Latent TGF-ß binding protein 3 identifies a second heart field in zebrafish.

The four-chambered mammalian heart develops from two fields of cardiac progenitor cells distinguished by their spatiotemporal patterns of differentiation and contributions to the definitive heart. The first heart field differentiates earlier in lateral plate mesoderm, generates the linear heart tube and ultimately gives rise to the left ventricle. The second heart field (SHF) differentiates later in pharyngeal mesoderm, elongates the heart tube, and gives rise to the outflow tract and much of the right ventricle. Because hearts in lower vertebrates contain a rudimentary outflow tract but not a right ventricle, the existence and function of SHF-like cells in these species has remained a topic of speculation. Here we provide direct evidence from Cre/Lox-mediated lineage tracing and loss-of-function studies in zebrafish, a lower vertebrate with a single ventricle, that latent TGF-ß binding protein 3 (ltbp3) transcripts mark a field of cardiac progenitor cells with defining characteristics of the anterior SHF in mammals. Specifically, ltbp3(+) cells differentiate in pharyngeal mesoderm after formation of the heart tube, elongate the heart tube at the outflow pole, and give rise to three cardiovascular lineages in the outflow tract and myocardium in the distal ventricle. In addition to expressing Ltbp3, a protein that regulates the bioavailability of TGF-ß ligands, zebrafish SHF cells co-express nkx2.5, an evolutionarily conserved marker of cardiac progenitor cells in both fields. Embryos devoid of ltbp3 lack the same cardiac structures derived from ltbp3(+) cells due to compromised progenitor proliferation. Furthermore, small-molecule inhibition of TGF-ß signalling phenocopies the ltbp3-morphant phenotype whereas expression of a constitutively active TGF-ß type I receptor rescues it. Taken together, our findings uncover a requirement for ltbp3-TGF-ß signalling during zebrafish SHF development, a process that serves to enlarge the single ventricular chamber in this species.

Zebrafish second heart field development relies on progenitor specification in anterior lateral plate mesoderm and nkx2.5 function.

Second heart field (SHF) progenitors perform essential functions during mammalian cardiogenesis. We recently identified a population of cardiac progenitor cells (CPCs) in zebrafish expressing latent TGFß-binding protein 3 (ltbp3) that exhibits several defining characteristics of the anterior SHF in mammals. However, ltbp3 transcripts are conspicuously absent in anterior lateral plate mesoderm (ALPM), where SHF progenitors are specified in higher vertebrates. Instead, ltbp3 expression initiates at the arterial pole of the developing heart tube. Because the mechanisms of cardiac development are conserved evolutionarily, we hypothesized that zebrafish SHF specification also occurs in the ALPM. To test this hypothesis, we Cre/loxP lineage traced gata4(+) and nkx2.5(+) ALPM populations predicted to contain SHF progenitors, based on evolutionary conservation of ALPM patterning. Traced cells were identified in SHF-derived distal ventricular myocardium and in three lineages in the outflow tract (OFT). We confirmed the extent of contributions made by ALPM nkx2.5(+) cells using Kaede photoconversion. Taken together, these data demonstrate that, as in higher vertebrates, zebrafish SHF progenitors are specified within the ALPM and express nkx2.5. Furthermore, we tested the hypothesis that Nkx2.5 plays a conserved and essential role during zebrafish SHF development. Embryos injected with an nkx2.5 morpholino exhibited SHF phenotypes caused by compromised progenitor cell proliferation. Co-injecting low doses of nkx2.5 and ltbp3 morpholinos revealed a genetic interaction between these factors. Taken together, our data highlight two conserved features of zebrafish SHF development, reveal a novel genetic relationship between nkx2.5 and ltbp3, and underscore the utility of this model organism for deciphering SHF biology.

Tbx1 is required for second heart field proliferation in zebrafish.

