Lung surfactant in the pathophysiology and treatment of lung fibrosis

The main function of the lung is gas exchange. This function is essential for life and optimized by combining a large surface area in contact with air and a thin barrier for gas (O2/CO2) diffusion. An array of conductive tubes (conductive airways: trachea and bronchi) leads atmospheric air through convection to the distal (alveolar) side of the lung, where gas exchange happens by diffusion (through a pressure gradient). At the alveoli level, many physical forces also play a vital role during the respiratory cycle. To the elastic recoil of the lung, due to the pleura and chest recoil forces, elastic properties of the connective tissue of the alveolar septa should also be taken into account. In addition, alveoli should also overcome interfacial forces from the air-liquid interface they are exposed to. This is mainly the surface tension at the liquid lining layer or hypophase. This force is primarily counteracted by lung surfactant, a specific mix of lipids and proteins synthesized and secreted by specialized cells, the alveolar epithelial type 2 cell (AE2C). On the other hand, the focus of this work is a specific lung disease, lung fibrosis. is a severe disease characterized by chronic inflammation, myofibroblast accumulation, and excessive extracellular matrix deposition, resulting in the damage of lung structure and respiratory failure. In Europe alone, approximately 40,000 new cases are diagnosed each year; however, its prognosis is overall poor, with a median survival of 3–4 years. We hypothesized that: 1) surfactant dysfunction contributes to the early stages and progression of the disease and; 2) we could target lung surfactant dysfunction as a therapeutic approach. With the use of animal models, we concluded that lung surfactant dysfunction is an early event, where AE2C injury plays a key role and alveolar macrophages play an important role during the progression of the disease. In addition, knowing the timing of lung surfactant dysfunction, we performed surfactant replacement therapies in order to prevent or slow down the onset or progression of the disease. From the results presented in this work, we can conclude that exogenous surfactant has an anti-fibrotic effect, increasing the surface area of opened alveoli and reducing the septal wall thickness. We could observe the same effect when transplanting healthy AE2C in diseased animal lungs, in order to replace the inactivated lung surfactant with healthy synthesizing cells

Resting time after phorbol 12-myristate 13-acetate in THP-1 derived macrophages provides a non-biased model for the study of NLRP3 inflammasome

Background: The activation of NLRP3 inflammasome in macrophages has been proven to play a crucial role in the development of cardiovascular diseases. THP-1 monocytes can be differentiated to macrophages by incubation with phorbol-12-myristate 13-acetate (PMA), providing a suitable model for in vitro studies. However, PMA has been shown to have effects on the levels of IL-1 beta, the main mediator of NLRP3 inflammasome, while the effects on the other mediators of the inflammasome have not been reported before. Methods: THP-1 monocytes were incubated without (THP-1), with 5ng/ml PMA for 48h (PMA48h) or with 5ng/ml PMA for 48h plus 24h in fresh medium (PMArest). Morphological changes and the expression of macrophage surface markers (CD14, CD11b, CD36 and CD204) were evaluated by flow cytometry. Changes in intracellular levels of inflammasome components (NLRP3, ASC, pro-caspase-1, pro-IL1 beta) were analyzed by western blot and release of mature IL-1 beta in cell supernatant was analyzed by ELISA. ASC speck formation was determined by immunofluorescence. Results: After 48h incubation with PMA or subsequent rest in fresh medium, cells became adherent, and the differential expression of CD36, CD11b, CD14 and CD204 compared to THP-1 cells confirmed that PMArest resemble macrophages from a molecular point of view. Changes in the levels were detected in PMA48h group for all the NLRP3-related proteins, with increase of NLRP3 and pro-IL-1 beta and secretion of mature IL-1 beta. In PMArest, no pro-IL-1 beta and lower amounts of mature IL-1 beta were detected. No ASC speck was found in PMA treated groups, but the addition of a second stimulus to PMArest resulted in ASC speck formation, together with IL-1 beta production, confirming the responsiveness of the model. Conclusion: Differentiation of THP-1 with 5ng/ml PMA followed by 24h resting period provides a model that morphologically and molecularly resembles macrophages. However, even at low concentrations, PMA induces production of IL-1 beta. The 24h rest period provides for down-regulation of pro-IL-1 beta in PMArest group, without affecting its ability to respond to a second stimulus through activation of inflammasome

