The glycosphingolipid, α-galactosylceramide (αGalCer), when presented by CD1d on antigen-presenting cells, efficiently activates invariant natural killer T (iNKT) cells. Thereby, it modulates immune responses against tumors, microbial and viral infections, and autoimmune diseases. Recently, the production of αGalCer by Bacteroidetes from the human gut microbiome was elucidated. Using hydrophilic interaction chromatography coupled to MS2, we screened murine intestinal tracts to identify and quantify αGalCers, and we investigated the αGalCer response to different dietary and physiologic conditions. In both the cecum and the colon of mice, we found 1-15 pmol of αGalCer per milligram of protein; in contrast, mice lacking microbiota (germ-free mice) and fed identical diet did not harbor αGalCer. The identified αGalCer contained a β(R)-hydroxylated hexadecanoyl chain N-linked to C18-sphinganine, which differed from what has been reported with Bacteroides fragilis Unlike β-anomeric structures, but similar to αGalCers from B. fragilis, the synthetic form of the murine αGalCer induced iNKT cell activation in vitro. Last, we observed a decrease in αGalCer production in mice exposed to conditions that alter the composition of the gut microbiota, including Western type diet, colitis, and influenza A virus infection. Collectively, this study suggests that αGalCer is produced by commensals in the mouse intestine and reveals that stressful conditions causing dysbiosis alter its synthesis. The consequences of this altered production on iNKT cell-mediated local and systemic immune responses are worthy of future studies.

• Klíčová slova
• Western diet, bacteria, cluster of differentiation 1d, experimental colitis, glycolipids, immunology, influenza A virus, invariant natural killer T cells, mass spectrometry, sphingolipids,
• MeSH
• Bacteroides fragilis chemie   imunologie   MeSH
• dieta * MeSH
• galaktosylceramidy genetika   imunologie   MeSH
• inbrední kmeny myší MeSH
• myši MeSH
• tlusté střevo imunologie   metabolismus   mikrobiologie   MeSH
• zánět imunologie   mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alpha-galactosylceramide MeSH Prohlížeč
• galaktosylceramidy MeSH

Combined immunotherapy constitutes a novel, advanced strategy in cancer treatment. In this study, we investigated immunotherapy in the mouse TC-1/A9 model of human papillomavirus type 16 (HPV16)-associated tumors characterized by major histocompatibility complex class I (MHC-I) downregulation. We found that the induction of a significant anti-tumor response required a combination of DNA vaccination with the administration of an adjuvant, either the synthetic oligodeoxynucleotide ODN1826, carrying immunostimulatory CpG motifs, or α-galactosylceramide (α-GalCer). The most profound anti-tumor effect was achieved when these adjuvants were applied in a mix with a one-week delay relative to DNA immunization. Combined immunotherapy induced tumor infiltration with various subsets of immune cells contributing to tumor regression, of which cluster of differentiation (CD) 8⁺ T cells were the predominant subpopulation. In contrast, the numbers of tumor-associated macrophages (TAMs) were not markedly increased after immunotherapy but in vivo and in vitro results showed that they could be repolarized to an anti-tumor M1 phenotype. A blockade of T cell immunoglobulin and mucin-domain containing-3 (Tim-3) immune checkpoint had a negligible effect on anti-tumor immunity and TAMs repolarization. Our results demonstrate a benefit of combined immunotherapy comprising the activation of both adaptive and innate immunity in the treatment of tumors with reduced MHC-I expression.

• Klíčová slova
• CpG ODN, DNA immunization, MHC-I, cancer immunotherapy, tumor-associated macrophages, α-galactosylceramide,
• MeSH
• adjuvancia imunologická terapeutické užití   MeSH
• CD8-pozitivní T-lymfocyty imunologie   MeSH
• down regulace MeSH
• experimentální nádory terapie   MeSH
• galaktosylceramidy imunologie   MeSH
• imunoterapie metody   MeSH
• makrofágy imunologie   MeSH
• MHC antigeny I. třídy imunologie   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• oligodeoxyribonukleotidy imunologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• adjuvancia imunologická MeSH
• galaktosylceramidy MeSH
• MHC antigeny I. třídy MeSH
• oligodeoxyribonukleotidy MeSH

