RESUMEN
This study aimed to evaluate the effect of three hydrofluoric acid (HF) etching periods on the micro-tensile bond strength between two CAD-CAM ceramic systems [Vita Suprinity (VS) and feldspathic CEREC blocs (CB)] and a composite resin. The ceramics were categorized into six groups based on the surface conditioning protocol used, as follows: G1: CB-HF 5% for 20 s; G2: CB-HF 5% for 40 s; G3: CB-HF 5% for 60 s; G4: VS-HF 5% for 20 s; G5: VS-HF 5% for 40 s; G6: VS-HF 5% for 60 s. Scotchbond Universal was applied onto the pretreated ceramic surfaces and covered with Filtek Z350 XT composite resin. After 24 h, the specimens were cut into microbars (n = 16) and a micro-tensile bond strength test (µTBS) was carried out. An optical microscope was used to examine the fractured microbars. The results showed statistically significant differences between the factors tested (p < 0.01). Moreover, the mean MPa of G1(17.27), G2(13.03), G3(12.82), G4(15.83), G5(21.66), and G6(14.50) was seen to significantly differ. The predominant failure type observed was adhesive, and all three periods of HF etching produced satisfactory bonding between the composite resin and CB. An etching time of 40 s provided the highest µTBS value for VS.
Asunto(s)
Recubrimiento Dental Adhesivo , Porcelana Dental , Cerámica , Resinas Compuestas , Diseño Asistido por Computadora , Ácido Fluorhídrico , Ensayo de Materiales , Cementos de Resina , Propiedades de Superficie , Resistencia a la TracciónRESUMEN
OBJECTIVES: To evaluate, in vitro, the fracture load of IPS e.max CAD occlusal veneers at thicknesses of either 0.3 or 0.6 mm luted to enamel or dentin. MATERIALS AND METHODS: Fifty human molars were randomly distributed into five groups (n = 10): G1 - healthy teeth (control); G2-0.3-mm-thick veneers luted to enamel; G3-0.6-mm-thick veneers luted to enamel; G4-0.3-mm-thick veneers luted to dentin; and G5-0.6-mm-thick veneers luted to dentin. After the luting procedures, the specimens were immersed in distilled water at 37°C for 24 hours and then subjected to mechanical loading (106 cycles at 200 N load). The specimens were subjected to a fracture load test in a universal testing machine. Two-way ANOVA and Tukey's test (α = 0.05) were used to analyze data. RESULTS: Only the thickness factor was significant (P = .002). Values of fracture load followed by distinct letters represent significant differences (P < .05): G1 (3204 N ± 730)ab ; G2 (3144 N ± 729)ab ; G3 (2489 N ± 606)b ; G4 (3591 N ± 776)a ; and G5 (2770 N ± 598)ab . CONCLUSION: IPS e.max ultrathin occlusal veneers luted to enamel or dentin obtained fracture load comparable to that of the healthy tooth. CLINICAL SIGNIFICANCE: IPS e.max CAD ultrathin occlusal veneers at 0.3 or 0.6-mm-thick seem to provide good perspectives in relation to the clinical use.
Asunto(s)
Porcelana Dental , Coronas con Frente Estético , Cerámica , Esmalte Dental , Análisis del Estrés Dental , Dentina , Humanos , Ensayo de MaterialesRESUMEN
Type 3 immunity encompasses innate and adaptive immune responses mediated by cells that produce the signature cytokines IL-17A and IL-17F. This class of effector immunity is particularly adept at controlling infections by pyogenic extracellular bacteria at epithelial barriers. Since mastitis results from infections by bacteria such as streptococci, staphylococci and coliform bacteria that cause neutrophilic inflammation, type 3 immunity can be expected to be mobilized at the mammary gland. In effect, the main defenses of this organ are provided by epithelial cells and neutrophils, which are the main terminal effectors of type 3 immunity. In addition to theoretical grounds, there is observational and experimental evidence that supports a role for type 3 immunity in the mammary gland, such as the production of IL-17A, IL-17F, and IL-22 in milk and mammary tissue during infection, although their respective sources remain to be fully identified. Moreover, mouse mastitis models have shown a positive effect of IL-17A on the course of mastitis. A lot remains to be uncovered before we can safely harness type 3 immunity to reinforce mammary gland defenses through innate immune training or vaccination. However, this is a promising way to find new means of improving mammary gland defenses against infection.
