The presence or absence of autoantibodies against citrullinated proteins (ACPAs) distinguishes two main groups of rheumatoid arthritis (RA) patients with different etiologies, prognoses, disease severities, and, presumably, disease pathogenesis. The heterogeneous responses of RA patients to various biologics, even among ACPA-positive patients, emphasize the need for further stratification of the patients. We used high-density protein array technology for fingerprinting of ACPA reactivity. Identification of the proteome recognized by ACPAs may be a step to stratify RA patients according to immune reactivity. Pooled plasma samples from 10 anti-CCP-negative and 15 anti-CCP-positive RA patients were assessed for ACPA content using a modified protein microarray containing 1631 different natively folded proteins citrullinated in situ by protein arginine deiminases (PADs) 2 and PAD4. IgG antibodies from anti-CCP-positive RA plasma showed high-intensity binding to 87 proteins citrullinated by PAD2 and 99 proteins citrullinated by PAD4 without binding significantly to the corresponding native proteins. Curiously, the binding of IgG antibodies in anti-CCP-negative plasma was also enhanced by PAD2- and PAD4-mediated citrullination of 29 and 26 proteins, respectively. For only four proteins, significantly more ACPA binding occurred after citrullination with PAD2 compared to citrullination with PAD4, while the opposite was true for one protein. We demonstrate that PAD2 and PAD4 are equally efficient in generating citrullinated autoantigens recognized by ACPAs. Patterns of proteins recognized by ACPAs may serve as a future diagnostic tool for further subtyping of RA patients.

• MeSH
• autoantigeny krev   imunologie   MeSH
• biologické markery krev   MeSH
• čipová analýza proteinů MeSH
• citrulin metabolismus   MeSH
• citrulinace MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• peptidylarginindeiminasa typu 2 metabolismus   MeSH
• peptidylarginindeiminasa typu 4 metabolismus   MeSH
• protilátky proti citrulinovaným peptidům imunologie   MeSH
• revmatoidní artritida imunologie   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• tandemová hmotnostní spektrometrie MeSH
• vysokoúčinná kapalinová chromatografie MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Omics-based methods are increasingly used in current ecotoxicology. Therefore, a large number of observations for various toxic substances and organisms are available and may be used for identifying modes of action, adverse outcome pathways, or novel biomarkers. For these purposes, good statistical analysis of toxicogenomic data is vital. In contrast to established ecotoxicological techniques, concentration-response modeling is rarely used for large datasets. Instead, statistical hypothesis testing is prevalent, which provides only a limited scope for inference. The present study therefore applied automated concentration-response modeling for 3 different ecotoxicotranscriptomic and ecotoxicometabolomic datasets. The modeling process was performed by simultaneously applying 9 different regression models, representing distinct mechanistic, toxicological, and statistical ideas that result in different curve shapes. The best-fitting models were selected by using Akaike's information criterion. The linear and exponential models represented the best data description for more than 50% of responses. Models generating U-shaped curves were frequently selected for transcriptomic signals (30%), and sigmoid models were identified as best fit for many metabolomic signals (21%). Thus, selecting the models from an array of different types seems appropriate, because concentration-response functions may vary because of the observed response type, and they also depend on the compound, the organism, and the investigated concentration and exposure duration range. The application of concentration-response models can help to further tap the potential of omics data and is a necessary step for quantitative mixture effect assessment at the molecular response level.

• MeSH
• dánio pruhované růst a vývoj   metabolismus   MeSH
• ekosystém * MeSH
• embryo nesavčí účinky léků   metabolismus   MeSH
• genomika * MeSH
• látky znečišťující životní prostředí toxicita   MeSH
• lineární modely MeSH
• metabolomika * MeSH
• rychlé screeningové testy MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• tetrachlorethylen toxicita   MeSH
• transkriptom účinky léků   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Interleukin 27 (IL-27), a member of the IL-12 family, is important for T cell differentiation; however, little is known about its effect on dendritic cells (DCs). IL-27 can activate multiple signaling cascades, including the JAK/STAT pathway, and depending on the setting it can both promote and antagonize inflammatory responses. An anti-inflammatory function of IL-27 has been reported in several autoimmune diseases; however, in type 1 diabetes (T1D), an autoimmune disease where autoreactive cytotoxic T cells attack insulin-producing beta cells, IL-27 has been shown to have a dual role and contradictory effects. Here, we show impaired IL-27 signaling in a large cohort of T1D patients (n = 51) compared to age- and gender-matched healthy donors. Increased expression of the IL-27 receptor subunit IL-27Ralpha mRNA in purified myeloid DCs (mDCs), detected by gene expression microarrays was mirrored by enhanced signal transduction in T1D mDCs in response to IL-27 stimulation. Higher STAT phosphorylation in T1D patients was also accompanied by elevated expression of the inhibitory molecules PD-L1, PD-L2 and PD-1, which may suggest not only immunomodulatory mechanisms of IL-27 in T1D but also a compensatory effort of T1D dendritic cells against the ongoing inflammation.

