Citrulline is a non-proteinogenic amino acid that forms as by-product in nitric oxide (NO) synthesis from arginine and may act in concert with NO as an independent signaling molecule that involves in the mechanism of vascular smooth muscle vasodilation. In this study we examined the effects of citrulline on pulmonary artery smooth muscles. Experimental design comprised outward potassium currents measurements in enzymatically isolated rat pulmonary artery smooth muscle (PASMc) cells using whole-cell patch clamp technique, isometric contractile force recordings on rat pulmonary artery rings and method of molecular docking simulation. Citrulline in a concentration 10-9-10-5 M relaxed phenylephrine (PHE)-preactivated SM of rat pulmonary artery in a dose-dependent manner (EC50 0,67 μM). This citrulline-induced relaxation was dependent on an intact endothelium. Bath application of citrulline (10-8-10-5 M) on isolated PASMc induced a significant increase in the amplitude of outward potassium current (Ik). The adenosine antagonist caffeine (10-6 M) effectively blocked both the citrulline-induced relaxation response and Ik increment. Molecular docking modeling suggests that caffeine blocking the potent activity of citrulline results from competitive interactions at the A2 adenosine receptor binding site. In summary, our data suggest that citrulline, released with NO at low concentrations, can effectively interact with adenosine receptors in smooth muscle cells, causing their relaxation, indicating surprising interaction between NO and adenosine pathways.
- MeSH
- arteria pulmonalis účinky léků metabolismus MeSH
- citrulin * farmakologie metabolismus MeSH
- krysa rodu rattus MeSH
- potkani Wistar MeSH
- purinergní receptory P1 metabolismus MeSH
- simulace molekulového dockingu * MeSH
- svaly hladké cévní metabolismus účinky léků MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
INTRODUCTION: To date, there is not generally accepted and universal indicator of activity, and functional integrity of the small intestine in patients with coeliac disease. The aim of our study was to investigate whether serum concentrations of the non-essential amino acids citrulline and ornithine might have this function. METHODS: We examined serum citrulline and ornithine concentrations in a subgroup of patients with proven coeliac disease and healthy controls (blood donors). RESULTS: A total of 94 patients with coeliac disease (29 men, mean age 53 ± 18 years; 65 women, mean age 44 ± 14 years) and 35 healthy controls (blood donors) in whom coeliac disease was serologically excluded (10 men, mean age 51 ± 14 years; 25 women, mean age 46 ± 12 years) were included in the study. Significantly lower concentrations of serum ornithine were found in patients with coeliac disease (mean 65 ± 3 μmol/L; median 63 μmol/L, IQR 34 μmol/L, p < 0.001). No statistically nor clinically significant differences were found in the citrulline concentrations between the study and control group. CONCLUSIONS: Serum ornithine (but not citrulline) may be useful for assessing the functional status of the small intestine in uncomplicated coeliac disease. Further studies involving more detailed analysis of dietary and metabolic changes in patients will be needed to reach definitive conclusions.
- MeSH
- celiakie * MeSH
- citrulin * metabolismus MeSH
- dieta MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- ornithin metabolismus 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
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
Citrullination is a post-translational protein modification, which is associated with inflammation in general and is thought to play an important pathogenic role in rheumatoid arthritis (RA). Here a mass spectrometry-based proteomics approach was applied to identify citrullination sites in synovial fluid fibrinogen from four RA patients. In general, high disease activity correlated with increased number of identified citrullination sites and higher relative citrulline occupancy. Altogether, 23 sites were identified, of which 9 have not been previously reported to be citrullinated in vivo. Citrullination at site α84, α123, α129, α547, α573, α591, β334 and γ134 was identified in more than one patient, and these positions were therefore regarded as hotspots. Following citrullination of fibrinogen in vitro using human recombinant peptidylarginine deiminase 2 (PAD2), a total of 46 citrullination sites were identified, including 6 hitherto unreported in vitro citrullination sites. Twenty-two out of the 23 citrullination sites identified in vivo were also detected in vitro, supporting the validity of the identifications. SIGNIFICANCE: This work provides information about previously uncharacterized citrullination sites in synovial fluid fibrinogen from rheumatoid arthritis patients. Detection of these novel citrullination sites may prove to have diagnostic or prognostic value in RA and enhance our understanding of the immune pathogenesis.
- MeSH
- citrulin metabolismus MeSH
- fibrinogen metabolismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- peptidylarginindeiminasa typu 2 MeSH
- posttranslační úpravy proteinů * MeSH
- proteomika MeSH
- revmatoidní artritida metabolismus patologie MeSH
- senioři MeSH
- synoviální membrána metabolismus patologie MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
LL-37, the only human cathelicidin that is released during inflammation, is a potent regulator of immune responses by facilitating delivery of oligonucleotides to intracellular TLR-9, thereby enhancing the response of human plasmacytoid dendritic cells (pDCs) to extracellular DNA. Although important for pathogen recognition, this mechanism may facilitate development of autoimmune diseases. In this article, we show that citrullination of LL-37 by peptidyl-arginine deiminases (PADs) hindered peptide-dependent DNA uptake and sensing by pDCs. In contrast, carbamylation of the peptide (homocitrullination of Lys residues) had no effect. The efficiency of LL-37 binding to oligonucleotides and activation of pDCs was found to be inversely proportional to the number of citrullinated residues in the peptide. Similarly, preincubation of carbamylated LL-37 with PAD2 abrogated the peptide's ability to bind DNA. Conversely, LL-37 with Arg residues substituted by homoarginine, which cannot be deiminated, elicited full activity of native LL-37 regardless of PAD2 treatment. Taken together, the data showed that citrullination abolished LL-37 ability to bind DNA and altered the immunomodulatory function of the peptide. Both activities were dependent on the proper distribution of guanidinium side chains in the native peptide sequence. Moreover, our data suggest that cathelicidin/LL-37 is citrullinated by PADs during NET formation, thus affecting the inflammatory potential of NETs. Together this may represent a novel mechanism for preventing the breakdown of immunotolerance, which is dependent on the response of APCs to self-molecules (including cell-free DNA); overactivation may facilitate development of autoimmunity.
- MeSH
- autoimunita imunologie MeSH
- biologický transport MeSH
- buněčné linie MeSH
- citrulin metabolismus MeSH
- citrulinace fyziologie MeSH
- dendritické buňky imunologie MeSH
- DNA imunologie metabolismus MeSH
- imunologická tolerance imunologie MeSH
- kationické antimikrobiální peptidy metabolismus MeSH
- lidé MeSH
- myši MeSH
- peptidylarginindeiminasy metabolismus MeSH
- RAW 264.7 buňky MeSH
- volné cirkulující nukleové kyseliny imunologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
- MeSH
- adalimumab farmakologie MeSH
- antirevmatika farmakologie MeSH
- citrulin * imunologie metabolismus účinky léků MeSH
- cyklické peptidy * imunologie metabolismus účinky léků MeSH
- indukce remise MeSH
- klinické zkoušky, fáze III jako téma MeSH
- lidé MeSH
- multicentrické studie jako téma MeSH
- randomizované kontrolované studie jako téma MeSH
- revmatoidní artritida * farmakoterapie MeSH
- statistika jako téma MeSH
- Check Tag
- lidé MeSH
Přehledový článek shrnuje specifické metody umělé výživy v oblasti rozvoje intenzivní péče. Tato oblast je do určité míry specifická a indikace farmakonutrientů odlišná v klasické umělé výživě. Jsou popsány farmakonutrienty ze skupiny aminokyselin a polyenových mastných kyselin. Komponenty nutriční farmakologie, které jsou založeny na mimořádně vysokých dávkách čistých nutričních substrátů, jsou však užitečnou a spolehlivou složkou v ovlivnění některých mechanismů, kterými jsou např. fluidokoagulační rovnováha, inflamatorní reakce, vazomotorika. Klíčová slova: farmakonutrice – inflamatorní mediátory – polyenové mastné kyseliny – citrulin – arginin – střevní bariéra
The review article covers specific methods of artificial nutrition in current advances in intensive care. This area of care is somewhat specific, and indications for pharmaconutrients are different from classical artificial nutrition. The pharmaconutrients of amino acid and polyenoic fatty acid groups are described. The components of nutritional pharmacology, based on exceedingly high doses of pure nutritional substrates, are a useful and safe means of modifying selected mechanisms, such as fluidocoagulation, inflammatory reactions or vasomotorics. Keywords: pharmaconutrition – inflammatory mediators – polyunsaturated fatty acids – citruline – arginine – bowel barrier
- Klíčová slova
- farmakonutrice, inflamatorní mediátory, farmakonutrient, nutriční substrát, nutriční farmakologie, orgánově specifická výživa,
- MeSH
- aminokyseliny * terapeutické užití MeSH
- arginin metabolismus nedostatek terapeutické užití MeSH
- citrulin metabolismus nedostatek terapeutické užití MeSH
- energetický metabolismus účinky léků MeSH
- fyziologie výživy MeSH
- glutamin terapeutické užití MeSH
- kyseliny mastné omega-6 metabolismus terapeutické užití MeSH
- lidé MeSH
- mediátory zánětu metabolismus MeSH
- nutriční podpora * metody MeSH
- omega-3 mastné kyseliny metabolismus terapeutické užití MeSH
- péče o pacienty v kritickém stavu * MeSH
- taurin metabolismus nedostatek terapeutické užití MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
- přehledy MeSH
Citrulline ureidase (CTU, EC3.5.1.20) degrades citrulline into ornithine, carbon dioxide, and ammonia. Here, we present the report on expression of recombinant CTU in Escherichia coli. The soluble and active recombinant CTU was expressed in the periplasmic space with the vector pET-22b and the His-tagged CTU was purified with Ni-Affinity Chromatography. The yield of soluble recombinant protein was significantly increased when 1% sorbitol was supplemented in medium. By using phenylisothiocyanate (PITC) pre-column derivatization HPLC, the enzyme activity of recombinant CTU was determined via measuring of the substrate citrulline and the corresponding products. Our results could be useful in the study of CTU biochemical characteristics, enzymatic preparation of ornithine and development of an enzymatic detection method of citrulline.
- MeSH
- Bacteria MeSH
- bakteriologické techniky metody využití MeSH
- chromatografie afinitní metody využití MeSH
- citrulin * izolace a purifikace metabolismus MeSH
- enzymatické testy metody využití MeSH
- Escherichia coli enzymologie imunologie izolace a purifikace MeSH
- Francisella tularensis * enzymologie izolace a purifikace metabolismus MeSH
- hydrolasy izolace a purifikace MeSH
- ornithin * izolace a purifikace metabolismus MeSH
- rekombinantní proteiny genetika izolace a purifikace metabolismus MeSH
- sorbitol diagnostické užití MeSH
- statistika jako téma MeSH
- tularemie diagnóza etiologie mikrobiologie MeSH
- ureasa izolace a purifikace metabolismus MeSH
- vysokoúčinná kapalinová chromatografie metody využití MeSH
- Publikační typ
- práce podpořená grantem MeSH
Diabetes is associated with dire peripheral sequelae including foot ulcers and amputations. A recent article by Wong et al. demystifies this connection by demonstrating that the neutrophil defense mechanism of extruding decondensed chromatin, termed NETosis, mediates delayed wound healing in diabetes and provides a therapeutic strategy for this indication.
- MeSH
- citrulin metabolismus MeSH
- extracelulární pasti * účinky léků MeSH
- hojení ran * účinky léků MeSH
- hydrolasy genetika metabolismus MeSH
- komplikace diabetu patologie MeSH
- lidé MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
