Depletion and functional impairment of circulating plasmacytoid dendritic cells (pDCs) are characteristic attributes of HIV-1-infection. The mechanism of dysfunction of pDCs is unclear. Here, we studied the development of phenotype of pDCs in a cohort of HIV-1-infected individuals monitored before the initiation and during a 9-month follow up with antiretroviral therapy (ART). Using polychromatic flow cytometry, we detected significantly higher pDC-surface expression of the HIV-1 receptor CD4, regulatory receptor BDCA-2, Fcγ receptor CD32, pDC dysfunction marker TIM-3, and the marker of killer pDC, TRAIL, in treatment-naïve HIV-1-infected individuals before initiation of ART when compared to healthy donors. After 9 months of ART, all of these markers approached but did not reach the expression levels observed in healthy donors. We found that the rate of decline in HIV-1 RNA level over the first 3 months of ART negatively correlated with the expression of TIM-3 on pDCs. We conclude that immunogenic phenotype of pDCs is not significantly restored after sustained suppression of HIV-1 RNA level in ART-treated patients and that the level of the TIM-3 expressed on pDCs in treatment naïve patients could be a predictive marker of the rate of decline in the HIV-1 RNA level during ART.

• Klíčová slova
• BDCA-2, HIV-1, T cell Ig and mucin-domain containing molecule 3 (TIM-3), Toll-like receptors 7 and 9 (TLR7/9), antiretroviral therapy (ART), innate and adaptive immune responses, pDC dysfunction, plasmacytoid dendritic cells (pDCs),
• MeSH
• biologické markery MeSH
• buněčný receptor 2 viru hepatitidy A genetika   MeSH
• CD4-pozitivní T-lymfocyty účinky léků   imunologie   metabolismus   MeSH
• dendritické buňky imunologie   metabolismus   MeSH
• dospělí MeSH
• exprese genu * MeSH
• HIV infekce farmakoterapie   genetika   imunologie   virologie   MeSH
• HIV-1 * imunologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• počet CD4 lymfocytů MeSH
• RNA virová MeSH
• virová nálož MeSH
• vysoce aktivní antiretrovirová terapie MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• biologické markery MeSH
• buněčný receptor 2 viru hepatitidy A MeSH
• HAVCR2 protein, human MeSH Prohlížeč
• RNA virová MeSH

Recent studies have reported that the crosslinking of regulatory receptors (RRs), such as blood dendritic cell antigen 2 (BDCA-2) (CD303) or ILT7 (CD85g), of plasmacytoid dendritic cells (pDCs) efficiently suppresses the production of type I interferons (IFN-I, α/β/ω) and other cytokines in response to toll-like receptor 7 and 9 (TLR7/9) ligands. The exact mechanism of how this B cell receptor (BCR)-like signaling blocks TLR7/9-mediated IFN-I production is unknown. Here, we stimulated BCR-like signaling by ligation of RRs with BDCA-2 and ILT7 mAbs, hepatitis C virus particles, or BST2 expressing cells. We compared BCR-like signaling in proliferating pDC cell line GEN2.2 and in primary pDCs from healthy donors, and addressed the question of whether pharmacological targeting of BCR-like signaling can antagonize RR-induced pDC inhibition. To this end, we tested the TLR9-mediated production of IFN-I and proinflammatory cytokines in pDCs exposed to a panel of inhibitors of signaling molecules involved in BCR-like, MAPK, NF-ĸB, and calcium signaling pathways. We found that MEK1/2 inhibitors, PD0325901 and U0126 potentiated TLR9-mediated production of IFN-I in GEN2.2 cells. More importantly, MEK1/2 inhibitors significantly increased the TLR9-mediated IFN-I production blocked in both GEN2.2 cells and primary pDCs upon stimulation of BCR-like or phorbol 12-myristate 13-acetate-induced protein kinase C (PKC) signaling. Triggering of BCR-like and PKC signaling in pDCs resulted in an upregulation of the expression and phoshorylation of c-FOS, a downstream gene product of the MEK1/2-ERK pathway. We found that the total level of c-FOS was higher in proliferating GEN2.2 cells than in the resting primary pDCs. The PD0325901-facilitated restoration of the TLR9-mediated IFN-I production correlated with the abrogation of MEK1/2-ERK-c-FOS signaling. These results indicate that the MEK1/2-ERK pathway inhibits TLR9-mediated type I IFN production in pDCs and that pharmacological targeting of MEK1/2-ERK signaling could be a strategy to overcome immunotolerance of pDCs and re-establish their immunogenic activity.

