slova
• Bordetella, adenylate cyclase toxin, cyclic AMP, dedifferentiation, macrophage,
• MeSH
• adenylátcyklasový toxin škodlivé účinky   metabolismus   MeSH
• alveolární makrofágy účinky léků   metabolismus   MeSH
• Bordetella pertussis chemie   MeSH
• buněčná diferenciace účinky léků   MeSH
• interakce hostitele a patogenu MeSH
• lidé MeSH
• monocyty účinky léků   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adenylátcyklasový toxin MeSH

Synovial fluid (SF)-derived monocyte-macrophage (MON-Mϕ)-lineage cells in knee osteoarthritis (KOA) remain poorly understood. We analyzed SF samples from 420 patients with KOA with effusion. The MON-Mϕ cells accounted for 47.4% (median; range 7.1%-94.4%) of CD45+ cells and consisted of four subpopulations that correlated with the distribution and activation of other immune cells. The most abundant subpopulation was that of inactive CD11b+CD14-CD16- myeloid dendritic cells (mDCs; cDC2), which exhibited low cytokine production, low T lymphocyte stimulation, and high migratory ability. Other major subpopulations included CD11b+CD14+CD16- monocyte-like cells and CD11b+CD14+CD16+ macrophages, which share a similar transcriptomic profile. A subpopulation of CD11b-CD14-CD16- mDCs (cDC1) was less common. A higher proportion of CD11b+CD14-CD16- mDCs was linked to early-stage KOA and mild joint pain. Dendritic cells were rarely present in KOA synovium. This study revealed the considerable complexity of SF-derived MON-Mϕ subpopulations and highlighted the role of inactive mDCs in KOA.

• Klíčová slova
• CP: Immunology, early-stage OA, inactive myeloid dendritic cells, joint immune microenvironment, knee osteoarthritis effusion, knee synovitis, monocytes, synovial fluid cells, synovial fluid-derived macrophages, synovial inflammation, synovial tissue,
• MeSH
• artróza kolenních kloubů * patologie   metabolismus   imunologie   MeSH
• buněčný rodokmen MeSH
• dendritické buňky * metabolismus   imunologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• makrofágy * metabolismus   imunologie   MeSH
• monocyty * metabolismus   imunologie   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• synoviální tekutina * metabolismus   imunologie   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Circulating inflammatory monocytes are attracted to infected mucosa and differentiate into macrophage or dendritic cells endowed with enhanced bactericidal and antigen presenting capacities. In this brief Perspective we discuss the newly emerging insight into how the cAMP signaling capacity of Bordetella pertussis adenylate cyclase toxin manipulates the differentiation of monocytes and trigger dedifferentiation of the alveolar macrophages to facilitate bacterial colonization of human airways.

• Klíčová slova
• Bordetella pertussis, adenylate cyclase toxin, dedifferentiation, macrophages, monocytes,
• MeSH
• adenylátcyklasový toxin farmakologie   fyziologie   MeSH
• alveolární makrofágy cytologie   účinky léků   MeSH
• AMP cyklický fyziologie   MeSH
• biologické modely MeSH
• Bordetella pertussis fyziologie   MeSH
• buněčná diferenciace MeSH
• dediferenciace buněk účinky léků   MeSH
• dýchací soustava účinky léků   imunologie   mikrobiologie   MeSH
• fagocytóza MeSH
• interakce hostitele a patogenu imunologie   MeSH
• lidé MeSH
• monocyty cytologie   účinky léků   MeSH
• myši MeSH
• prezentace antigenu účinky léků   MeSH
• přirozená imunita účinky léků   MeSH
• slizniční imunita účinky léků   MeSH
• systémy druhého messengeru účinky léků   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• adenylátcyklasový toxin MeSH
• AMP cyklický MeSH

