Francisella tularensis influences several host molecular/signaling pathways during infection. Ubiquitination and deubiquitination are among the most important regulatory mechanisms and respectively occur through attachment or removal of the ubiquitin molecule. The process is necessary not only to mark molecules for degradation, but also, for example, to the activation of signaling pathways leading to pro-inflammatory host response. Many intracellular pathogens, including Francisella tularensis, have evolved mechanisms of modifying such host immune responses to escape degradation. Here, we describe that F. tularensis interferes with the host's ubiquitination system. We show increased total activity of deubiquitinating enzymes (DUBs) in human macrophages after infection, while confirm reduced enzymatic activities of two specific DUBs (USP10 and UCH-L5), and demonstrate increased activity of USP25. We further reveal the enrichment of these three enzymes in exosomes derived from F. tularensis-infected cells. The obtained results show the regulatory effect on ubiquitination mechanism in macrophages during F. tularensis infection.

• MeSH
• deubikvitinasy metabolismus   MeSH
• Francisella tularensis * MeSH
• gramnegativní bakteriální infekce * metabolismus   MeSH
• lidé MeSH
• makrofágy MeSH
• signální transdukce MeSH
• thiolesterasa ubikvitinu metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Three-dimensional cell culture systems are increasingly used for biological and anticancer drug screening as they mimic the structure and microenvironment of tumors more closely than conventional two-dimensional cell models. In this study, the growth kinetics of colon adenocarcinoma-derived spheroids (HT-29 cell line) cultivated in liquid marble micro-bioreactors and nonadherent PDMS-coated well plates was investigated in detail and enabled precise control of the spheroid size by the seed cell density and cultivation time. The therapeutic effect of 5-fluorouracil and irinotecan hydrochloride in 2D monolayer cell culture and 3D tumor spheroids revealed an unexpected twist in their efficacy due to different ability to penetrate through 3D microtissue. For 5-fluorouracil, the inhibitory concentration IC50 after 48 h exposure increased from 11.3 μM for a 2D cell culture to 707.7 μM for a 3D spheroid. In the case of irinotecan, IC50 increased from 24.9 μM to 77.8 μM. Despite its higher molar weight, irinotecan appeared to penetrate the 3D spheroid structure more efficiently than 5-fluorouracil. While 5-fluorouracil mainly caused a suppression of spheroid growth from the outside, irinotecan affected the entire spheroid and caused its originally compact structure to disintegrate. The acquired results highlight the need to screen cancer chemotherapeutics on 3D tumor models, as contrasting results can be obtained compared to standard 2D cell cultures.

• MeSH
• adenokarcinom * MeSH
• buněčné sféroidy MeSH
• cytostatické látky * farmakologie   MeSH
• fluoruracil farmakologie   MeSH
• irinotekan farmakologie   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádorové mikroprostředí MeSH
• nádory tračníku * farmakoterapie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Yeast glucan particles (GPs) are promising agents for the delivery of biologically active compounds as drugs. GPs possess their own biological activities and can act synergistically with their cargo. This study aimed to determine how incorporating artemisinin, ellagic acid, (-)-epigallocatechin gallate, morusin, or trans-resveratrol into GPs affects their anti-inflammatory and antioxidant potential in vitro. Two different methods - slurry evaporation and spray drying - were used to prepare composites (GPs + bioactive compound) and the anti-inflammatory and antioxidative properties of the resultant products were compared. Several of the natural compounds showed the beneficial effects of being combined with GPs. The materials prepared by spray drying showed greater activity than those made using a rotary evaporator. Natural compounds incorporated into yeast GPs showed greater anti-inflammatory potential in vitro than simple suspensions of these compounds as demonstrated by their inhibition of the activity of transcription factors NF-κB/AP-1 and the secretion of the pro-inflammatory cytokine TNF-α.