BACKGROUND: The mammalian outflow tract (OFT) and primitive right ventricle arise by accretion of newly differentiated cells to the arterial pole of the heart tube from multi-potent progenitor cells of the second heart field (SHF). While mounting evidence suggests that the genetic pathways regulating SHF development are highly conserved in zebrafish, this topic remains an active area of investigation. RESULTS: Here, we extend previous observations demonstrating that zebrafish tbx1 (van gogh, vgo) mutants show ventricular and OFT defects consistent with a conserved role in SHF-mediated cardiogenesis. Surprisingly, we reveal through double in situ analyses that tbx1 transcripts are excluded from cardiac progenitor cells and differentiated cardiomyocytes, suggesting a non-autonomous role in SHF development. Further, we find that the diminutive ventricle in vgo animals results from a 25% decrease in cardiomyocyte number that occurs subsequent to heart tube stages. Lastly, we report that although SHF progenitors are specified in the absence of Tbx1, they fail to be maintained due to compromised SHF progenitor cell proliferation. CONCLUSIONS: These studies highlight conservation of Tbx1 function in zebrafish SHF biology.

Leptin receptor expression in blood mononuclear cells of lactating women is associated with infant body weight: Potential role as a molecular biomarker.

BACKGROUND: Molecular biomarkers of maternal leptin resistance associated with infant weight are needed. OBJECTIVES: To evaluate gene expression of leptin receptor (LEPR), suppressor of cytokine signalling 3 (SOCS3) and insulin receptor in peripheral blood mononuclear cells (PBMCs) of lactating women and their relationship with infant body weight and adiposity. METHODS: At day 10 postpartum, maternal gene expression in PBMCs as well as leptin and insulin concentrations in plasma and milk were assessed (n = 68). Infant weight and BMI z-scores, skinfolds and arm circumference were obtained at 10 days and/or at 3 months old. RESULTS: In mothers with pre-pregnancy overweight or obesity (OW/OB), LEPR expression was reduced (p = 0.013) whereas plasma and milk leptin and milk insulin concentrations were elevated. LEPR expression was positively related with infant weight z-score (Beta (95% CI): 0.40 (0.17, 0.63), p = 0.001) but not with leptin concentrations. SOCS3 expression was positively related with infant weight z-score (Beta (95% CI): 0.28 (0.04, 0.51), p = 0.024) and arm circumference (Beta (95% CI): 0.57 (0.32, 0.82), p < 0.001). Relationships remained significant after adjusting for maternal and infant confounders. CONCLUSIONS: LEPR and SOCS3 gene expression in PBMCs are novel maternal molecular biomarkers that reflect leptin resistance and are associated with infant body weight and adiposity.

A biosecurity survey on Iberian pig farms in Spain: Farmers' attitudes towards the implementation of measures and investment.

The Iberian pork industry has proven to be very important in South-Western Europe, not only due its contribution to the economy of the area, but also because of the peculiarities of the farms situated there. These farms raise Iberian pigs in systems ranging from fully outdoor rearing systems to intensive-rearing farms similar to the conventional pig production models. The implementation of biosecurity measures in Iberian pig farms can be complex and, at times, it can be perceived somewhat negatively by farmers, especially those who run more extensive pig production systems. This research attempts to understand the attitudes of Iberian pig farmers in the South-West of Spain towards the adoption of and investment in biosecurity measures. For that purpose, data obtained from 104 online surveys taken by Iberian pig farmers in 2019 was analysed. A correspondence and a cluster analysis were performed, whereby three farm groups were identified according to their features (land area, breeding conditions, availability of facilities, etc.), the existence/use of biosecurity facilities and the adoption of biosecurity measures. The results have shown that most of the analysed farms have the necessary infrastructure to comply with the biosecurity principles and farmers perform basic biosecurity practices. However, farms that are more extensive, mainly those finishing their pigs outdoors (acorn-feed), need to improve their biosecurity. It is necessary therefore to implement an improvement plan following the current regulations in collaboration with policy makers with the purpose of improving the levels of biosecurity and increase disease risk awareness.

