BACKGROUND & AIMS: Acute liver failure (ALF) is defined as rapid onset coagulopathy and encephalopathy in patients without a prior history of liver disease. We performed untargeted and targeted serum proteomics to delineate processes occurring in adult patients with ALF and to identify potential biomarkers. METHODS: Sera of 319 adult patients with ALF (∼50% acetaminophen [APAP]-related cases) were randomly selected from admission samples of the multicenter USA Acute Liver Failure Study Group consortium and subdivided into discovery/validation cohorts. They were analyzed using untargeted proteomics with mass spectroscopy and a serum cytokine profiling and compared with 30 healthy controls. The primary clinical outcome was 21-day transplant-free survival. Single-cell RNAseq data mapped biomarkers to cells of origin; functional enrichment analysis provided mechanistic insights. Novel prognostic scores were compared with the model for end-stage liver disease and ALFSG prognostic index scores. RESULTS: In the discovery cohort, 117 proteins differed between patients with ALF and healthy controls. There were 167 proteins associated with APAP-related ALF, with the majority being hepatocyte-derived. Three hepatocellular proteins (ALDOB, CAT, and PIGR) robustly and reproducibly discriminated APAP from non-APAP cases (AUROCs ∼0.9). In the discovery cohort, 37 proteins were related to 21-day outcome. The key processes associated with survival were acute-phase response and hepatocyte nuclear factor 1α signaling. SERPINA1 and LRG1 were the best individual discriminators of 21-day transplant-free survival in both cohorts. Two models of blood-based proteomic biomarkers outperformed the model for end-stage liver disease and ALFSG prognostic index and were reproduced in the validation cohort (AUROCs 0.83-0.86) for 21-day transplant-free survival. CONCLUSIONS: Proteomics and cytokine profiling identified new, reproducible biomarkers associated with APAP etiology and 21-day outcome. These biomarkers may improve prognostication and understanding of the etiopathogenesis of ALF but need to be independently validated. IMPACT AND IMPLICATIONS: Acute liver failure (ALF) is a sudden, and severe condition associated with high fatality. More sensitive and specific prognostic scores are urgently needed to facilitate decision-making regarding liver transplantation in patients with ALF. Our proteomic analysis uncovered marked differences between acetaminophen and non-acetaminophen-related ALF. The identification of routinely measurable biomarkers that are associated with 21-day transplant-free survival and the derivation of novel prognostic scores may facilitate clinical management as well as decisions for/against liver transplantation. Further studies are needed to quantify less abundant proteins. Although we used two cohorts, our findings still need to be independently and prospectively validated.
- Klíčová slova
- ALF subtyping, Acetaminophen, Acute liver injury, Proteomic profiling,
- Publikační typ
- časopisecké články MeSH
Despite the importance of drug release testing of parenteral depot formulations, the current in vitro methods still require ameliorations in biorelevance. We have investigated here the use of muscle tissue components to better mimic the intramuscular administration. For convenient handling, muscle tissue was used in form of a freeze-dried powder, and a reproducible process of incorporation of tested microspheres to an assembly of muscle tissue of standardized dimensions was successfully developed. Microspheres were prepared from various grades of poly(lactic-co-glycolic acid) (PLGA) or ethyl cellulose, entrapping flurbiprofen, lidocaine, or risperidone. The deposition of microspheres in the muscle tissue or addition of only isolated lipids into the medium accelerated the release rate of all model drugs from microspheres prepared from ester-terminated PLGA grades and ethyl cellulose, however, not from the acid-terminated PLGA grades. The addition of lipids into the release medium increased the solubility of all model drugs; nonetheless, also interactions of the lipids with the polymer matrix (ad- and absorption) might be responsible for the faster drug release. As the in vivo drug release from implants is also often faster than in simple buffers in vitro, these findings suggest that interactions with the tissue lipids may play an important role in these still unexplained observations.
