PURPOSE OF REVIEW: RTX toxin action often defines the outcome of bacterial infections. Here, we discuss the progress in understanding the impacts of RTX toxin activities on host immunity. RECENT FINDINGS: Bordetella pertussis CyaA activity paralyzes sentinel phagocytic cells by elevating cellular cAMP levels and blocks differentiation of infiltrating monocytes into bactericidal macrophages, promoting also de-differentiation of resident alveolar macrophages into monocyte-like cells. Vibrio cholerae multifunctional autoprocessing repeats-in-toxins (MARTX), through Rho inactivating and α/β-hydrolase (ABH) domain action blocks mitogen-activated protein kinase signaling in epithelial cells and dampens the inflammatory responses of intestinal epithelia by blocking immune cell recruitment. The action of actin crosslinking effector domain and Ras/Rap1-specific endopeptidase (RRSP) domains of MARTX compromises the phagocytic ability of macrophages. Aggregatibacter actinomycetemcomitans LtxA action triggers neutrophil elastase release into periodontal tissue, compromising the epithelial barrier and promoting bacterial spreads into deeper tissue. SUMMARY: Action of RTX toxins enables bacterial pathogens to cope with the fierce host immune defenses. RTX toxins often block phagocytosis and bactericidal reactive oxygen species and NO production. Some RTX toxins can reprogram the macrophages to less bactericidal cell types. Autophagy is hijacked for example by the activity of the V. cholerae ABH effector domain of the MARTX protein. Subversion of immune functions by RTX toxins thus promotes bacterial survival and proliferation in the host.

• MeSH
• adaptivní imunita MeSH
• Bacteria metabolismus   patogenita   MeSH
• bakteriální toxiny toxicita   MeSH
• buňky NK účinky léků   MeSH
• dendritické buňky účinky léků   MeSH
• epitelové buňky účinky léků   MeSH
• lidé MeSH
• makrofágy účinky léků   MeSH
• monocyty účinky léků   MeSH
• virulence MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The preparation of dendritic cells (DCs) for adoptive cellular immunotherapy (ACI) requires the maturation of ex vivo-produced immature(i) DCs. This maturation ensures that the antigen presentation triggers an immune response towards the antigen-expressing cells. Although there is a large number of maturation agents capable of inducing strong DC maturation, there is still only a very limited number of these agents approved for use in the production of DCs for ACI. In seeking novel DC maturation agents, we used differentially activated human mast cell (MC) line LAD2 as a cellular adjuvant to elicit or modulate the maturation of ex vivo-produced monocyte-derived iDCs. We found that co-culture of iDCs with differentially activated LAD2 MCs in serum-containing media significantly modulated polyinosinic:polycytidylic acid (poly I:C)-elicited DC maturation as determined through the surface expression of the maturation markers CD80, CD83, CD86, and human leukocyte antigen(HLA)-DR. Once iDCs were generated in serum-free conditions, they became refractory to the maturation with poly I:C, and the LAD2 MC modulatory potential was minimized. However, the maturation-refractory phenotype of the serum-free generated iDCs was largely overcome by co-culture with thapsigargin-stimulated LAD2 MCs. Our data suggest that differentially stimulated mast cells could be novel and highly potent cellular adjuvants for the maturation of DCs for ACI.

• MeSH
• adjuvancia imunologická farmakologie   MeSH
• buněčná diferenciace účinky léků   MeSH
• buněčné kultury metody   MeSH
• dendritické buňky cytologie   účinky léků   imunologie   MeSH
• imunoterapie adoptivní * MeSH
• kokultivační techniky MeSH
• lidé MeSH
• mastocyty cytologie   účinky léků   imunologie   MeSH
• monocyty cytologie   účinky léků   imunologie   MeSH
• prezentace antigenu účinky léků   imunologie   MeSH
• thapsigargin farmakologie   MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Wear particles from automotive friction brake pads of various sizes, morphology, and chemical composition are significant contributors towards particulate matter. Knowledge concerning the potential adverse effects following inhalation exposure to brake wear debris is limited. Our aim was, therefore, to generate brake wear particles released from commercial low-metallic and non-asbestos organic automotive brake pads used in mid-size passenger cars by a full-scale brake dynamometer with an environmental chamber simulating urban driving and to deduce their potential hazard in vitro. The collected fractions were analysed using scanning electron microscopy via energy-dispersive X-ray spectroscopy (SEM-EDS) and Raman microspectroscopy. The biological impact of the samples was investigated using a human 3D multicellular model consisting of human epithelial cells (A549) and human primary immune cells (macrophages and dendritic cells) mimicking the human epithelial tissue barrier. The viability, morphology, oxidative stress, and (pro-)inflammatory response of the cells were assessed following 24 h exposure to ~ 12, ~ 24, and ~ 48 µg/cm2 of non-airborne samples and to ~ 3.7 µg/cm2 of different brake wear size fractions (2-4, 1-2, and 0.25-1 µm) applying a pseudo-air-liquid interface approach. Brake wear debris with low-metallic formula does not induce any adverse biological effects to the in vitro lung multicellular model. Brake wear particles from non-asbestos organic formulated pads, however, induced increased (pro-)inflammatory mediator release from the same in vitro system. The latter finding can be attributed to the different particle compositions, specifically the presence of anatase.