Mutants of Ralstonia (Pseudomonas) solanacearum sensitive to antimicrobial peptides are altered in their lipopolysaccharide structure and are avirulent in tobacco

Ralstonia solanacearum K60 was mutagenized with the transposon Tn5, and two mutants, M2 and M88, were isolated. Both mutants were selected based on their increased sensitivity to thionins, and they had the Tn5 insertion in the same gene, 34 bp apart. Sequence analysis of the interrupted gene showed clear homology with the rfaF gene from Escherichia coli and Salmonella typhimurium (66% similarity), which encodes a heptosyltransferase involved in the synthesis of the lipopolysaccharide (LPS) core. Mutants M2 and M88 had an altered LPS electrophoretic pattern, consistent with synthesis of incomplete LPS cores. For these reasons, the R. solanacearum gene was designated rfaF. The mutants were also sensitive to purified lipid transfer proteins (LTPs) and to an LTP-enriched, cell wall extract from tobacco leaves. Mutants M2 and M88 died rapidly in planta and failed to produce necrosis when infiltrated in tobacco leaves or to cause wilting when injected in tobacco stems. Complemented strains M2* and M88* were respectively obtained from mutants M2 and M88 by transformation with a DNA fragment harboring gene rfaF. They had a different degree of wild-type reconstituted phenotype. Both strains retained the rough phenotype of the mutants, and their LPS electrophoretic patterns were intermediate between those of the wild type and those of the mutants

On Top of the Alveolar Epithelium: Surfactant and the Glycocalyx

Gas exchange in the lung takes place via the air-blood barrier in the septal walls of alveoli. The tissue elements that oxygen molecules have to cross are the alveolar epithelium, the interstitium and the capillary endothelium. The epithelium that lines the alveolar surface is covered by a thin and continuous liquid lining layer. Pulmonary surfactant acts at this air-liquid interface. By virtue of its biophysical and immunomodulatory functions, surfactant keeps alveoli open, dry and clean. What needs to be added to this picture is the glycocalyx of the alveolar epithelium. Here, we briefly review what is known about this glycocalyx and how it can be visualized using electron microscopy. The application of colloidal thorium dioxide as a staining agent reveals differences in the staining pattern between type I and type II alveolar epithelial cells and shows close associations of the glycocalyx with intraalveolar surfactant subtypes such as tubular myelin. These morphological findings indicate that specific spatial interactions between components of the surfactant system and those of the alveolar epithelial glycocalyx exist which may contribute to the maintenance of alveolar homeostasis, in particular to alveolar micromechanics, to the functional integrity of the air-blood barrier, to the regulation of the thickness and viscosity of the alveolar lining layer, and to the defence against inhaled pathogens. Exploring the alveolar epithelial glycocalyx in conjunction with the surfactant system opens novel physiological perspectives of potential clinical relevance for future research

Improved Alveolar Dynamics and Structure After Alveolar Epithelial Type II Cell Transplantation in Bleomycin Induced Lung Fibrosis

