Aldazabal, J. (Javier)

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    Contractile force assessment methods for in vitro skeletal muscle tissues.
    (eLIFE Sciences Publ. LTD., 2022) Vallejo-Illarramendi, A. (Ainara); Aldazabal, J. (Javier); Vesga-Castro, C. (Camila); Paredes-Puente, J. (Jacobo)
    Over the last few years, there has been growing interest in measuring the contractile force (CF) of engineered muscle tissues to evaluate their functionality. However, there are still no standards available for selecting the most suitable experimental platform, measuring system, culture protocol, or stimulation patterns. Consequently, the high variability of published data hinders any comparison between different studies. We have identified that cantilever deflection, post deflection, and force transducers are the most commonly used configurations for CF assessment in 2D and 3D models. Additionally, we have discussed the most relevant emerging technologies that would greatly complement CF evaluation with intracellular and localized analysis. This review provides a comprehensive analysis of the most significant advances in CF evaluation and its critical parameters. In order to compare contractile performance across experimental platforms, we have used the specific force (sF, kN/m2 ), CF normalized to the calculated cross-sectional area (CSA). However, this parameter presents a high variability throughout the different studies, which indicates the need to identify additional parameters and complementary analysis suitable for proper comparison. We propose that future contractility studies in skeletal muscle constructs report detailed information about construct size, contractile area, maturity level, sarcomere length, and, ideally, the tetanusto-twitch ratio. These studies will hopefully shed light on the relative impact of these variables on muscle force performance of engineered muscle constructs. Prospective advances in muscle tissue engineering, particularly in muscle disease models, will require a joint effort to develop standardized methodologies for assessing CF of engineered muscle tissues.
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    Molecular and Cellular Mechanisms of Delayed Fracture Healing in Mmp10 (Stromelysin 2) Knockout Mice
    (Wiley, 2021) Calvo, I.A. (Isabel A.); Granero-Moltó, F. (Froilán); Paramo, J.A. (José Antonio); Ripalda-Cemboráin, P. (Purificación); Montiel-Terrón, V. (Verónica); Aldazabal, J. (Javier); Orbe, J. (Josune); Rodriguez, J.A. (José Antonio); Lopez, T. (Tania); Valdés-Fernández, J. (José); Muiños-López, E. (Emma); Romero-Torrecilla, J.A. (Juan Antonio); Prosper-Cardoso, F. (Felipe); Saez, B. (Borja)
    The remodeling of the extracellular matrix is a central function in endochondral ossification and bone homeostasis. During secondary fracture healing, vascular invasion and bone growth requires the removal of the cartilage intermediate and the coordinate action of the collagenase matrix metalloproteinase (MMP)-13, produced by hypertrophic chondrocytes, and the gelatinase MMP-9, produced by cells of hematopoietic lineage. Interfering with these MMP activities results in impaired fracture healing characterized by cartilage accumulation and delayed vascularization. MMP-10, Stromelysin 2, a matrix metalloproteinase with high homology to MMP-3 (Stromelysin 1), presents a wide range of putative substrates identified in vitro, but its targets and functions in vivo and especially during fracture healing and bone homeostasis are not well defined. Here, we investigated the role of MMP-10 through bone regeneration in C57BL/6 mice. During secondary fracture healing, MMP-10 is expressed by hematopoietic cells and its maximum expression peak is associated with cartilage resorption at 14 days post fracture (dpf). In accordance with this expression pattern, when Mmp10 is globally silenced, we observed an impaired fracture-healing phenotype at 14 dpf, characterized by delayed cartilage resorption and TRAP-positive cell accumulation. This phenotype can be rescued by a non-competitive transplant of wild-type bone marrow, indicating that MMP-10 functions are required only in cells of hematopoietic linage. In addition, we found that this phenotype is a consequence of reduced gelatinase activity and the lack of proMMP-9 processing in macrophages. Our data provide evidence of the in vivo function of MMP-10 during endochondral ossification and defines the macrophages as the lead cell population in cartilage removal and vascular invasion
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    Murine femur micro-computed tomography and biomechanical datasets for an ovariectomy-induced osteoporosis model.
    (Nature, 2021-09) Ortuzar, N. (Naiara); Lopez-Linares, K. (Karen); Alcorta-Sevillano, N. (Natividad); Aldazabal, J. (Javier); Macias, I. (Iratxe); Infante, A. (Arantza); Bulnes, S. (Susana); Lafuente, J.V. (José Vicente); Rodriguez, C.I. (Clara I.); Stephens, M. (Maialen)
    The development of new effective and safer therapies for osteoporosis, in addition to improved diagnostic and prevention strategies, represents a serious need in the scientific community. Micro-CT image-based analyses in association with biomechanical testing have become pivotal tools in identifying osteoporosis in animal models by assessment of bone microarchitecture and resistance, as well as bone strength. Here, we describe a dataset of micro-CT scans and reconstructions of 15 whole femurs and biomechanical tests on contralateral femurs from C57BL/6JOlaHsd ovariectomized (OVX), resembling human post-menopausal osteoporosis, and sham operated (sham) female mice. Data provided for each mouse include: the acquisition images (.tiff), the reconstructed images (.bmp) and an.xls file containing the maximum attenuations for each reconstructed image. Biomechanical data include an.xls file with the recorded load-displacement, a movie with the filmed test and an.xls file collecting all biomechanical results.
