De uitdagingen van de ontwikkeling van vaccins tegen Betacoronaviruses: Antibody afhankelijke versterking en Sendai Virus als een mogelijke vaccin Vector
Om een effectief en veilig vaccin tegen bètacoronavirussen te ontwerpen, is het noodzakelijk om hun evolutionair conservatieve antigene determinanten te gebruiken die de combinatie van sterke humorale en celgemedieerde immuunresponsen zullen opwekken. Het richten op dergelijke determinanten minimaliseert het risico van antilichaamafhankelijke versterking van virale infectie. Dit fenomeen werd waargenomen in dierproeven met experimentele vaccins tegen SARS-CoV-1 en MERS-CoV die waren ontwikkeld op foundation van geïnactiveerd coronavirus of vectorconstructen die het spike-eiwit (S) van het virion tot expressie brengen.
De substitutie en glycosylering van bepaalde aminozuren in de antigene determinanten van het S-eiwit, evenals de conformationele veranderingen, kunnen tot hetzelfde impact leiden in een nieuw experimenteel vaccin tegen SARS-CoV-2. Het gebruik van meer conservatieve structurele en aanvullende virale eiwitten voor de antigene determinanten van het vaccin zal dit probleem helpen voorkomen. Deze evaluation schetst benaderingen voor het ontwikkelen van vaccins tegen het nieuwe SARS-CoV-2-coronavirus die zijn gebaseerd op niet-pathogene virale vectoren.
Voor een efficiënte preventie van infecties veroorzaakt door respiratoire pathogenen is het vermogen van het vaccin om mucosale immuniteit in de luchtwegen te stimuleren belangrijk. Een dergelijk vaccin kan worden ontwikkeld met behulp van een niet-pathogene Sendai-virusvector, omdat het intranasaal kan worden toegediend en een mucosale immuunrespons kan induceren die de antivirale barrière in de luchtwegen versterkt en een betrouwbare bescherming biedt tegen infectie.
Detectie van mutaties in gyrB met behulp van denaturerende hogedrukvloeistofchromatografie ( DHPLC ) tussen Salmonella enterica serovar Typhi en Paratyphi A.
Fluorochinolonresistentie wordt gemedieerd door mutaties in het chinolonenresistentiebepalende gebied (QRDR) van de topoisomerasegenen. Denaturerende hogedrukvloeistofchromatografie (DHPLC) werd geëvalueerd voor de detectie van klinisch belangrijke mutaties in gyrB bij Salmonella.
Salmonella Typhi en S. Paratyphi A, gekarakteriseerd voor mutatie in QRDR van gyrA, parC en parE, werden onderzocht op mutatie in gyrB door DHPLC en gevalideerd door sequencing.
De DHPLC-analyse was in staat om de testmutant te onderscheiden van isolaten met wildtype gyrB en mutanten te onderscheiden van andere mutanten op foundation van piekprofiel en verschuiving in retentietijd. Er werden drie sequentievarianten gedetecteerd op codon 464 en er werd ook een nieuwe mutatie Ser→Thr gedetecteerd . gyrB-mutatie was geassocieerd met niet-klassieke chinolonresistentie (NALS-CIPDS) in 34 isolaten van alleen S. Typhi en was verschillend van klassieke chinolonresistentie geassocieerd met gyrA-mutaties (NALR-CIPDS).
Opstelling van een protocol voor moleculaire diagnose van β-thalassemie-mutaties in Tunesië met behulp van denaturerende high-performance vloeistofchromatografie ( DHPLC ).
ACHTERGROND
β-thalassemie is een van de meest voorkomende autosomaal recessieve aandoeningen wereldwijd . Het vertoont een grote moleculaire heterogeniteit als gevolg van meer dan 200 oorzakelijke mutaties in het β-globine-gen. In Tunesië vertegenwoordigt β- thalassemie de meest voorkomende monogene hemoglobinestoornis met 2,21% van de dragers.