Maturation of blood cells depends on dramatic changes of expression profiles of specific genes. Although these changes have been extensively studied, their functional outcomes often remain unclear. In this study, we explored the identity and function of an unknown protein that was greatly overexpressed in v-myb-transformed BM2 monoblasts undergoing differentiation to macrophage-like cells. We identified this protein as vimentin, the intermediate filament protein. We show that an increased level of vimentin protein results from activation of the vimentin gene promoter occurring in monoblastic cells induced to differentiate by multiple agents. Furthermore, our studies reveal that the vimentin gene promoter is stimulated by Myb and Jun proteins, the key transcriptional regulators of myeloid maturation. Silencing of vimentin gene expression using siRNA markedly suppressed the ability of BM2 cells to form macrophage polykaryons active in phagocytosis and producing reactive oxygen species. Taken together, these findings document that up-regulation of vimentin gene expression is important for formation of fully active macrophage-like cells and macrophage polykaryons.

• MeSH
• 2D gelová elektroforéza MeSH
• buněčná diferenciace * MeSH
• fibroblasty MeSH
• geny jun genetika   MeSH
• hematopoéza genetika   MeSH
• hmotnostní spektrometrie MeSH
• křepelky a křepelovití MeSH
• kur domácí MeSH
• makrofágy cytologie   fyziologie   MeSH
• monocyty cytologie   fyziologie   MeSH
• onkogenní proteiny v-myb genetika   MeSH
• promotorové oblasti (genetika) genetika   MeSH
• protoonkogenní proteiny c-jun fyziologie   MeSH
• regulace genové exprese MeSH
• tetradekanoylforbolacetát MeSH
• transformované buněčné linie MeSH
• transkripční faktory genetika   MeSH
• upregulace MeSH
• vimentin genetika   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• onkogenní proteiny v-myb MeSH
• protoonkogenní proteiny c-jun MeSH
• tetradekanoylforbolacetát MeSH
• transkripční faktory MeSH
• vimentin MeSH

BACKGROUND: The PLAUR gene encodes the urokinase-like plasminogen activator receptor (uPAR) and may undergo alternative splicing. Excluding cassette exons 3, 5 and 6 from the transcript results in truncated protein variants whose precise functions have not been elucidated yet. The PLAUR gene is one of several expressed in myeloid cells, where uPAR participates in different cellular processes, including the contact activation system and kallikrein-kinin system, which play an important role in hereditary angioedema (HAE) pathogenesis. A hypothesis about the PLAUR splicing pattern impact on HAE severity was tested. METHODS AND RESULTS: The RT-PCR quantified by capillary electrophoresis was used. Although no significant difference in alternative transcript frequency was observed between healthy volunteers and HAE patients, a significant increase in all cassette exon inclusion variants was revealed during monocyte-to-macrophage differentiation. CONCLUSIONS: PLAUR alternative splicing in monocytes and macrophages neither was different between HAE patients and healthy controls, nor reflected disease severity. However, the results showed an PLAUR splicing pattern was changing during monocyte-to-macrophage differentiation, but the significance of these changes is unknown and awaits future clarification.

• Klíčová slova
• Differentiation, HAE, Macrophages, Monocyte, PLAUR, uPAR,
• MeSH
• alternativní sestřih genetika   MeSH
• hereditární angioedémy * genetika   patologie   MeSH
• leukocyty MeSH
• lidé MeSH
• makrofágy patologie   MeSH
• monocyty * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• PLAUR protein, human MeSH Prohlížeč