• MeSH
• antigeny CD274 genetika   metabolismus   MeSH
• dendritické buňky imunologie   MeSH
• diabetes mellitus 1. typu genetika   imunologie   metabolismus   MeSH
• dospělí MeSH
• fosforylace MeSH
• interleukiny genetika   metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• receptory interleukinů genetika   metabolismus   MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• senioři MeSH
• signální transdukce * MeSH
• stanovení celkové genové exprese metody   MeSH
• studie případů a kontrol MeSH
• transkripční faktor STAT3 metabolismus   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

Perfect maternal diabetes compensation is crucial for the outcome of the baby. However, little is known how hyperglycaemia influences the specific immune response. Furthermore, babies of type 1 diabetes (T1D) mothers have less risk of development T1D than babies with a T1D father. This study aimed to analyze the effect of maternal hyperglycaemia on newborns with focus on the response to diabetes-associated autoantigens. Populations: (1) Newborns of T1D mothers split into groups according to maternal diabetes compensation during the 3rd trimester: perfect (n = 15) or acceptable (n = 25) compensation. (2) newborns with T1D father (n = 12) (3) newborns with a mother treated for either gestational or type 2 diabetes (n = 10) (4) control newborns (n = 25). Spontaneous as well as diabetes-associated autoantigen-stimulated production of 23 cytokines and chemokines were tested using protein microarray. In addition, the influence of glucose on cytokine and chemokine responsiveness was analyzed in vitro. The study groups differed in their spontaneous as well as stimulated cytokine and chemokine spectra. A prominent Th1 response (high IFN-gamma) from autoantigen stimulation was observed especially in babies of T1D fathers (P = 0.001) and also in mothers with perfect diabetes compensation during the 3rd trimester (P = 0.016) in comparison with control newborns. By contrast, cord blood mononuclear cells cultivated in vitro in high glucose concentration decreased the diabetogenic stimulated Th1 cytokine response. Maternal 'sweet' as well as 'autoimmune environment' may both lead to lower occurrence of T1D within their offspring. Further studies will reveal the exact immunological mechanism of this observation.

• MeSH
• autoantigeny imunologie   MeSH
• čipová analýza proteinů MeSH
• cytokiny biosyntéza   metabolismus   účinky léků   MeSH
• diabetes mellitus 1. typu imunologie   MeSH
• dospělí MeSH
• fetální krev imunologie   metabolismus   MeSH
• glukosa farmakologie   MeSH
• glutamát dekarboxyláza farmakologie   MeSH
• hyperglykemie imunologie   MeSH
• leukocyty mononukleární imunologie   účinky léků   MeSH
• lidé MeSH
• novorozenec MeSH
• těhotenství při diabetu imunologie   MeSH
• těhotenství MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• novorozenec MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• práce podpořená grantem MeSH

BACKGROUND: Autoreactive T cells have a crucial role in type 1 diabetes (T1D) pathogenesis. OBJECTIVES: The aim of our study was to monitor the in vitro production of cytokines by peripheral blood mononuclear cells (PBMCs) after stimulation with diabetogenic autoantigens. Subjects: Ten T1D patients (tested at the time of diagnosis and 6 and 12 months later), 10 first-degree relatives of the T1D patients, and 10 controls underwent the study. METHODS: PBMCs were stimulated with glutamic acid decarboxylase 65 (GAD65) amino acids (a.a.) 247-279, 509-528, and 524-543; proinsulin a.a. 9-23; and tyrosine phosphatase (islet antigen-2)/R2 a.a. 853-872. Interleukin (IL)-2, IL-4, IL-5, IL-6, IL-10, IL-13, interferon (IFN)-gamma, tumor necrosis factor beta, transforming growth factor beta1, and granulocyte colony-stimulating factor (GCSF) were analyzed by protein microarray. RESULTS: Differences in cytokine(s) poststimulatory and mainly in basal production were observed in all groups. The most prominent findings were in controls, the higher basal levels of IL-2, IL-4, IL-5, IL-13, and GCSF were observed when compared with relatives (p < 0.05, for all). After stimulation in controls, there was a significant decrease in IL-2, IL-13, GCSF, and IFN-gamma (p < 0.05, for all). The group of relatives was the most variable in poststimulatory production. A strong correlation between cytokines production was found but groups differed in this aspect. CONCLUSION: By multiplex analysis, it may be possible, for example, to define the risk immunological response pattern among relatives or to monitor the immune response in patients on immune modulation therapy.