Idiopathic pulmonary fibrosis (IPF) is a progressively and ultimately fatal lung disease. Previously it has been shown that intratracheal administration of alveolar epithelial type II cells (AE2C) in the animal model of bleomycin-induced pulmonary fibrosis is able to reverse fibrosis and restore surfactant protein levels. However, to date, it has not been evaluated whether these changes involve any improvement in alveolar dynamics. Consequently, the aim of the present work was to study lung physiology after AE2C transplantation at different time points during the development of injury and fibrosis. Lung fibrosis was induced by intratracheal instillation of bleomycin (4U/kg) in rat lungs. The animals were transplanted with AE2C (2.5 x 10(6) cells/animal) 3 or 7 days after bleomycin instillation. Assessments were done at day 7 and 14 after the induction of fibrosis to plot time dependent changes in lung physiology and mechanics. To assess the pressures and rates at which closed alveoli reopens invasive pulmonary tests using a small-animal mechanical ventilator (Flexivent (R), Scireq, Canada) including de-recruitability tests and forced oscillation technique as well as quasi-static pressure volume loops were performed. Afterwards lungs were fixed by vascular perfusion and subjected to design-based stereological evaluation at light and electron microscopy level. AE2C delivered during the lung injury phase (3 days) of the disease are only able to slightly recover the volume of AE2C and volume fraction of LB in AE2C. However, it did not show either positive effects regarding ventilated alveolar surface nor any increase of lung compliance. On the other hand, when AE2C are delivered at the beginning of the fibrotic phase (7 days after bleomycin instillation), an increased ventilated alveolar surface to control levels and reduced septal wall thickness can be observed. Moreover, transplanted animals showed better lung performance, with increased inspiratory capacity and compliance. In addition, a detailed analysis of surfactant active forms [mainly tubular myelin, lamellar body (LB)-like structures and multilamellar vesicles (MLV)], showed an effective recovery during the pro-fibrotic phase due to the healthy AE2C transplantation. In conclusion, AE2C transplantation during fibrogenic phases of the disease improves lung performance, structure and surfactant ultrastructure in bleomycin-induced lung fibrosis

Twisted bilayered graphenes at magic angles and Casimir interactions: correlation-driven effects

Twisted bilayered graphenes at magic angles are systems housing long ranged periodicity of Moir\'e pattern together with short ranged periodicity associated with the individual graphenes. Such materials are a fertile ground for novel states largely driven by electronic correlations. Here we find that the ubiquitous Casimir force can serve as a platform for macroscopic manifestations of the quantum effects stemming from the magic angle bilayered graphenes properties and their phases determined by electronic correlations. By utilizing comprehensive calculations for the electronic and optical response, we find that Casimir torque can probe anisotropy from the Drude conductivities in nematic states, while repulsion in the Casimir force can help identify topologically nontrivial phases in magic angle twisted bilayered graphenes.Comment: 9 pages, 6 figures (main), 7 pages, 7 figures (supplementary); provisionally accepted for publication in 2D Material

Evaluacion del sistema de drenaje en canal abierto en el cultivo de banano (Musa AAA) en la finca La bomba

El presente estudio se realizó en los suelos de la finca Bomba", corregimiento de Orihueca, Municipio de Ciénaga, Departamento del Magdalena. El corregimiento de Orihueca se encuentra enmarcado en los siguientes límites: por el Norte con el río Rio Frío, por el este con la carretera que de Ciénaga conduce a Fundación, por el Oeste con terrenos que dan a la Ciénaga Grande de Santa Marta y por el Sur con la Quebrada Orihueca. La finca La Bomba limita por el Norte con, la parcela Osario Serrano Alberto y con Inversiones urbanas rurales, al sur con la finca de Dávila - y Pedro, por el Este con las fincas de Navarro y Antonio Torres, por el Oeste con camino en medio. La zona bananera está entre las siguientes o coordenadas: Longitud Oeste 74 21' - 74 24"; latitud o Norte: 11 01' - 10 22'. La finca presenta un área aproximada de 46,5 Has, el relieve es plano con pendiente de 0 - 1%, con una altura de 15 m.s.n.m una precipitación promedio anual de 1.31e mm, una o temperatura media de 28 C y la humedad relativa es de 20,57, los suelos son permeables, con un PH variable predominando el ligeramente ácido. La zona se encuentra influenciada por los vientos alisios que soplan durante los meses de Diciembre a Marzo alcanzando velocidades de 10 1 m/seg. La investigación se inició en el mes de enero de 1.993 y - finalizó en enero de 1.994. Durante este período se llevó cabo el trabajo de campo que partió como primera medida un reconocimiento del terreno, las instalaciones que poseían la finca y el estado en que se encontraba. Para evaluar los resultados y establecer el diseño del drenaje propuesto se tuvo en cuenta los siguientes parámetros: Profundidad efectiva, clase de textura del suelo, conductividad hidráulica, retención de humedad, salinidad del suelo, salinidad del agua de riego, profundidad del nivel freático, profundidad de la capa impermeable, infiltración, precipitación promedia, evaporación y temperatura de la zora. Los resultados señalan que para establecer un correcto diseño de drenaje fue necesario además de las evaluaciones de los parámetros anteriores, establecer el espaciamiento entre drenes terciarios para lo cual se utilizó la fórmula de Houghoutd, encontrándose que dicho espaciamiento depende no solo de las características físicas de los suelos, sino de la profundidad en la cual se encuentra ubicada la capa impermeable del suelo que para nuestro caso es de 17,42 m. El diseño actual y el diseño propuesto presentan igual número de canales secundarios y primarios con 10 y uno respectivamente, el diseño propuesto presenta además canales terciarios ubicados a 50 metros de separación entre ellos; necesarios para una evacuación rápida y eficiente de las aguas de escorrentía después de una fuerte precipitación. Es claramente notorio que en el diseño propuesto las áreas para los canales secundarios y primarios son más reducidas que las áreas de los canales del diseño actual, trayendo como consecuencia un sobre diseño en el drenaje externo, reflejándose esto en un incremento de los costos de excavación además por los ingresos dejados de percibir al no cultivarse las áreas que actualmente se encuentran ocupadas por los canales del diseño actual