One of the gene therapy strategies in oncology is immunization with cancer cells that express various cytokines. We used a thymidine-kinase deficient (cTK-) cell line designated 123IA, which had been derived from HPV16-transformed mouse (C57BL/6) cells MK16/I/III/ABC (MK16). To obtain genetically modified cells, 123IA cells were transfected with bicistronic plasmid vectors carrying the herpes simplex type 1 thymidine kinase (HSV TK) gene and either the gene for the mouse B7.1 (CD80) co-stimulatory molecule or the gene for the monocyte-chemoattractant protein 1 (MCP-1). For control purposes, a plasmid vector carrying only the HSV TK gene was used. The transfected cells were cultivated in medium supplemented with hypoxanthine, aminopterin and thymidine. For comparative purposes we also used B9 cells, which express the granulocyte-macrophage colony stimulation factor (GM-CSF) and had been derived from 123A cells by transduction with the recombinant adeno-associated virus carrying the HSV TK gene and the mouse GM-CSF gene. All of the cell lines isolated were found to be sensitive to minute amounts of ganciclovir, revealing the production of HSV TK, and to express the respective transgenes. When inoculated into 5-week-old female syngeneic mice, cells expressing either GM-CSF or B7.1 were non-oncogenic. On the other hand, nearly all mice inoculated with MCP-1-producing cells developed tumours, though considerably later than animals inoculated with the same dose of the parental MK16 cells. Animals injected with GM-CSF- or B7.1-producing cells were protected against challenge with the parental MK16 cells. When another mouse (C57BL/6) HPV16-transformed oncogenic cell line, TC-1, which differs from the MK16 cells in a number of properties such as MHC class I and B7.1 expression, was used for the challenge, the protective effect was much less pronounced.

• MeSH
• antigeny CD80 genetika   MeSH
• buněčné linie MeSH
• chemokin CCL2 genetika   MeSH
• faktor stimulující granulocyto-makrofágové kolonie genetika   MeSH
• ganciklovir farmakologie   MeSH
• geny MHC třídy I MeSH
• imunizace MeSH
• lidský papilomavirus 16 genetika   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• onkogenní proteiny virové genetika   MeSH
• Papillomavirus E7 - proteiny MeSH
• protinádorové vakcíny imunologie   MeSH
• represorové proteiny genetika   MeSH
• transfekce MeSH
• transformované buněčné linie MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antigeny CD80 MeSH
• Ccl2 protein, mouse MeSH Prohlížeč
• chemokin CCL2 MeSH
• E6 protein, Human papillomavirus type 16 MeSH Prohlížeč
• faktor stimulující granulocyto-makrofágové kolonie MeSH
• ganciklovir MeSH
• oncogene protein E7, Human papillomavirus type 16 MeSH Prohlížeč
• onkogenní proteiny virové MeSH
• Papillomavirus E7 - proteiny MeSH
• protinádorové vakcíny MeSH
• represorové proteiny MeSH

Bordetella pertussis infects human upper airways and deploys an array of immunosuppressive virulence factors, among which the adenylate cyclase toxin (CyaA) plays a prominent role in disarming host phagocytes. CyaA binds the complement receptor-3 (CR3 aka αMβ2 integrin CD11b/CD18 or Mac-1) of myeloid cells and delivers into their cytosol an adenylyl cyclase enzyme that hijacks cellular signaling through unregulated conversion of cytosolic ATP to cAMP. We found that the action of as little CyaA as 22 pM (4 ng/mL) blocks macrophage colony-stimulating factor (M-CSF)-driven transition of migratory human CD14+ monocytes into macrophages. Global transcriptional profiling (RNAseq) revealed that exposure of monocytes to 22 pM CyaA for 40 hours in culture with 20 ng/mL of M-CSF led to upregulation of genes that exert negative control of monocyte to macrophage differentiation (e.g., SERPINB2, DLL1, and CSNK1E). The sustained CyaA action yielded downregulation of numerous genes involved in processes crucial for host defense, such as myeloid cell differentiation, chemotaxis of inflammatory cells, antigen presentation, phagocytosis, and bactericidal activities. CyaA-elicited signaling also promoted deacetylation and trimethylation of lysines 9 and 27 of histone 3 (H3K9me3 and H3K27me3) and triggered the formation of transcriptionally repressive heterochromatin patches in the nuclei of CyaA-exposed monocytes. These effects were partly reversed by the G9a methyltransferase inhibitor UNC 0631 and by the pleiotropic HDAC inhibitor Trichostatin-A, revealing that CyaA-elicited epigenetic alterations mediate transcriptional reprogramming of monocytes and play a role in CyaA-triggered block of monocyte differentiation into bactericidal macrophage cells.IMPORTANCETo proliferate on host airway mucosa and evade elimination by patrolling sentinel cells, the whooping cough agent Bordetella pertussis produces a potently immunosubversive adenylate cyclase toxin (CyaA) that blocks opsonophagocytic killing of bacteria by phagocytes like neutrophils and macrophages. Indeed, chemotactic migration of CD14+ monocytes to the infection site and their transition into bactericidal macrophages, thus replenishing the exhausted mucosa-patrolling macrophages, represents one of the key mechanisms of innate immune defense to infection. We show that the cAMP signaling action of CyaA already at a very low toxin concentration triggers massive transcriptional reprogramming of monocytes that is accompanied by chromatin remodeling and epigenetic histone modifications, which block the transition of migratory monocytes into bactericidal macrophage cells. This reveals a novel layer of toxin action-mediated hijacking of functional differentiation of innate immune cells for the sake of mucosal pathogen proliferation and transmission to new hosts.