• MeSH
• autoprotilátky genetika   MeSH
• čipová analýza proteinů MeSH
• cytokiny chemická syntéza   MeSH
• diabetes mellitus 1. typu genetika   imunologie   MeSH
• dítě MeSH
• financování organizované MeSH
• glutamát dekarboxyláza genetika   chemie   MeSH
• interleukiny genetika   MeSH
• lidé MeSH
• mladiství MeSH
• molekulární sekvence - údaje MeSH
• předškolní dítě MeSH
• rodina MeSH
• sekvence aminokyselin MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH

Type 1 diabetes (T1D) is an autoimmune disease characterized by the lack of insulin due to an autoimmune destruction of pancreatic beta cells. Here, we report a unique case of a family with naturally conceived quadruplets in which T1D was diagnosed in two quadruplets simultaneously. At the same time, the third quadruplet was diagnosed with the pre-diabetic stage. Remarkably, all four quadruplets were positive for anti-islet cell antibodies, GAD65 and IA-A2. Monozygotic status of the quadruplets was confirmed by testing 14 different short tandem repeat polymorphisms. Serological examination confirmed that all quadruplets and their father suffered from a recent enteroviral infection of EV68-71 serotype. To assess the nature of the molecular pathological processes contributing to the development of diabetes, immunocompetent cells isolated from all family members were characterized by gene expression arrays, immune-cell enumerations and cytokine-production assays. The microarray data provided evidence that viral infection, and IL-27 and IL-9 cytokine signalling contributed to the onset of T1D in two of the quadruplets. The propensity of stimulated immunocompetent cells from non-diabetic members of the family to secrete high level of IFN-α further corroborates this conclusion. The number of T regulatory cells as well as plasmacytoid and/or myeloid dendritic cells was found diminished in all family members. Thus, this unique family is a prime example for the support of the so-called 'fertile-field' hypothesis proposing that genetic predisposition to anti-islet autoimmunity is 'fertilized' and precipitated by a viral infection leading to a fully blown T1D.

• MeSH
• autoimunita MeSH
• čtyřčata MeSH
• diabetes mellitus 1. typu genetika   imunologie   MeSH
• lidé MeSH
• předškolní dítě MeSH
• regulační T-lymfocyty imunologie   metabolismus   MeSH
• Check Tag
• lidé MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH

Východiska: Akutní lymfoblastická leukémie je nejčastěji diagnostikovanou malignitou dětských pacientů v české republice. V posledních desetiletích byl učiněn nebývalý pokrok v její léčbě, k němuž bezesporu napomohla i řada poznatků týkající se molekulárních charakteristik onemocnění. Analýza expresních profilů pomocí „Arrays“ (čipů) se stala jednou z významných metod při identifikaci potenciálních markerů onemocnění, které by umožnily bližší charakterizaci nádorových buněk a přispěly k porozumění procesu vzniku leukémie i stratifikace léčby. Metody a výsledky:V rámci studie byl sledován vliv methotrexátu na expresi genů p53-signální dráhy u maligních blastů izolovaných z kostní dřeně dětských pacientů s akutní leukémií. Změny exprese genů byly detekovány pomocí „GEArray Q series Human p53 Signalling Pathway Gene Array“. Porovnáním získaných expresních profilů jednotlivých pacientů byly pozorovány odlišné hladiny exprese sledovaných genů u kontrolních buněk, taktéž změny transkripční aktivity vlivem methotrexátu byly různorodé. Kromě jiných došlo vlivem methotrexátu k výrazným změnám transkripce genů uplatňující se při apoptóze či regulaci buněčného cyklu. Změny exprese některých genů byly nalezeny u více pacientů, řada ostatních se však projevila individuálně, nezávisle na typu onemocnění. Závěry: Byly detekovány změny v expresi řady genů p53-signální dráhy u maligních blastů kultivovaných ex vivo s methotrexátem. Odlišné hladiny exprese genů potvrzují značnou heterogenitu dětských leukémií, projevující se v našem souboru v expresních profilech a ve způsobu buněčné odpovědi na přítomnost methotrexátu, jejichž důsledkem může být různá míra citlivosti k aplikované látce a také různá odpověď pacienta na terapii.

