Celiac disease (CeD) manifests with autoimmune intestinal inflammation from gluten and genetic predisposition linked to human leukocyte antigen class-II (HLA-II) gene variants. Antigen-presenting cells facilitate gluten exposition through the interaction of their surface major histocompatibility complex (MHC) with the T cell receptor (TCR) on T lymphocytes. This fundamental mechanism of adaptive immunity has broadened upon recognition of extracellular exosomal MHC, raising awareness of an alternative means for antigen presentation. This study demonstrates that conditioned growth media (CGM) previously exposed to monocyte-derived dendritic cells from CeD significantly downregulates the CD3+ lineage marker of control T cells. Such increased activation was reflected in their elevated IL-2 secretion. Exosome localization motif identification and quantification within HLA-DQA1 and HLA-DQB1 transcripts highlighted their significant prevalence within HLA-DQB1 alleles associated with CeD susceptibility. Flow cytometry revealed the strong correlation between HLA-DQ and the CD63 exosomal marker in T cells exposed to CGM from MoDCs sourced from CeD patients. This resulted in lower concentrations of CD25+ CD127- T cells, suggestive of their compromised induction to T-regulatory cells associated with CeD homeostasis. This foremost comparative study deciphered the genomic basis and extracellular exosomal effects of HLA transfer on T lymphocytes in the context of CeD, offering greater insight into this auto-immune disease.
- Klíčová slova
- autoimmunity, exosome, gluten, major histocompatibility complex II, monocyte-derived dendritic cells,
- MeSH
- alely MeSH
- celiakie * MeSH
- gluteny genetika MeSH
- HLA-DQ antigeny genetika MeSH
- lidé MeSH
- regulační T-lymfocyty MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- gluteny MeSH
- HLA-DQ antigeny MeSH
The autoimmune condition, Celiac Disease (CeD), displays broad clinical symptoms due to gluten exposure. Its genetic association with DQ variants in the human leukocyte antigen (HLA) system has been recognised. Monocyte-derived mature dendritic cells (MoDCs) present gluten peptides through HLA-DQ and co-stimulatory molecules to T lymphocytes, eliciting a cytokine-rich microenvironment. Having access to CeD associated families prevalent in the Czech Republic, this study utilised an in vitro model to investigate their differential monocyte profile. The higher monocyte yields isolated from PBMCs of CeD patients versus control individuals also reflected the greater proportion of dendritic cells derived from these sources following lipopolysaccharide (LPS)/ peptic-tryptic-gliadin (PTG) fragment stimulation. Cell surface markers of CeD monocytes and MoDCs were subsequently profiled. This foremost study identified a novel bio-profile characterised by elevated CD64 and reduced CD33 levels, unique to CD14++ monocytes of CeD patients. Normalisation to LPS stimulation revealed the increased sensitivity of CeD-MoDCs to PTG, as shown by CD86 and HLA-DQ flow cytometric readouts. Enhanced CD86 and HLA-DQ expression in CeD-MoDCs were revealed by confocal microscopy. Analysis highlighted their dominance at the CeD-MoDC membrane in comparison to controls, reflective of superior antigen presentation ability. In conclusion, this investigative study deciphered the monocytes and MoDCs of CeD patients with the identification of a novel bio-profile marker of potential diagnostic value for clinical interpretation. Herein, the characterisation of CD86 and HLA-DQ as activators to stimulants, along with robust membrane assembly reflective of efficient antigen presentation, offers CeD targeted therapeutic avenues worth further exploration.
- Klíčová slova
- CD33, CD64, CD86, MHCDQ, autoimmunity, major histocompatibility complex II, monocyte, monocyte-derived dendritic cells,
- MeSH
- autoimunitní nemoci imunologie MeSH
- biologické markery metabolismus MeSH
- buněčná membrána metabolismus MeSH
- CD antigeny metabolismus MeSH
- celiakie epidemiologie imunologie MeSH
- cytotoxické T-lymfocyty imunologie MeSH
- dendritické buňky imunologie MeSH
- dospělí MeSH
- gliadin škodlivé účinky MeSH
- HLA-DQ antigeny metabolismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- lipopolysacharidy MeSH
- mladiství MeSH
- mladý dospělý MeSH
- monocyty metabolismus MeSH
- náchylnost k nemoci MeSH
- prezentace antigenu imunologie MeSH
- rodina MeSH
- rodokmen MeSH
- senioři MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika epidemiologie MeSH
- Názvy látek
- biologické markery MeSH
- CD antigeny MeSH
- gliadin MeSH
- HLA-DQ antigeny MeSH
- lipopolysacharidy MeSH
BACKGROUND: CD16 was previously suggested to be a new marker of basophils that is subject to downregulation by FcεRI crosslinking. Certain compounds, including supraoptimal concentrations of the PKC inhibitors, bisindolylmaleimides, decouple the release of granules containing CD203c, CD63 and histamine, and may thus help to identify the mechanisms related to the CD16 externalization. OBJECTIVE: We hypothesized that CD16 is differentially expressed on the surface of basophils in patients with birch pollen or insect venom allergy and is subject to a regulation in response to allergens. We also employed CD203c and CD63 externalization decoupling by bisindolylmaleimides. METHODS: We performed a basophil activation test coupled with CD16 and histamine detection using cells isolated from patients with allergy to birch pollen or insect venom and negative controls. We employed two PKC inhibitors, bisindolylmaleimide II and Ro 31-8220 at their supraoptimal concentrations and, after difficulties reproducing previously published data, we analyzed the fluorescence of these inhibitors alone. We identified the CD16 isoforms by sequencing nested RT-PCR amplicons from flow cytometry sorted basophils and by cleaving the CD16b GPI anchor using a phospholipase C. RESULTS: We provide the first evidence that CD16a is expressed as a surface antigen on a small subpopulation of human basophils in patients with respiratory and insect venom allergy, and this antigen shows increased surface expression following allergen challenge or FcεRI crosslinking. We rejected the apparent decoupling of the surface expression of basophil activation markers following the administration of bisindolylmaleimides. CONCLUSIONS & CLINICAL RELEVANCE: The inclusion of αCD16 in negative selection cocktails selects against a subset of basophils that are CD16+ or CD16dim . Using CD16dim basophils and unstained leucocytes, we show that previous studies with supraoptimal concentrations of bisindolylmaleimides are likely flawed and are not associated with the differential expression of CD203c and CD63.