- Klíčová slova
- PLGA, biorelevant,, depot microspheres,, in vitro release,, intramuscular,,
- MeSH
- celulosa analogy a deriváty MeSH
- flurbiprofen aplikace a dávkování MeSH
- kopolymer kyseliny glykolové a mléčné MeSH
- léky s prodlouženým účinkem * MeSH
- lidokain aplikace a dávkování MeSH
- mikrosféry MeSH
- nosiče léků MeSH
- parenterální infuze * MeSH
- pomocné látky MeSH
- prasata MeSH
- příprava léků MeSH
- risperidon aplikace a dávkování MeSH
- svaly metabolismus MeSH
- techniky in vitro MeSH
- uvolňování léčiv MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- celulosa MeSH
- ethyl cellulose MeSH Prohlížeč
- flurbiprofen MeSH
- kopolymer kyseliny glykolové a mléčné MeSH
- léky s prodlouženým účinkem * MeSH
- lidokain MeSH
- nosiče léků MeSH
- pomocné látky MeSH
- risperidon MeSH
Current in vitro drug-release testing of the sustained-release parenterals represents the in vivo situation insufficiently. In this work, a thin agarose hydrogel layer surrounding the tested dosage form was proposed to mimic the tissue. The method was applied on implantable formulations of different geometries (films, microspheres, and cylindrical implants); prepared from various polymers (several Resomer® grades or ethyl cellulose) and loaded with different model drugs: flurbiprofen, lidocaine or risperidone. The hydrogel layer did not possess any retarding effect on the released drug and acted as a physical restriction to swelling and/or plastic deformation of the tested dosage forms. This led to a different surface area available for drug-release compared with testing in release medium alone and correspondingly to significantly different release profiles of the majority of the formulations obtained between the two methods (e.g. t50% = 18 days in pure release medium vs. t50% = 26 days in gel-setup for risperidone loaded Resomer® 503 H films or t50% = 7 days vs. t50% = 19 days for risperidone loaded Resomer® 503 H microspheres). The limited space for swelling and the rigidity of the agarose gel might mimic the tight encapsulation of the dosage form in the tissue better than the conventional liquid medium.
- Klíčová slova
- Agarose, Biorelevant, Drug release, Hydrogel, Implants, In-vitro, PLGA,
- MeSH
- kopolymer kyseliny glykolové a mléčné MeSH
- kyselina mléčná * MeSH
- kyselina polyglykolová * MeSH
- mikrosféry MeSH
- sefarosa MeSH
- uvolňování léčiv MeSH
- velikost částic MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- kopolymer kyseliny glykolové a mléčné MeSH
- kyselina mléčná * MeSH
- kyselina polyglykolová * MeSH
- sefarosa MeSH
Concomitant intake of alcoholic beverages with sustained-release oral formulations may potentially lead to dose dumping. Alcohol-resistance testing is currently a requirement for the manufacturers by regulatory authorities. Silk fibroin produced by silkworm Bombyx mori is suggested in this work as a potential alternative to a narrow spectrum of alcohol-resistant excipients. Oxycodone HCl, tramadol HCl, and flurbiprofen were selected as model drugs and formulated with regenerated silk fibroin either in the form of an amorphous solid dispersion or as a physical mixture and compressed into tablets. Preliminary compactability and tampering-resistance studies were performed. The ethanol-resistance was tested in media containing 5%, 10%, 20%, or 40% (v/v) ethanol concentration. Drug release profiles were compared using f2 similarity factor. Good mechanical tampering-resistance (tensile strength of 14.6 MPa at 400 MPa compression pressure) was obtained for tablets compressed from physical mixture. Tablets compressed from amorphous solid dispersion had lower tensile strength (2.2 MPa) but showed chemical tampering-resistance to extraction by pure ethanol (7.1% of oxycodone HCl after 24 h). Drug release is controlled predominantly by swelling and diffusion. With an increasing ethanol concentration in release medium, the tablets swelled less, resulting in a slower release. This trend was similar for all tested drugs and for both physical states formulations. No dose dumping occurred in the presence of ethanol; therefore, silk fibroin could be considered as an alternative alcohol-resistant excipient for sustained release application.
- Klíčová slova
- alcohol-induced dose dumping, ethanol-resistant, opioids, silk fibroin, sustained-release,
- MeSH
- fibroiny chemie MeSH
- léky s prodlouženým účinkem * MeSH
- pomocné látky * MeSH
- tablety MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- fibroiny MeSH
- léky s prodlouženým účinkem * MeSH
- pomocné látky * MeSH
- tablety MeSH
BACKGROUND: In recent years, many achievements have been realized in the therapy of inflammatory bowel disease (IBD) although its etiology remains unknown. Thus IBD treatment is symptomatic and targets general inflammatory mechanisms. Oral formulations containing 5-aminosalicylic acid (5-ASA) have become the standard therapy for mild-to-moderate IBD. OBJECTIVE: This article is a review of recently published research dealing with new 5-ASA dosage forms. Thus promising candidates for IBD treatment evaluated in vitro are reported; systems tested in vivo in trinitrobenzene sulfonic acid (TNBS)-induced colitis in rats are mentioned; and 5-ASA formulations used in clinical studies are presented. Moreover, all oral dosage forms containing 5-ASA or its prodrugs are reviewed; their characteristics and utilization in IBD treatment are discussed. CONCLUSION: In several clinical studies, it has been shown that multiparticulates such as pellets offer more advantages as compared with single unit forms, that is, coated tablets. Prolonged presence close to the site of the action, improved drug bioavailability, and easier administration of large drug doses belong to the benefits of pellets.