Backgrounds: Acute lymphoblastic leukemia is the most frequent malignancy diagnosed in children in the Czech Republic. During last few decades, considerable progress in the treatment had been made and it was similarly contributed by many findings on the molecular basis of the disease. Expression profiling analysis using Macro or Micro Arrays has become an important technique to identify potential markers of the disorder, which might characterise malignant cells and help in the understanding of development of leukaemia and treatment stratification. Methods and Results: In the present study, the effect of methotrexate on the expression of p53-signalling pathway genes was investigated in malignant blasts isolated from the bone marrow of patients with acute leukemia. Expression variations were detected using „GEArray Q series Human p53 Signalling Pathway Gene Array“. Having drawn a comparison between expression profiles, variable gene expression levels in the control cells; likewise various transcription activity alterations were observed. Significant changes in the transcription were found among genes involved in the of apoptosis or cell cycle regulation. Regarding some genes, changes in expression were observed in more that one patient. However, the expression levels of most other genes varied individually, independently on the subtype of the disease. Conclusions: Changes in the expression of p53-signalling pathway genes were detected in the malignant blasts cultivated ex vivo with methotrexate. Different levels of the transcription in our series confirm heterogeneity of childhood leukemias, where patients with the same diagnosis do not need to share identical gene expression profiles even the manner of cellular response on the presence of methotrexate, resulting in the various level of perceptiveness to the medication and in the final response to the therapy.

• MeSH
• C-DNA MeSH
• exprese genu genetika   účinky léků   MeSH
• financování organizované MeSH
• geny p53 genetika   imunologie   účinky léků   MeSH
• hematologické nádory diagnóza   etiologie   genetika   MeSH
• hodnocení léčiv MeSH
• kostní dřeň MeSH
• lékařská onkologie metody   trendy   MeSH
• lékové transportní systémy metody   využití   MeSH
• leukemie diagnóza   etiologie   genetika   MeSH
• lidé MeSH
• methotrexát aplikace a dávkování   terapeutické užití   MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů metody   využití   MeSH
• Check Tag
• lidé MeSH

Rhodococci are bacteria which can survive under various extreme conditions, in the presence of toxic compounds, and in other hostile habitats. Their tolerance of unfavorable conditions is associated with the structure of their cell wall and their large array of enzymes, which degrade or detoxify harmful compounds. Their physiological and biotechnological properties, together with tools for their genetic manipulation, enable us to apply them in biotransformations, biodegradation and bioremediation. Many such biotechnological applications cause stresses that positively or negatively affect their efficiency. Whereas numerous reviews on rhodococci described their enzyme activities, the optimization of degradation or production processes, and corresponding technological solutions, only a few reviews discussed some specific effects of stresses on the physiology of rhodococci and biotechnological processes. This review aims to comprehensively describe individual stress responses in Rhodococcus strains, the interconnection of different types of stresses and their consequences for cell physiology. We examine here the responses to (1) environmental stresses (desiccation, heat, cold, osmotic and pH stress), (2) the presence of stress-inducing compounds (metals, organic compounds and antibiotics) in the environment (3) starvation and (4) stresses encountered during biotechnological applications. Adaptations of the cell envelope, the formation of multicellular structures and stresses induced by the interactions of hosts with pathogenic rhodococci are also included. The roles of sigma factors of RNA polymerase in the global regulation of stress responses in rhodococci are described as well. Although the review covers a large number of stressful conditions, our intention was to provide an overview of the selected stress responses and their possible connection to biotechnological processes, not an exhaustive survey of the scientific literature. The findings on stress responses summarized in this review and the demonstration of gaps in current knowledge may motivate researchers working to fill these gaps.

