Upper mesophotic reef fish assemblages at Bahía de Banderas, Mexico.

There is no information on the species associated with the mesophotic reefs of Banderas Bay, located in the central Mexican Pacific. This study analysed the reef fish assemblage from three depths (50, 60 and 70 m) in three sampling sites of the southern submarine canyon of the Bay: Los Arcos, Bajo de Emirio and Majahuitas. Several analyses were performed to test the hypothesis that there are important differences in fish abundance and species composition between sites and depths. Twenty-two species of bony fishes grouped in 14 families were recorded. PERMANOVA results showed that there were no significant differences in fish diversity parameters between sites, indicating a certain uniformity in their distribution. However, nine species were exclusive to one site and depth (five singleton species with only one individual recorded and four unique species recorded only once). On the other hand, there were significant differences between depths, mainly between 50 and 70 m. Diversity decreases with depth and species composition changes. SIMPER, Shade Plot and NMDS analysis show the most representative species at each depth, with at least half of the species (11) recorded only at 50 m and four species at the deeper levels (60 - 70 m). The observed assemblage includes several of the most caught species in the shallow water artisanal fishery, which is the most traditional and common type of fishery in the Bay. In addition, the Pomacanthuszonipectus (Cortés angelfish) is of particular interest, as it has a special protection status in the official Mexican standard (NOM-059-SEMARNAT, 2010) due to its use as an ornamental species in aquaria. We hypothesised that the mesophotic zone may serve as a refuge for these fishes, so we propose that the information obtained is an important basis for new research aimed at the sustainable management of fisheries in the area.

Identification of RP1 as the genetic cause of retinitis pigmentosa in a multi-generational pedigree using Extremely Low-Coverage Whole Genome Sequencing (XLC-WGS).

MOTIVATION: Next-generation sequencing (NGS) technologies are decisive for discovering disease-causing variants, although their cost limits their utility in a clinical setting. A cost-mitigating alternative is an extremely low coverage whole-genome sequencing (XLC-WGS). We investigated its use to identify causal variants within a multi-generational pedigree of individuals with retinitis pigmentosa (RP). Causing progressive vision loss, RP is a group of genetically heterogeneous eye disorders with approximately 60 known causal genes. RESULTS: We performed XLC-WGS in seventeen members of this pedigree, including three individuals with a confirmed diagnosis of RP. Sequencing data were processed using Illumina's DRAGEN pipeline and filtered using Illumina's genotype quality score metric (GQX). The resulting variants were analyzed using Expert Variant Interpreter (eVai) from enGenome as a prioritization tool. A nonsense known mutation (c.1625C > G; p.Ser542*) in exon 4 of the RP1 gene emerged as the most likely causal variant. We identified two homozygous carriers of this variant among the three sequenced RP cases and three heterozygous individuals with sufficient coverage of the RP1 locus. Our data show the utility of combining pedigree information with XLC-WGS as a cost-effective approach to identify disease-causing variants.

Analysis of the MCTP Amino Acid Sequence Reveals the Conservation of Putative Calcium- and Lipid-Binding Pockets Within the C2 Domains In Silico.

MCTPs (Multiple C2 Domains and Transmembrane region Proteins) are evolutionarily and structurally related to other C2 proteins, which are central to exocytosis and membrane trafficking; however, their specific function has been little studied. MCTPs are associated with endosomes and the endoplasmic reticulum and possess three C2 domains (C2A-C2C) and two transmembrane regions (TMRs) well conserved in different species. Here, we generated structural models of the MCTP C2 domains of C. elegans and analyzed their putative function by docking, which revealed that these domains possess Ca2+- and lipid-binding pockets, suggesting that MCTPs play a significant, calcium-dependent role in membrane physiology.

MCTP-1 modulates neurotransmitter release in C. elegans.

Multiple C2 and Transmembrane Domain Proteins (MCTPs) are putative calcium sensors. Proteins that contain C2 domains play essential roles in membrane trafficking and exocytosis; however, MCTPs functions in neurotransmitter release are not known. Here we report that in C. elegans mctp-1 is under the control of two promoters - one active in the nervous system and the second in the spermatheca. We generated and characterized a loss of function amt1 mutant and compared it to a previously published loss of function mutant (av112). Loss of mctp-1 function causes defects in egg-laying, crawling velocity, and thrashing rates. Both amt1 and av112 mutants are hyposensitive to the acetylcholinesterase blocker aldicarb, suggesting that MCTP-1 may play a role in synaptic vesicle release.