- MeSH
- antiflogistika nesteroidní aplikace a dávkování MeSH
- idiopatické střevní záněty farmakoterapie patofyziologie MeSH
- klinické zkoušky jako téma MeSH
- lékové transportní systémy metody MeSH
- lidé MeSH
- mesalamin aplikace a dávkování MeSH
- příprava léků metody MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Názvy látek
- antiflogistika nesteroidní MeSH
- mesalamin MeSH
Topical delivery of 5-aminosalicylic acid (5-ASA) to the colonic mucosa is important in order to achieve effective drug concentration in the site of inflammation and to minimize its systemic availability. 5-ASA loaded pellets were prepared by an extrusion/spheronization method. Mucoadhesive biopolymer chitosan was incorporated into the pellets, and drug delivery to the colon was controlled by the pH-sensitive polymer Eudragit® FS. Dissolution profiles of coated pellets revealed no drug release at pH 1.2 within 2h and release as intended in the simulated distal ileum and colon. In vivo, chitosan-core drug loaded pellets (AMCh) showed 2.5-fold higher drug metabolite concentration than after chitosan free pellets (AM) administration in the inflamed colonic tissue. Additionally, AMCh demonstrated decreased in AUC in colitis group (1507 ± 400 ng h/ml) compared with AM (1907 ± 122 ng h/ml). In terms of therapeutic efficiency, administration of pellets markedly decreased the colon/body weight ratio (colitis: 0.0355 ± 0.0028; AM 0.0092 ± 0.0033; AMCh 0.0086 ± 0.0022) and myeloperoxidase activity (colitis: 3212 ± 294 U/g tissue; AM 796 ± 211 U/g; AMCh 552 ± 319 U/g). Bioadhesive chitosan pellets showed additional beneficial properties for colonic 5-ASA delivery in the treatment of inflammatory bowel disease by increasing the drug concentration locally.
- MeSH
- antiflogistika aplikace a dávkování chemie MeSH
- biopolymery aplikace a dávkování chemie farmakokinetika MeSH
- chitosan aplikace a dávkování chemie farmakokinetika MeSH
- implantované léky aplikace a dávkování chemie farmakokinetika MeSH
- kolitida farmakoterapie metabolismus MeSH
- kolon účinky léků metabolismus MeSH
- koncentrace vodíkových iontů MeSH
- krysa rodu Rattus MeSH
- lékové transportní systémy metody MeSH
- mesalamin aplikace a dávkování chemie farmakokinetika MeSH
- peroxidasa metabolismus MeSH
- potkani Wistar MeSH
- rozpustnost MeSH
- velikost částic MeSH
- zánět farmakoterapie metabolismus MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- antiflogistika MeSH
- biopolymery MeSH
- chitosan MeSH
- implantované léky MeSH
- mesalamin MeSH
- peroxidasa MeSH
Freshwater pulmonate snails from three locations in Lake Furesø north of Copenhagen were screened for infection with furcocercariae (by shedding in the laboratory) and recovered parasite larvae were diagnosed by molecular methods (by performing PCR of rDNA and sequencing the internal transcribed spacer [ITS] region). Overall prevalence of infection in snails was 2%. Recovered cercariae from Lymnaea stagnalis (Linnaeus) were diagnosed as Diplostomum pseudospathaceum Niewiadomska, 1984 (prevalence 4%) and cercariae from Radix balthica (Linnaeus) as D. mergi (Dubois, 1932) (prevalence 2%). Pathogen-free rainbow trout were then exposed to isolated cercariae and infection success and site location of metacercariae in these fish were determined. Infection experiments confirmed that both species could infect rainbow trout with the eye lens as infection site for the metacercarial stage although infection success differed. Combination of molecular and biological assays may contribute to improvement of our knowledge on diagnosis, distribution and biology of diplostomids in fish.