• MeSH
• biodegradace MeSH
• biotechnologie MeSH
• biotransformace MeSH
• fyziologický stres MeSH
• Rhodococcus * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

BACKGROUND: The genome of Pseudomonas aeruginosa contains at least three genes encoding eukaryotic-type Ser/Thr protein kinases, one of which, ppkA, has been implicated in P. aeruginosa virulence. Together with the adjacent pppA phosphatase gene, they belong to the type VI secretion system (H1-T6SS) locus, which is important for bacterial pathogenesis. To determine the biological function of this protein pair, we prepared a pppA-ppkA double mutant and characterised its phenotype and transcriptomic profiles. RESULTS: Phenotypic studies revealed that the mutant grew slower than the wild-type strain in minimal media and exhibited reduced secretion of pyoverdine. In addition, the mutant had altered sensitivity to oxidative and hyperosmotic stress conditions. Consequently, mutant cells had an impaired ability to survive in murine macrophages and an attenuated virulence in the plant model of infection. Whole-genome transcriptome analysis revealed that pppA-ppkA deletion affects the expression of oxidative stress-responsive genes, stationary phase σ-factor RpoS-regulated genes, and quorum-sensing regulons. The transcriptome of the pppA-ppkA mutant was also analysed under conditions of oxidative stress and showed an impaired response to the stress, manifested by a weaker induction of stress adaptation genes as well as the genes of the SOS regulon. In addition, expression of either RpoS-regulated genes or quorum-sensing-dependent genes was also affected. Complementation analysis confirmed that the transcription levels of the differentially expressed genes were specifically restored when the pppA and ppkA genes were expressed ectopically. CONCLUSIONS: Our results suggest that in addition to its crucial role in controlling the activity of P. aeruginosa H1-T6SS at the post-translational level, the PppA-PpkA pair also affects the transcription of stress-responsive genes. Based on these data, it is likely that the reduced virulence of the mutant strain results from an impaired ability to survive in the host due to the limited response to stress conditions.

• MeSH
• bakteriální proteiny genetika   MeSH
• bakteriální RNA genetika   MeSH
• buněčné linie MeSH
• delece genu MeSH
• fenotyp MeSH
• makrofágy mikrobiologie   MeSH
• mikrobiální viabilita MeSH
• myši MeSH
• oligopeptidy biosyntéza   MeSH
• oxidační stres MeSH
• protein-serin-threoninkinasy genetika   MeSH
• Pseudomonas aeruginosa genetika   růst a vývoj   patogenita   MeSH
• regulace genové exprese u bakterií MeSH
• salát (hlávkový) mikrobiologie   MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• testy genetické komplementace MeSH
• transkriptom MeSH
• virulence MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Signal transduction pathways in both prokaryotes and eukaryotes utilize protein phosphorylation as a key regulatory mechanism. Recent studies have proven that eukaryotic-type serine/threonine protein kinases (Hank's type) are widespread in many bacteria, although little is known regarding the cellular processes they control. In this study, we have attempted to establish the role of a single eukaryotic-type protein kinase, StkP of Streptococcus pneumoniae, in bacterial survival. Our results indicate that the expression of StkP is important for the resistance of S. pneumoniae to various stress conditions. To investigate the impact of StkP on this phenotype, we compared the whole-genome expression profiles of the wild-type and DeltastkP mutant strains by microarray technology. This analysis revealed that StkP positively controls the transcription of a set of genes encoding functions involved in cell wall metabolism, pyrimidine biosynthesis, DNA repair, iron uptake, and oxidative stress response. Despite the reduced transformability of the stkP mutant, we found that the competence regulon was derepressed in the stkP mutant under conditions that normally repress natural competence development. Furthermore, the competence regulon was expressed independently of exogenous competence-stimulating peptide. In summary, our studies show that a eukaryotic-type serine/threonine protein kinase functions as a global regulator of gene expression in S. pneumoniae.

• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• delece genu MeSH
• eukaryotické buňky enzymologie   MeSH
• fenotyp MeSH
• financování organizované MeSH
• koncentrace vodíkových iontů MeSH
• mikrobiální viabilita genetika   účinky léků   MeSH
• mutace MeSH
• osmotický tlak MeSH
• oxidační stres MeSH
• peroxid vodíku farmakologie   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• protein-serin-threoninkinasy genetika   metabolismus   MeSH
• regulace genové exprese u bakterií MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• Streptococcus pneumoniae enzymologie   genetika   růst a vývoj   MeSH
• testy genetické komplementace MeSH
• vysoká teplota MeSH