• MeSH
• biologické modely * MeSH
• buňky A549 MeSH
• cytokiny metabolismus   MeSH
• dendritické buňky účinky léků   metabolismus   ultrastruktura   MeSH
• kokultivační techniky MeSH
• látky znečišťující vzduch toxicita   MeSH
• lidé MeSH
• makrofágy účinky léků   metabolismus   ultrastruktura   MeSH
• motorová vozidla MeSH
• oxidační stres účinky léků   MeSH
• pevné částice toxicita   MeSH
• plíce účinky léků   metabolismus   patologie   MeSH
• povrchové vlastnosti MeSH
• velikost částic MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Germ-free animals have been used to define the vital role of commensal bacteria on the maturation of the host immune system. However, the role of bacterial residues in diet in this setting is poorly understood. Here we investigated the effect of bacterial contamination in sterile diet on the level of allergic sensitization in germ-free mice. Sterile grain-based diets ST1 and R03 were tested for the level of bacterial contamination. ST1 contained higher amount of bacterial DNA, approximately ten times more endotoxin, and induced higher, TLR4-dependent, cytokine production in dendritic cells compared to R03. In a germ-free mouse model of sensitization to the major birch pollen allergen Bet v 1, feeding on ST1 for at least two generations was associated with decreased production of allergen-specific IgE and IgG1 antibodies in sera in comparison to R03. Furthermore, reduced levels of allergen-specific and ConA-induced cytokines IL-4, IL-5 and IL-13 accompanied by increased levels of IFN-γ were detected in splenocytes cultures of these mice. Our results show that contamination of experimental diet with bacterial residues, such as endotoxin, significantly affects the development of allergic sensitization in germ-free mice. Therefore, careful selection of sterile food is critical for the outcomes of germ-free or gnotobiotic experimental models of immune-deviated diseases.

• MeSH
• alergie imunologie   patologie   MeSH
• antigeny rostlinné imunologie   MeSH
• buněčná diferenciace účinky léků   MeSH
• chov MeSH
• cytokiny biosyntéza   MeSH
• dendritické buňky účinky léků   MeSH
• dieta * MeSH
• DNA bakterií analýza   MeSH
• endotoxiny toxicita   MeSH
• epitopy imunologie   MeSH
• gnotobiologické modely MeSH
• HEK293 buňky MeSH
• imunizace * MeSH
• imunoglobulin A imunologie   MeSH
• imunoglobulin G imunologie   MeSH
• kontaminace DNA MeSH
• kultivované buňky MeSH
• lidé MeSH
• ligandy MeSH
• mitogeny farmakologie   MeSH
• myši inbrední BALB C MeSH
• slezina patologie   MeSH
• toll-like receptor 4 metabolismus   MeSH
• trávicí systém imunologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Filamentous hemagglutinin (FHA) is an important adhesin of the whooping cough agent Bordetella pertussis and is contained in most acellular pertussis vaccines. Recently, FHA was proposed to exert an immunomodulatory activity through induction of tolerogenic IL-10 secretion from dendritic cells. We have re-evaluated the cytokine-inducing activity of FHA, placing specific emphasis on the role of the residual endotoxin contamination of FHA preparations. We show that endotoxin depletion did not affect the capacity of FHA to bind primary human monocyte-derived dendritic cells, while it abrogated the capacity of FHA to elicit TNF-α and IL-10 secretion and strongly reduced its capacity to trigger IL-6 production. The levels of cytokines induced by the different FHA preparations correlated with their residual contents of B. pertussis endotoxin. Moreover, FHA failed to trigger cytokine secretion in the presence of antibodies that block TLR2 and/or TLR4 signaling. The TLR2 signaling capacity appeared to be linked to the presence of endotoxin-associated components in FHA preparations and not to the FHA protein itself. These results show that the endotoxin-depleted FHA protein does not induce cytokine release from human dendritic cells.