• MeSH
• antiflogistika farmakologie   MeSH
• antioxidancia farmakologie   MeSH
• artemisininy farmakologie   MeSH
• cytokiny MeSH
• flavonoidy chemie   farmakologie   MeSH
• glukany chemie   farmakologie   MeSH
• katechin analogy a deriváty   chemie   farmakologie   MeSH
• kyselina ellagová chemie   farmakologie   MeSH
• monocyty účinky léků   MeSH
• NF-kappa B MeSH
• resveratrol chemie   farmakologie   MeSH
• Saccharomyces cerevisiae - proteiny MeSH
• Saccharomyces cerevisiae MeSH
• Publikační typ
• časopisecké články MeSH

The goal of this work was to assess the usability of yeast glucan particles (GPs) as carriers for curcumin and determine the beneficial effect of a pharmacological composite of curcumin in GPs on dextran sulfate sodium induced colitis in rats. The assessment of the anti-inflammatory effect of particular substances was evaluated on the basis of the calculated disease activity index and by assessment of cytokines and enzymes from the gut tissue - tumor necrosis factor α (TNF-α), transforming growth factor β1, interleukin (IL)-1β, IL-6, IL-10, IL-17, catalase, superoxide dismutase 2, myeloperoxidase (MPO), and matrix metalloproteinase 9. Composites of GPs with incorporated curcumin showed promising results with the capability to lower symptoms of colitis and significantly decrease the production of pro-inflammatory cytokines TNF-α, IL-1β, IL-6, and the activity of MPO, as well. The anti-inflammatory effect of the composites was greater than those of pure GPs or curcumin.

• MeSH
• antiflogistika aplikace a dávkování   terapeutické užití   MeSH
• cytokiny metabolismus   MeSH
• glukany aplikace a dávkování   terapeutické užití   MeSH
• interleukiny metabolismus   MeSH
• kolitida chemicky indukované   farmakoterapie   MeSH
• krysa rodu rattus MeSH
• kurkumin aplikace a dávkování   terapeutické užití   MeSH
• nosiče léků terapeutické užití   MeSH
• potkani Wistar MeSH
• Saccharomyces cerevisiae metabolismus   MeSH
• síran dextranu MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Yeast glucan particles are porous polysaccharide cell walls extracted from Saccharomyces cerevisiae. Being mildly immunogenic, they are efficiently phagocytosed and have therefore been proposed as possible vehicles for drug delivery. Using curcumin as a model poorly water-soluble drug, a systematic comparison of three different physical loading methods - incipient wetness impregnation, slurry evaporation, and spray drying - was carried out and their influence on the particle morphology, encapsulation efficiency, amorphous drug content and release kinetics was evaluated. It was found that yeast glucan particles can contain up to 30% wt. of curcumin in the amorphous form when prepared by slurry evaporation. The dissolution of curcumin from glucan particles lead to a supersaturated solution in asimilar way as amorphous solid dispersions do, despite the fact that glucan particles themselves do not dissolve. Bi-phasic dissolution tests revealed up to 4-fold acceleration of curcumin dissolution rate from amorphous glucan particles compared to its crystalline form. Crucially, glucan particles were shown to retain the ability to be recognised and phagocytosed even after drug encapsulation.

• MeSH
• beta-glukany chemie   MeSH
• farmaceutická chemie metody   MeSH
• kinetika MeSH
• krystalizace MeSH
• kurkumin aplikace a dávkování   chemie   MeSH
• lékové transportní systémy * MeSH
• nosiče léků chemie   MeSH
• příprava léků metody   MeSH
• rozpustnost MeSH
• uvolňování léčiv MeSH
• voda chemie   MeSH
• Publikační typ
• časopisecké články MeSH

Glucan particles (GPs) from Saccharomyces cerevisiae consist mainly of β-1,3-d-glucan. Curcumin is a phenolic compound of plant origin. A 24 h incubation with a mixture of GPs and curcumin increased the expression of the Nrf2 protein and increased the activation of the Nrf2-ARE system significantly.

• MeSH
• antioxidancia * chemie   metabolismus   farmakologie   MeSH
• buňky Hep G2 MeSH
• faktor 2 související s NF-E2 metabolismus   MeSH
• glukany chemie   MeSH
• kurkumin * chemie   farmakologie   MeSH
• lidé MeSH
• oxidační stres účinky léků   MeSH
• příprava léků MeSH
• Saccharomyces cerevisiae chemie   MeSH
• THP-1 buňky MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

