Résumé
OBJECTIVE@#The aim of this study is to explore the potential modulatory role of quercetin against Endotoxin or lipopolysaccharide (LPS) induced septic cardiac dysfunction.@*METHODS@#Specific pathogen-free chicken embryos ( n = 120) were allocated untreated control, phosphate buffer solution (PBS) vehicle, PBS with ethanol vehicle, LPS (500 ng/egg), LPS with quercetin treatment (10, 20, or 40 nmol/egg, respectively), Quercetin groups (10, 20, or 40 nmol/egg). Fifteen-day-old embryonated eggs were inoculated with abovementioned solutions via the allantoic cavity. At embryonic day 19, the hearts of the embryos were collected for histopathological examination, RNA extraction, real-time polymerase chain reaction, immunohistochemical investigations, and Western blotting.@*RESULTS@#They demonstrated that the heart presented inflammatory responses after LPS induction. The LPS-induced higher mRNA expressions of inflammation-related factors (TLR4, TNFα, MYD88, NF-κB1, IFNγ, IL-1β, IL-8, IL-6, IL-10, p38, MMP3, and MMP9) were blocked by quercetin with three dosages. Quercetin significantly decreased immunopositivity to TLR4 and MMP9 in the treatment group when compared with the LPS group. Quercetin significantly decreased protein expressions of TLR4, IFNγ, MMP3, and MMP9 when compared with the LPS group. Quercetin treatment prevented LPS-induced increase in the mRNA expression of Claudin 1 and ZO-1, and significantly decreased protein expression of claudin 1 when compared with the LPS group. Quercetin significantly downregulated autophagy-related gene expressions (PPARα, SGLT1, APOA4, AMPKα1, AMPKα2, ATG5, ATG7, Beclin-1, and LC3B) and programmed cell death (Fas, Bcl-2, CASP1, CASP12, CASP3, and RIPK1) after LPS induction. Quercetin significantly decreased immunopositivity to APOA4, AMPKα2, and LC3-II/LC3-I in the treatment group when compared with the LPS group. Quercetin significantly decreased protein expressions of AMPKα1, LC3-I, and LC3-II. Quercetin significantly decreased the protein expression to CASP1 and CASP3 by immunohistochemical investigation or Western blotting in treatment group when compared with LPS group.@*CONCLUSION@#Quercetin alleviates cardiac inflammation induced by LPS through modulating autophagy, programmed cell death, and myocardiocytes permeability.
Sujets)
Embryon de poulet , Animaux , Quercétine/usage thérapeutique , Lipopolysaccharides/toxicité , Matrix metalloproteinase 9 , Caspase-3 , Matrix metalloproteinase 3 , Récepteur de type Toll-4 , Claudine-1 , Inflammation/métabolisme , Apoptose , ARN messager , Autophagie , Facteur de transcription NF-kappa BRésumé
SUMMARY: The domestic chicken is a species of bird that has been extensively studied in regard to its biology and as a model organism for science. The reproduction of the species is by the laying of fertilized eggs, which in a period of 21 days will develop a chick inside. Several methods have been described to develop embryos ex-ovo, allowing the observation and manipulation of the organism. This work has the propose to standardize a method that allows the development of the embryos inside the artificial incubation system, which has a low cost and is easy to make. In this work, 100 chicken eggs were used to study the effects of humidity, mineral supplementation, and the preincubation time of the egg on the incubation ex-ovo of the embryos. Embryo development was documented through the different days. Pulverized eggshell was selected as an optimal source to provide calcium, magnesium, phosphorus, and other minerals to the developing embryo. By providing 900-1200 mg of pulverized eggshell, 40 mL of the 0.001 % solution of benzalkonium chloride, and a preincubation time of approximately 56 h, the embryos were able to develop until 19 days, and even though they did not reach hatching, the incubation conditions that allowed the survival and development of embryos until late stages were achieved. Thus, due to the conditions established for calcium, humidity and preincubation time, in the present work, the chicks reached 19 days of development.
El pollo doméstico es una especie de ave que ha sido ampliamente estudiada en cuanto a su biología y como organismo modelo para la ciencia. La reproducción de la especie es por la puesta de huevos fecundados, que en un período de 21 días desarrollarán un polluelo en su interior. Se han descrito varios métodos para desarrollar embriones ex-ovo, permitiendo la observación y manipulación del organismo. Este trabajo tuvo como objetivo estandarizar un método que permita el desarrollo de los embriones dentro del sistema de incubación artificial, el cual tiene un bajo costo y es fácil de realizar. En este trabajo se utilizaron 100 huevos de gallina para estudiar los efectos de la humedad, la suplementación mineral y el tiempo de preincubación del huevo sobre la incubación ex-ovo de los embriones. El desarrollo embrionario se documentó a través de los diferentes días. Se seleccionó la cáscara de huevo pulverizada como una fuente óptima para proporcionar calcio, magnesio, fósforo y otros minerales al embrión en desarrollo. Al suministrar 900-1200 mg de cáscara de huevo pulverizada, 40 mL de la solución de cloruro de benzalconio al 0.001 % y un tiempo de preincubación de aproximadamente 56 h, los embriones lograron desarrollarse hasta los 19 días, y aunque no llegaron a eclosionar, los embriones lograron desarrollarse hasta los 19 días. Se lograron condiciones de incubación que permitieron la supervivencia y desarrollo de los embriones hasta etapas tardías. Así, debido a las condiciones establecidas de calcio, humedad y tiempo de preincubación, en el presente trabajo los pollitos alcanzaron los 19 días de desarrollo.
Sujets)
Animaux , Embryon de poulet , Poulets/croissance et développement , Développement embryonnaire , Oiseaux/embryologie , Techniques de cultureRésumé
Commercial inactivated avian influenza H5 vaccine is used as an essential control strategy for avian influenza disease in Egypt. Since the initial outbreaks of highly pathogenic avian influenza H5N8, the virus has diverged with new genotypes and variant viruses continuing to emerge which mainly stand behind vaccination failure. In the present work, four different commercial avian influenza vaccines were inoculated in specific pathogenic free chickens for assessing its efficacy against local highly pathogenic avian influenza H5N8 virus isolated in 2018 and 2020. Two hundred and forty specific pathogenic free chickens were clustered into four groups; each group was inoculated with the corresponding vaccine (60 specific pathogenic free chickens/vaccine). Sixty specific pathogenic free chicks were kept as control unvaccinated group. Sera collected from vaccinated chicken groups at 3rd and 4th week post vaccination were examined for calculating neutralizing antibodies using heterologous highly pathogenic avian influenza H5N8 2018 and 2020. At 4th week post vaccination, vaccinated chickens were challenged; moreover, oropharyngeal swabs were collected from challenged vaccinated chickens to calculate the viral shedding. Our findings revealed the groups vaccinated with vaccine code no 1 and 2 that contains two vaccine strains (H5N1 and H5N8) of local origin exhibited the highest hemagglutination inhibition titer, protection (percent) and reduction in viral shedding titer when examined by highly pathogenic avian influenza H5N8 2018 while, vaccine code no 3 induced lower antibody response, protection (percent) and reduction in viral shedding, but still within satisfactory level when compared to previous groups. When highly pathogenic avian influenza H5N8 2020 was used, it was found the seroconversion rate, protection (percent) and mean titer of reduction of viral shedding decreased in comparison to those recorded for highly pathogenic avian influenza H5N8 2018. Vaccine code no 4 was impotent to either highly pathogenic avian influenza 2018 or 2020. Accordingly, it was recommended to update vaccine strain according to epidemiological condition and used the predominant circulating strain isolate in challenge test(AU)
La vacuna comercial inactivada H5 se utiliza como estrategia esencial de control de la enfermedad de la gripe aviar en Egipto. Desde los brotes iniciales de la gripe aviar altamente patógena H5N8, el virus ha variado al aparecer continuamente nuevos genotipos y variantes virales, que son los principales responsables del fracaso de la vacunación. En el presente trabajo, cuatro vacunas comerciales diferentes contra la gripe aviar se inocularon en pollos libres de patógenos específicos para evaluar su eficacia contra cepas del virus local de la gripe aviar altamente patógeno H5N8 aisladas en 2018 y 2020. Se agruparon 240 pollos pollos libres de patógenos específicos en cuatro grupos, cada uno fue inoculado con la vacuna correspondiente (60 pollos pollos libres de patógenos específicos/vacuna). Sesenta pollos SPF se mantuvieron como grupo control sin vacunar. Los sueros de los pollos vacunados recogidos en la 3ª y 4ª semana después de la vacunación se examinaron para calcular los anticuerpos neutralizantes contra la gripe aviar heteróloga H5N8 2018 y 2020. En la cuarta semana después de la vacunación, los pollos vacunados fueron retados; además, se recogieron hisopados orofaríngeos de los pollos vacunados retados para calcular la diseminación viral. Nuestros resultados revelaron que los grupos vacunados con las vacunas con códigos nº 1 y 2, que contienen dos cepas vacunales (H5N1 y H5N8) de origen local, mostraron el mayor título de inhibición de la hemaglutinación, protección (por ciento) y reducción del título de excreción viral cuando se evaluaron contra la gripe aviar altamente patógena H5N8 2018, mientras que la vacuna con código nº 3 indujo menor respuesta de anticuerpos, protección (por ciento) y reducción de la excreción viral, pero todavía dentro de un nivel satisfactorio en comparación con los grupos anteriores. Al utilizar la vacuna contra la gripe aviar altamente patógena H5N8 2020, se observó que la tasa de seronconversión, la protección (por ciento) y el título medio de reducción de la excreción viral disminuyeron en comparación con los registrados para la gripe aviar altamente patógena H5N8 2018. La vacuna con código nº 4 no fue potente para la gripe aviar altamente patógena de 2018 o de 2020. Por consiguiente, se recomendó actualizar la cepa de la vacuna de acuerdo con las condiciones epidemiológicas y utilizar el aislamiento de la cepa circulante predominante en la prueba de reto(AU)
Sujets)
Animaux , Embryon de poulet , Tests sérologiques/méthodes , Vaccins antigrippaux/usage thérapeutique , Grippe chez les oiseaux/prévention et contrôleRésumé
RESUMEN: El uso de plataformas virtuales se muestra como un nuevo recurso didáctico que posibilita el proceso de enseñanza-aprendizaje de forma dinámica. A grandes rasgos, permite el acceso a imágenes digitales en alta resolución mediante el uso de computadores, smartphones y/o tabletas. Portanto, este trabajo presenta nuestra metodología en el campo de la embriología de aves domésticas y la experiencia adquirida en el desarrollo de recursos para la enseñanza por medio de las tecnologías de la información y comunicación, de gran utilidad hoy en día en medio de la pandemia ocasionada por el nuevo coronavirus.
SUMMARY: Online platforms are a new didactic resource that enable an active teaching-learning process. In general, they allows access to high resolution digital images through the use of computers, smartphones and / or tablets. Therefore, this study presents our methodology in the field of domestic bird embryology and the experience acquired in the development of teaching resources through information and communication technologies, which are very useful nowadays, particularly in the midst of the pandemic caused by the new coronavirus.