• MeSH
• bakteriální adheziny imunologie   MeSH
• dendritické buňky účinky léků   imunologie   MeSH
• faktory virulence rodu Bordetella imunologie   MeSH
• interleukin-10 sekrece   MeSH
• kultivované buňky MeSH
• lidé MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Crosslinking of regulatory immunoreceptors (RR), such as BDCA-2 (CD303) or ILT7 (CD85g), of plasmacytoid dendritic cells (pDCs) efficiently suppresses production of type-I interferon (IFN)-α/β and other cytokines in response to Toll-like receptor (TLR) 7/9 ligands. This cytokine-inhibitory pathway is mediated by spleen tyrosine kinase (Syk) associated with the ITAM-containing adapter of RR. Here we demonstrate by pharmacological targeting of Syk that in addition to the negative regulation of TLR7/9 signaling via RR, Syk also positively regulates the TLR7/9 pathway in human pDCs. Novel highly specific Syk inhibitor AB8779 suppressed IFN-α, TNF-α and IL-6 production induced by TLR7/9 agonists in primary pDCs and in the pDC cell line GEN2.2. Triggering of TLR9 or RR signaling induced a differential kinetics of phosphorylation at Y352 and Y525/526 of Syk and a differential sensitivity to AB8779. Consistent with the different roles of Syk in TLR7/9 and RR signaling, a concentration of AB8779 insufficient to block TLR7/9 signaling still released the block of IFN-α production triggered via the RR pathway, including that induced by hepatitis B and C viruses. Thus, pharmacological targeting of Syk partially restored the main pDC function-IFN-α production. Opposing roles of Syk in TLR7/9 and RR pathways may regulate the innate immune response to weaken inflammation reaction.

• MeSH
• cytokiny metabolismus   MeSH
• dendritické buňky účinky léků   metabolismus   MeSH
• fosforylace účinky léků   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• kinasa Syk antagonisté a inhibitory   metabolismus   MeSH
• lidé MeSH
• přirozená imunita MeSH
• signální transdukce účinky léků   fyziologie   MeSH
• toll-like receptory agonisté   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Tolerogenic dendritic cells (tDCs) may offer an intervention therapy in autoimmune diseases or transplantation. Stable immaturity and tolerogenic function of tDCs after encountering inflammatory environment are prerequisite for positive outcome of immunotherapy. However, the signaling pathways regulating their stable tolerogenic properties are largely unknown. In this study, we demonstrated that human monocyte-derived tDCs established by using paricalcitol (analogue of vitamin D2), dexamethasone and monophosphoryl lipid A exposed for 24h to LPS, cytokine cocktail, polyI:C or CD40L preserved reduced expression of co-stimulatory molecules, increased levels of inhibitory molecules ILT-3, PDL-1 and TIM-3, increased TLR-2, increased secretion of IL-10 and TGF-β, reduced IL-12 and TNF-α secretion and reduced T cell stimulatory capacity. tDCs further induced IL-10-producing T regulatory cells that suppressed the proliferation of responder T cells. In the inflammatory environment, tDCs maintained up-regulated indoleamine 2, 3 dioxygenase but abrogated IκB-α phosphorylation and reduced transcriptional activity of p65/RelA, RelB and c-Rel NF-κB subunits except p50. Mechanistically, p38 MAPK, ERK1/2, mTOR, STAT3 and mTOR-dependent glycolysis regulated expression of ILT-3, PDL-1 and CD86, secretion of IL-10 and T cell stimulatory capacity of tDCs in the inflammatory environment. Stability of tDCs in the inflammatory environment is thus regulated by multiple signaling pathways.