- MeSH
- ekologie MeSH
- fylogeneze MeSH
- hlemýždi parazitologie MeSH
- infekce červy třídy Trematoda parazitologie přenos veterinární MeSH
- jezera parazitologie MeSH
- metacerkárie MeSH
- mezerníky ribozomální DNA chemie genetika MeSH
- molekulární sekvence - údaje MeSH
- nemoci ryb parazitologie přenos MeSH
- Oncorhynchus mykiss parazitologie MeSH
- organismy bez specifických patogenů MeSH
- polymerázová řetězová reakce veterinární MeSH
- roční období MeSH
- ryby MeSH
- sekvence nukleotidů MeSH
- sekvenční analýza DNA veterinární MeSH
- sladká voda parazitologie MeSH
- Trematoda genetika izolace a purifikace fyziologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Dánsko MeSH
- Názvy látek
- mezerníky ribozomální DNA MeSH
Preparation of coated pellets intended for rutin colon delivery, their evaluation in vitro and in vivo in experimental colitis in rats was the purpose of this study. Pellets were obtained using extrusion/spheronization and coated with three types of coatings (caffeic acid/hypromellose/alginic acid; sodium alginate/hypromellose/zinc acetate; sodium alginate/chitosan). Dissolution using buffers of pH values, β-glucosidase and times corresponding to gastrointestinal tract (GIT) was provided. Pellets coated with alginate/chitosan showed low rutin dissolution (12-14%) in upper GIT conditions and fast release (87-89%) under colon conditions; that is a good presumption of intended rutin release. After colitis induction and development, the rats were treated with pellets and rutin solution administered orally, solution also rectally. Colon/body weight ratio, myeloperoxidase activity and histological evaluation were performed. Rutin was able to promote colonic healing at the dose of 10mg/kg: colon/body weight ratio decreased and myeloperoxidase activity was significantly suppressed. Pellets coated with alginate/chitosan applied orally and rutin solution administered rectally showed the best efficacy. The combination of rutin as natural product, mucoadhesive chitosan degraded in the colon and sodium alginate as the main coating substance in the form of pellets create a promising preparation for therapy of this severe illness.
- MeSH
- algináty chemie MeSH
- antiflogistika aplikace a dávkování chemie farmakologie MeSH
- aplikace orální MeSH
- časové faktory MeSH
- chitosan chemie MeSH
- deriváty hypromelózy MeSH
- farmaceutická chemie MeSH
- farmaceutická technologie metody MeSH
- gastrointestinální látky aplikace a dávkování chemie farmakologie MeSH
- implantované léky MeSH
- kolitida chemicky indukované farmakoterapie patologie MeSH
- kolon účinky léků patologie MeSH
- koncentrace vodíkových iontů MeSH
- krysa rodu Rattus MeSH
- kyselina glukuronová chemie MeSH
- kyselina trinitrobenzensulfonová MeSH
- kyseliny hexuronové chemie MeSH
- kyseliny kávové chemie MeSH
- methylcelulosa analogy a deriváty chemie MeSH
- modely nemocí na zvířatech MeSH
- octan zinečnatý chemie MeSH
- potkani Wistar MeSH
- příprava léků MeSH
- pufry MeSH
- rozpustnost MeSH
- rutin aplikace a dávkování chemie farmakologie MeSH
- stabilita léku MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- algináty MeSH
- antiflogistika MeSH
- caffeic acid MeSH Prohlížeč
- chitosan MeSH
- deriváty hypromelózy MeSH
- gastrointestinální látky MeSH
- implantované léky MeSH
- kyselina glukuronová MeSH
- kyselina trinitrobenzensulfonová MeSH
- kyseliny hexuronové MeSH
- kyseliny kávové MeSH
- methylcelulosa MeSH
- octan zinečnatý MeSH
- pufry MeSH
- rutin MeSH
Primary cilia are microtubule-based organelles that are important for signaling and sensing in eukaryotic cells. Unlike the thoroughly studied motile cilia, the three-dimensional architecture and molecular composition of primary cilia are largely unexplored. Yet, studying these aspects is necessary to understand how primary cilia function in health and disease. We developed an enabling method for investigating the structure of primary cilia isolated from MDCK-II cells at molecular resolution by cryo-electron tomography. We show that the textbook '9 + 0' arrangement of microtubule doublets is only present at the primary cilium base. A few microns out, the architecture changes into an unstructured bundle of EB1-decorated microtubules and actin filaments, putting an end to a long debate on the presence or absence of actin filaments in primary cilia. Our work provides a plethora of insights into the molecular structure of primary cilia and offers a methodological framework to study these important organelles.
- MeSH
- buněčné kultury MeSH
- buňky MDCK MeSH
- Chlamydomonas metabolismus ultrastruktura MeSH
- cilie metabolismus ultrastruktura MeSH
- elektronová kryomikroskopie MeSH
- exprese genu MeSH
- lidé MeSH
- mikrofilamenta metabolismus ultrastruktura MeSH
- mikrotubuly metabolismus ultrastruktura MeSH
- proteiny asociované s mikrotubuly genetika metabolismus MeSH
- psi MeSH
- tomografie elektronová MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- psi MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- proteiny asociované s mikrotubuly MeSH