In this work, novel amorphous solid dispersions based on yeast glucan particles were produced. Yeast glucan particles are hollow and porous, and they are mainly composed of amorphous polysaccharides. We hypothesized that these particles are suitable candidates for the amorphization of drugs with low water solubility. Model drugs ibuprofen and curcumin were successfully encapsulated in glucan particles by spray drying. Different spray-drying parameters were tested to evaluate the influence of atomizing droplet size and initial solid content on encapsulation efficiency. It was shown that higher solid content and, more significantly, larger droplet sizes lead to higher encapsulation efficiencies. The encapsulation efficiency of ibuprofen (10 wt%) into glucan particles was considerably improved from 41.3 ± 0.5% to 64.3 ± 0.2% by increasing initial solid content and droplet size with the two-fluid nozzle. The spray drying process was further optimized by using the ultrasonic nozzle and it was possible to achieve complete encapsulation of ibuprofen and curcumin without any precipitation of the active compound outside of the glucan particles. Overall, it was possible to produce completely amorphous composites with outstanding wettability and dispersion properties, and with significantly faster dissolution rates when compared to the micronized crude drug.

• MeSH
• aerosoly MeSH
• beta-glukany chemie   izolace a purifikace   MeSH
• ibuprofen chemie   MeSH
• kinetika MeSH
• kurkumin chemie   MeSH
• nosiče léků * MeSH
• příprava léků MeSH
• rozpustnost MeSH
• Saccharomyces cerevisiae chemie   MeSH
• ultrazvuk * MeSH
• uvolňování léčiv MeSH
• velikost částic MeSH
• vysoušení * MeSH
• Publikační typ
• časopisecké články MeSH

Nespecifické střevní záněty (např. Crohnova choroba) jsou onemocnění se stále narůstající incidencí, jejichž současná farmakoterapie není zcela efektivní. Existuje ovšem celá řada přírodních aktivních látek s pozitivním vlivem při terapii střevních zánětů, které jsou intenzivně zkoumány. Prozatím ale nebyla testována možná terapeutická synergie mezi těmito látkami a glukany, přírodními sacharidy s dobře zdokumentovaným imunomodulačním účinkem. Glukanové částice mohou sloužit jako nosiče biologicky aktivních látek, což zjednoduší aplikaci ve vodě nerozpustných látek do organismu a zároveň skýtá možnost razantního zesílení protizánětlivého účinku. Na základě předchozích in vivo a klinických studií byly vytipovány chemicky různorodé přírodní látky, které jsou slibné pro terapii chronických zánětlivých onemocnění a lze je enkapsulovat do glukanových částic. Cílem projektu je pomocí in vitro a in vivo studií ověřit, zda je vhodné použít glukanové mikročástice jako nosiče těchto protizánětlivých látek k terapii střevních zánětů a optimalizovat složení kompozitu glukan-aktivní látka.; Inflammatory Bowel Diseases (IBD), e.g. Crohn’s Disease, are disorders with constantly increasing incidence and not completely effective pharmacotherapy. Number of active natural compounds with positive effect in therapy of IBD is intensively studied. Therapeutic synergy between them and glucans - natural saccharides with well-documented immunomodulatory effect - was not tested so far. Glucan microparticles can serve as carriers for biologically active compounds to simplify the administration of water insoluble drugs into organism and to open the possibility of significant amplification of anti-inflammatory effect as well. Based on previous in vivo and clinical trials, chemically diverse natural compounds, which may be encapsulated into glucan particles, were chosen as promising for therapy of chronic inflammatory disorders. The aim of project is to prove via in vitro and in vivo studies the suitability of utilization the glucan microparticles as vectors of these anti-inflammatory compounds in the therapy of IBD and to optimize the composition of glucan-active compound composite.

• MeSH
• antiflogistika MeSH
• biologické faktory MeSH
• glukany terapeutické užití   MeSH
• idiopatické střevní záněty farmakoterapie   MeSH
• intravitální mikroskopie MeSH
• lidé MeSH
• mikropartikule MeSH
• nosiče léků MeSH
• techniky in vitro MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• hodnotící studie MeSH

• Konspekt
• Farmacie. Farmakologie
• NLK Obory
• gastroenterologie
• farmacie a farmakologie
• NLK Publikační typ
• závěrečné zprávy o řešení grantu AZV MZ ČR