Humoral deficiencies represent a broad group of disorders. The aim of the study was to compare the levels of antibodies against pneumococcal capsular polysaccharides (anti-PCP) and natural anti-galactosyl (anti-Gal) antibodies in (1) patients with chronic lymphocytic leukaemia (CLL), (2) patients with common variable immunodeficiency (CVID), and (3) a healthy population and to explore their diagnostic and prognostic potential. Serum immunoglobulin levels and levels of anti-Gal IgG, IgA, and IgM and anti-PCP IgG and IgG2 were determined in 59 CLL patients, 30 CVID patients, and 67 healthy controls. Levels of IgG, IgA, IgM, anti-Gal IgA, anti-Gal IgM, and anti-PCP IgA were lower in CLL and CVID patients than in healthy controls (p value for all parameters < 0.0001). Decrease in the levels of IgA, IgM, anti-Gal IgA, and anti-PCP IgA was less pronounced in the CLL group than in the CVID group. IgA decline, anti-Gal IgA, anti-PCP IgA, and anti-PCP IgG2 were negatively correlated with CLL stage. We devise the evaluation of anti-Gal antibodies to be a routine test in humoral immunodeficiency diagnostics, even in cases of immunoglobulin substitution therapy. Significant reductions, mainly in anti-Gal IgA, IgM, and anti-PCP IgA levels, may have prognostic importance in CLL patients.

• MeSH
• autoprotilátky krev   MeSH
• bakteriální pouzdra imunologie   MeSH
• běžná variabilní imunodeficience diagnóza   imunologie   MeSH
• biologické markery krev   MeSH
• dospělí MeSH
• galaktosylceramidy imunologie   MeSH
• humorální imunita MeSH
• imunoglobulin A krev   MeSH
• imunoglobulin G krev   MeSH
• imunoglobulin M krev   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• pneumokokové infekce diagnóza   imunologie   MeSH
• prognóza MeSH
• protilátky bakteriální krev   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Streptococcus pneumoniae imunologie   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• autoprotilátky MeSH
• biologické markery MeSH
• galaktosylceramidy MeSH
• imunoglobulin A MeSH
• imunoglobulin G MeSH
• imunoglobulin M MeSH
• protilátky bakteriální MeSH

Ceramide (Cer) release from glucosylceramides (GlcCer) is critical for the formation of the skin permeability barrier. Changes in β-glucocerebrosidase (GlcCer'ase) activity lead to diminished Cer, GlcCer accumulation and structural defects in SC lipid lamellae; however, the molecular basis for this impairment is not clear. We investigated impaired GlcCer-to-Cer processing in human Cer membranes to determine the physicochemical properties responsible for the barrier defects. Minor impairment (5-25%) of the Cer generation from GlcCer decreased the permeability of the model membrane to four markers and altered the membrane microstructure (studied by X-ray powder diffraction and infrared spectroscopy), in agreement with the effects of topical GlcCer in human skin. At these concentrations, the accumulation of GlcCer was a stronger contributor to this disturbance than the lack of human Cer. However, replacement of 50-100% human Cer by GlcCer led to the formation of a new lamellar phase and the maintenance of a rather good barrier to the four studied permeability markers. These findings suggest that the major cause of the impaired water permeability barrier in complete GlcCer'ase deficiency is not the accumulation of free GlcCer but other factors, possibly the retention of GlcCer bound in the corneocyte lipid envelope.

• MeSH
• aplikace lokální MeSH
• ceramidy metabolismus   MeSH
• difrakce rentgenového záření MeSH
• elektrická impedance MeSH
• glukosylceramidy aplikace a dávkování   metabolismus   farmakologie   MeSH
• indomethacin farmakokinetika   MeSH
• kůže chemie   účinky léků   metabolismus   MeSH
• lidé MeSH
• membránové lipidy chemie   metabolismus   MeSH
• permeabilita buněčné membrány * účinky léků   MeSH
• permeabilita MeSH
• spektroskopie infračervená s Fourierovou transformací MeSH
• theofylin farmakokinetika   MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ceramidy MeSH
• glukosylceramidy MeSH
• indomethacin MeSH
• membránové lipidy MeSH
• theofylin MeSH

CD4(+)CD25(+)Foxp3(+) T regulatory cells (Tregs) and CD1d-restricted invariant natural killer T (iNKT) cells are two cell types that are known to regulate immune reactions. Depletion or inactivation of Tregs using specific anti-CD25 antibodies in combination with immunostimulation is an attractive modality especially in anti-tumour immunotherapy. However, CD25 is not expressed exclusively on Tregs but also on subpopulations of activated lymphocytes. Therefore, the modulatory effects of the specific anti-CD25 antibodies can also be partially attributed to their interactions with the effector cells. Here, the effector functions of iNKT cells were analysed in combination with anti-CD25 mAb PC61. Upon PC61 administration, α-galactosylceramide (α-GalCer)-mediated activation of iNKT cells resulted in decreased IFN-γ but not IL-4 production. In order to determine whether mutual interactions between Tregs and iNKT cells take place, we compared IFNγ production after α-GalCer administration in anti-CD25-treated and "depletion of regulatory T cell" (DEREG) mice. Since no profound effects on IFNγ induction were observed in DEREG mice, deficient in FoxP3(+) Tregs, our results indicate that the anti-CD25 antibody acts directly on CD25(+) effector cells. In vivo experiments demonstrated that although both α-GalCer and PC61 administration inhibited TC-1 tumour growth in mice, no additive/synergic effects were observed when these substances were used in combination therapy.

• MeSH
• antigeny CD1d imunologie   metabolismus   MeSH
• ELISA MeSH
• experimentální nádory farmakoterapie   imunologie   patologie   MeSH
• exprese genu účinky léků   imunologie   MeSH
• forkhead transkripční faktory imunologie   metabolismus   MeSH
• galaktosylceramidy aplikace a dávkování   imunologie   farmakologie   MeSH
• heparin-vázající růstový faktor podobný EGF MeSH
• interferon gama genetika   imunologie   metabolismus   MeSH
• interleukin-4 genetika   imunologie   metabolismus   MeSH
• Kaplanův-Meierův odhad MeSH
• mezibuněčné signální peptidy a proteiny genetika   imunologie   metabolismus   MeSH
• monoklonální protilátky aplikace a dávkování   imunologie   farmakologie   MeSH
• myši inbrední C57BL MeSH
• myši transgenní MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• NKT buňky účinky léků   imunologie   metabolismus   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• protokoly protinádorové kombinované chemoterapie terapeutické užití   MeSH
• průtoková cytometrie MeSH
• receptor interleukinu-2 - alfa-podjednotka imunologie   metabolismus   MeSH
• regulační T-lymfocyty účinky léků   imunologie   metabolismus   MeSH
• tumor burden účinky léků   imunologie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alpha-galactosylceramide MeSH Prohlížeč
• antigeny CD1d MeSH
• forkhead transkripční faktory MeSH
• Foxp3 protein, mouse MeSH Prohlížeč
• galaktosylceramidy MeSH
• Hbegf protein, mouse MeSH Prohlížeč
• heparin-vázající růstový faktor podobný EGF MeSH
• interferon gama MeSH
• interleukin-4 MeSH
• mezibuněčné signální peptidy a proteiny MeSH
• monoklonální protilátky MeSH
• PC61 monoclonal antibody MeSH Prohlížeč
• receptor interleukinu-2 - alfa-podjednotka MeSH

Sphingolipid ceramide N-deacylase (SCDase, EC 3.5.1.69) is a hydrolytic enzyme isolated from Pseudomonas sp. TK 4. In addition to its primary deacylation function, this enzyme is able to reacylate lyso-sphingolipids under specific conditions. We immobilised this enzyme on magnetic macroporous cellulose and used it to semisynthesise C17:0 glucosylceramide and C17:0 sulphatide, which are required internal standards for quantification of the corresponding glycosphingolipids (GSL) by tandem mass spectrometry. A high rate of conversion was achieved for both lipids (80% for C17:0 sulphatide and 90% for C17:0 glucosylceramide). In contrast to synthesis with a soluble form of the enzyme, use of immobilised SCDase significantly reduced the contamination of the sphingolipid products with other isoforms, so further purification was not necessary. Our method can be effectively used for the simple preparation of specifically labelled sphingolipids of high isoform purity for application in mass spectrometry.

• MeSH
• amidohydrolasy chemie   MeSH
• bakteriální proteiny chemie   MeSH
• enzymy imobilizované chemie   MeSH
• glukosylceramidy chemická syntéza   chemie   MeSH
• glykosfingolipidy analýza   MeSH
• hmotnostní spektrometrie metody   normy   MeSH
• hydrolýza MeSH
• Pseudomonas enzymologie   MeSH
• referenční standardy MeSH
• stereoizomerie MeSH
• sulfoglykosfingolipidy chemická syntéza   chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• amidohydrolasy MeSH
• bakteriální proteiny MeSH
• enzymy imobilizované MeSH
• glukosylceramidy MeSH
• glykosfingolipidy MeSH
• sphingolipid ceramide N-deacylase MeSH Prohlížeč
• sulfoglykosfingolipidy MeSH

Gaucher disease (GD), deficiency of acid glucosylceramidase (GlcCer-ase) is characterized by deficient degradation of beta-glucosylceramide (GlcCer). It is well known that, in GD, the lysosomal accumulation of uncleaved GlcCer is limited to macrophages, which are gradually converted to storage cells with well known cytology--Gaucher cells (GCs). On the basis of previous studies of the disorder and of a comparison with other lysosomal enzymopathies affecting degradation of the GlcCer-based glycosphingolipid series, it is hypothesized that in other cell types (i.e. non-macrophage cells) the uncleaved GlcCer, in GlcCer-ase deficiency, is transferred to other cell compartments, where it may be processed and even accumulated to various degrees. The consequence of the abnormal extralysosomal load may differ according to the cell type and compartment targeted and may be influenced by genetically determined factors, by a number of acquired conditions, including the current metabolic situation. The sequelae of the uncleaved GlcCer extralysosomal transfer may range from probably innocent or positive stimulatory, to the much more serious, in which it interferes with a variety of cell functions, and in extreme cases, can lead to cell death. This alternative processing of uncleaved GlcCer may help to explain tissue alterations seen in GD that have, so far, resisted explanation based simply on the presence of GCs. Paralysosomal alternative processing may thus go a long way towards filling a long-standing gap in the understanding of the molecular pathology of the disorder. The impact of this alternative process will most likely be inversely proportional to the level of residual GlcCer-ase activity. Lysosomal sequestration of GlcCer in these cells is either absent or in those exceptional cases where it does occur, it is exceptional and rudimentary. It is suggested that paralysosomal alternative processing of uncleaved GlcCer is the main target for enzyme replacement therapy. The mechanism responsible for GlcCer transfer remains to be elucidated. It may also help in explaining the so far unclear origin of glucosylsphingosine (GlcSph) and define the mutual relation between these two processes.

• MeSH
• biologické modely MeSH
• biologický transport MeSH
• glukosylceramidasa nedostatek   terapeutické užití   MeSH
• glukosylceramidy metabolismus   MeSH
• lidé MeSH
• lyzozomy chemie   metabolismus   MeSH
• makrofágy metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• glukosylceramidasa MeSH
• glukosylceramidy MeSH

• MeSH
• aorta cytologie   metabolismus   patologie   MeSH
• cerebrosidy metabolismus   MeSH
• cholesterol dietní metabolismus   MeSH
• dieta aterogenní MeSH
• histocytochemie MeSH
• koronární cévy cytologie   metabolismus   patologie   MeSH
• metabolismus lipidů MeSH
• morčata MeSH
• sfingomyeliny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• morčata MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cerebrosidy MeSH
• cholesterol dietní MeSH
• sfingomyeliny MeSH

• MeSH
• cerebrosidy biosyntéza   MeSH
• Gaucherova nemoc metabolismus   MeSH
• lidé MeSH
• lipidózy metabolismus   MeSH
• metabolismus lipidů * MeSH
• nervový systém metabolismus   MeSH
• Niemannova-Pickova nemoc metabolismus   MeSH
• sfingolipidy biosyntéza   MeSH
• sfingomyeliny biosyntéza   MeSH
• xantomatóza metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cerebrosidy MeSH
• sfingolipidy MeSH
• sfingomyeliny MeSH

• MeSH
• antigeny MeSH
• časové faktory MeSH
• cerebrosidy metabolismus   MeSH
• králíci MeSH
• metabolismus lipidů MeSH
• modely nemocí na zvířatech MeSH
• poranění mozku imunologie   MeSH
• tvorba protilátek MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antigeny MeSH
• cerebrosidy MeSH