The effect of CTLA-4Ig, a CD28/B7 antagonist, on the lung inflammation and T cell subset profile during murine hypersensitivity pneumonitis.

Hypersensitivity pneumonitis (HP) is an inflammatory lung disease characterized by an influx of activated T cells to the lung, in which the CD28/B7 costimulatory signals are essential for the T cell activation and the outcome of the inflammatory response. In this study, we investigated the effect of the CD28/B7 antagonist, CTLA-4Ig, on the lung inflammation and the T cell subset profile in experimental Saccharopolyspora recivirgula (SR)-induced HP. C57BL/6 mice were treated with SR or saline during two and three weeks and in addition of CTLA-4Ig was administrated after either the second or third week and mice were sacrificed seven days later. The extent of the lung inflammation was quantified by histopathology and the lung T cell subsets (Treg, Th17, γδT and NKT) were analyzed by flow cytometry. Mice treated with CTLA-4Ig showed a significant decrease in the extent of lung damage (p<0.05), and exhibited a decreased number of inflammatory cells in the bronchoalveolar lavage (BAL) with diminished CD4/CD8 T cell ratio. Also, a significant increase in the percentage of lung γδT (p<0.01) and NKT (p<0.05) cells was observed in two weeks SR-treated mice with the administration of CTLA-4Ig/SR. At 3 weeks, SR-treated mice showed an increased percentage of regulatory T cells but no significantly differences were found in the percentage of Th17 cells when compared with CTLA-4Ig/SR-treated mice. Our findings suggest that the treatment with CTLA-4Ig affects the HP progression and the lung T cell subset kinetics in mice.

Overexpression of MMP9 in macrophages attenuates pulmonary fibrosis induced by bleomycin.

Pulmonary fibrosis is a common response to a variety of lung injuries, characterized by fibroblast/myofibroblast expansion and abnormal accumulation of extracellular matrix. An increased expression of matrix metalloprotease 9 (MMP9) in human and experimental lung fibrosis has been documented, but its role in the fibrotic response is unclear. We studied the effect of MMP9 overexpression in bleomycin-driven lung fibrosis using transgenic mice expressing human MMP9 in alveolar macrophages (hMMP9-TG). At 8 weeks post-bleomycin, the extent of fibrotic lesions and OH-proline content were significantly decreased in the TG mice compared to the WT mice. The decreased fibrosis in hMMP9-TG mice was preceded by a significant reduction of neutrophils and lymphocytes in bronchoalveolar lavage (BAL) at 1 and 4 weeks post-bleomycin, respectively, as well as by significantly less TIMP-1 than the WT mice. From a variety of cytokines/chemokines investigated, we found that BAL levels of insulin-like growth factor binding protein-3 (IGFBP3) as well as the immunoreactive protein in the lungs were significantly lower in hMMP9-TG mice compared with WT mice despite similar levels of gene expression. Using IGFBP-3 substrate zymography we found that BAL from TG mice at 1 week after bleomycin cleaved IGFBP-3. Further, we demonstrated that MMP9 degraded IGFBP-3 into lower molecular mass fragments. These findings suggest that increased activity of MMP9 secreted by alveolar macrophages in the lung microenvironment may have an antifibrotic effect and provide a potential mechanism involving IGFBP3 degradation.

Local and circulating microchimerism is associated with hypersensitivity pneumonitis.