- Klíčová slova
- FcγRIIIA, IgG-mediated anaphylaxis, basophil activation test, bisindolylmaleimide, flow cytometry artifact,
- MeSH
- alergie imunologie patologie MeSH
- antigeny CD63 imunologie MeSH
- bazofily imunologie patologie MeSH
- dospělí MeSH
- fosfodiesterasy imunologie MeSH
- GPI-vázané proteiny imunologie MeSH
- indoly chemie MeSH
- jedy členovců toxicita MeSH
- kousnutí a bodnutí hmyzem imunologie patologie MeSH
- lidé středního věku MeSH
- lidé MeSH
- maleimidy chemie MeSH
- pyrofosfatasy imunologie MeSH
- receptory IgG imunologie MeSH
- senioři MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- antigeny CD63 MeSH
- bisindolylmaleimide MeSH Prohlížeč
- CD63 protein, human MeSH Prohlížeč
- ENPP3 protein, human MeSH Prohlížeč
- FCGR3B protein, human MeSH Prohlížeč
- fosfodiesterasy MeSH
- GPI-vázané proteiny MeSH
- indoly MeSH
- jedy členovců MeSH
- maleimidy MeSH
- pyrofosfatasy MeSH
- receptory IgG MeSH
Pimecrolimus (Elidel, SDZ ASM 981) is an anti-inflammatory and immunomodulatory 33-epichloro-derivative of macrolactam ascomycin, with low potential for affecting systemic immune responses compared with other calcineurin inhibitors, cyclosporin A and tacrolimus. Despite numerous studies focused on the mechanism of pimecrolimus action on mast cells, only the single report has addressed pimecrolimus effects on other typical FcεRI-expressing cells, the basophils. Patients allergic to birch pollen (n = 20), hymenopteran venoms (n = 23) and 10 non-allergic volunteers were examined. Primary human basophils pre-treated or not with 0.5-50 μMol pimecrolimus were exposed to various concentrations of recombinant Bet v 1a allergen, bee or wasp venom extracts and anti-IgE for 20 min, and then examined for the expression of CD45, CD193, CD203c, CD63 and CD164 using flow cytometry. The externalization of basophil activation markers (CD63 and CD164) was equally inhibited through pimecrolimus in cells activated by recombinant pollen allergen, hymenopteran venom extracts and anti-IgE. Although the individual response rate was subject to strong variation, importantly, pre-treatment with pimecrolimus lowered the number of activated basophils in response to any of the stimuli in the basophils from all patients. The inhibition was concentration-dependent; approximately half of the basophils were inhibited in the presence of 2.5 mMol pimecrolimus. Pimecrolimus is a valuable new tool for the inhibition of hyper-reactive basophils in patients with pollen allergy and a history of anaphylactic reactions to bee or wasp venoms. Further research should address short-term use of pimecrolimus in vivo in a wide spectrum of allergic diseases.
- MeSH
- alergeny imunologie MeSH
- alergie farmakoterapie imunologie MeSH
- antialergika farmakologie MeSH
- antiflogistika nesteroidní farmakologie MeSH
- bazofily účinky léků imunologie MeSH
- bříza imunologie MeSH
- dospělí MeSH
- imunoglobulin E imunologie MeSH
- kultivované buňky MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- protilátky anti-idiotypické imunologie MeSH
- receptory IgE imunologie MeSH
- senioři MeSH
- sezónní alergická rýma farmakoterapie imunologie MeSH
- sršňovití imunologie MeSH
- takrolimus analogy a deriváty farmakologie MeSH
- včely imunologie MeSH
- vosí jedy imunologie MeSH
- zvířata MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- senioři MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- alergeny MeSH
- anti-IgE antibodies MeSH Prohlížeč
- antialergika MeSH
- antiflogistika nesteroidní MeSH
- imunoglobulin E MeSH
- pimecrolimus MeSH Prohlížeč
- protilátky anti-idiotypické MeSH
- receptory IgE MeSH
- takrolimus MeSH
- vosí jedy MeSH