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    Technological advances in ocular trabecular meshwork in vitro models for glaucoma research
    (Wiley, 2022) Moreno-Montañes, J. (Javier); Aldazabal, J. (Javier); Extramiana, L. (Leire); Bikuna-Izagirre, M. (María); Paredes-Puente, J. (Jacobo); Carnero, E. (Elena)
    Glaucoma is the leading cause of irreversible blindness worldwide and ischaracterized by the progressive degeneration of the optic nerve. Intraocularpressure (IOP), which is considered to be the main risk factor for glaucomadevelopment, builds up in response to the resistance (resistance to what?) providedby the trabecular meshwork (TM) to aqueous humor (AH) outflow. Although theTMand its relationship to AH outflow have remained at the forefront of scientificinterest, researchers remain uncertain regarding which mechanisms drive thedeterioration of the TM. Current tissue‐engineering fabrication techniques havecome up with promising approaches to successfully recreate the TM. Nonetheless,more accurate models are needed to understand the factors that make glaucomaarise. In this review, we provide a chronological evaluation of the technologicalmilestones that have taken place in the field of glaucoma research, and we conduct acomprehensive comparison of available TM fabrication technologies. Additionally,we also discuss AH perfusion platforms, since they are essential for the validation ofthese scaffolds, as well as pressure–outflow relationship studies and the discovery ofnew IOP‐reduction therapies.
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    Three-dimensional bioprinting scaffolding for nasal cartilage defects: A systematic review.
    (Springer, 2021) Garcés, J.P. (Juan P.); Chiesa-Estomba, C.M. (Carlos M.); Delgado, A. (Alba); Hernaez-Moya, R. (Raquel); Aldazabal, J. (Javier); Rodino, C. (Claudia); González-Fernández, I. (Iago); Altuna, X. (Xabier); Izeta, A. (Ander); Aiastui, A. (Ana); Paredes-Puente, J. (Jacobo)
    In recent years, three-dimensional (3D)-printing of tissue-engineered cartilaginous scaffolds is intended to close the surgical gap and provide bio-printed tissue designed to fit the specific geometric and functional requirements of each cartilage defect, avoiding donor site morbidity and offering a personalizing therapy. METHODS: To investigate the role of 3D-bioprinting scaffolding for nasal cartilage defects repair a systematic review of the electronic databases for 3D-Bioprinting articles pertaining to nasal cartilage bio-modelling was performed. The primary focus was to investigate cellular source, type of scaffold utilization, biochemical evaluation, histological analysis, in-vitro study, in-vivo study, animal model used, length of research, and placement of experimental construct and translational investigation. RESULTS: From 1011 publications, 16 studies were kept for analysis. About cellular sources described, most studies used primary chondrocyte cultures. The cartilage used for cell isolation was mostly nasal septum. The most common biomaterial used for scaffold creation was polycaprolactone alone or in combination. About mechanical evaluation, we found a high heterogeneity, making it difficult to extract any solid conclusion. Regarding biological and histological characteristics of each scaffold, we found that the expression of collagen type I, collagen Type II and other ECM components were the most common patterns evaluated through immunohistochemistry on in-vitro and in-vivo studies. Only two studies made an orthotopic placement of the scaffolds. However, in none of the studies analyzed, the scaffold was placed in a subperichondrial pocket to rigorously simulate the cartilage environment. In contrast, scaffolds were implanted in a subcutaneous plane in almost all of the studies included. CONCLUSION: The role of 3D-bioprinting scaffolding for nasal cartilage defects repair is growing field. Despite the amount of information collected in the last years and the first surgical applications described recently in humans. Further investigations are needed due to the heterogeneity on mechanical evaluation parameters, the high level of heterotopic scaffold implantation and the need for quantitative histological data.
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    Gelatin blends enhance performance of electrospun polymeric scaffolds in comparison to coating protocols.
    (MDPI, 2022-04) Aldazabal, J. (Javier); Bikuna-Izagirre, M. (María); Paredes-Puente, J. (Jacobo)
    The electrospinning of hybrid polymers is a versatile fabrication technique which takes advantage of the biological properties of natural polymers and the mechanical properties of synthetic polymers. However, the literature is scarce when it comes to comparisons of blends regarding coatings and the improvements offered thereby in terms of cellular performance. To address this, in the present study, nanofibrous electrospun scaffolds of polycaprolactone (PCL), their coating and their blend with gelatin were compared. The morphology of nanofibrous scaffolds was analyzed under field emission scanning electron microscopy (FE-SEM), indicating the influence of the presence of gelatin. The scaffolds were mechanically tested with tensile tests; PCL and PCL gelatin coated scaffolds showed higher elastic moduli than PCL/gelatin meshes. Viability of mouse embryonic fibroblasts (MEF) was evaluated by MTT assay, and cell proliferation on the scaffold was confirmed by fluorescence staining. The positive results of the MTT assay and cell growth indicated that the scaffolds of PCL/gelatin excelled in comparison to other scaffolds, and may serve as good candidates for tissue engineering applications.
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    Nanofibrous PCL-based human trabecular meshwork for aqueous humor outflow studies
    (2023) Moreno-Montañes, J. (Javier); Aldazabal, J. (Javier); Extramiana, L. (Leire); Bikuna-Izagirre, M. (María); Paredes-Puente, J. (Jacobo); Carnero, E. (Elena)
    Primary open-angle glaucoma is characterized by the progressive degeneration of the optic nerve, with the high intraocular pressure (IOP) being one of the main risk factors. The human trabecular meshwork (HTM), specifically the juxtacanalicular tissue (JCT), is responsible for placing resistance to the aqueous humor (AH) outflow and the resulting IOP control. Currently, the lack of a proper in vitro JCT model and the complexity of three-dimensional models impede advances in understanding the relationship between AH outflow and HTM degeneration. Therefore, we design an in vitro JCT model using a polycaprolactone (PCL) nanofibrous scaffold, which supports cells to recapitulate the functional JCT morphology and allow the study of outflow physiology. Mechanical and morphological characterizations of the electrospun membranes were performed, and human trabecular meshwork cells were seeded over the scaffolds. The engineered JCT was characterized by scanning electron microscopy, quantitative real-time polymerase chain reaction, and immunochemistry assays staining HTM cell markers and proteins. A pressure-sensitive perfusion system was constructed and used for the investigation of the outflow facility of the polymeric scaffold treated with dexamethasone (a glucocorticoid) and netarsudil (a novel IOP lowering the rho inhibitor). Cells in the in vitro model exhibited an HTM-like morphology, expression of myocilin, fibronectin, and collagen IV, genetic expression, outflow characteristics, and drug responsiveness. Altogether, the present work develops an in vitro JCT model to better understand HTM cell biology and the relationship between the AH outflow and the HTM and allow further drug screening of pharmacological agents that affect the trabecular outflow facility.
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    Hydrogen embrittlement susceptibility of R4 and R5 high-strength mooring steels in cold and warm seawater
    (MDPI AG, 2018) Arredondo, A. (Alberto); Aldazabal, J. (Javier); Artola-Beobide, G.(Garikoitz); Fernández-Calvo, A.I. (Ana Isabel)
    Hydrogen embrittlement susceptibility ratios calculated from slow strain rate tensile tests have been employed to study the response of three high-strength mooring steels in cold and warm synthetic seawater. The selected nominal testing temperatures have been 3 ◦C and 23 ◦C in order to resemble sea sites of offshore platform installation interest, such as the North Sea and the Gulf of Mexico, respectively. Three scenarios have been studied for each temperature: free corrosion, cathodic protection and overprotection. An improvement on the hydrogen embrittlement tendency of the steels has been observed when working in cold conditions. This provides a new insight on the relevance of the seawater temperature as a characteristic to be taken into account for mooring line design in terms of hydrogen embrittlement assessment.
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    Ex vivo maturation of 3D-Printed, chondrocyte-laden, polycaprolactone-based scaffolds prior to transplantation improves engineered cartilage substitute properties and integration
    (Sage, 2022-10) Chiesa-Estomba, C.M. (Carlos M.); Delgado, A. (Alba); Fernández-Blanco, G. (Gonzalo); Hernaez-Moya, R. (Raquel); Aldazabal, J. (Javier); Rodino, C. (Claudia); Izeta, A. (Ander); Aiastui, A. (Ana); Paredes-Puente, J. (Jacobo)
    The surgical management of nasal septal defects due to perforations, malformations, congenital cartilage absence, traumatic defects, or tumors would benefit from availability of optimally matured septal cartilage substitutes. Here, we aimed to improve in vitro maturation of 3-dimensional (3D)-printed, cell-laden polycaprolactone (PCL)-based scaffolds and test their in vivo performance in a rabbit auricular cartilage model.
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    Hydrogen Assisted Fracture of 30MnB5 High Strength Steel: A Case Study
    (2020) Aldazabal, J. (Javier); Artola-Beobide, G.(Garikoitz)
    When steel components fail in service due to the intervention of hydrogen assisted cracking, discussion of the root cause arises. The failure is frequently blamed on component design, working conditions, the manufacturing process, or the raw material. This work studies the influence of quench and tempering and hot-dip galvanizing on the hydrogen embrittlement behavior of a high strength steel. Slow strain rate tensile testing has been employed to assess this influence. Two sets of specimens have been tested, both in air and immersed in synthetic seawater, at three process steps: in the delivery condition of the raw material, after heat treatment and after heat treatment plus hot-dip galvanizing. One of the specimen sets has been tested without further manipulation and the other set has been tested after applying a hydrogen effusion treatment. The outcome, for this case study, is that fracture risk issues only arise due to hydrogen re-embrittlement in wet service.