Pectin activation of MAP kinase and gene expression is WAK2 dependent.

The angiosperm extracellular matrix, or cell wall, is composed of a complex array of cellulose, hemicellulose, pectins and proteins, the modification and regulated synthesis of which are essential for cell growth and division. The wall associated kinases (WAKs) are receptor-like proteins that have an extracellular domain that bind pectins, the more flexible portion of the extracellular matrix, and are required for cell expansion as they have a role in regulating cellular solute concentrations. We show here that both recombinant WAK1 and WAK2 bind pectin in vitro. In protoplasts pectins activate, in a WAK2-dependent fashion, the transcription of vacuolar invertase, and a wak2 mutant alters the normal pectin regulation of mitogen-activated protein kinases. Microarray analysis shows that WAK2 is required for the pectin activation of numerous genes in protoplasts, many of which are involved in cell wall biogenesis. Thus, WAK2 plays a major role in signaling a diverse array of cellular events in response to pectin in the extracellular matrix.

Carbon nanotube integration with a CMOS process.

This work shows the integration of a sensor based on carbon nanotubes using CMOS technology. A chip sensor (CS) was designed and manufactured using a 0.30 µm CMOS process, leaving a free window on the passivation layer that allowed the deposition of SWCNTs over the electrodes. We successfully investigated with the CS the effect of humidity and temperature on the electrical transport properties of SWCNTs. The possibility of a large scale integration of SWCNTs with CMOS process opens a new route in the design of more efficient, low cost sensors with high reproducibility in their manufacture.

Side effects of cyclosporine-A treatment in rats: gingival overgrowth and early hyperglycemia.

Gingival overgrowth is an adverse side effect of cyclosporine A (CsA) in the treatment of transplanted patients. The purpose of this study was to evaluate the effects of CsA on new-onset diabetes mellitus and gingival overgrowth in rats, by measuring collagen, nitric oxide and microvascular permeability. Blood glucose level, collagen, nitric oxide level and vascular permeability were determined. Blood glucose level increased significantly from 6.5 +/- 0.9 for the control group to 15 +/- 1.2, 17 +/- 1.2 and 21.6 +/- 1.6 mM/L at 1, 4 or 8 weeks of CsA treatment, respectively. Collagen (ug HO Proline/mg p) increased significantly from 2.5 +/- 0.5 for the control group to 4.2 +/- 0.8, 5.9 +/- 0.6 and 7.3 +/- 0.8 at 1, 4 or 8 weeks of CsA treatment, respectively. Vascular permeability was 10.3 +/- 1.2 for the control group and 15 +/- 1; 17.2 +/- 1.3, and 22.1 +/- 2.1 ug EB/g T; at 1, 4 or 8 weeks of CsA treatment, respectively Nitric oxide level was 3.5 +/- .9 umol/mg P for the control group and 4 +/- 0.2, 8.2 +/- 0.9 and 11 +/- 1 for 1, 2 or 8 weeks of CsA treatment, respectively. These findings appear to indicate that the development of significant gingival changes induced by CsA is related to new-onset of diabetes mellitus during the immunosuppressive treatment.

Antidiabetic efficacy of bradykinin antagonist R-954 on glucose tolerance test in diabetic type 1 mice.