Efficiënte en betrouwbare screeningsmethoden voor mutaties zijn essentieel om geschikte preventieprogramma’s voor risicoparen op te zetten. Het doel van de huidige studie is om een efficiënte methode te ontwikkelen op foundation van de denaturerende hogedrukvloeistofchromatografie (DHPLC) waarbij het hele β-globinegen (HBB) wordt gescreend op mutaties die ongeveer 90% van het spectrum beslaan.
METHODEN
We hebben de validatie uitgevoerd van een DHPLC-assay voor directe genotypering van 11 bekende β-thalassemie-mutaties in de Tunesische bevolking.
RESULTATEN
De DHPLC-assay werd vastgesteld op foundation van de analyse van 62 gearchiveerde β-thalassemiemonsters die eerder waren gegenotypeerd en vervolgens werden gevalideerd met volledige overeenstemming op 50 exams met blinde gerandomiseerde monsters die eerder waren gegenotypeerd met DNA-sequencing en met 96% consistentie op 40 monsters als een prospectieve studie.
CONCLUSIES
In vergelijking met andere genotyperingstechnieken kan de DHPLC-methode voldoen aan de vereisten van directe genotypering van bekende β-thalassemie-mutaties in Tunesië en kan het worden toegepast als een krachtig hulpmiddel voor de genetische screening van prenatale en postnatale individuen.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation. It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1. The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation. It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1. The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation (2). It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1 (2). The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation (2). It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1 (2). The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: Based on its helical structure, LIF (Leukemia Inhibitory Factor) is considered a member of the Interleukin-6 family of cytokines. Functionally, it has been implicated in a many physiological processes including development, hematopoiesis, bone metabolism, and inflammation. Some cell types known to express LIF include activated T cells, monocytes, astrocytes, osteoblasts, keratinocytes, regenerating skeletal muscle, mast cells, and fibroblasts.
Description: Leukemia Inhibitory Factor also called LIF is a lymphoid factor that promotes long-term maintenance of embryonic stem cells by suppressing spontaneous differentiation. Leukemia Inhibitory Factor has several functions such as cholinergic neuron differentiation, control of stem cell pluripotency, bone & fat metabolism, mitogenesis of factor dependent cell lines & promotion of megakaryocyte production in vivo. Human and mouse LIF exhibit a 78% identity in its amino acid sequence. Human LIF is as active on human cells as is it is on mouse cells, though mouse LIF is about 1000 fold less active on human cells, than human LIF.
Description: Leukemia inhibitory factor (LIF) is a member of Interleukin 6 family. This protein is mainly expressed in the trophectoderm of the developing embryo, with its receptor LIFR expressed throughout the inner cell mass. LIF has the capacity to induce terminal differentiation in leukemic cells. Its activities include the induction of hematopoietic differentiation in normal and myeloid leukemia cells, the induction of neuronal cell differentiation, and the stimulation of acute-phase protein synthesis in hepatocytes. LIF is used in mouse embryonic stem cell culture, because that removal of LIF pushes stem cells toward differentiation, but they retain their proliferative potential or pluripotency. It is also used in phase II clinical trial, which can assist embryo implantation in women who have failed to become pregnant despite assisted reproductive technologies (ART). Mature mouse LIF shares 78 % a.a. sequence identity with Human LIF.
Description: Leukemia Inhibitory Factor also called LIF is a lymphoid factor that promotes long-term maintenance of embryonic stem cells by suppressing spontaneous differentiation. Leukemia Inhibitory Factor has several functions such as cholinergic neuron differentiation, control of stem cell pluripotency, bone & fat metabolism, mitogenesis of factor dependent cell lines & promotion of megakaryocyte production in vivo. Human and mouse LIF exhibit a 78% identity in its amino acid sequence. Human LIF is as active on human cells as is it is on mouse cells, though mouse LIF is about 1000 fold less active on human cells, than human LIF. Recombinant mouse LIF produced in E. coli is a single, non-glycosylated, polypeptide chain containing 180 amino acids and having a molecular mass of 19.86 kDa.
Description: Leukemia Inhibitory Factor also called LIF is a lymphoid factor that promotes long-term maintenance of embryonic stem cells by suppressing spontaneous differentiation. Leukemia Inhibitory Factor has several functions such as cholinergic neuron differentiation, control of stem cell pluripotency, bone & fat metabolism, mitogenesis of factor dependent cell lines & promotion of megakaryocyte production in vivo. Human and mouse LIF exhibit a 78% identity in its amino acid sequence. Human LIF is as active on human cells as is it is on mouse cells, though mouse LIF is about 1000 fold less active on human cells, than human LIF. Recombinant mouse LIF produced in E. coli is a single, non-glycosylated, polypeptide chain containing 180 amino acids and having a molecular mass of 19.86 kDa.
Description: Description of target: Leukemia inhibitory factor, or LIF, is an interleukin 6 class cytokine that affects cell growth by inhibiting differentiation. When LIF levels drop, the cells differentiate. The LIF was mapped gene to 22q11-q12.2 by Southern analysis of a series of mouse/human somatic cell hybrids and by in situ hybridization to the chromosomes of 2 normal males and some individuals with chromosomal rearrangements. The gene maps between the Philadelphia translocation BCR1 and the breakpoint of the translocation in cell line GM2324 at 22q12.2. LIF derives its name from its ability to induce the terminal differentiation of myeloid leukemic cells, thus preventing their continued growth. Other properties attributed to the cytokine include: the growth promotion and cell differentiation of different types of target cells, influence on bone metabolism, cachexia, neural development, embryogenesis and inflammation.;Species reactivity: Mouse;Application: ELISA;Assay info: ;Sensitivity: <10pg/ml
Description: Description of target: LIF has the capacity to induce terminal differentiation in leukemic cells. Its activities include the induction of hematopoietic differentiation in normal and myeloid leukemia cells, the induction of neuronal cell differentiation, and the stimulation of acute-phase protein synthesis in hepatocytes.;Species reactivity: Mouse;Application: ;Assay info: Assay Methodology: Quantitative Sandwich ELISA;Sensitivity: 0.039 ng/mL
Ontwikkeling van hoge resolutie DNA Melting Analyse voor gelijktijdige detectie van Potato Mop-Prime Virus en Zijn Vector , Spongospora subterranea, in de bodem
In deze studie werd een reeks duplex reverse transcriptie (RT)-PCR-gemedieerde hoge resolutie DNA-smelt (HRM)-analyses voor gelijktijdige detectie van aardappelmop-virus (PMTV) en zijn protistische vector, Spongospora subterranea f.sp. subterranea (Sss), ontwikkeld.
De aantasting van grond door PMTV werd gedetecteerd met behulp van een op tabak gebaseerd lokaassysteem . Totaal RNA geëxtraheerd uit de bodem leidde tot succesvolle RT-PCR-gelelektroforese-detectie van zowel PMTV als Sss. Om efficiëntere detectie te vergemakkelijken, werden nieuw ontworpen primerparen voor PMTV RNA-soorten (dwz RNA-Rep, -CP en -TGB) samen met de bestaande Sss-primers geanalyseerd met behulp van real-time RT-PCR.
De resulterende amplicons vertoonden smeltprofielen die gemakkelijk konden worden onderscheiden. Onder duplex RT-PCR-formaat leidden alle PMTV- en Sss-primercombinaties tot succesvolle detectie van respectieve PMTV-RNA-soorten en Sss in de monsters door DNA- smeltanalyses met hoge resolutie (HRM) . Toen de duplex HRM-assay werd toegepast op bodemmonsters die waren verzameld van zes velden op vier verschillende locaties in New Brunswick, Canada, werd een positieve detectie van PMTV en/of Sss gevonden in 63-100% monsters verzameld uit velden waarin met PMTV geïnfecteerde knollen waren waargenomen.
De monsters van velden waar noch PMTV- noch Sss-geïnfecteerde knollen waren waargenomen, resulteerden daarentegen in een negatieve detectie door de check. Aastabak bioassay voor PMTV en Sss leverde vergelijkbare resultaten op. Tussen 63% -83% en 100% van de grondmonsters die werden verzameld op PMTV-besmette velden leidden tot respectievelijk PMTV- en Sss-infecties in de lokaasplanten; terwijl er geen PMTV- of Sss-geïnfecteerde planten werden verkregen uit grondmonsters die waren verzameld op PMTV/Sss-vrije velden.
Versnelling van de morfogenetische cyclus van de virale vector Aedes aegypti larven voor snellere larvicide bioassays
Elke bioassay om nieuwe chemisch gesynthetiseerde larviciden of fytolarviciden te testen tegen Culicidae en meer schadelijke muggensoorten, zoals Aede s aegypti en Aedes albopictus , die specifiek knokkelkoorts, gele koorts, chikungunya- viruskoorts overbrengen, evenals het Zika-virus of Anopheles gambiae , een vector voor malaria en philariasis, vereist duizenden goed ontwikkelde larven, bij voorkeur in het vierde stadium. De natuurlijke morfogenetische cyclus van Aedes spp., in het veld of in het laboratorium, kan bij kamertemperatuur (bijv. 25°C) tot 19 dagen duren vanaf het eerste permanente contact tussen levensvatbare eieren en wateren het laatste stadium van larvale groei of metamorfose tot vliegende volwassenen. Een versnelde sequentiële vervelling is dus wenselijk voor snellere bioassays van larviciden.
We hebben dit doel bereikt in Aedes aegypti met zeer beperkte strategische en goedkope toevoegingen aan voedsel, zoals kokoswater, melk of caseïne, gistextract en in mindere mate glycerol. Het van nature rijke kokoswater was uitstekend geschikt om snel de populatie van instar IV-larven te bereiken , de meest geavanceerde vóór de verpopping, wat ongeveer een week bespaart voor daaropvolgende larvicide bioassays. Verdunde melk, als een andere voedselbron, zorgde voor een nog snellere uiteindelijke vervelling en volwassenen zijn nuttig voor taxonomische doeleinden van muggen.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation. It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1. The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation. It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1. The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation (2). It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1 (2). The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: LIF is a multifunctional secreted glycoprotein that exists in both soluble and matrix-bound forms. It displays biologic activities ranging from the differentiation of myeloid leukemic cells into macrophage lineage to effects on bone metabolism, inflammation, neural development, embryogenesis, and the maintenance of implantation (2). It is now clear that LIF is related in both structure and mechanism of action to the interleukin IL-6 family of cytokines, which also includes IL-11, ciliary neurotrophic factor, oncostatin M, and cardiotrophin 1 (2). The actions of these cytokines are mediated through specific cell-surface receptors that consist of a unique chain and the shared signal transducing subunit gp130.
Description: Based on its helical structure, LIF (Leukemia Inhibitory Factor) is considered a member of the Interleukin-6 family of cytokines. Functionally, it has been implicated in a many physiological processes including development, hematopoiesis, bone metabolism, and inflammation. Some cell types known to express LIF include activated T cells, monocytes, astrocytes, osteoblasts, keratinocytes, regenerating skeletal muscle, mast cells, and fibroblasts.
Description: Leukemia Inhibitory Factor also called LIF is a lymphoid factor that promotes long-term maintenance of embryonic stem cells by suppressing spontaneous differentiation. Leukemia Inhibitory Factor has several functions such as cholinergic neuron differentiation, control of stem cell pluripotency, bone & fat metabolism, mitogenesis of factor dependent cell lines & promotion of megakaryocyte production in vivo. Human and mouse LIF exhibit a 78% identity in its amino acid sequence. Human LIF is as active on human cells as is it is on mouse cells, though mouse LIF is about 1000 fold less active on human cells, than human LIF.