Brenneria quercina and Serratia spp. isolated from Spanish oak trees: molecular characterization and PCR development

Brenneria quercina has been reported as one of the causal agents of oak decline in Spain. To investigate the bacterial variability of this pathogen from different Spanish oak forests, a collection of 38 bacterial isolates from seven geographic locations and from different oak species was analysed by sequencing 16S rDNA and rep-PCR fingerprinting. All Spanish isolates of B. quercina were grouped by rep-PCR into a homogenous cluster that differed significantly from B. quercina reference strains from California. 16S rDNA analysis revealed that 34 out of 38 isolates were Brenneria . However, four isolates belonged to the genus Serratia , suggesting that this bacterium could cause cankers in oak trees. The information obtained by rep-PCR fingerprint analysis was used to develop PCR primers for the sensitive and specific detection of B. quercina from infected plant tissues. Pathogenicity tests performed with Brenneria and Serratia isolates showed that both were able to grow and cause cankers in oak trees

Bioinspired design of triboceramics: Learning from the anisotropic microfracture response of dental enamel under sliding contact

[EN] In the quest for novel ceramics for tribological applications via bioinspired design, the differences in the fracture modes that arise upon scratching relevant locations of ceramic-like tooth enamel are investigated. It is found that fracture initiates from weak rod-sheath interfaces at relatively low loads, independent of the sliding direction. However, the geometry and propagation of the cracks depends on the orientation of the interfaces relative to the maximum tensile stress: scratching along the occlusal surface propagates approximately sinusoidal cracks, parallel to the sliding direction, while scratching along the cross-section produces straight cracks that propagate normal (scratch parallel to occlusal surface) or parallel (scratch perpendicular to occlusal surface) to the sliding direction. The formation of cracks is hindered in scratching near the enamel-dentine junction. Implications for the microstructural design of triboceramics (bulks and coatings) with improved durability are discussed.The authors wish to thank Dr Florencio Monje Gil for kindly providing tooth specimens from his clinic (CICOM, Centro de Implantologia Cirugia Oral y Maxilofacial, Badajoz, Spain). This study was supported by Junta de Extremadura, Spain, and FEDER/ERDF funds (grants IB16139 and GR18149), and the Spanish Ministry of Science and Innovation (grant PID2019-105377RB-I00). E. P.-C. gratefully acknowledges support from the Spanish Ministry of Economy and Competitiveness (MINECO) under grant FJCI-2015-27228.Sanchez-Gonzalez, E.; Rodriguez-Rojas, F.; Pinilla-Cienfuegos, E.; Borrero-Lopez, O.; Ortiz, AL.; Guiberteau, F. (2020). Bioinspired design of triboceramics: Learning from the anisotropic microfracture response of dental enamel under sliding contact. Ceramics International. 46(18):27983-27989. https://doi.org/10.1016/j.ceramint.2020.07.2922798327989461