• Klíčová slova
• B cell-like receptor signaling, MEK1/2, blood dendritic cell antigen 2, c-FOS, plasmacytoid dendritic cells, regulatory receptors, toll-like receptors 7 and 9 (TLR7/9), type I interferon,
• MeSH
• B-lymfocyty imunologie   MeSH
• buněčné linie MeSH
• dendritické buňky fyziologie   MeSH
• extracelulárním signálem regulované MAP kinasy metabolismus   MeSH
• interferon typ I metabolismus   MeSH
• lidé MeSH
• MAP kinasa-kinasa 1 metabolismus   MeSH
• MAP kinasa-kinasa 2 metabolismus   MeSH
• MAP kinasový signální systém MeSH
• NF-kappa B metabolismus   MeSH
• proteinkinasa C metabolismus   MeSH
• protoonkogenní proteiny c-fos metabolismus   MeSH
• receptory antigenů B-buněk genetika   metabolismus   MeSH
• toll-like receptor 9 metabolismus   MeSH
• vápníková signalizace MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• extracelulárním signálem regulované MAP kinasy MeSH
• interferon typ I MeSH
• MAP kinasa-kinasa 1 MeSH
• MAP kinasa-kinasa 2 MeSH
• NF-kappa B MeSH
• proteinkinasa C MeSH
• protoonkogenní proteiny c-fos MeSH
• receptory antigenů B-buněk MeSH
• toll-like receptor 9 MeSH

The innate immune cells sense microbial infection and self-ligands by pathogen recognition receptors (PRRs), such as toll-like receptors (TLRs) and regulatory receptors (RRs), associated with immunoreceptor tyrosine-based activation motif (ITAM). Rapid activation and concerted action of PRRs signaling and feedback inhibitory mechanisms must be engaged to ensure the host defense functions and to prevent cytotoxicity associated with excessive activation. ITAM-associated RRs can generate stimulatory or, paradoxically, inhibitory signals. The network of ITAM-associated RR, together with TLR-signaling pathways, are responsible for immunogenic or tolerogenic responses of macrophages and dendritic cells to their microenvironment. In macrophages, TLR4 signaling is inhibited by low-avidity ligation of ITAM-associated receptors, while high-avidity ligation of ITAM-associated receptors results in potentiation of TLR4 signaling together with resistance to extracellular cytokine microenvironment signals. In contrast to macrophages, TLR7/9 signaling in plasmacytoid DCs (pDCs) is inhibited by high-avidity ligation of ITAM-associated RR, while low-avidity ligation does not show any effect. Surprisingly, interference of ITAM-associated receptor signaling with TLR pathways has not been reported in conventional dendritic cells. Here, we present an overview of molecular mechanisms acting at the crossroads of TLR and ITAM-signaling pathways and address the question of how the high-avidity engagement of the ITAM-associated receptors in pDCs inhibits TLR7/9 signaling. Cellular context and spatiotemporal engagement of ITAM- and TLR-signaling pathways are responsible for different outcomes of macrophage versus pDC activation. While the cross-regulation of cytokine and TLR signaling, together with antigen presentation, are the principal functions of ITAM-associated RR in macrophages, the major role of these receptors in pDCs seems to be related to inhibition of cytokine production and reestablishment of a tolerogenic state following pDC activation. Pharmacologic targeting of TLR and ITAM signaling could be an attractive new therapeutic approach for treatment of chronic infections, cancer, and autoimmune and inflammatory diseases related to pDCs.

• Klíčová slova
• B cell receptor-like signaling, conventional dendritic cells, immunoreceptor tyrosine-based activation motif-associated receptor, macrophage, plasmacytoid dendritic cell, regulatory receptors, toll-like receptors,
• Publikační typ
• časopisecké články MeSH
• přehledy 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
• Názvy látek
• cytokiny MeSH
• inhibitory proteinkinas MeSH
• kinasa Syk MeSH
• toll-like receptory MeSH