In the shadow of SARS-CoV-2, influenza seems to be an innocent virus, although new zoonotic influenza viruses evolved by mutations may lead to severe pandemics. According to WHO, there is an urgent need for better antiviral drugs. Blocking viral hemagglutinin with multivalent N-acetylneuraminic acid derivatives is a promising approach to prevent influenza infection. Moreover, dual inhibition of both hemagglutinin and neuraminidase may result in a more powerful effect. Since both viral glycoproteins can bind to neuraminic acid, we have prepared three series of amphiphilic self-assembling 2-thio-neuraminic acid derivatives constituting aggregates in aqueous medium to take advantage of their multivalent effect. One of the series was prepared by the azide-alkyne click reaction, and the other two by the thio-click reaction to yield neuraminic acid derivatives containing lipophilic tails of different sizes and an enzymatically stable thioglycosidic bond. Two of the three bis-octyl derivatives produced proved to be active against influenza viruses, while all three octyl derivatives bound to hemagglutinin and neuraminidase from H1N1 and H3N2 influenza types.
- MeSH
- chřipka lidská * farmakoterapie MeSH
- hemaglutininové glykoproteiny viru chřipky metabolismus MeSH
- hemaglutininy farmakologie MeSH
- kyselina N-acetylneuraminová farmakologie metabolismus MeSH
- kyseliny neuraminové MeSH
- lidé MeSH
- neuraminidasa metabolismus MeSH
- virus chřipky A, podtyp H1N1 * MeSH
- virus chřipky A, podtyp H3N2 MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
The pertussis agent Bordetella pertussis produces a number of virulence factors, of which the filamentous hemagglutinin (FhaB) plays a role in B. pertussis adhesion to epithelial and phagocytic cells. Moreover, FhaB was recently found to play a crucial role in nasal cavity infection and B. pertussis transmission to new hosts. The 367 kDa FhaB protein translocates through an FhaC pore to the outer bacterial surface and is eventually processed to a ~220 kDa N-terminal FHA fragment by the SphB1 protease. A fraction of the mature FHA then remains associated with bacterial cell surface, while most of FHA is shed into the bacterial environment. Previously reported indirect evidence suggested that FHA, or its precursor FhaB, may bind the β2 integrin CD11b/CD18 of human macrophages. Therefore, we assessed FHA binding to various cells producing or lacking the integrin and show that purified mature FHA does not bind CD11b/CD18. Further results then revealed that the adhesion of B. pertussis to cells does not involve an interaction between the bacterial surface-associated FhaB and/or mature FHA and the β2 integrin CD11b/CD18. In contrast, FHA binding was strongly inhibited at micromolar concentrations of heparin, corroborating that the cell binding of FHA is ruled by the interaction of its heparin-binding domain with sulfated glycosaminoglycans on the cell surface.
- MeSH
- antigeny CD18 MeSH
- bakteriální adheze MeSH
- bakteriální adheziny metabolismus MeSH
- Bordetella pertussis * metabolismus MeSH
- faktory virulence rodu Bordetella MeSH
- glykosaminoglykany MeSH
- hemaglutininy metabolismus MeSH
- heparin MeSH
- integriny MeSH
- lidé MeSH
- makrofágový antigen 1 MeSH
- pertuse * MeSH
- proteasy MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Leucine-rich repeat-containing G-protein-coupled receptor 4 (LGR4) is produced in a broad spectrum of mouse embryonic and adult tissues and its deficiency results in embryonal or perinatal lethality. The LGR4 function was mainly related to its potentiation of canonical Wnt signaling; however, several recent studies associate LGR4 with additional signaling pathways. To obtain a suitable tool for studying the signaling properties of Lgr4, we generated a tagged variant of the Lgr4 receptor using gene targeting in the mouse oocyte. The modified Lgr4 allele expresses the Lgr4 protein fused with a triple hemagglutinin (3HA) tag located at the extracellular part of the protein. The allele is fully functional, enabling tracking of Lgr4 expression in the mouse tissues. We also show that via surface labeling, the 3HA tag allows direct isolation and analysis of living Lgr4-positive cells obtained from the small intestinal crypts. Finally, the HA tag-specific antibody can be employed to characterize the biochemical features of Lgr4 and to identify possible biding partners of the protein in cells derived from various mouse tissues.