Geranylovaný flavanon diplakon je látka izolovaná z Paulownia tomentosa (Thunb.) Steud. (Paulowniaceae) vykazující protizánětlivé a antioxidační vlastnosti a také vysokou lipofilitu a nízkou rozpustnost ve vodě. Jako modelová molekula byl proto použit k inkorporaci do glukanových částic (GP) s cílem zvýšit jeho potenciální biodostupnost. GP jsou v principu duté schránky připravené čištěním kvasinek Saccharomyces cerevisiae pro získání buněčné stěny, obsahující převážně β-(1→3)/β-(1→6) glukan. Cílem práce je porovnat antiflogistické působení kompozitů diplakonu a glukanového nosiče s působením samotné látky, samotných glukanových částic a fyzické směsi čistých glukanových částic s čistým diplakonem. Na buněčné linii odvozené z lidských leukemických monocytů THP1-XBlueTM-MD2-CD14 byla simulována zánětlivá reakce stimulací buněk lipopolysacharidem (LPS) z Escherichia coli. Kompozity GP a diplakonu signifikantně snížily aktivitu prozánětlivých transkripčních faktorů nukleárního faktoru κB (NF-κB) a aktivátorového proteinu 1 (AP-1) v porovnání s čistou látkou.

Geranylated flavanone diplacone is a flavanone isolated from Paulownia tomentosa (Thunb.) Steud. (Paulowniaceae) with anti-inflammatory and antioxidant properties, nevertheless showing high lipophilicity and low solubility in water. Diplacone was therefore used as a model molecule for incorporation into glucan particles (GPs). GPs are prepared by intensive washing of yeast (Saccharomyces cerevisiae) leading to hollow shells consisting of β-(1→3)/β-(1→6) glucan mainly. The aim of this study was to compare anti-inflammatory potential of GPs-diplacone composites with the compound itself, GPs themselves and the physical mixture of GPs and diplacone. The cell line THP1-XBlueTM-MD2-CD14 derived from human leukemic monocytes was stimulated with lipopolysaccharide (LPS) from Escherichia coli to trigger inflammatory reaction. The composites of GPs with diplacone significantly decreased the activity of pro-inflammatory transcription factors nuclear factor κB (NF-κB) and activator protein 1 (AP-1).

• Klíčová slova
• diplakon,
• MeSH
• enkapsulace buněk MeSH
• glukany terapeutické užití   MeSH
• klinická studie jako téma MeSH
• lidé MeSH
• zánět * farmakoterapie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH

Natural compounds offer a wide spectrum of potential active substances, but often they have a poor bioavailability. To increase the bioavailability and bioactivity of the natural anti-inflammatory molecules curcumin and diplacone, we used glucan particles (GPs), hollow shells from Saccharomyces cerevisiae composed mainly of β-1,3-d-glucan. Their indigestibility and relative stability in the gut combined with their immunomodulatory effects makes them promising carriers for such compounds. This study aimed to determine how curcumin and diplacone, either alone or incorporated in GPs, affect the immunomodulatory activity of the latter by assessing the respiratory burst response and the secretion of pro-inflammatory cytokines by primary porcine innate immune cells. Incorporating curcumin and diplacone into GPs by controlled evaporation of the organic solvent substantially reduced the respiratory burst response mediated by GPs. Incorporated curcumin in GPs also reduced GPs mediated secretion of IL-1β and TNF-α by innate immune cells. The obtained results indicate a potentially beneficial effect of the incorporation of curcumin or diplacone into GPs against inflammation.

• MeSH
• antiflogistika chemie   farmakologie   MeSH
• beta-glukany chemie   izolace a purifikace   farmakologie   MeSH
• flavanony chemie   farmakologie   MeSH
• imunologické faktory chemie   izolace a purifikace   farmakologie   MeSH
• interleukin-1beta metabolismus   MeSH
• kultivované buňky MeSH
• kurkumin chemie   farmakologie   MeSH
• leukocyty mononukleární účinky léků   imunologie   metabolismus   MeSH
• neutrofily účinky léků   imunologie   metabolismus   MeSH
• nosiče léků * MeSH
• příprava léků MeSH
• respirační vzplanutí účinky léků   MeSH
• rozpouštědla chemie   MeSH
• Saccharomyces cerevisiae chemie   MeSH
• Sus scrofa MeSH
• TNF-alfa metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH