Identification of latexin by a proteomic analysis in rat normal articular cartilage

<p>Abstract</p> <p>Background</p> <p>Osteoarthritis (OA) is characterized by degeneration of articular cartilage. Animal models of OA induced are a widely used tool in the study of the pathogenesis of disease. Several proteomic techniques for selective extraction of proteins have provided protein profiles of chondrocytes and secretory patterns in normal and osteoarthritic cartilage, including the discovery of new and promising biomarkers. In this proteomic analysis to study several proteins from rat normal articular cartilage, two-dimensional electrophoresis and mass spectrometry (MS) were used. Interestingly, latexin (LXN) was found. Using an immunohistochemical technique, it was possible to determine its localization within the chondrocytes from normal and osteoarthritic articular cartilage.</p> <p>Results</p> <p>In this study, 147 proteins were visualized, and 47 proteins were identified by MS. A significant proportion of proteins are involved in metabolic processes and energy (32%), as well as participating in different biological functions including structural organization (19%), signal transduction and molecular signaling (11%), redox homeostasis (9%), transcription and protein synthesis (6%), and transport (6%). The identified proteins were assigned to one or more subcellular compartments.</p> <p>Among the identified proteins, we found some proteins already recognized in other studies such as OA-associated proteins. Interestingly, we identified LXN, an inhibitor of mammalian carboxypeptidases, which had not been described in articular cartilage. Immunolabeling assays for LXN showed a granular distribution pattern in the cytoplasm of most chondrocytes of the middle, deep and calcified zones of normal articular cartilage as well as in subchondral bone. In osteoarthritic cartilage, LXN was observed in superficial and deep zones.</p> <p>Conclusions</p> <p>This study provides the first proteomic analysis of normal articular cartilage of rat. We identified LXN, whose location was demonstrated by immunolabeling in the chondrocytes from the middle, deep and calcified zones of normal articular cartilage, and superficial and deep zones of osteoarthritic cartilage.</p

LATE-NC staging in routine neuropathologic diagnosis : an update

An international consensus report in 2019 recommended a classification system for limbic-predominant age-related TDP-43 encephalopathy neuropathologic changes (LATE-NC). The suggested neuropathologic staging system and nomenclature have proven useful for autopsy practice and dementia research. However, some issues remain unresolved, such as cases with unusual features that do not fit with current diagnostic categories. The goal of this report is to update the neuropathologic criteria for the diagnosis and staging of LATE-NC, based primarily on published data. We provide practical suggestions about how to integrate available genetic information and comorbid pathologies [e.g., Alzheimer's disease neuropathologic changes (ADNC) and Lewy body disease]. We also describe recent research findings that have enabled more precise guidance on how to differentiate LATE-NC from other subtypes of TDP-43 pathology [e.g., frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS)], and how to render diagnoses in unusual situations in which TDP-43 pathology does not follow the staging scheme proposed in 2019. Specific recommendations are also made on when not to apply this diagnostic term based on current knowledge. Neuroanatomical regions of interest in LATE-NC are described in detail and the implications for TDP-43 immunohistochemical results are specified more precisely. We also highlight questions that remain unresolved and areas needing additional study. In summary, the current work lays out a number of recommendations to improve the precision of LATE-NC staging based on published reports and diagnostic experience.Peer reviewe

Cuantificación por inmunomicroscopía electrónica del efecto terapéutico del EGF en úlceras del pie diabético

Introducción: la Inmunomicroscopía electrónica cuantitativa se aplicó recientemente en el estudio de la cuantificación de las distribuciones de determinadas proteínas en diferentes organelos celulares en fibroblastos de Úlceras de pie diabético tratados con el Factor de crecimiento epidérmico en humanos. Objetivo: el presente se enfoca en los resultados relacionados con una molécula clave [el antígeno nuclear de proliferación celular] en la señalización inducida por el Factor de crecimiento epidérmico. Desarrollo: las muestras de Úlceras de pie diabético se analizaron por la inmunomicroscopía electrónica cuantitativa. Las referencias se obtuvieron de la Base de datos Pubmed. En concordancia con una afectación funcional de la señalización mediada por el Factor de crecimiento epidérmico en el tejido de granulación de los individuos diabéticos, se observó poca detección del antígeno nuclear de proliferación celular en los fibroblastos. No obstante, el tratamiento de las Úlceras de pie diabético con el Factor de crecimiento epidérmico indujo una activación temprana del antígeno nuclear de proliferación celular en el núcleo de los fibroblastos de las Úlceras de pie diabético. Se observó, además, un incremento en el inmunomarcaje del antígeno nuclear de proliferación celular en las mitocondrias de los fibroblastos en tiempos tardíos después de la inoculación del Factor de crecimiento epidérmico. Conclusiones: esta investigación demostró la utilidad y el valor de la cuantificación de las distribuciones de inmunomarcaje en organelos celulares para el estudio de las vías de señalización intracelulares de relevancia terapéutica

The bactericidal effect of silver nanoparticles.

Abstract Nanotechnology is expected to open new avenues to fight and prevent disease using atomic scale tailoring of materials. Among the most promising nanomaterials with antibacterial properties are metallic nanoparticles, which exhibit increased chemical activity due to their large surface to volume ratios and crystallographic surface structure. The study of bactericidal nanomaterials is particularly timely considering the recent increase of new resistant strains of bacteria to the most potent antibiotics. This has promoted research in the well known activity of silver ions and silver-based compounds, including silver nanoparticles. The present work studies the effect of silver nanoparticles in the range of 1-100 nm on Gram-negative bacteria using high angle annular dark field (HAADF) scanning transmission electron microscopy (STEM). Our results indicate that the bactericidal properties of the nanoparticles are size dependent, since the only nanoparticles that present a direct interaction with the bacteria preferentially have a diameter of ∼1-10 nm

Regulation of α5 and αV Integrin Expression by GDF-5 and BMP-7 in Chondrocyte Differentiation and Osteoarthritis.

The Integrin β1 family is the major receptors of the Extracellular matrix (ECM), and the synthesis and degradation balance of ECM is seriously disrupted during Osteoarthritis (OA). In this scenario, integrins modify their pattern expression and regulate chondrocyte differentiation in the articular cartilage. Members of the Transforming growth factor beta (Tgf-β) Superfamily, such as Growth differentiation factor 5 (Gdf-5) and Bone morphogenetic protein 7 (Bmp-7), play a key role in joint formation and could regulate the integrin expression during chondrocyte differentiation and osteoarthritis progression in an experimental OA rat model. Decrease of α5 integrin expression in articular cartilage was related with chondrocyte dedifferentiation during OA progression, while increase of α1, α2, and α3 integrin expression was related with fibrous areas in articular cartilage during OA. Hypertrophic chondrocytes expressed αV integrin and was increased in the articular cartilage of rats with OA. Integrin expression during chondrocyte differentiation was also analyzed in a micromass culture system of mouse embryo mesenchymal cells, micromass cultures was treated with Gdf-5 or Bmp-7 for 4 and 6 days, respectively. Gdf-5 induced the expression of the α5 sub-unit, while Bmp-7 induced the expression of the αV sub-unit. This suggests a switch in signaling for prehypertrophic chondrocyte differentiation towards hypertrophy, where Gdf-5 could maintain the articular chondrocyte phenotype and Bmp-7 would induce hypertrophy. Decrease of Ihh expression during late stages of OA in rat model suggest that the ossification in OA rat knees and endochondral ossification could be activated by Bmp-7 and αV integrin in absence of Ihh. Thus, chondrocyte phenotype in articular cartilage is similar to prehypetrophic chondrocyte in growth plate, and is preserved due to the presence of Indian hedgehog (Ihh), Gdf-5 and α5 integrin to maintain articular cartilage and prevent hypertrophy