RATIONALE: Hypersensitivity pneumonitis (HP) is a lymphocytic alveolitis provoked by exposure to a variety of antigens. However, the disease occurs in only a subset of exposed individuals, suggesting that additional factors may be involved. Microchimerism has been implicated in the pathogenesis of autoimmune diseases, especially in those showing increased incidence after childbearing age. OBJECTIVES: To evaluate the presence of circulating and local microchimeric cells in female patients with HP. METHODS: Male microchimerism was examined in 103 patients with HP, 30 with idiopathic pulmonary fibrosis (IPF), and 43 healthy women. All of them had given birth to at least one son, with no twin siblings, blood transfusions, or transplants. Microchimerism was examined by dot blot hybridization (peripheral blood), and by fluorescence in situ hybridization in bronchoalveolar lavage cells and lungs. MEASUREMENTS AND MAIN RESULTS: Blood microchimerism was found in 33% of the patients with HP in comparison with 10% in those with IPF (p = 0.019) and 16% in healthy women (p = 0.045). Patients with HP with microchimerism showed a significant reduction of diffusing capacity of carbon monoxide (Dl(CO); 53.5 +/- 11.9% vs. 65.2 +/- 19.7%; p = 0.02) compared with patients with HP without microchimerism. In bronchoalveolar lavage cells, microchimerism was detected in 9 of 14 patients with HP compared with 2 of 10 patients with IPF (p = 0.047). Cell sorting revealed that microchimeric cells were either macrophages or CD4+ or CD8+ T cells. Male microchimeric cells were also found in the five HP lungs examined by fluorescence in situ hybridization. CONCLUSIONS: Our findings (1) demonstrate that patients with HP exhibit increased frequency of fetal microchimerism, (2) confirm the multilineage capacity of microchimeric cells, and (3) suggest that microchimeric cells may increase the severity of the disease.

Paradoxical effect of salbutamol in a model of acute organophosphates intoxication in guinea pigs: role of substance P release.

Organophosphates induce bronchoobstruction in guinea pigs, and salbutamol only transiently reverses this effect, suggesting that it triggers additional obstructive mechanisms. To further explore this phenomenon, in vivo (barometric plethysmography) and in vitro (organ baths, including ACh and substance P concentration measurement by HPLC and immunoassay, respectively; intracellular Ca2+) measurement in single myocytes) experiments were performed. In in vivo experiments, parathion caused a progressive bronchoobstruction until a plateau was reached. Administration of salbutamol during this plateau decreased bronchoobstruction up to 22% in the first 5 min, but thereafter airway obstruction rose again as to reach the same intensity as before salbutamol. Aminophylline caused a sustained decrement (71%) of the parathion-induced bronchoobstruction. In in vitro studies, paraoxon produced a sustained contraction of tracheal rings, which was fully blocked by atropine but not by TTX, omega-conotoxin (CTX), or epithelium removal. During the paraoxon-induced contraction, salbutamol caused a temporary relaxation of approximately 50%, followed by a partial recontraction. This paradoxical recontraction was avoided by the M2- or neurokinin-1 (NK1)-receptor antagonists (methoctramine or AF-DX 116, and L-732138, respectively), accompanied by a long-lasting relaxation. Forskolin caused full relaxation of the paraoxon response. Substance P and, to a lesser extent, ACh released from tracheal rings during 60-min incubation with paraoxon or physostigmine, respectively, were significantly increased when salbutamol was administered in the second half of this period. In myocytes, paraoxon did not produce any change in the intracellular Ca2+ basal levels. Our results suggested that: 1) organophosphates caused smooth muscle contraction by accumulation of ACh released through a TTX- and CTX-resistant mechanism; 2) during such contraction, salbutamol relaxation is functionally antagonized by the stimulation of M2 receptors; and 3) after this transient salbutamol-induced relaxation, a paradoxical contraction ensues due to the subsequent release of substance P.

Bleomycin-induced pulmonary fibrosis is attenuated in gamma-glutamyl transpeptidase-deficient mice.

To investigate repair mechanisms in bleomycin-induced pulmonary fibrosis, we used mice deficient in gamma-glutamyl transpeptidase (GGT-/-), a key enzyme in glutathione (GSH) and cysteine metabolism. Seventy-two hours after bleomycin (0.03 U/g), GGT-/- mice displayed a different inflammatory response to wild-type mice as judged by a near absence of neutrophils in lung tissue and bronchoalveolar lavage and a less pronounced rise in matrix metalloproteinase-9. Inflammation in GGT-/- mice consisted mainly of lymphocytes and macrophages. At 1 month, lungs from bleomycin-treated GGT-/- mice exhibited minimal areas of fibrosis compared with wild-type mice(light microscopy fibrosis index: 510 +/- 756 versus 1975 +/- 817, p < 0.01). Lung collagen content revealed a significant increase in bleomycin-treated wild-type (15.1 +/- 3.8 versus 8.5 +/- 0.7 microg hydroxy(OH)-proline/mg dry weight, p < 0.01) but not in GGT-/- (10.4 +/- 1.7 versus 8.8 +/- 0.8). Control lungs from GGT-/- showed a significant reduction of cysteine (0.03 +/- 0.005 versus 0.055 +/- 0.001, p < 0.02) and GSH levels (1.24 +/- 0.055 versus 1.79 +/- 0.065, p < 0.002). These values decreased after 72 hours of bleomycin in both GGT-/- and wild-type but reached their respective control values after 1 month. Supplementation with N-acetyl cysteine partially ameliorated the effects of GGT deficiency. These findings suggest that increased neutrophils and matrix metalloproteinase-9 during the early inflammatory response and adequate thiol reserves are key elements in the fibrotic response after bleomycin-induced pulmonary injury.