- MeSH
- alely MeSH
- genový targeting metody MeSH
- hemaglutininy genetika MeSH
- lidé MeSH
- myši MeSH
- oocyty metabolismus MeSH
- receptory spřažené s G-proteiny genetika MeSH
- signální dráha Wnt genetika MeSH
- těhotenství MeSH
- tkáňová distribuce MeSH
- vazba proteinů genetika MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
In recent decades, it has become clear that most of human proteins are glycosylated and that protein glycosylation plays an important role in health and diseases. At present, simple, fast and inexpensive methods are sought for clinical applications and particularly for improved diagnostics of various diseases, including cancer. We propose a label- and reagent-free electrochemical method based on chronopotentiometric stripping (CPS) analysis and a hanging mercury drop electrode for the detection of interaction of sialylated protein biomarker a prostate specific antigen (PSA) with two important lectins: Sambucus nigra agglutinin (SNA) and Maackia amurensis agglutinin (MAA). Incubation of PSA-modified electrode with specific SNA lectin resulted in an increase of CPS peak H of the complex as compared to this peak of individual PSA. By adjusting polarization current and temperature, PSA-MAA interaction can be either eliminated or distinguished from the more abundant PSA-SNA complex. CPS data were in a good agreement with the data obtained by complementary methods, namely surface plasmon resonance and fluorescent lectin microarray. It can be anticipated that CPS will find application in glycomics and proteomics.
Transplantace ledviny od žijícího dárce inkompatibilního v krevní skupině představuje nový nástroj, jak zvýšit dostupnost transplantací. Většina evropských protokolů je založena na desenzitizačním protokolu s rituximabem, trojkombinace imunosupresiv, intravenózních imunoglobulinů a specifické imunoadsorpce, při které jsou z plazmy odstraňovány konkrétní hemaglutininy. Program transplantací ledvin od inkompatibilních dárců v krevní skupině byl v našem centru založen v roce 2011. Dosud bylo provedeno 21 těchto transplantací. Nejvyšší titr hemaglutininů před rituximabem byl 1 : 64 a korespondující počty imunoadsorpcí před transplantací od 2 do 9 výkonů. Po transplantaci byly imunoadsorpce prováděny podle aktuálního titru hemaglutininů 1–2krát u 5 nemocných. V posledních 2 letech jsou tyto transplantace nabízeny inkompatibilním příjemcům, kteří nenaleznou uplatnění v systému párových výměn ledvin anebo se účastní párové výměny s cílem dosáhnout vyšší míry shody v HLA antigenech. Hlavní komplikací bylo častější pooperační krvácení a infekce močových cest u rizikových nemocných. Ve většině protokolárních biopsií byla popsána pozitivita C4d barvení. Nebyla pozorována ztráta štěpu ani úmrtí nemocných.
Living donor AB0 incompatible kidney transplantation represents a new tool how to improve the access to transplantation. Majority of European protocols are based on desensitization with rituximab, triple drug immunosuppression, intravenous immunoglobulins and specific immunoadsoption (IA) which eliminates isohaemaglutinins. AB0i kidney transplant program was initiated in our centre in 2011 and 21 patients have received grafts from incompatible donors until recently. Highest accepted isohaemaglutinins titers before rituximab were 1 : 64 and corresponding pretransplant immunoadsorption procedures varied from 2 to 9. In 5 patients 1–2 IA procedures were performed also after transplantation. With the advent of paired exchange program the AB0i transplantation is offered to patients with unsuccessful matching run or with aim to improve HLA match between donor and recipient. The main complications were postoperative bleeding and urinary tract infections in patients at risk. Majority of protocol biopsies exhibited positivity of C4d staining. Neither graft loss nor patient death were noticed.
- Klíčová slova
- AB0 inkompatibilita, imunoadsorpce, glykosorb,
- MeSH
- ABO systém krevních skupin imunologie MeSH
- dospělí MeSH
- hemaglutininy krev MeSH
- imunosorpční techniky * MeSH
- imunosupresiva terapeutické užití MeSH
- krvácení komplikace MeSH
- lidé středního věku MeSH
- lidé MeSH
- nekompatibilita krevních skupin * imunologie terapie MeSH
- perioperační péče MeSH
- pooperační komplikace MeSH
- příprava pacienta k transplantaci metody MeSH
- rejekce štěpu prevence a kontrola MeSH
- senioři MeSH
- transplantace ledvin * MeSH
- výsledek terapie MeSH
- žijící dárci MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- senioři MeSH
- Publikační typ
- přehledy MeSH
Hemagglutinin (HA) is an antigenic glycoprotein, which is placed on the surface of the influenza viruses. It is responsible for binding the virus to the host cell, that is being infected. The name „hemagglutinin“ comes from the ability of protein to cause erythrocytes to agglutinate („clump together“). The process is like this: Hemagglutinin (HA) binds to the monosaccharide sialic acid which is present on the surface of its target host cells. The cell membrane then engulfs the virus through endocytosis and followed by formation of endosome. The cell then attempts to begin digesting the contents of the endosome by acidifying its interior and transforming it into a lysosome. When the pH decrease to 6.0, the HA molecule becomes partially unfold, and release a hydrophobic portion of peptide chain that was previously hidden. This so-called „fusion peptide“ acts like a molecular grapple hook for lock on the endosomal membrane. The rest of the HA molecule refolds into a new structure and pulls the endosomal membrane right up next to the viral membrane, causing the two to fuse together. When it happened, the viral RNA genome enters into the cell‘s cytoplasm.