• Klíčová slova
• Bordetella pertussis, RTX toxins, cyclic AMP, differentiation, epigenetics, macrophages, monocytes,
• MeSH
• adenylátcyklasový toxin * metabolismus   MeSH
• Bordetella pertussis * patogenita   enzymologie   MeSH
• buněčná diferenciace * účinky léků   MeSH
• faktor stimulující kolonie makrofágů MeSH
• kultivované buňky MeSH
• lidé MeSH
• makrofágy * účinky léků   cytologie   MeSH
• monocyty * účinky léků   cytologie   fyziologie   MeSH
• přeprogramování buněk * MeSH
• restrukturace chromatinu * účinky léků   MeSH
• signální transdukce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• adenylátcyklasový toxin * MeSH
• faktor stimulující kolonie makrofágů MeSH

Adherence and phagocytosis of invasive and noninvasive Neisseria meningitidis strains was investigated using light, fluorescence and electron microscopy. Invasive strains were isolated from the cerebrospinal fluid and/or blood of the patients with invasive meningococcal disease and noninvasive strains from the nasopharynx and/or larynx of healthy carriers. Adherence/endocytosis was studied on monkey kidney cells (the LLC-MK2 cell line) and phagocytosis on mouse monocytes and human macrophages (the P388D1 and U-937 cell lines, respectively). Although invasive and noninvasive meningococci isolated in the same cluster showed identical genotype and phenotype markers, they were found to interact differently with epithelial cells as well as with monocytes/macrophages. Invasive isolates displayed higher adherence to the surface of LLC-MK2 cells compared to noninvasive ones. Phagocytosis by P388D1 cells of noninvasive strains was effective and the bacteria were damaged by cytolysis. In contrast, invasive bacteria frequently persisted in "coiling" vacuoles and in effect could destroy the host cell. This is the first demonstration of coiling phagocytosis induced by meningococci. Efficiency of phagocytosis by U-937 cells was significantly higher for the noninvasive than invasive strains. Different behaviour of invasive and noninvasive strains of N. meningitidis observed during 4 hours of interactions with epithelial cells and monocytes/macrophages reflects well the higher pathogenic potential of invasive bacteria.