Interleucinas IL-1ß, IL-6 e IL-10 durante la perfusión y la reperfusión pulmonar / Interleukines IL-1ß, IL-6 e IL-10 during lung perfusión and reperfusion

Introducción: La perfusión y reperfusión de órganos provocan daño celular. La síntesis y la liberación de citocinas se afectan como respuesta al daño celular. Objetivo: Evaluar los cambios que ocurren en la concentración de la interleucinas 1ß, 6 y 10 presentes en la solución de perfusión durante la perfusión y reperfusión pulmonar. Material y métodos: Se utilizaron los bloques pulmonares de 21 ratones Balb-c, estos bloques fueron divididos al azar en tres grupos de estudio (n = 7 en cada grupo). Grupo 1: los bloques fueron perfundidos in situ a través de la arteria pulmonar con solución de Krebs-Henseleit a 4ºC con un flujo de perfusión de 0.2 mL/min únicamente durante tiempo necesario para exaguinarlos; Grupos 2: Los bloques fueron perfundidos in situ durante 15' y 30' a través de la arteria pulmonar con solución de Krebs-Henseleit a 4ºC con un flujo de perfusión de 0.2 ml/min y Grupo 3: Los bloques fueron exanguinados mediante perfusión in situ a través de la arteria pulmonar con solución de Krebs-Henseleit a 4ºC con un flujo de perfusión de 0.2 mL/min, se preservaron durante 24 horas a 4ºC sumergidos en la misam solución y transcurrido el tiempo de isquemia, fueron reperfundidos ex vivo durante 15' y 30' con solución de Krebs-Henseleit a 4 ºC con un flujo de perfusión de 0.2 mL/min. Durante las perfusiones y las reperfusiones se recolectó la solución de perfución pulmonar y se determinó la concentración de las interleucinas mediante la técnica de ELISA. Resultados: Durante la perfusión y reperfusión pulmonar, las concentraciones de IL-1ß en la solución de perfusión, fueron menores que las concentraciones de IL-6 e IL-10, siendo ésta última, la interleucina detectada en mayor concentración. Las concentraciones de IL-1ß e IL-6 no se modificaron ni por el timepo de perfusión ni por efecto de la reperfusión mientras que la concentración de IL-10 disminuyó por efecto de la reperfusión

Interleucinas 1ß, 6 y 10 en homogeneizados de corazón y de pulmón postperfusión y reperfusión del bloque cardiopulmonar / Interleukines 1ß, 6 and 10 in heart and lung homogenates postperfusion and reperfusion of the cardiopulmonary block

Introducción: Como consecuencia de la perfusión y de la reperfusión de órganos existe daño tisular. La síntesis y la liberación de citocinas se afectan como respuesta al daño celular. Objetivo: En este trabajo, evaluamos los cambios que ocurren en la concentración de interleucinas 1ß, 6 y 10 en homogeneizados de corazón y de pulmón como respuesta al daño ocasionado por el efecto de la perfusión y de la reperfusión cardiopulmonar. Material y métodos: utilizamos los bloqueos cardiopulmonares de 21 ratones Balb-C, estos bloques fueron divididos al azar en tres grupos de estudio (n= 7 en cada grupo). Grupo 1: Los bloques fueron perfundidos in situ a través de la arteria pulmonar con solución de Krebs-Henseleit a 4 ºC con un flujo de perfusión de 0.2 mL/min únicamente durante el tiempo necesario para exanguinarlos. Grupo 2: los bloques fueron perfundidos in situ durante 30' a través de la arteria pulmonar con solución de Krebs-Henseleit a 4 ºC con un flujo de perfusión de 0.2 mL/min. Grupo 3: los bloques fueron exanguinados mediante perfusión in situ a través de la arteria pulmonar con solución de Krebs-Henseleit a 4 ºC con un flujo de perfusión de 0.2 mL/min, se preservaron durante 24 horas a 4 ºC sumergidos en la misma solución a transcurrido el tiempo de isquemia, fueron reperfundidos ex vivo durante 30' con solución de Krebs-Henseleit a ºC con un flujo de perfusión de 0.2 mL/min. Concluidas las perfusiones y las reperfusiones preparamos homogeneizados de corazón y de pulmón. Determinamos la concentración de las interleucinas presentes en cada homogeneizado tisular mediante la técnica de ELISA. Resultados. La concentración de IL-6 fue similar en los homogeneizados de corazón y de pulmón y no se alteró por efecto de la reperfusión, mientras que las concentraciones de IL-1ß e IL-10 parecen actuar de manera inversa entre ambos órganos y durante todo el estudio