• MeSH
• buněčná diferenciace fyziologie   MeSH
• dendritické buňky účinky léků   metabolismus   MeSH
• dexamethason farmakologie   MeSH
• ergokalciferoly farmakologie   MeSH
• glykolýza účinky léků   MeSH
• kultivované buňky MeSH
• lidé MeSH
• mitogenem aktivované proteinkinasy metabolismus   MeSH
• NF-kappa B metabolismus   MeSH
• signální transdukce MeSH
• TOR serin-threoninkinasy metabolismus   MeSH
• transkripční faktor STAT3 metabolismus   MeSH
• zánět metabolismus   patologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Transmission of pathogens by ticks is greatly supported by tick saliva released during feeding. Dendritic cells (DC) act as immunological sentinels and interconnect the innate and adaptive immune system. They control polarization of the immune response towards Th1 or Th2 phenotype. We investigated whether salivary cystatins from the hard tick Ixodes scapularis, sialostatin L (Sialo L) and sialostatin L2 (Sialo L2), influence mouse dendritic cells exposed to Borrelia burgdorferi and relevant Toll-like receptor ligands. METHODS: DCs derived from bone-marrow by GM-CSF or Flt-3 ligand, were activated with Borrelia spirochetes or TLR ligands in the presence of 3 μM Sialo L and 3 μM Sialo L2. Produced chemokines and IFN-β were measured by ELISA test. The activation of signalling pathways was tested by western blotting using specific antibodies. The maturation of DC was determined by measuring the surface expression of CD86 by flow cytometry. RESULTS: We determined the effect of cystatins on the production of chemokines in Borrelia-infected bone-marrow derived DC. The production of MIP-1α was severely suppressed by both cystatins, while IP-10 was selectively inhibited only by Sialo L2. As TLR-2 is a major receptor activated by Borrelia spirochetes, we tested whether cystatins influence signalling pathways activated by TLR-2 ligand, lipoteichoic acid (LTA). Sialo L2 and weakly Sialo L attenuated the extracellular matrix-regulated kinase (Erk1/2) pathway. The activation of phosphatidylinositol-3 kinase (PI3K)/Akt pathway and nuclear factor-κB (NF-κB) was decreased only by Sialo L2. In response to Borrelia burgdorferi, the activation of Erk1/2 was impaired by Sialo L2. Production of IFN-β was analysed in plasmacytoid DC exposed to Borrelia, TLR-7, and TLR-9 ligands. Sialo L, in contrast to Sialo L2, decreased the production of IFN-β in pDC and also impaired the maturation of these cells. CONCLUSIONS: This study shows that DC responses to Borrelia spirochetes are affected by tick cystatins. Sialo L influences the maturation of DC thus having impact on adaptive immune response. Sialo L2 affects the production of chemokines potentially engaged in the development of inflammatory response. The impact of cystatins on Borrelia growth in vivo is discussed.

• MeSH
• Borrelia burgdorferi imunologie   MeSH
• cystatiny farmakologie   MeSH
• dendritické buňky účinky léků   fyziologie   MeSH
• klíště fyziologie   MeSH
• kyseliny teichoové MeSH
• lipopolysacharidy MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• signální transdukce fyziologie   MeSH
• sliny chemie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Th17 cells constitute a subset of CD4(+) T lymphocytes that play a crucial role in protection against extracellular bacteria and fungi. They are also associated with tissue injury in autoimmune and inflammatory diseases. Here, we report that serpin from the tick Ixodes ricinus, IRS-2, inhibits Th17 differentiation by impairment of the interleukin-6 (IL-6)/STAT-3 signaling pathway. Following activation, mature dendritic cells produce an array of cytokines, including the pleiotropic cytokine IL-6, which triggers the IL-6 signaling pathway. The major transcription factor activated by IL-6 is STAT-3. We show that IRS-2 selectively inhibits production of IL-6 in dendritic cells stimulated with Borrelia spirochetes, which leads to attenuated STAT-3 phosphorylation and finally to impaired Th17 differentiation. The results presented extend the knowledge about the effect of tick salivary serpins on innate immunity cells and their function in driving adaptive immune responses.

• MeSH
• Borrelia imunologie   MeSH
• buněčná diferenciace účinky léků   MeSH
• buňky Th17 účinky léků   fyziologie   MeSH
• dendritické buňky účinky léků   fyziologie   MeSH
• interleukin-6 antagonisté a inhibitory   metabolismus   MeSH
• klíště MeSH
• myši inbrední C57BL MeSH
• serpiny metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• transkripční faktor STAT3 antagonisté a inhibitory   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• alveolární makrofágy enzymologie   imunologie   účinky léků   MeSH
• chronická obstrukční plicní nemoc * farmakoterapie   imunologie   patofyziologie   MeSH
• dendritické buňky imunologie   účinky léků   MeSH
• eozinofily imunologie   účinky léků   MeSH
• lidé MeSH
• mediátory zánětu diagnostické užití   imunologie   MeSH
• neutrofily enzymologie   imunologie   účinky léků   MeSH
• zánět * diagnóza   imunologie   MeSH
• Check Tag
• lidé MeSH