Description: Leukemia inhibitory factor (LIF) is a member of Interleukin 6 family. This protein is mainly expressed in the trophectoderm of the developing embryo, with its receptor LIFR expressed throughout the inner cell mass. LIF has the capacity to induce terminal differentiation in leukemic cells. Its activities include the induction of hematopoietic differentiation in normal and myeloid leukemia cells, the induction of neuronal cell differentiation, and the stimulation of acute-phase protein synthesis in hepatocytes. LIF is used in mouse embryonic stem cell culture, because that removal of LIF pushes stem cells toward differentiation, but they retain their proliferative potential or pluripotency. It is also used in phase II clinical trial, which can assist embryo implantation in women who have failed to become pregnant despite assisted reproductive technologies (ART). Mature mouse LIF shares 78 % a.a. sequence identity with Human LIF.
Description: Leukemia Inhibitory Factor also called LIF is a lymphoid factor that promotes long-term maintenance of embryonic stem cells by suppressing spontaneous differentiation. Leukemia Inhibitory Factor has several functions such as cholinergic neuron differentiation, control of stem cell pluripotency, bone & fat metabolism, mitogenesis of factor dependent cell lines & promotion of megakaryocyte production in vivo. Human and mouse LIF exhibit a 78% identity in its amino acid sequence. Human LIF is as active on human cells as is it is on mouse cells, though mouse LIF is about 1000 fold less active on human cells, than human LIF. Recombinant mouse LIF produced in E. coli is a single, non-glycosylated, polypeptide chain containing 180 amino acids and having a molecular mass of 19.86 kDa.
Description: Leukemia Inhibitory Factor also called LIF is a lymphoid factor that promotes long-term maintenance of embryonic stem cells by suppressing spontaneous differentiation. Leukemia Inhibitory Factor has several functions such as cholinergic neuron differentiation, control of stem cell pluripotency, bone & fat metabolism, mitogenesis of factor dependent cell lines & promotion of megakaryocyte production in vivo. Human and mouse LIF exhibit a 78% identity in its amino acid sequence. Human LIF is as active on human cells as is it is on mouse cells, though mouse LIF is about 1000 fold less active on human cells, than human LIF. Recombinant mouse LIF produced in E. coli is a single, non-glycosylated, polypeptide chain containing 180 amino acids and having a molecular mass of 19.86 kDa.
Description: Description of target: Leukemia inhibitory factor, or LIF, is an interleukin 6 class cytokine that affects cell growth by inhibiting differentiation. When LIF levels drop, the cells differentiate. The LIF was mapped gene to 22q11-q12.2 by Southern analysis of a series of mouse/human somatic cell hybrids and by in situ hybridization to the chromosomes of 2 normal males and some individuals with chromosomal rearrangements. The gene maps between the Philadelphia translocation BCR1 and the breakpoint of the translocation in cell line GM2324 at 22q12.2. LIF derives its name from its ability to induce the terminal differentiation of myeloid leukemic cells, thus preventing their continued growth. Other properties attributed to the cytokine include: the growth promotion and cell differentiation of different types of target cells, influence on bone metabolism, cachexia, neural development, embryogenesis and inflammation.;Species reactivity: Mouse;Application: ELISA;Assay info: ;Sensitivity: <10pg/ml
Description: Description of target: LIF has the capacity to induce terminal differentiation in leukemic cells. Its activities include the induction of hematopoietic differentiation in normal and myeloid leukemia cells, the induction of neuronal cell differentiation, and the stimulation of acute-phase protein synthesis in hepatocytes.;Species reactivity: Mouse;Application: ;Assay info: Assay Methodology: Quantitative Sandwich ELISA;Sensitivity: 0.039 ng/mL