Asunto(s)
Inmunidad Adaptativa , Inmunidad Innata , Interleucina-17/inmunología , Mamíferos/inmunología , Glándulas Mamarias Animales/inmunología , Animales , FemeninoRESUMEN
The cytokine IL-17A has been shown to play critical roles in host defense against bacterial and fungal infections at different epithelial sites, but its role in the defense of the mammary gland (MG) has seldom been investigated, although infections of the MG constitute the main pathology afflicting dairy cows. In this study, we showed that IL-17A contributes to the defense of the MG against Escherichia coli infection by using a mouse mastitis model. After inoculation of the MG with a mastitis-causing E. coli strain, the bacterial load increased rapidly, triggering an intense influx of leukocytes into mammary tissue and increased concentrations of IL-6, IL-22, TNF-α, and IL-10. Neutrophils were the first cells that migrated intensely to the mammary tissue, in line with an early production of CXCL2. Depletion of neutrophils induced an increased mammary bacterial load. There was a significant increase of IL-17-containing CD4(+) αß T lymphocyte numbers in infected glands. Depletion of IL-17A correlated with an increased bacterial colonization and IL-10 production. Intramammary infusion of IL-17A at the onset of infection was associated with markedly decreased bacterial numbers, decreased IL-10 production, and increased neutrophil recruitment. Depletion of CD25(+) regulatory T cells correlated with a decreased production of IL-10 and a reduced bacterial load. These results indicate that IL-17A is an important effector of MG immunity to E. coli and suggest that an early increased local production of IL-17A would improve the outcome of infection. These findings point to a new lead to the development of vaccines against mastitis.
Asunto(s)
Infecciones por Escherichia coli/inmunología , Interleucina-17/inmunología , Mastitis/inmunología , Animales , Citocinas/análisis , Citocinas/biosíntesis , Citocinas/inmunología , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Escherichia coli , Femenino , Citometría de Flujo , Inmunohistoquímica , Glándulas Mamarias Animales/inmunología , Ratones , Ratones Endogámicos C57BLRESUMEN
Retinoic acid (RA) is of major importance during vertebrate embryonic development and its levels need to be strictly regulated otherwise congenital malformations will develop. Through the action of specific nuclear receptors, named RAR/RXR, RA regulates the expression of genes that eventually influence proliferation and tissue patterning. RA has been described as crucial for different stages of mammalian lung morphogenesis, and as part of a complex molecular network that contributes to precise organogenesis; nonetheless, nothing is known about its role in avian lung development. The current report characterizes, for the first time, the expression pattern of RA signaling members (stra6, raldh2, raldh3, cyp26a1, rarα, and rarß) and potential RA downstream targets (sox2, sox9, meis1, meis2, tgfß2, and id2) by in situ hybridization. In the attempt of unveiling the role of RA in chick lung branching, in vitro lung explants were performed. Supplementation studies revealed that RA stimulates lung branching in a dose-dependent manner. Moreover, the expression levels of cyp26a1, sox2, sox9, rarß, meis2, hoxb5, tgfß2, id2, fgf10, fgfr2, and shh were evaluated after RA treatment to disclose a putative molecular network underlying RA effect. In situ hybridization analysis showed that RA is able to alter cyp26a1, sox9, tgfß2, and id2 spatial distribution; to increase rarß, meis2, and hoxb5 expression levels; and has a very modest effect on sox2, fgf10, fgfr2, and shh expression levels. Overall, these findings support a role for RA in the proximal-distal patterning and branching morphogenesis of the avian lung and reveal intricate molecular interactions that ultimately orchestrate branching morphogenesis.
Asunto(s)
Pollos/metabolismo , Pollos/fisiología , Pulmón/metabolismo , Organogénesis/fisiología , Tretinoina/metabolismo , Animales , Regulación del Desarrollo de la Expresión Génica/fisiología , Hibridación in Situ/métodos , Transducción de Señal/fisiología , Factores de Transcripción/metabolismoRESUMEN
Lung organogenesis is guided by epithelial-mesenchymal interactions that coordinate cellular events responsible for the formation of the respiratory system. Several signaling pathways have been implicated in this process; among them, sonic hedgehog (Shh) signaling has emerged as a crucial regulator of branching morphogenesis in the mammalian lung. Canonical Shh signaling requires the presence of patched (Ptch) and smoothened (Smo) transmembrane receptors in order to induce the activation of glioblastoma (Gli) zinc finger transcription factors that are the true effectors of the pathway. Signal transduction is finely regulated by Ptch1, Gli, and Hhip (hedgehog-interacting protein). The present work characterizes, for the first time, the expression pattern of shh, ptch1, smo, gli1, and hhip in early stages of the embryonic chick lung. In situ hybridization studies revealed that these genes are expressed in the same cellular compartments as their mammalian counterparts, although their proximo-distal distribution is slightly changed. Moreover, the molecular interactions between fibroblast growth factor (FGF) and Shh signaling pathway were assessed, in vitro, by grafting beads soaked in SU5402 (an FGF receptor inhibitor). In the chick lung, Shh signaling seems to have some features that are species specific since shh is not a downstream target of FGF signaling. Nonetheless and despite the observed differences, these findings suggest a role for Shh signaling in the epithelial-mesenchymal interactions that control chick lung morphogenesis.
Asunto(s)
Pollos , Proteínas Hedgehog/análisis , Proteínas Hedgehog/metabolismo , Pulmón/embriología , Pulmón/metabolismo , Transducción de Señal , Animales , Proteínas Hedgehog/biosíntesisRESUMEN
Lung development is a very complex process that relies on the interaction of several signaling pathways that are controlled by precise regulatory mechanisms. Recently, microRNAs (miRNAs), small non-coding regulatory RNAs, have emerged as new players involved in gene expression regulation controlling several biological processes, such as cellular differentiation, apoptosis and organogenesis, in both developmental and disease processes. Failure to correctly express some specific miRNAs or a component of their biosynthetic machinery during embryonic development is disastrous, resulting in severe abnormalities. Several miRNAs have already been identified as modulators of lung development. Regarding the spatial distribution of the processing machinery of miRNAs, only two of its members (dicer1 and argonaute) have been characterized. The present work characterizes the expression pattern of drosha, dgcr8, exportin-5 and dicer1 in early stages of the embryonic chick lung by whole mount in situ hybridization and cross-section analysis. Overall, these genes are co-expressed in dorsal and distal mesenchyme and also in growing epithelial regions. The expression pattern of miRNA processing machinery supports the previously recognized regulatory role of this mechanism in epithelial and mesenchymal morphogenesis.
Asunto(s)
Pulmón/embriología , Pulmón/metabolismo , MicroARNs/metabolismo , Procesamiento Postranscripcional del ARN/genética , Animales , Proteínas Aviares/genética , Proteínas Aviares/metabolismo , Embrión de Pollo , Regulación del Desarrollo de la Expresión Génica , MicroARNs/genéticaRESUMEN
Mastitis remains a major disease of cattle with a strong impact on the dairy industry. There is a growing interest in understanding how cell mediated immunity contributes to the defence of the mammary gland against invading mastitis causing bacteria. Cytokines belonging to the IL-17 family, and the cells that produce them, have been described as important modulators of the innate immunity, in particular that of epithelial cells. We report here that expression of IL-17A and IL-17F genes, encoding two members of the IL-17 family, are induced in udder tissues of cows experimentally infected with Escherichia coli. The impact of IL-17A on the innate response of bovine mammary epithelial cells was investigated using a newly isolated cell line, the PS cell line. We first showed that PS cells, similar to primary bovine mammary epithelial cells, were able to respond to agonists of TLR2 and to LPS, provided CD14 was added to the culture medium. We then showed that secretion of CXCL8 and transcription of innate immunity related-genes by PS cells were increased by IL-17A, in particular when these cells were stimulated with live E. coli bacteria. Together with data from the literature, these results support the hypothesis that IL-17A and IL-17 F could play an important role in mediating of host-pathogen interactions during mastitis.
Asunto(s)
Infecciones por Escherichia coli/veterinaria , Escherichia coli/fisiología , Regulación de la Expresión Génica , Inmunidad Innata , Interleucina-17/genética , Mastitis Bovina/genética , Mastitis Bovina/inmunología , Animales , Bovinos , Línea Celular , Células Epiteliales/inmunología , Células Epiteliales/microbiología , Infecciones por Escherichia coli/genética , Infecciones por Escherichia coli/inmunología , Infecciones por Escherichia coli/microbiología , Femenino , Interleucina-17/metabolismo , Glándulas Mamarias Animales/inmunología , Glándulas Mamarias Animales/microbiología , Mastitis Bovina/microbiologíaRESUMEN
OBJECTIVES: The aims of this study were to compare laboratory indices of spontaneous bacterial peritonitis (SBP) and noninfected ascites in children with chronic liver disease and to determine the infectious agents involved in SBP. METHODS: The medical records of 90 children with chronic liver disease and ascites studied between January 2005 and August 2011 were reviewed for laboratory data of diagnostic significance in SBP. Standard laboratory tests included blood cell count, coagulation indices, liver and renal function tests, C-reactive protein (CRP), serum sodium concentration, serum albumin, and serum cultures. Ascitic fluid obtained from 152 paracentesis procedures was assayed for cytology, Gram stains, neutrophil counts, and bacteriological cultures. RESULTS: The SBP group manifested significantly lower albumin levels and elevated CRP levels, prothrombin times, international normalized ratios, and leukocyte number (P<0.05 in each case). CRP was shown to be an independent variable in the prediction of SBP. Values of serum creatinine, sodium concentration, urea, total bilirubin and differential leukocyte shift were comparable in SBP and noninfected ascites. Streptococcus pneumoniae was the most prevalent infectious agent in the ascitic fluid (44%). CONCLUSIONS: CRP may be useful in early detection and monitoring of SBP in children with liver disease.
Asunto(s)
Ascitis/metabolismo , Líquido Ascítico/microbiología , Infecciones Bacterianas/metabolismo , Proteína C-Reactiva/metabolismo , Cirrosis Hepática/metabolismo , Peritonitis/metabolismo , Streptococcus pneumoniae , Adolescente , Ascitis/microbiología , Infecciones Bacterianas/microbiología , Infecciones Bacterianas/patología , Niño , Preescolar , Femenino , Humanos , Lactante , Relación Normalizada Internacional , Recuento de Leucocitos , Cirrosis Hepática/patología , Masculino , Paracentesis , Peritonitis/microbiología , Peritonitis/patología , Tiempo de Protrombina , Albúmina Sérica/metabolismoRESUMEN
Mastitis caused by Escherichia coli and Staphylococcus aureus is a major pathology of dairy cows. To better understand the differential response of the mammary gland to these two pathogens, we stimulated bovine mammary epithelial cells (bMEC) with either E. coli crude lipopolysaccharide (LPS) or with S. aureus culture supernatant (SaS) to compare the transcriptomic profiles of the initial bMEC response. By using HEK 293 reporter cells for pattern recognition receptors, the LPS preparation was found to stimulate TLR2 and TLR4 but not TLR5, Nod1 or Nod2, whereas SaS stimulated TLR2. Biochemical analysis revealed that lipoteichoic acid, protein A and α-hemolysin were all present in SaS, and bMEC were found to be responsive to each of these molecules. Transcriptome profiling revealed a core innate immune response partly shared by LPS and SaS. However, LPS induced expression of a significant higher number of genes and the fold changes were of greater magnitude than those induced by SaS. Microarray data analysis suggests that the activation pathways and the early chemokine and cytokine production preceded the defense and stress responses. A major differential response was the activation of the type I IFN pathway by LPS but not by SaS. The higher upregulation of chemokines (Cxcl10, Ccl2, Ccl5 and Ccl20) that target mononuclear leucocytes by LPS than by SaS is likely to be related to the differential activation of the type I IFN pathway, and could induce a different profile of the initial recruitment of leucocytes. The MEC responses to the two stimuli were different, as LPS was associated with NF-κB and Fas signaling pathways, whereas SaS was associated with AP-1 and IL-17A signaling pathways. It is noteworthy that at the protein level secretion of TNF-α and IL-1ß was not induced by either stimulus. These results suggest that the response of MEC to diffusible stimuli from E. coli and S. aureus contributes to the onset of the response with differential leucocyte recruitment and distinct inflammatory and innate immune reactions of the mammary gland to infection.
Asunto(s)
Infecciones por Escherichia coli/inmunología , Inmunidad Innata , Péptidos y Proteínas de Señalización Intracelular/genética , Glándulas Mamarias Animales/inmunología , Mastitis Bovina/inmunología , Infecciones Estafilocócicas/inmunología , Receptores Toll-Like/genética , Animales , Bovinos , Células Cultivadas , Quimiocinas/genética , Quimiocinas/metabolismo , Citocinas/genética , Citocinas/metabolismo , Ensayo de Inmunoadsorción Enzimática/veterinaria , Células Epiteliales/inmunología , Células Epiteliales/microbiología , Escherichia coli/fisiología , Infecciones por Escherichia coli/genética , Infecciones por Escherichia coli/microbiología , Femenino , Perfilación de la Expresión Génica/veterinaria , Regulación de la Expresión Génica , Células HEK293 , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Lipopolisacáridos/fisiología , Glándulas Mamarias Animales/microbiología , Mastitis Bovina/genética , Mastitis Bovina/microbiología , Proteínas Adaptadoras de Señalización NOD/genética , Proteínas Adaptadoras de Señalización NOD/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos/veterinaria , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/veterinaria , Transducción de Señal , Infecciones Estafilocócicas/genética , Infecciones Estafilocócicas/microbiología , Staphylococcus aureus/fisiología , Receptores Toll-Like/metabolismoRESUMEN
Breast cancer (BC) is a complex disease and obesity is a well-known risk factor for its development, especially after menopause. Several studies have shown Single Nucleotide Polymorphisms (SNPs) linked to overweight and obesity, such as: rs1121980 (T/C) and rs9939609 (A/T) in Fat Mass and Obesity Associated gene (FTO) and rs17782313 (T/C) in Melanocortin 4 Receptor gene (MC4R). Thus, we aimed to investigate the association between these obesity-related SNPs and BC risk. One hundred BC patients and 148 healthy women from Santa Catarina, Brazil entered the study. SNPs were genotyped using Taqman assays. For statistical analyses SNPStats and SPSS softwares were used. Association analyses were performed by logistic regression and were adjusted for age and Body mass index (BMI). Multiple SNPs inheritance models (log-additive, dominant, recessive, codominant) were performed to determine odds ratios (ORs), assuming 95 % confidence interval (CI) and P value = 0.05 as the significance limit. When analyzed alone, FTO rs1121980 and rs9939609 did not show significant associations with BC development, however MC4R rs17782313 showed increased risk for BC even after adjustments (P-value = 0.032). Interestingly, the interaction of FTO and MC4R polymorphisms showed a powerful association with BC. We observed a 4.59-fold increased risk for woman who have the allele combination C/T/C (FTO rs1121980/FTO rs9939609/MC4R rs17782313) (P-value = 0.0011, adjusted for age and BMI). We found important and unpublished associations between these obesity-related genes and BC risk. These associations seem to be independent of their effect on BMI, indicating a direct role of the interaction between FTO and MC4R polymorphisms in BC development.
Asunto(s)
Neoplasias de la Mama/genética , Epistasis Genética , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Obesidad/genética , Proteínas/genética , Receptor de Melanocortina Tipo 4/genética , Alelos , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato , Antropometría , Neoplasias de la Mama/patología , Estudios de Casos y Controles , Femenino , Frecuencia de los Genes/genética , Humanos , Menopausia , Polimorfismo de Nucleótido Simple/genética , Factores de Riesgo , Transducción de Señal/genéticaRESUMEN
Linalool is a monoterpene alcohol and constituent of several Brazilian aromatic medicinal plants, popularly used against hypertension. Cardiovascular effects induced by linalool were evaluated. In normotensive rats, (+/-)-linalool [1, 5, 10, and 20 mg/kg body weight (BW); intravenous (i.v.)]-induced hypotension was associated with tachycardia, which was attenuated by atropine (2 mg/kg BW) and N(G)-nitro-L-arginine methyl ester (20 mg/kg BW), but was not modified after indomethacin (5 mg/kg BW) administration. In hypertensive rats, linalool [200 mg/kg BW; oral (v.o.)] reduced blood pressure without changing the heart rate. In intact rings of rat mesenteric artery precontracted with 10 microM phenylephrine, linalool (from 6.4 x 10(-6) to 6.4 x 10(-3) M) induced relaxations in a concentration-dependent manner [E(max) = (115 +/- 13)%] that were not changed after atropine administration [E(max) = (105 +/- 2)%], and were not different from those obtained in endothelium-denuded rings precontracted with phenylephrine [E(max) = (108 +/- 7)%] or 80 mM KCl [E(max) = (113 +/- 7)%] or tetraethylammonium incubation [E(max) = (105 +/- 12)%]. Linalool (1.9 x 10(-3) M) antagonized the contractions induced by CaCl2 (3 x 10(-6)-10(-2) M) (maximal inhibition, 81%). Furthermore, linalool inhibited the contractions induced by 10 microM phenylephrine or 20 mM caffeine. In conclusion, these results demonstrate that linalool reduces blood pressure probably due to a direct effect on the vascular smooth muscle leading to vasodilation.
Asunto(s)
Sistema Cardiovascular/efectos de los fármacos , Hipertensión/tratamiento farmacológico , Monoterpenos/farmacología , Monoterpenos Acíclicos , Animales , Aorta/efectos de los fármacos , Aorta/fisiología , Presión Sanguínea/efectos de los fármacos , Frecuencia Cardíaca/efectos de los fármacos , Técnicas In Vitro , Masculino , Monoterpenos/uso terapéutico , Ratas , Ratas WistarRESUMEN
T-lymphocytes are key mediators of adaptive cellular immunity and knowledge about distinct subsets of these cells in healthy and infected mammary gland secretions remains limited. In this study, we used a multiplex cytometry panel to show that staphylococcal mastitis causes the activation of CD4+, CD8+ and γδ T-cells found in bovine milk. We also highlight remarkable differences in the proportions of naïve and memory T-cells subsets found in blood and milk. These observations will contribute to a better understanding of cell-mediated immune mechanisms in the udder and to the development of new therapeutic and preventive strategies targeting mastitis.
Asunto(s)
Mastitis Bovina , Leche , Humanos , Femenino , Animales , Bovinos , Staphylococcus aureus , Subgrupos de Linfocitos T , Diferenciación Celular , Glándulas Mamarias AnimalesRESUMEN
BACKGROUND: Mammary gland (MG) infections (mastitis) are frequent diseases of dairy cows that affect milk quality, animal welfare and farming profitability. These infections are commonly associated with the bacteria Escherichia coli and Staphylococcus aureus. Different in vitro models have been used to investigate the early response of the MG to bacteria, but the role of the teat in mastitis pathogenesis has received less attention. In this study, we used punch-excised teat tissue as an ex vivo model to study the immune mechanisms that arise early during infection when bacteria have entered the MG. RESULTS: Cytotoxicity and microscopic analyses showed that bovine teat sinus explants have their morphology and viability preserved after 24 h of culture and respond to ex vivo stimulation with TLR-agonists and bacteria. LPS and E. coli trigger stronger inflammatory response in teat when compared to LTA and S. aureus, leading to a higher production of IL-6 and IL-8, as well as to an up-regulation of proinflammatory genes. We also demonstrated that our ex vivo model can be applied to frozen-stored explants. CONCLUSIONS: In compliance with the 3Rs principle (replacement, reduction and refinement) in animal experimentation, ex vivo explant analyses proved to be a simple and affordable approach to study MG immune response to infection. This model, which better reproduces organ complexity than epithelial cell cultures or tissue slices, lends itself particularly well to studying the early phases of the MG immune response to infection.
RESUMEN
Staphylococcus aureus is a prevalent pathogen for mastitis in dairy ruminants and is responsible for both clinical and subclinical mastitis. Mammary epithelial cells (MEC) represent not only a physical barrier against bacterial invasion but are also active players of the innate immune response permitting infection clearance. To decipher their functions in general and in animals showing different levels of genetic predisposition to Staphylococcus in particular, MEC from ewes undergoing a divergent selection on milk somatic cell count were stimulated by S. aureus. MEC response was also studied according to the stimulation condition with live bacteria or culture supernatant. The early MEC response was studied during a 5 h time course by microarray to identify differentially expressed genes with regard to the host genetic background and as a function of the conditions of stimulation. In both conditions of stimulation, metabolic processes were altered, the apoptosis-associated pathways were considerably modified, and inflammatory and immune responses were enhanced with the upregulation of il1a, il1b, and tnfa and several chemokines known to enhance neutrophil (cxcl8) or mononuclear leukocyte (ccl20) recruitment. Genes associated with oxidative stress were increased after live bacteria stimulation, whereas immune response-related genes were higher after supernatant stimulation in the early phase. Only 20 genes were differentially expressed between Staphylococcus spp-mastitis resistant and susceptible animals without any clearly defined role on the control of infection. To conclude, this suggests that MEC may not represent the cell type at the origin of the difference of mastitis susceptibility, at least as demonstrated in our genetic model. Supernatant or heat-killed S. aureus produce biological effects that are essentially different from those induced by live bacteria.
Asunto(s)
Células Epiteliales/metabolismo , Perfilación de la Expresión Génica , Predisposición Genética a la Enfermedad , Glándulas Mamarias Animales/patología , Mastitis/veterinaria , Ovinos/genética , Staphylococcus aureus/fisiología , Animales , Análisis por Conglomerados , Células Epiteliales/microbiología , Células Epiteliales/patología , Femenino , Redes Reguladoras de Genes/genética , Glándulas Mamarias Animales/microbiología , Mastitis/genética , Mastitis/microbiología , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ovinos/microbiología , Fracciones Subcelulares/metabolismoRESUMEN
Escherichia coli is a frequent cause of clinical mastitis in dairy cows. It has been shown that a prompt response of the mammary gland after E. coli entry into the lumen of the gland is required to control the infection, which means that the early detection of bacteria is of prime importance. Yet, apart from lipopolysaccharide (LPS), little is known of the bacterial components which are detected by the mammary innate immune system. We investigated the repertoire of potential bacterial agonists sensed by the udder and bovine mammary epithelial cells (bMEC) during E. coli mastitis by using purified or synthetic molecular surrogates of bacterial agonists of identified pattern-recognition receptors (PRRs). The production of CXCL8 and the influx of leucocytes in milk were the readouts of reactivity of stimulated cultured bMEC and challenged udders, respectively. Quantitative PCR revealed that bMEC in culture expressed the nucleotide oligomerization domain receptors NOD1 and NOD2, along with the Toll-like receptors TLR1, TLR2, TLR4, and TLR6, but hardly TLR5. In line with expression data, bMEC proved to react to the cognate agonists C12-iE-DAP (NOD1), Pam3CSK4 (TLR1/2), Pam2CSK4 (TLR2/6), pure LPS (TLR4), but not to flagellin (TLR5). As the udder reactivity to NOD1 and TLR5 agonists has never been reported, we tested whether the mammary gland reacted to intramammary infusion of C12-iE-DAP or flagellin. The udder reacted to C12-iE-DAP, but not to flagellin, in line with the reactivity of bMEC. These results extend our knowledge of the reactivity of the bovine mammary gland to bacterial agonists of the innate immune system, and suggest that E. coli can be recognized by several PRRs including NOD1, but unexpectedly not by TLR5. The way the mammary gland senses E. coli is likely to shape the innate immune response and finally the outcome of E. coli mastitis.
Asunto(s)
Proteínas Adaptadoras de Señalización CARD/genética , Células Epiteliales/inmunología , Infecciones por Escherichia coli/veterinaria , Proteínas de Escherichia coli/metabolismo , Glándulas Mamarias Animales/inmunología , Mastitis Bovina/inmunología , Receptores Toll-Like/genética , Animales , Proteínas Adaptadoras de Señalización CARD/agonistas , Proteínas Adaptadoras de Señalización CARD/metabolismo , Bovinos , Células Cultivadas , Células Epiteliales/microbiología , Escherichia coli/inmunología , Infecciones por Escherichia coli/inmunología , Infecciones por Escherichia coli/microbiología , Femenino , Inmunidad Innata , Glándulas Mamarias Animales/microbiología , Mastitis Bovina/microbiología , Reacción en Cadena de la Polimerasa/veterinaria , Receptores Toll-Like/agonistas , Receptores Toll-Like/metabolismoRESUMEN
Dendritic cells are sentinels of the immune system responsible for the initiation of adaptive immune mechanisms. In that respect, the study of these cells is essential for a full understanding of host response to infectious agents and vaccines. In ruminants, the large blood volume facilitates the isolation of abundant monocytes and their derivation to other antigen-presenting cells such as dendritic cells and macrophages. However, the available protocols for the production of bovine monocyte-derived dendritic cells (moDCs) rely mostly on time-consuming and costly techniques such as density gradient centrifugation and magnetic sorting of cells. In this study, we describe a simplified protocol for the production of bovine moDC using conventional and serum-free media. We also employ moDC produced by this approach to carry out a flow cytometry-based antigen presentation assay adapted to blood fresh or frozen cells. The experimental strategies described here might enable the setup of studies involving a large number of individuals, requiring a large number of dendritic cells, or relying on the utilization of cryopreserved blood cells. These simplified protocols might contribute to the elucidation of cell-mediated immune responses in bovine.