Reparación de defectos de pared tóracoabdominal de perros con bioprótesis de pericardio bovino / Repair of thoracoabdominal wall defects dogs with a bovine pericardial bioprothesis

Objetivos. Evaluar las características de resistencia in vitro de las bioprótesis de tejido pericárdio y su empleo en la reconstrucción de defectos de pared tóracoabdominal. Material y métodos. Se prepararon prótesis mediante su preservación por 15 días en cinco concentraciones de glutaraldehído (0.5 a 10 por ciento). Como controles se usaron solución salina y solución de Hanks. Se evaluaron la fuerza tansil de ruptura y el porcentaje de elogación en los tejidos preservados así como las de muestras de malla de polipropileno (Marlex) y poliéster (Mersilene) utilizando un equipo automático Instrom 1011. Además, se evaluó la capacidad de la prótesis de glutaraldehído al 0.5 por ciento para reparar defectos experimentalmente producidos en pared abdominal (n= 12), pared torácica (n= 6), diafragma (n= 6) y esternón (n= 7) de perros mestizos. Resultados. La mayor fuerza tensil se observó con glutaraldehído al 0.5 por ciento (11.7 N/mm²) y fue significativamente mayor a las de las otras concentraciones de glutaraldehído y a las mallas. El porcentaje de elongación del glutaraldehído al 0.5 por ciento fue de 56 por ciento, similar a la salina y significativamente menor que el del glutaraldehído al 1,25 y 5 por ciento y al polipropileno. La tolerancia fue adecuada en todos los animales: histológicamente hubo formación de cicatriz resistente. No hubo infección o rechazo en ningún animal. Conclusión. Las bioprótesis de pericardio mostraron ser un material de suficiente resistencia para ser utilizada quirúrgicamente en la reparación de defectos de pared tóracoabdominal

Criopreservación de traquea: estudio experimental / Cryopreservation of trachea: experimental study

El propósito de este estudio fue de evaluar la viabilidad de segmentos de tráquea implantados en el omento mayor después de haber sido sometidos o no a criopreservación. Se utilizaron dos grupos de cinco perros adultos mestizos. En el grupo I los segmentos traqueales fueron sometidos a criopreservación en nitrógeno líquido a -90oC por 15 días, tras los cuales se implantó en el omento mayor de los perros receptores. En el grupo II, los segmentos fueron implantados en el omento inmediatamente después de haber sido obtenidos de los perros donadores. Todos los segmentos fueron estudiados 15 días después de su implantación. Los cambios histológicos fueron evaluados al azar en microscopía de luz, recibiendo una gradación numérica para cada uno de los cambios observados. Macroscópicamente todos los segmentos recuperados presentaron aspecto normal. Histológicamente al evaluar el cartílago, se encontró que el grupo II hubo gradación significativamente mayor; mientras que los cartílagos tratados con criopreservación no mostraron alteración alguna (p=0.018)fuera de esto no se encontró ninguna otra diferencia. Al perfundirlos con solución de tinta china a través de la arteria gastroepiploica derecha, se observó marcro y microscópicamente que todos los segmentos traqueales lograron ser revascularizados adecuadamente. Concluimos que la criopreservación de un segmento traqueal por 15 días en nitrógeno líquido, no altera la integridad histológica del mismo. Además el implante de un segmento traqueal en el omento mayor permite mantener su viabilidad gracias a la revascularización que se produce a partir del mismo omento