Autoimunitní hemolytická anémie je skupina patologických stavů, charakterizovaných výskytem autoprotilátek proti antigenům erytrocytů. Autoimunitní hemolytickou anémii klasifikujeme jednak podle asociace s jinými onemocněními (primární vs. sekundární autoimunitní hemolytická anémie), jednak podle vlastností patologických autoprotilátek. Klinické projevy jsou závislé na intenzitě autoprotilátkami zprostředkovaného zrychleného zániku erytrocytů a na schopnosti erytropoézy tento zánik kompenzovat, průběh může být akutní, event. chronický se střídáním období relapsů a remisí, ale i bezpříznakový pouze s laboratorními projevy. Výskyt autoprotilátek může představovat komplikaci případné transfúzní substituce a vyžadovat složitější postupy při předtransfúzním vyšetření
Autoimmune haemolytic anaemia is a group of pathological conditions characterised by the presence of autoantibodies against erythrocyte antigens. Autoimmune haemolytic anaemia is classified according to associations with other diseases (primary vs. secondary autoimmune haemolytic anaemia), and also based on properties of the pathological autoantibodies. The clinical symptoms are dependent on the intensity of the autoantibodies-induced shortened survival of erythrocytes and on the bone marrow erythropoietic capacity. The condition can be both acute and chronic, with periodical phases of relapses and remissions, but it can also be asymptomatic, detectable only by laboratory findings. The presence of the autoantibodies can complicate blood transfusion and thus call for more complex pre-transfusion testings.
- Klíčová slova
- klasifikace AIHA, autoprotilátky proti erytrocytům, diagnostika AIHA, komplikace transfúze při AIHA, poléková AIHA,
- MeSH
- aglutininy MeSH
- autoimunitní hemolytická anemie diagnóza etiologie klasifikace MeSH
- autoprotilátky diagnostické užití MeSH
- erytrocyty imunologie patologie MeSH
- hematologické testy využití MeSH
- imunologické testy využití MeSH
- krevní transfuze metody MeSH
- léčivé přípravky MeSH
- lidé MeSH
- nežádoucí účinky léčiv MeSH
- Check Tag
- lidé MeSH
BACKGROUND: Fibrinogen-related proteins with lectin activity are believed to be part of the tick innate immune system. Several fibrinogen-related proteins have been described and characterised mainly on the basis of their cDNA sequences while direct biochemical evidence is missing. One of them, the haemolymph lectin Dorin M from the tick Ornithodoros moubata was isolated and characterised in more depth. RESULTS: Several fibrinogen-related proteins were detected in the haemolymph of ixodid ticks Dermacentor marginatus, Rhipicephalus appendiculatus, R. pulchellus, and R. sanguineus. These proteins were recognised by sera directed against the tick lectin Dorin M and the haemagglutination activity of the ticks R. appendiculatus and D. marginatus. Cross-reactivity of the identified proteins with antibodies against the fibrinogen domain of the human ficolin was also shown. The carbohydrate-binding ability of tick haemolymph was confirmed by haemagglutination activity assays, and this activity was shown to be inhibited by neuraminic acid and sialylated glycoproteins as well as by N-acetylated hexosamines. The fibrinogen-related proteins were shown to be glycosylated and they were localised in salivary glands, midguts, and haemocytes of D. marginatus. Hemelipoglycoprotein was also recognised by sera directed against the fibrinogen-related proteins in all three Rhipicephalus species as well as in D. marginatus. However, this protein does not contain the fibrinogen domain and thus, the binding possibly results from the structure similarity between hemelipoglycoprotein and the fibrinogen domain. CONCLUSIONS: The presence of fibrinogen-related proteins was shown in the haemolymph of four tick species in high abundance. Reactivity of antibodies directed against ficolin or fibrinogen-related proteins with proteins which do not contain the fibrinogen domain points out the importance of sequence analysis of the identified proteins in further studies. Previously observed expression of fibrinogen-related proteins in haemocytes together with the results of this study suggest involvement of fibrinogen-related proteins in tick immunity processes. Thus, they have potential as targets for anti-tick vaccines and as antimicrobial proteins in pharmacology. Research on fibrinogen-related proteins could reveal further details of tick innate immunity processes.