Sujets)
Animaux , Interface utilisateur , Embryon de poulet/anatomie et histologie , Embryologie/enseignement et éducation , Enseignement à distance , Oiseaux/embryologie , Internet , Technologie de l'informationRésumé
The vascular network expansion and functioning are important factors affecting normal intra-uterine fetal development. This study addressed the previously reported antiangiogenic potential of beta-2-glycoprotein I (β2GPI) in vivo in the chick embryo model of angiogenesis. The effects of two naturally occurring β2GPI forms on the development of the chorioallantoic membrane (CAM) vessels and the chicken embryo were investigated. β2GPI monomers and dimers were obtained by fractioned purification and characterized using SDS-PAGE, immunoblot, and ELISA. The egg exposure was performed by injection of small volumes of 2.5 µg/mL solutions of the β2GPI subfractions. Angiogenesis was evaluated through quantitative measurements of vascular architecture parameters in the captured CAM images, using computational analysis of texture contrasts and computer vision techniques. Quantitative information was assigned to the CAM vasculature modifications. In vivo, the β2GPI dimer completely halted the formation of CAM vessels and led to embryo death after 48 h of exposure. The β2GPI monomer allowed the embryo to develop up to the 10th day, despite early changes of CAM vessels. The impaired normal vessel growth proceeded as a self-limited effect. The β2GPI monomer-exposed eggs showed reduced vascularization on the 6th day of incubation, but embryos were viable on the 10th day of incubation, with ingurgitated CAM vessels implying sequelae of the angiogenesis inhibition. Both subfractions impaired CAM vasculature development. The β2GPI dimer proved to be largely more harmful than the β2GPI monomer. β2GPI modification by cleavage or dimerization may play a role in angiogenesis control in vivo.
Sujets)
Poulets , Chorioallantoïde , Embryon de poulet , Néovascularisation physiologique , Inhibiteurs de l'angiogenèse/pharmacologie , bêta 2-Glycoprotéine IRésumé
To rapidly and accurately manipulate genome such as gene deletion, insertion and site mutation, the whole genome of a very virulent strain Md5 of Marek's disease virus (MDV) was inserted into bacterial artificial chromosome (BAC) through homogeneous recombination. The recombinant DNA was electroporated into DH10B competent cells and identified by PCR and restriction fragment length polymorphism analysis. An infectious clone of Md5BAC was obtained following transfection into chicken embryo fibroblast (CEF) cells. Furthermore, a lorf10 deletion mutant was constructed by two step Red-mediated homologous recombination. To confirm the specific role of gene deletion, the lorf10 was reinserted into the original site of MDV genome to make a revertant strain. All the constructs were rescued by transfection into CEF cells, respectively. The successful packaging of recombinant viruses was confirmed by indirect immunofluorescence assay. The results of growth kinetics assay and plaques area measurement showed that the lorf10 is dispensable for MDV propagation in vitro. Overall, this study successfully constructed an infectious BAC clone of MDV and demonstrated its application in genome manipulation; the knowledge gained from our study could be further applied to other hepesviruses.
Sujets)
Animaux , Embryon de poulet , Poulets , Chromosomes artificiels de bactérie , ADN recombiné , Herpèsvirus aviaire de type 2/génétique , Maladie de MarekRésumé
Avian coronavirus (AvCoV) infects a range of tissues in chickens and several other avian species. Although the virus can be isolated in chicken embryos, only a few strains of the 6 genotypes/33 lineages can grow in cell lines, with the Beaudette strain (GI-1 lineage) being the most used for in vitro studies. Considering the differences between cell lines and chicken embryos as habitats for AvCoV, this study aimed to assess the diversity of the genes coding for the nonstructural protein 3 (nsp3) and spike envelope protein (S) after serial passages in BHK-21 and Vero cells. After 14 passages of an embryo-adapted Beaudette strain, the virus loads fluctuated in both cell lines, with the highest loads being 8.72 log genome copies/µL for Vero and 6.36 log genome copies/µL for BHK-21 cells. No polymorphisms were found for nsp3; regarding S, not only aa substitutions (Vero: 8th passage A150S, and 14th S150A; BHK-21: 4th S53F, 8th F53Y, and 8th S95R), but also minor variants could be detected on chromatograms with fluctuating intensities. As the regions of these aa substitutions are within the receptor-binding domain of S, it can be speculated that differences in cell receptors between Vero and BHK-21 cells and the speed of cell death led to the selection of different dominant strains, while the stability of nsp3 supports its function as a protease involved in AvCoV replication. In conclusion, AvCoV quasispecies evolution is influenced by the biological model under consideration, and a gradual transition is seen for minor and major variants.(AU)
O Coronavírus aviário AvCoV infecta uma variedade de tecidos de galinhas e de outras espécies aviárias. Apesar de este vírus poder ser isolado em ovos embrionados de galinha, apenas alguns dos 6 genótipos / 33 linhagens podem crescer em cultivo celular, sendo a cepa Beuadette (linhagem GI-11) a mais utilizada para estudos in vitro. Considerando as diferentes linhagens celulares e ovos embrionados como habitats para o AvCoV, este estudo teve por objetivo estudar a diversidade de genes que codificam para a proteína não-estrutural 3 (nsp3) e espícula (S) após passagens seriadas em células BHK-21 e VERO. Após 14 passagens, de uma amostra Beuadette adaptada a ovos embrionados, os títulos virais variaram em ambas as células, com os maiores títulos sendo de 8,72 log cópias genômicas/µL para Vero e 6,36 cópias genômicas/µL para BHK-21. Nenhum polimorfismo foi encontrando para nsp3. Considerando a proteína S, não somente foram encontradas substituições de aminoácidos (Vero: 8a passagem A150S e 14a passagem S150A; BHK-21: 4a passagem S53F, 8a passagem F53Y e S95R), mas também, variantes subconsensuais foram detectadas pelos cromatogramas com intensidades flutuantes. Uma vez que as regiões destes aa se encontram no domínio de ligação de receptor de S, pode-se especular que diferenças em receptores celulares entre Vero e BHK-21, além da velocidade da morte celular, levaram à seleção de diferentes cepas dominantes, enquanto que a estabilidade de nsp3 concorda com sua função como protease com papel na replicação de AvCoV. Como conclusão, a evolução de quase-espécies de AvCoV é influenciada pelo modelo biológico sob consideração e uma transição gradual é vista para variantes dominantes e subdominantes.(AU)
Sujets)
Embryon de poulet , Protéines virales non structurales , Infections à coronavirus/médecine vétérinaire , Glycoprotéine de spicule des coronavirus , GammacoronavirusRésumé
La angiogénesis es el proceso por el cual se forman nuevos vasos sanguíneos a partir de otros ya existentes. Para que esto se lleve a cabo de forma correcta debe existir un balance entre los factores proangiogénicos y los factores antiangiogénicos dentro del microambiente tisular. Por otra parte, la existencia de productos químicos naturales como los polifenoles, que son capaces de adquirirse en la dieta, inducen a estos factores a intervenir en el proceso de angiogénesis. Se administraron los polifenoles en filtros de metilcelulosa sobre la membrana alantocoriónica de huevos White Leghorn, manteniendo el posterior desarrollo normal del feto. Se utilizaron 15 fetos de pollo fijados en formalina tamponada, a los cuales se extrajo el corazón. El procesamiento de las muestras de corazón se realizó a través de técnicas histológicas, histoquímicas e inmunohistoquímica. Finalmente se evaluó la presencia del VEGF y la capacidad de formar vasos sanguíneos bajo el tratamiento con los polifenoles. La inmunorreactividad fue cuantificada mediante Image J®. Los resultados indican que Ácido cafeico y Pinocembrina disminuyen la densidad microvascular y la expresión de VEGF en corazones de fetos de pollo tratados con estos polifenoles. Tanto el Ácido Cafeico como la Pinocembrina cumplen un rol inhibitorio en el proceso de angiogénesis fisiológica en corazón de pollo, pudiendo modular las vías de señalización mediadas por los VEGFR o modulando la disponibilidad de VEGF. Estos polifenoles podrían utilizarse para el estudio de otros tejidos asociados a angiogénesis patológica.