RESUMEN
Aim: This case report describes a protocol for restoring a crown fracture of an unerupted permanent incisor in a child. Background: Crown fractures are an important concern in pediatric dentistry due to the negative impact on oral health-related quality of life (OHRQoL) in children and adolescents resulting from functional limitations as well as consequences related to social and emotional well-being. Case description: An enamel and dentin fracture of the crown of unerupted tooth 11 due to direct trauma is being presented in a 7-year-old girl. The restorative treatment involved minimally invasive dentistry, including computer-aided design (CAD)/computer-aided manufacturing (CAM) technology and direct resin restoration. Conclusion: The treatment decision was essential for maintaining pulp vitality and continued root development, as well as ensuring esthetic and functional results. Clinical significance: Crown fracture of an unerupted incisor may occur in childhood, requiring a long-term clinical and radiographic follow-up. Predictable, positive, and reliable esthetic outcomes can be achieved using CAD/CAM technology combined with adhesive protocols. How to cite this article: Kamanski D, Tavares JG, Weber JBB, et al. Crown Fracture of an Unerupted Incisor in a Young Child: Case Report and Restorative Protocol. Int J Clin Pediatr Dent 2022;15(5):636-641.
RESUMEN
Breast cancer (BC) is one of the leading causes of cancer mortality in women worldwide, and therefore, novel biomarkers for early disease detection are critically needed. We performed herein an untargeted plasma metabolomic profiling of 55 BC patients and 55 healthy controls (HC) using ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC/Q-TOF-MS). Pre-processed data revealed 2494 ions in total. Data matrices' paired t-tests revealed 792 ions (both positive and negative) which presented statistically significant changes (FDR < 0.05) in intensity levels between cases versus controls. Metabolites identified with putative names via MetaboQuest using MS/MS and mass-based approaches included amino acid esters (i.e., N-stearoyl tryptophan, L-arginine ethyl ester), dipeptides (ile-ser, met-his), nitrogenous bases (i.e., uracil derivatives), lipid metabolism-derived molecules (caproleic acid), and exogenous compounds from plants, drugs, or dietary supplements. LASSO regression selected 16 metabolites after several variables (TNM Stage, Grade, smoking status, menopausal status, and race) were adjusted. A predictive conditional logistic regression model on the 16 LASSO selected ions provided a high diagnostic performance with an area-under-the-curve (AUC) value of 0.9729 (95% CI 0.96−0.98) on all 55 samples. This study proves that BC possesses a specific metabolic signature that could be exploited as a novel metabolomics-based approach for BC detection and characterization. Future studies of large-scale cohorts are needed to validate these findings.
RESUMEN
Species of Erythroxylum genus are popularly used as anti-inflammatories and in the treatment of renal and respiratory disorders. Although it has been reported that species from the Erythroxylum genus induce cardiovascular effects, E. passerinum had not been studied specifically in this respect. However, previous phytochemical studies of E. passerinum demonstrated the presence of compounds which can have potential activity on the cardiovascular system. In this study, phytochemical screening of the ethanol extract of E. passerinum (EEEP) detected polyphenols, but not alkaloids. EEEP caused hypotension, bradycardia and vasorelaxation in rats. The vasorelaxation was attenuated by Nw-nitro-L-arginine methyl ester (L-NAME) or L-NAME + indomethacin (INDO), but not by INDO alone. Vasorelaxation was also significantly attenuated after endothelium removal or after incubation with high K+, 4-aminopyridine, glibenclamide or tetraethylammonium, but was not affected by pre-contraction with serotonin. Thus, EEEP induces hypotension and endothelium-dependent and independent vasorelaxation, which seems to involve the nitric oxide and K+-channels.