- MeSH
- anatomické struktury zvířat chemie MeSH
- Dermacentor chemie MeSH
- fibrinogen imunologie MeSH
- glykoproteiny imunologie metabolismus MeSH
- hemaglutininy imunologie metabolismus MeSH
- hmyzí proteiny imunologie metabolismus MeSH
- lektiny imunologie metabolismus MeSH
- lidé MeSH
- metabolismus sacharidů MeSH
- Ornithodoros chemie MeSH
- protilátky imunologie MeSH
- Rhipicephalus chemie MeSH
- vazba proteinů MeSH
- zkřížené reakce MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Klíčová slova
- Progenika,
- MeSH
- aglutininy diagnostické užití MeSH
- biotechnologie MeSH
- diagnostické techniky a postupy MeSH
- diagnostické techniky molekulární metody využití MeSH
- genetické testování MeSH
- krev MeSH
- krevní transfuze MeSH
- lidé MeSH
- molekulární biologie MeSH
- společnosti farmaceutické MeSH
- společnosti MeSH
- Check Tag
- lidé MeSH
- Geografické názvy
- Španělsko MeSH
BACKGROUND: Tick carrier proteins are able to bind, transport, and store host-blood heme, and thus they function also as antioxidants. Nevertheless, the role of carrier proteins in ticks is not fully understood. Some of them are found also in tick males which do not feed on hosts to such an extent such as females (there are differences in male feeding in different tick species) and thus they are not dealing with such an excess of heme; some of the carrier proteins were found in salivary glands where the processing of blood and thus release of heme does not occur. Besides, the carrier proteins bind relatively low amounts of heme (in one case only two molecules of heme per protein) compared to their sizes (above 200 kDa). The main aim of this study is the biochemical characterization of a carrier protein from the ornate sheep tick Dermacentor marginatus, hemelipoglycoprotein, with emphasis on its size in native conditions, its glycosylation and identification of its modifying glycans, and examining its carbohydrate-binding specificity. RESULTS: Hemelipoglycoprotein from D. marginatus plasma was purified in native state by immunoprecipitation and denatured using electroelution from SDS-PAGE separated plasma. The protein (290 kDa) contains two subunits with molecular weights 100 and 95 kDa. It is glycosylated by high-mannose and complex N-glycans HexNAc(2)Hex(9), HexNAc(2)Hex(10), HexNAc(4)Hex(7), and HexNAc(4)Hex(8). The purified protein is able to agglutinate red blood cells and has galactose- and mannose-binding specificity. The protein is recognized by antibodies directed against plasma proteins with hemagglutination activity and against fibrinogen-related lectin Dorin M from the tick Ornithodoros moubata. It forms high-molecular weight complexes with putative fibrinogen-related proteins and other unknown proteins under native conditions in tick plasma. Feeding does not increase its amounts in male plasma. The hemelipoglycoprotein was detected also in hemocytes, salivary glands, and gut. In salivary glands, the protein was present in both glycosylated and nonglycosylated forms. CONCLUSION: A 290 kDa hemelipoglycoprotein from the tick Dermacentor marginatus, was characterized. The protein has two subunits with 95 and 100 kDa, and bears high-mannose and complex N-linked glycans. In hemolymph, it is present in complexes with putative fibrinogen-related proteins. This, together with its carbohydrate-binding activity, suggests its possible involvement in tick innate immunity. In fed female salivary glands, it was found also in a form corresponding to the deglycosylated protein.
- MeSH
- Dermacentor chemie metabolismus MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- glykoproteiny chemie izolace a purifikace metabolismus MeSH
- glykosylace MeSH
- hemaglutininy chemie izolace a purifikace metabolismus MeSH
- hemoproteiny chemie izolace a purifikace metabolismus MeSH
- imunoprecipitace MeSH
- lipoproteiny chemie izolace a purifikace metabolismus MeSH
- metabolismus sacharidů MeSH
- molekulová hmotnost MeSH
- Ornithodoros imunologie MeSH
- ovce MeSH
- podjednotky proteinů chemie izolace a purifikace metabolismus MeSH
- protilátky imunologie MeSH
- transportní proteiny chemie izolace a purifikace metabolismus MeSH
- vazba proteinů MeSH
- zkřížené reakce MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