Angiogenesis is the process by which new blood vessels are formed from other existing ones. A balance between proangiogenic factors and anti-angiogenic factors within the microenvironment must exist for the process to be carried out correctly. Similarly, the existence of natural chemicals such as polyphenols, which are capable of being acquired in the diet, induce these factors in the angiogenic process. Polyphenols were administered in the methylcellulose filters on the of chorioallantoic membrane of White Leghorn eggs, maintaining the normal posterior development of the fetus. 15 chicken fetuses were fixed in buffered formalin, obtaining the hearts to histological processing, performing histological, histochemical and immunohistochemical techniques. VEGF levels and the ability of the blood vessels growing under the stimulation of the polyphenols were evaluated. Immunoreactivity was quantified by Image J. The results indicate that caffeic acid and pinocembrin decreased microvascular density and VEGF expression in hearts stimulated with these polyphenols. Both the caffeic and pinocembrin acids play an inhibitory role in the physiological angiogenesis process in the chicken heart, which decrease the microvascular density and could act by modulating the signaling pathways mediated by the VEGFR or by modulating the availability of VEGF. The use of these polyphenols could be useful in studies of other tissues associated with pathological angiogenesis.
Sujets)
Animaux , Acides caféiques/pharmacologie , Néovascularisation physiologique/effets des médicaments et des substances chimiques , Embryon de poulet , Polyphénols/pharmacologieRésumé
La exposición prenatal al alcohol es causa de alteraciones somáticas, cognitivas y conductuales que se agrupan bajo el término de trastorno del espectro alcohólico fetal (TEAF). La evolución a largo plazo de los sujetos afectados a menudo es desfavorable, especialmente a nivel académico y adaptativo social. En el perfil neuropsicológico es característica la disfunción ejecutiva a menudo asociada a trastornos de la conducta que evolucionan en muchos casos hacia la delincuencia a partir de la adolescencia y en la edad adulta. Se han descrito también déficits de las habilidades sociales y la empatía. La exposición prenatal al alcohol constituye la causa más frecuente de trastorno del neurodesarrollo adquirido y prevenible.
Prenatal exposure to alcohol is the cause of cognitive and behavioural disorders grouped under the term fetal alcohol spectrum disorders (FASD). The long-term evolution of subjects with FASD is often unfavourable, especially in social and academic fields. Executive dysfunction is a hallmark deficit for children with FASD with increased rates of externalizing behaviours, such as aggressiveness and frequently delinquency in adolescence and adulthood. Deficits in social skills, empathy and communication ability are frequent observed among FASD. Prenatal exposure to alcohol is the most frequent cause of acquired and preventable neurodevelopmental disorder.
Sujets)
Humains , Animaux , Femelle , Grossesse , Incapacités de développement/diagnostic , Troubles du spectre de l'alcoolisation foetale/diagnostic , Pronostic , Troubles du comportement social/étiologie , Embryon de poulet , Incapacités de développement/complications , Incapacités de développement/physiopathologie , Incertitude , Erreurs de diagnostic , Troubles du spectre de l'alcoolisation foetale/physiopathologieRésumé
RESUMEN: Los niveles de VEGF y su unión a sus receptores son etapas claves en la regulación de la angiogénesis. El ácido acetilsalicílico (AAS), ampliamente utilizado en tratamiento post infarto al miocardio ha mostrado poseer un efecto antiangiogénico en modelos tumorales. Este efecto potencialmente contraproducente requiere ser estudiado en miocardio. El objetivo del presente trabajo es cuantificar el efecto de AAS y de ácido salicílico (AS) sobre la vascularización en membrana alantocoriónica (MAC) y sobre los niveles de VEGF-A y VEGFR2 en miocardio de embriones de pollo. Para ello, treinta fetos de pollo White Leghorn fueron instilados a los 10 días de gestación con 60 µL de DMSO 0,1 % (control) o conteniendo además 0,3 µmol de AAS o AS. A las 48 horas se realizó procesamiento histológico de MAC para recuento de vasos sanguíneos y de tejido cardíaco para cuantificar VEGF-A y VEGFR2 por inmunohistoquímica. La inmunorreactividad fue cuantificada mediante Image J. Tanto AAS como AS disminuyeron la densidad microvascular de MAC. En miocardio, AAS aunque no AS, disminuyó la concentración de VEGFR2. No hubo efecto sobre VEGF-A. En nuestro modelo experimental, fetos de pollo a los 10 días de gestación también se observó el efecto inhibidor de AAS sobre la angiogénesis en MAC. La disminución de VEGFR2 en cardiomiocitos sugiere que AAS también afecta la angiogénesis en miocardio sano, modificando la disponibilidad del receptor a VEGF. Estos hallazgos nos permiten postular que AAS podría interferir con la regeneración de tejido, en situaciones como post infarto al miocardio.
SUMMARY: The VEGF levels and its binding to its receptors are key stages in the regulation of angiogenesis. Acetylsalicylic acid (ASA), widely used in post-myocardial infarction treatment, has been shown to have an anti-angiogenic effect in tumor models. This potentially counterproductive effect requires to be studied in myocardium. The aim of this study is to quantify the effect of ASA and salicylic acid (SA) on the vascularization in chick allantochorionic membrane (CAM) and on the levels of VEGF-A and VEGFR2 in myocardium of chicken embryos. Thirty White Leghorn chicken fetuses were instilled at 10 days of gestation with 60 mL of 0.1 % DMSO (control) or also containing 0.3 mmol of ASA or SA. After 48 hours, CAM histological processing was performed to count blood vessels and heart tissue to quantify VEGFA and VEGFR2 by immunohistochemistry. Immunoreactivity was quantified by Image J. Both ASA and SA decreased CAM microvascular density. In myocardium, AAS, although not SA, decreased the concentration of VEGFR2. There was no effect on VEGF-A. In our experimental model, chicken fetuses at 10 days of gestation, the inhibitory effect of ASA on angiogenesis in CAM were also observed. The decrease in VEGFR2 in cardiomyocytes suggests that ASA also affects angiogenesis in healthy myocardium, modifying the availability of the receptor to VEGF. These findings allow us to postulate that ASA could interfere with tissue regeneration, when it is required, as post myocardial infarction.
Sujets)
Animaux , Embryon de poulet , Acide acétylsalicylique/pharmacologie , Acide salicylique/pharmacologie , Récepteur-2 au facteur croissance endothéliale vasculaire/effets des médicaments et des substances chimiques , Facteur de croissance endothéliale vasculaire de type A/effets des médicaments et des substances chimiques , Coeur/effets des médicaments et des substances chimiques , Immunohistochimie , Néovascularisation physiologique/effets des médicaments et des substances chimiques , Récepteur-2 au facteur croissance endothéliale vasculaire/analyse , Récepteur-2 au facteur croissance endothéliale vasculaire/métabolisme , Facteur de croissance endothéliale vasculaire de type A/analyse , Facteur de croissance endothéliale vasculaire de type A/métabolismeRésumé
RESUMEN: El proceso angiogénico se define como el proceso en el que los vasos sanguíneos generan brotes dando como resultado neovascularidad. Un desbalance en el proceso angiogénico contribuye a numerosos desórdenes inflamatorios, infecciosos, isquémicos, inmunológicos y malignos. En el territorio maxilofacial se pueden encontrar patologías neoplásicas benignas de desarrollo local con un marcado componente angiogénico que determinan su crecimiento y agresividad. Sin embargo, existe escasa evidencia de cómo tratarlas en base al control de la angiogénesis. Terry & Jacoway (1994) desarrollaron un protocolo de tratamiento para lesiones neoplásicas benignas con un importante componente vascular que se utiliza actualmente. Este protocolo consiste en la infiltración intralesional de una suspensión de triamcinolona 10 mg/ml más una solución de anestésico local de uso odontológico como la lidocaína al 2 % asociada a epinefrina en una concentración de 1:200.000. Sin embargo, el uso de epinefrina podría disminuir la acción antiangiogénica de la triamcinolona al ser un vasoconstrictor. El objetivo de este estudio es comparar el efecto antiangiogénico, en la membrana alantocoriónica de pollo (MAC), de esta suspensión versus el efecto de la triamcinolona sin asociar a anestésicos locales. Los resultados del efecto antiangiogénico en la MAC de pollo, obtenidos en la investigación concluyeron que la suspensión de triamcinolona asociada a lidocaína con epinefrina es similar al de la suspensión de triamcinolona sin asociar a anestésicos locales. Además, se logró determinar que las suspensiones de triamcinolona sin asociar a anestésicos locales y las asociadas a anestésicos locales con o sin vasoconstrictor poseen un marcado efecto antiangiogénico, en la MAC de pollo, en comparación al grupo control.
SUMMARY: Angiogenesis is defined as the process through which new blood vessels form from previously existing vessels. Several inflammatory, infectious, ischemic, immunological and malignant disorders are caused by the lack of adequate angiogenesis balance. In the maxillofacial area, there are invasive benign neoplastic pathologies with a strong angiogenic component, which determines aggressive behavior and growth. Studies in the literature are scarce regarding treatment of these conditions based on angiogenesis control. Currently, the protocol used to treat these maxillofacial benign neoplastic lesions, was developed in 1994 by Terry & Jacoway and has a strong angiogenic component. Consequently lesions are treated via intra-lesion administration of triamcinolone 10 mg / mL, a solution used in dental local anesthetic, such as lidocaine 2 %, in conjunction with epinephrine at a concentration of 1:200,000. The objective of this study was to compare the antiangiogenic effect of this protocol in chicken chorioallantoic membrane (CAM) without the use of local anesthetic. The results of the antiangiogenic effect in the CAM obtained in this study concluded that the effect of the suspension of triamcinolone associated to lidocaine with epinephrine, is similar to the suspension of triamcinolone without associating local anesthetics. Furthermore, it was determined that suspensions of triamcinolone without local anesthetic, and those associated to local anesthetic with, and without vasoconstrictor have a strong antiangiogenic effect in CAM compared to the control group.
Sujets)
Animaux , Embryon de poulet , Triamcinolone/administration et posologie , Épinéphrine/administration et posologie , Inhibiteurs de l'angiogenèse/administration et posologie , Chorioallantoïde/effets des médicaments et des substances chimiques , Lidocaïne/administration et posologie , Anesthésiques locaux/administration et posologie , Néovascularisation pathologiqueRésumé
Fowl adenovirus serotype 4 (FAdV-4) strain SD1511 was isolated from chickens with severe inclusion body hepatitis and hydropericardium syndrome in Shandong Province, China. The isolate was cultured in primary chicken embryo kidney cells. A study of pathogenicity indicated that SD1511 readily infected 7-35-d-old chickens by intramuscular injection and intranasal and oral routes, causing 50%-100% mortality. The 35-d-old chickens suffered more severe infection than 7- and 21-d-old chickens with mortality highest in the intramuscular injection group. The serum from surviving chickens showed potent viral neutralizing capability. The complete genome of SD1511 was sequenced and analyzed. The strain was found to belong to the FAdV-4 cluster with more than 99% identity with the virulent FAdV-4 strains isolated in China in recent years except for some distinct variations, including deletions of open reading frame 27 (ORF27), ORF48, and part of ORF19. Our findings suggest that SD1511 might be used as a prototype strain for the study of pathogenesis and vaccine development.
Sujets)
Animaux , Anticorps neutralisants , Aviadenovirus/pathogénicité , Lignée cellulaire , Embryon de poulet/virologie , Poulets/virologie , Chine , Délétion de gène , Variation génétique , Génome , Génome viral , Génomique , Rein/virologie , Foie/virologie , Cadres ouverts de lecture , Maladies de la volaille/virologie , Sérogroupe , Charge virale , Virulence , Maladies virales/virologieRésumé
OBJECTIVE@#To compare the proliferation and capacity of differentiation to vascular endothelial cells and angiogenesis induction among stem cells from human exfoliated deciduous teeth (SHED), dental pulp stem cells (DPSC) and human bone marrow mesenchymal stem cells (BMSC) from orofacial bone.@*METHODS@#SHED and DPSC were isolated from pulp tissue of the patients. BMSC were isolated from orthognathic or alveolar surgical sites. The surface markers of the cells were detected by flowcytometry. Cell counting kit-8 (CCK-8) assays were conducted to detect the proliferation ability of the cells. The cells were induced into endothelial cells with conditional medium and then the induced cells were cultured in Matrigel medium. The expression of angiogenesis-related genes such as platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31), vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 1 (VEGFR1), vascular endothelial growth factor receptor 2 (VEGFR2) and von Willebrand Factor (vWF) were quantified by real-time PCR. The cells were cultured in chick embryo chorioallantoic membrane (CAM) and the vessels were counted after 5 days.@*RESULTS@#The cell surface markers CD73, CD90, CD105 and CD146 of all the stem cells were positive, CD34 and CD45 were negative. The CD146 positive rate of SHED and DPSC was higher than that of BMSC. SHED had a higher proliferation rate than DPSC and BMSC. After angiogenic induction for 14 d, 3 kinds of cells emanated pseudopodia formed grid structure long vasculature in Matrigel media. The total length of tube formation of induced BMSC (7 759.7 μm) and SHED (7 734.3 μm) was higher than DPSC (5 541.0 μm). The meshes number of induced SHED (70.7) was higher than DPSC (60) and BMSC (53.7) in Matrigel medium. The expression of CD31, VEGFR2 and vWF genes of SHED were higher than those of BMSC and DPSC. VEGFR1 gene expression of BMSC was higher than that of the other groups, and SHED was higher than DPSC. The expression of VEGF showed no difference among the cells. No deference was showed between the effect of the stem cells and negative control on new formed vessels in CAM. The total length of vessels of SHED (30.4 mm) was higher than that of the negative control (20.9 mm) and BMSC (28.0 mm).@*CONCLUSION@#SHED, DPSC and BMSC can differentiate into vascular endothelial cells. SHED showed a stronger angiogenesis differentiation and proliferation potential compared with DPSC and BMSC.
Sujets)
Animaux , Embryon de poulet , Humains , Différenciation cellulaire , Prolifération cellulaire , Cellules cultivées , Cellules endothéliales , Cellules souches mésenchymateuses , Facteur de croissance endothéliale vasculaire de type ARésumé
ABSTRACT Objective. The present study aimed to describe in detail the expression patterns of the gene Hey1, an effector of the Notch pathway, during the development of branchial arches and facial prominences. Materials and methods. Fertilized chicken (Gallus gallus) eggs obtained from a local egg farm were incubated at 37.5 -38.5ºC with 70% relative humidity until the embryos reached Hamilton-Hamburger stages HH14 through HH25. Digoxigenin-UTP labeled probes Hey1 were generated from linearized plasmids with either T3 polimerase for in vitro transcription. Whole-mount in situ hybridization was then performed. At least 3 replicates (n=3) were obtained for each stage. To confirm the results observed in whole embryos, sagittal and coronal cryosectioning was performed using a thickness of 10 µm. Results. During developmental stages HH14 and HH18, Hey1 gene expression was localized to the endoderm of branchial pouches. Hey1 gene expression was also observed in the epithelium that covers the maxillary and mandibular prominences during developmental stages HH19 and HH21, as well as in the nasal epithelium between HH19 and HH25. Transcripts were also detected in the epithelium that covers the frontonasal prominence during stage HH21. Conclusions. These expression patterns suggest the participation of this component of the Notch signaling pathway in craniofacial morphogenesis, possibly establishing pharyngeal segmentation patterns during early stages and/or regulating cell proliferation and differentiation during the late stages of facial development.
RESUMEN Objetivo. El presente estudio tuvo como objetivo describir detalladamente los patrones de expresión del gen Hey1, un efector de la vía Notch durante el desarrollo de arcos branquiales y prominencias faciales. Materiales y métodos. Se incubaron huevos fertilizados de pollo (Gallus gallus) obtenidos de una granja local entre 37.5-38.5ºC con humedad relativa del 70% hasta que los embriones alcanzaron los estadios HH14 hasta HH25 de Hamilton-Hamburger. Las sondas Hey1 marcadas con digoxigenina-UTP se generaron a partir de plásmidos linearizados con T3 polimerasa por transcripción in vitro. Luego se realizó hibridación in situ sobre embriones completos. Se obtuvieron al menos 3 repeticiones (n=3) para cada estadio. Para confirmar los resultados observados en embriones completos, se realizaron cortes sagitales y coronales de 10 µm. Resultados. Durante los estadios de desarrollo HH14 y HH18, la expresión del gen Hey1 se localizó en el endodermo de las bolsas branquiales. La expresión génica de Hey1 también se observó en el epitelio que cubre las prominencias maxilares y mandibulares durante las etapas de desarrollo HH19 y HH21, así como en el epitelio nasal entre HH19 y HH25. También se detectaron transcritos de Hey1 en el epitelio que cubre la prominencia frontonasal durante la etapa HH21. Conclusiones. Estos patrones de expresión sugieren la participación de este componente de la vía de señalización Notch en la morfogénesis craneofacial, posiblemente estableciendo patrones de segmentación faríngea durante las primeras etapas y / o regulando la proliferación y diferenciación celular durante las últimas etapas del desarrollo facial.
Sujets)
Région branchiale , Embryon de poulet , PouletsRésumé
In order to compare Wnt/ beta-catenin expression in mouse and chick facial primordia during development, we performed an immunohistochemical analysis of both protein expressions in E12 to E17 mouse embryos and in E3 and E10 chick embryos. During odontogenesis, from bud to bell stage, both proteins exhibit similar fixation patterns, with epithelial and mesenchymal immunoreactivity, consistant with literature data. Double labelling demonstrates that the same cells express both antigens, even in undifferentiated mesenchyme. The enamel knot, and the ameloblastic and odontoblastic layers are stained at the same manner. In the chick, Wnt and beta-catenin are diffusely present on craniofacial mesenchyme. In both species, premuscular blastemata express Wnt and b-catenin, but Wnt is specifically expressed on the perichondrium and ossification centers, suggesting a role independent from beta-catenin pathway.
Sujets)
Animaux , Embryon de poulet , Souris , Dent/embryologie , Embryon non mammalien/métabolisme , Protéines de type Wingless/métabolisme , bêta-Caténine/métabolisme , Immunohistochimie , Technique d'immunofluorescenceRésumé
Objective: To analyze the association between sociodemographic profile, health profile and the quality of life of caregivers of elderly people diagnosed with dementia. Method: A cross-sectional study was carried out in a geriatric outpatient unit with 35 elderly caregivers diagnosed with dementia. A socio-demographic and health characterization form was used as well as the WHOQOL-bref scale. To verify the association of the variables, the Spearman or Pearson correlation tests were applied, according to the normality of the data. Results: A moderate association between caregiver age and quality of life was found, as well as between total time of care and quality of life, which reveals that the care provided to the elderly with dementia has repercussions on the quality of life. The domains of the WHOQOL-bref with the highest means were Social and Physical Relations, and those with lower mean values were Psychological and Environmental. Conclusion: The identification of conditions that influence the quality of life of caregivers of elderly people with dementia allows actions to promote, protect and recover the health and well-being of those who provide care to be established, so that this care is both for themselves and for the elderly.
Objetivo: Analisar associação entre o perfil sociodemográfico, características do cuidado e a qualidade de vida de cuidadores de idosos diagnosticados com demência. Método: Estudo transversal, realizado em uma unidade ambulatorial geriátrica com 35 cuidadores de idosos diagnosticados com demência. Foi utilizado uma ficha de caracterização sociodemográfica e de saúde assim como a escala WHOQOL-bref. Para verificar a associação das variáveis idade, qualidade de vida e tempo total de cuidado foram aplicados os testes de "correlação de Spearman" ou "Pearson", conforme a normalidade dos dados. Resultados: Constatou-se associação moderada entre idade do cuidador e sua qualidade de vida, assim como tempo total de cuidado e qualidade de vida, o que mostra que o cuidado prestado ao idoso demenciado gera repercussão na qualidade de vida. Os domínios do WHOQOL-bref que tiveram melhores médias foram Relações Sociais e Físico, e os que apresentaram menores médias foram Psicológico e Meio Ambiente. Conclusão: A identificação de condições que influenciam a qualidade de vida dos cuidadores de idosos com demência permite o estabelecimento de ações de promoção, proteção e recuperação da saúde e do bem-estar de quem cuida, de forma que este cuidado transcenda para si e para o idoso.
Sujets)
Humains , Mâle , Femelle , Adulte d'âge moyen , Sujet âgé , Embryon de poulet , Qualité de vie , Santé des Anciens , Soins , Aidants , DémenceRésumé
Este estudo objetivou avaliar os efeitos da inoculação de proteína isolada de soja (P.I.S.) em ovos embrionados de matrizes semipesadas com diferentes idades. Foram utilizados 320 ovos embrionados de matrizes semipesadas da linhagem Rhode Island Red com 35 e 70 semanas de idade. Ao 17º dia de incubação, os ovos foram inoculados com 0,5mL de solução diretamente na cavidade alantoide. O delineamento experimental utilizado foi o inteiramente ao acaso, em esquema fatorial (2 x 4), constituído por dois fatores: idades das matrizes (matrizes jovens e matrizes velhas) e soluções inoculadas (ovo íntegro; 0,5% de solução salina; 1% de P.I.S. + 0,5% de solução salina; e 2% de P.I.S. + 0,5% de solução salina), totalizando oito tratamentos contendo 40 ovos cada. Os dados coletados foram analisados pelo teste de Tukey a 5% de significância. A inoculação de proteína isolada de soja apresentou efeito positivo sobre a relação pinto-ovo, porém sem proporcionar melhora nos rendimentos de incubação, no desenvolvimento dos órgãos e no desempenho de pintos na fase pré-inicial. A idade da matriz influenciou diretamente as variáveis avaliadas, em que ovos oriundos de aves velhas apresentaram maior peso, maior mortalidade embrionária na fase tardia, pintos mais pesados e pior conversão alimentar.(AU)
This study aimed to evaluate the influence of inoculation of isolated soy protein (P.I.S.) in embryonated eggs of semi-heavy with different ages. 320 embryonated eggs were used from semi-heavy breeders of Rhode Island Red lineage with 35 and 70 weeks of age. In the 17th day of incubation, the eggs were inoculated with 0.5 ml of solution in the allantoic cavity. The experimental method was completely randomized in factorial scheme (2 x 4), consisting of two factors: matrices ages (young and old matrices) and inoculation solutions (non-inoculated eggs; 0.5% of saline solution; 1% of P.I.S. + 0.5% of saline solution; and 2% of P.I.S. + 0.5% of saline solution, totalling eight treatments containing 40 eggs each. The inoculation of isolated soy protein showed positive effect on the chick-egg relation, albeit without improving incubation yields, organ development and chicks's performance in the pre-initial phase. The age of the breeder directly influenced the variables evaluated, with eggs from older breeder showing better weight, higher embryonic mortality in late phase, heavier chicks and worse feed conversion.(AU)
Sujets)
Animaux , Embryon de poulet/croissance et développement , Poulets , Oeufs/analyse , Développement embryonnaire , Protéines/analyse , Glycine maxRésumé
Abstract Very high gravity (VHG) technology was employed on industrial scale to produce ethanol from molasses (fermented) as well as by-products formation estimation. The effect of different Brix° (32, 36 and 40) air-flow rates (0.00, 0.20, 0.40, and 0.60 vvm) was studied on ethanol production. The maximum ethanol production was recorded to be 12.2% (v/v) at 40 Brix° with 0.2 vvm air-flow rate. At optimum level aeration and 40 Brix° VHG, the residual sugar level was recorded in the range of 12.5-18.5 g/L, whereas the viable cell count remained constant up to 50 h of fermentation and dry matter production increased with fermentation time. Both water and steam consumption reduced significantly under optimum conditions of Brix° and aeration rate with compromising the ethanol production. Results revealed VHG with continuous air flow is viable technique to reduce the ethanol production cost form molasses at commercial scale.