Cicatrización de heridas cutáneas y papel de los miofibroblastos

Objetivo: realizar una revisión de las características fundamentales de la respuesta de cicatrización de heridas (RCH) cutáneas agudas y crónicas. Materiales y Métodos: la información se obtuvo de la base de datos pubmed y de los trabajos de inves-tigación de los autores. Resultados: la RCH cutáneas se desarrolla en cuatro fases secuenciales: hemostasia, inflamación, proliferación y remodelación. Primero ocurre la activación de fibroblastos, acumulación de un infiltrado celular inflamatorio que incluye a los miofibroblastos y la alteración de la matriz extracelular local (MEC). Después ocurre proliferación de los miofibroblastos, angiogénesis y proliferación de las células epiteliales. Finalmente tiene lugar el cierre de la herida y el restablecimiento de la arquitectura normal. Las heridas crónicas no siguen el patrón normal de reparación y no ocurre la cicatrización. Esto conduce a condiciones patológicas como las úlceras del pie diabético. Los miofibroblastos desempeñan un papel clave y su evolución coincide con los eventos de la RCH. Primero ocurre la trans-diferenciación que involucra la conversión de los fibroblastos en reposo a miofibroblastos que proliferan, son fibrogénicos y contráctiles. Posteriormente ocurre la perpetuación del fenotipo activado que incluye respuestas de fibrogénesis, proliferación, contractibilidad, liberación de citoquinas proinflamatorias, quimotaxis y degradación de la MEC. La resolución involucra la remoción del exceso de MEC y de los miofibroblastos. La eliminación de estos ocurre por tres mecanismos fundamentales: apoptosis, senescencia y reversión al fenotipo de fibroblastos. Esto constituye un paso fundamental en la restitución de la integridad del tejido. Conclusiones: se presentó una revisión actualizada de la RCH fisiológica y patológica

Osteopontin expression and localization of Ca2+ deposits in early stages of osteoarthritis in a rat model

Calcium deposits have been related to articular cartilage (AC) degeneration and have been observed in late stages of osteoarthritis (OA). However, the role of those deposits, whether they induce the OA pathogenesis or they appear as a consequence of such process, is still unknown. In this work, we present the kinetics of expression and tissue localisation of osteopontin (OPN), a mineralisation biomarker, and calcium deposits in samples from (normal, sham) and osteoarthritic cartilage (in a rat model). Immunohisto-chemical and Western blot assays for OPN, as well as Alizarin red staining for calcium deposits were performed; superficial, middle, and deep zones of AC were analysed. An increased expression of OPN and calcium deposits was found in the osteoarthritic cartilage compared with that of control groups, particularly in the superficial zone of AC in early stages of OA. In addition, the expression and localisation of OPN and calcium deposits during the OA pathogenesis suggest that the pathological AC mineralisation starts in the superficial zone during OA pathogenesis

Regulation between α5 integrin and Ihh signaling.

<p><i>In situ</i> hybridization for Indian hedgehog (<i>Ihh</i>) gene (A–D), in healthy articular cartilage (A), during OA at day 5 (B), at day 10 (C), and at day 20 (D) showed decrease of <i>Ihh</i> gene expression in Osteoarthritis (OA). Immunofluorescence in mouse micromass cultures (E–H), without treatment (E, G) showed expression for Ihh (E) and integrin α5 (F); treatment micromasses with blocking antibodies (G, H) for integrin α5 inhibited Ihh expression (F), and treatment with blocking antibodies for Ihh inhibits integrin α5 expression (H).</p

Regulation of integrin expression by Growth differentiation factor (Gdf)-5 and Bone morphogenetic protein (Bmp)-7 during chondrogenesis.

<p>Mouse micromass cultures without treatment (A–E), Gdf-5 treatment (F–J), and Bmp-7 treatment (K–O). Immunohistochemical stain for β1 (A, F, K), α1 (B, G, L), α5 (C, H, M), and αV (D, I, N) integrins, IgG control for Immunohostochemestry (E, J, O). Statistical analysis for integrin immunohistochemistry data (P), percentage of positive area for integrin β1, integrin α1, integrin α5, integrin αV and IgG control stains in micromasses with different treatments (control, GDF-5, and BMP-7); mean values are shown with Standard deviations (SD) (<i>n</i> = three independent experiments). One-way Analysis of variance (ANOVA). ***<i>P</i> ˂0.0001, <i>**p</i> ˂0.001, <i>*p</i> ˂0.05. Micromass cultures with Gdf-5 treatment induce α5 integrin expression and Bmp-7 treatment induces αV integrin expression.</p