Insulin-dependent diabetes mellitus (type 1 diabetes) is an inflammatory autoimmune disease associated with many complications including nephropathy, retinopathy, neuropathy and hyperalgesia. Experimental evidence has shown that the bradykinin B1 receptor (BKB1-R) is involved in the development of type 1 diabetes and found to be upregulated alongside the disease. In the present study the effects of the selective BKB1-R antagonist the R-954 (Ac-Orn-[Oic(2), alpha-MePhe(5), D-beta Nal(7), Ile(8) ]des-Arg(9)-BK and the BKB1-R agonist des Arg(9)-BK (DBK) were studied on diabetic hyperglycemia. Diabetic type 1 was induced in C57 BL/KsJ mdb male mice by five consecutives doses of STZ (45mg/kg i.p.). A glucose tolerance test (GTT) was performed by an intraperitoneal administration of glucose, 8, 12 and 18days after the diabetes induction. The induction of type 1 diabetes provoked a significant hyperglycemia levels in diabetic mice at 12 and 18days after STZ. The administration of R-954 (400microg/kg i.p.) at 12 and 18days after STZ returned the glycemia levels of this animals to normal values. In addition the administration of DKB (300microg/kg i.p.) significantly potentiated the diabetes-induced hyperglycemia; this effect that was totally reversed by R-954. These results provide further evidence for the implication of BKB1-R in the type 1 diabetes mellitus (insulitis).

Side effects of cyclosporine-A treatment in rats: gingival overgrowth and early hyperglycemia

Gingival overgrowth is an adverse side effect of cyclosporine A (CsA) in the treatment of transplanted patients. The purpose of this study was to evaluate the effects of CsA on new-onset diabetes mellitus and gingival overgrowth in rats, by measuring collagen, nitric oxide and microvascular permeability. Blood glucose level, collagen, nitric oxide level and vascular permeability were determined. Blood glucose level increased significantly from 6.5 +/- 0.9 for the control group to 15+/- 1.2, 17 +/- 1.2 and 21.6+/- 1.6 mM/L at 1, 4 or 8 weeks of CsA treatment, respectively. Collagen (ug HO Proline/mg p) increased significantly from 2.5+/- 0.5 for the control group to 4.2+/- 0.8, 5.9+/- 0.6 and 7.3 +/- 0.8 at 1, 4 or 8 weeks of CsA treatment, respectively. Vascular permeability was 10.3+/- 1.2 for the control group and 15+/-1; 17.2 +/- 1.3, and 22.1+/- 2.1 ug EB/g T; at 1, 4 or 8 weeks of CsA treatment, respectively. Nitric oxide level was 3.5 +/- .9 umol/mg P for the control group and 4+/- 0.2, 8.2+/- 0.9 and 11+/-1 for 1, 2 or 8 weeks of CsA treatment, respectively. These findings appear to indicate that the development of significant gingival changes induced by CsA is related to new-onset of diabetes mellitus during the immunosuppressive treatment.

La hiperplasia gingival es un efecto colateral adverso del tratamiento con ciclosporina A (CsA) en pacientes transplantados. El proposito de este estudio fue evaluar el efecto de CsA en el inicio de diabetes mellitus, la concentracion de colageno, y de oxido nitrico y la permeabilidad capilar gingival. El nivel de glucosa en sangre de los animales controles fue: 6.5+/- 0.9, en tanto que los tratados con CsA fue: 15+/-1.2; 17+/- 1.1 y 21.6+/- 1.6 mM/L a las 1, 4 y 8 semanas respectivamente. El colageno (ug OH prolina/mg p) mostro un aumento significativo en los animales tratados con CsA respecto de los controles: 2.5+/- 0.5; 4.2+/- 0.8; 5.9+/- 0.6; 7.3+/- 0.8 respectivamente a las 1,4 y 8 semanas de tratamiento. Los valores de permeabilidad capilar (ug AE/ g T) fueron: en los animales control 10.3+./- 1.2; en los animales tratados con CsA, a las 1, 4 y 8 semanas 15+/- 1.0; 17.2 +/- 1.3 y 22.1+/- 2.1 respectivamente. Los valores de oxido nitrico (umol/mg p) en los animales control: 3.5+/-0.9; y en los animales tratados con CsA 4+/- 0.2; 8.2+/- 0.9 y 11.2 +/- 1.0 respectivamente. Estos resultados parecen indicar que el desarrollo de los significativos cambios gingivales inducidos por la administracion de CsA esta relacionado con la hiperglucemia temprana que se asocia al tratamiento con inmunosupresores.