Sujets)
Embryon de poulet , Éthanol/métabolisme , Microbiologie industrielle/méthodes , Mélasses/microbiologie , Saccharomyces cerevisiae/métabolisme , Fermentation , Surpesanteur , Microbiologie industrielle/instrumentation , Mélasses/analyse , Mélasses/économie , Saccharomyces cerevisiae/génétiqueRésumé
The aim of this study was to observe morphological changes of the cultured otocysts isolated from various stages of the chick embryo. Isolated otocysts were dissected from embryonic day, E2.5-4.5 of incubation (HH stage 16-26) according to stages of developing inner ear. Morphology of the chick otocyst exhibited an ovoid shape. The width and height of the otocyst were 0.2 mm and 0.3 mm, respectively. Elongation of a tube-like structure, the endolymphatic duct, was found at the dorsal aspect of the otocyst. The cultured otocyst is lined by the otic epithelium and surrounding periotic mesenchymal cells started to migrate outwards the lateral aspect of such epithelium. Notably, the acoustic-vestibular ganglion (AVG) was observed at the ventrolateral aspect of the otocyst. Appearance of AVG in vitro can be applied for studying chemical-induced ototoxicity and sensorineural hearing loss. It was concluded that the organ-cultured otocyst of the chick embryo could be used as a model to study sensory organ development of avian inner ear.
El objetivo de este estudio fue observar los cambios morfológicos de otocistos cultivados aislados en las diversas etapas del desarrollo del embrión de pollo. Otocistos aislados fueron obtenidos de embriones día, E2.5-4.5 de incubación (HH etapa 16-26) de acuerdo a las etapas de desarrollo del oído interno. El otocisto de pollo presentó una morfología ovoide. El ancho y la altura del otocisto fue de 0,2 mm y 0,3 mm, respectivamente. En la cara dorsal del otocisto se visualizó el alargamiento de una estructura similar a un tubo, el conducto endolinfático. El otocisto cultivado está revestido por epitelio ótico y células mesenquimatosas perióticas que comienzan a migrar hacia el exterior de la cara lateral en búsqueda del epitelio. En particular, el ganglio acústico-vestibular (GAV) fue observado en la parte ventrolateral del otocisto. La aparición de GAV in vitro puede ser aplicado para el estudio de la ototoxicidad inducida por productos químicos y la pérdida de audición neurosensorial. Se concluyó que el otocisto cultivado de embrión de pollo podría ser utilizado como un modelo para estudiar el desarrollo de órganos sensoriales del oído interno aviar.
Sujets)
Animaux , Embryon de poulet/anatomie et histologie , Oreille interne/embryologie , MorphogenèseRésumé
BACKGROUND: The marine environment is a rich source of bioactive natural products. Many of the marine bioactive compounds have been derived successfully from molluscs. Euchelus asper is a marine mollusc which is commonly found in the intertidal rocky regions of the Mumbai coast. The present study was focused on evaluating the anti-angiogenic and anti- proliferative activities of methanolic extract of Euchelus asper (EAME). METHODS: The anti-angiogenic activity of EAME (50-800 µg/mL) was assessed by chick chorio-allantoic membrane (CAM) model wherein multiple parameters in the CAM blood vessels were analysed through morphometric and histo-logical investigations. In vitro testing of EAME (5-20 µg/mL) included its cytotoxicity against three different cancer cell lines, its effect on cell proliferation by wound healing assay as well as their relevant molecular mechanisms. Statistical analysis was carried out by two-tailed student's t test for two unpaired groups. RESULTS: Analysis of CAM revealed that the extract is effective in reducing the branching points of the 1st order blood vessels or capillaries of CAM. Histological analysis of CAM showed significant decrease in capillary plexus and compartmentalization along with increase in mesodermal blood vessels, thus establishing its anti-angiogenicity. Further, EAME exhibited moderate but significant cytotoxicity against A549 non-small cell lung carcinoma cell line. We also demonstrated that the cytotoxicity of EAME in A549 was associated with its apoptotic activity by subG1 phase arrest. Lastly, EAME significantly reduced A549 proliferation by reducing the expression of Matrix metalloproteinase-2 (MMP-2) and Matrix metalloproteinase-9 (MMP-9). CONCLUSION: Overall, our study suggested that EAME has potential to inhibit tumour angiogenic and proliferative activity and may be a potential source for development of new anti-cancer pharmaceuticals.