• MeSH
• epitelové buňky mikrobiologie   MeSH
• fagocytóza MeSH
• farynx mikrobiologie   MeSH
• Haplorrhini MeSH
• lidé MeSH
• makrofágy mikrobiologie   MeSH
• meningokokové infekce krev   mozkomíšní mok   mikrobiologie   MeSH
• monocyty mikrobiologie   ultrastruktura   MeSH
• myši MeSH
• Neisseria meningitidis patogenita   MeSH
• U937 buňky MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Prenatal development of cord blood monocytes and tissue macrophages was studied in pig foetuses by immunophenotyping and functional assays. The function of peripheral blood monocytes was compared in germ-free and conventional piglets. First macrophages were identified by electron microscopy in foetal liver on the 25th day of gestation. Monoclonal antibodies against porcine CD45 and SWC3 antigens were used for flow cytometric identification of myelomonocytic cells in cell suspensions prepared from the yolk sac, foetal liver, spleen and cord blood. Leukocytes expressing the common myelomonocytic antigen SWC3 were found in all organs studied since the earliest stages of development. Opsonized zymosan ingestion assay was used to determine the phagocytic capacity of foetal mononuclear phagocytes isolated from cord blood, liver and spleen. In the foetal liver, avid phagocytosis of apoptic cells had been found to occur before cells were able to ingest zymosan in vitro. The first cells capable of ingesting zymosan particles were found on the 40th day of gestation in umbilical blood and 17 days later in foetal spleen and liver. Their relative proportion increased with age. Cord blood monocytes and peripheral blood monocytes in germ-free piglets had low oxidatory burst activity as shown by iodonitrophenyl tetrazolium reduction assay. A remarkable increase of oxidatory burst activity was observed in conventional piglets, probably due to activation of immune mechanisms by the microflora colonizing gastrointestinal tract.

• MeSH
• antigeny CD45 analýza   MeSH
• apoptóza MeSH
• elektronová mikroskopie MeSH
• fagocytóza MeSH
• fetální krev cytologie   MeSH
• gestační stáří MeSH
• imunofenotypizace MeSH
• játra cytologie   embryologie   MeSH
• makrofágy imunologie   fyziologie   MeSH
• MHC antigeny II. třídy analýza   MeSH
• monocyty imunologie   fyziologie   MeSH
• monoklonální protilátky MeSH
• prasata MeSH
• průtoková cytometrie MeSH
• slezina cytologie   embryologie   MeSH
• žloutkový vak cytologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antigeny CD45 MeSH
• MHC antigeny II. třídy MeSH
• monoklonální protilátky MeSH

Porcine cytomegalovirus (PCMV) causes lifelong latent infections in swine. The pathogen is occasionally associated with inclusion body rhinitis and pneumonia in piglets, reproductive disorders in pregnant sows and respiratory disease complex in older pigs. Immunosuppressive potential of PCMV infection is discussed. Macrophages were recognised as one of target cell types where propagation of virus occurs. The aim of present study was to set up model PCMV infection of monocyte derived macrophages (MDMs) in vitro for PCMV immunobiology research. Obtained results showed that PCMV is able to infect and propagate in MDMs. Possible immunosuppressive effect of PCMV on infected macrophages was evaluated by measurement of immune relevant gene expression in MDMs. Infection decreased expression of IL-8 and TNF-α (pro-inflammatory cytokines) and increased expression of IL-10 (anti-inflammatory cytokine) on mRNA transcription level. Obtained data support hypothesis that higher sensitivity of animals to coinfection with other swine pathogens and its more severe clinical manifestations could potentially be the consequence of PCMV infection.

• Klíčová slova
• Immune response, Macrophages, Porcine cytomegalovirus,
• MeSH
• cytokiny imunologie   MeSH
• cytomegalovirové infekce imunologie   veterinární   MeSH
• Cytomegalovirus fyziologie   MeSH
• exprese genu imunologie   MeSH
• interleukin-10 imunologie   MeSH
• interleukin-8 imunologie   MeSH
• makrofágy imunologie   virologie   MeSH
• nemoci prasat imunologie   virologie   MeSH
• prasata MeSH
• přirozená imunita MeSH
• TNF-alfa imunologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cytokiny MeSH
• interleukin-10 MeSH
• interleukin-8 MeSH
• TNF-alfa MeSH