Una opción más en la repación de la hernia inguinal / Another alternative for inguinal hernia repair

Objetivo: Brindar una opción más para la reparación de las hernias inguinales. Material y métodos: De abril de 1986 a abril de 1996 se realizaron 312 hernioplastias con colocación de material sintético; 271 fueron reparaciones primarias y 41 fueron reparaciones de hernias recidivantes. Todas eran hernias directas, 271 directas y 41 recividantes, clasificadas por el método de Gilbert. Se realizó abordaje anterior, utilizando el ligamento de Cooper y el tracto iliopúbico, formando la malla de polipropileno parte de la plastia misma, quedando doblada sobre ella misma (en un doblez) y no como parche o empastamiento sobre el defecto herniario. Resultados: Se presentó recidiva en 0.32 por ciento (un caso) en un paciente con hernia recidivante por tercera ocasión, a los 28 meses de su cirugía, coincidiendo con una resección transuretral de próstata. Las complicaciones se limitaron a ocho seromas y nueve hematomas subdérmicos (5.29 por ciento). El promedio de estancia hospitalaria fue de 28 horas. El seguimiento máximo de los pacientes fue de cinco años, con un promedio de cuatro años. Conclusiones: Creemos firmemente que esta es una excelente alternativa y este reporte así lo demuestra. Sin embargo, sólo a través del tiempo podremos contestar la pregunta de ¿cuál es el método ideal de reparación en una hernia inguinal? Es indispensable el uso de material sintético (polipropileno) en la hernioplastia inguinal para evitar recidivas tardías.

Tratamiento quirúrgico de la enfermedad por reflujo gastroesofágico / Surgical treatment for gastroesophageal reflux disease

Objetivo: Evaluar el tratamiento quirúrgico laparoscópico de la enfermedad por reflujo gastroesofágico, la efectividad y las complicaciones, por un mismo equipo quirúrgico. Diseño: Estudio retrospectivo y longitudinal. Material y método: Se analizaron los resultados de 129 pacientes con enfermedad por reflujo gastroesofágico operados mediante laparoscopia entre enero de 1992 y junio del 2001. Resultados: Fueron operados 89 hombres y 48 mujeres, con edad media de 41.5 años. En el 100 por ciento se llevó a cabo panendoscopia, en 78 por ciento manometrías y en 21 por ciento pHmetrías. Se efectuaron cuatro funduplicaturas a 360º y 125 funduplicaturas posteriores a 270º. El tiempo de seguimiento fue de uno a 12 meses. Se presentaron complicaciones mayores en dos pacientes y menores en 17. El tiempo quirúrgico promedio fue de 71 minutos. Sólo hubo una conversión. El tiempo promedio de estancia hospitalaria fue de 31.30 horas. Conclusión: El tratamiento quirúrgico por vía laparoscópica es seguro, eficiente y ofrece resultados a largo plazo sin modificaciones en la forma de vida.

Microcirugía videoscópica técnica asistida aplicada al diseño de procedimientos diversos utilizando a la rata como modelo experimental / Videoscopic microsurgery assisted technique applied to the design of procedures utilizing the rat as an experimental model

Presentamos un nuevo método de aplicación y entrenamiento para la realización de microcirugía, desarrollado en el departamento de Cirugía Experimental ®Karl Storz-Brimex II¼ del hospital ABC, al que hemos denominado ®microcirugía videoscópica asistida¼ que permite el refinamiento de la técnica microquirúrgica, siendo éste el eslabón de enlace entre el microscopio y la técnica microquirúrgica laparoendoscópica. Nuestra propuesta original de utilizar a la rata como el modelo idela para la enseñanza de la cirugía laparoscópica se ha visto reforzada por la implementación de este método y, de hecho, su aplicación clínica y algunos ejemplos de su utilización en diversos espacios anatómicos de la rata