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[Abstract] Gene transfer to mesenchymal stem cells constitutes a powerful approach to promote their differentiation into the appropriate cartilage phenotype. Although viral vectors represent gold standard vehicles, because of their high efficiency, their use is precluded by important concerns including an elevated immunogenicity and the possibility of insertional mutagenesis. Therefore, the development of new and efficient non-viral vectors is under active investigation. In the present study, we developed new non-viral carriers based on niosomes to promote the effective chondrogenesis of human MSCs. Two different niosome formulations were prepared by varying their composition on non-ionic surfactant, polysorbate 80 solely (P80), or combined with poloxamer 407 (P80PX). The best niosome formulation was proven to transfer a plasmid, encoding for the potent chondrogenic transcription factor SOX9 in hMSC aggregate cultures. Transfection of hMSC aggregates via nioplexes resulted in an increased chondrogenic differentiation with reduced hypertrophy. These results highlight the potential of niosome formulations for gene therapy approaches focused on cartilage repair.

[Abstract] Osteoarthritis (OA) is a heterogeneous disease in which the cross-talk between the cells from different tissues within the joint is affected as the disease progresses. Extracellular vesicles (EVs) are known to have a crucial role in cell-cell communication by means of cargo transfer. Subchondral bone (SB) resident cells and its microenvironment are increasingly recognised to have a major role in OA pathogenesis. The aim of this study was to investigate the EV production from OA SB mesenchymal stromal cells (MSCs) and their possible influence on OA chondrocytes. Small EVs were isolated from OA-MSCs, characterized and cocultured with chondrocytes for viability and gene expression analysis, and compared to small EVs from MSCs of healthy donors (H-EVs). OA-EVs enhanced viability of chondrocytes and the expression of chondrogenesis-related genes, although the effect was marginally lower compared to that of the H-EVs. miRNA profiling followed by unsupervised hierarchical clustering analysis revealed distinct microRNA sets in OA-EVs as compared to their parental MSCs or H-EVs. Pathway analysis of OA-EV miRNAs showed the enrichment of miRNAs implicated in chondrogenesis, stem cells, or other pathways related to cartilage and OA. In conclusion, OA SB MSCs were capable of producing EVs that could support chondrocyte viability and chondrogenic gene expression and contained microRNAs implicated in chondrogenesis support. These EVs could therefore mediate the cross-talk between the SB and cartilage in OA potentially modulating chondrocyte viability and endogenous cartilage regeneration.

[Resumen] La artrosis es una enfermedad heterogénea que afecta a las articulaciones móviles y se caracteriza por estrés celular y degradación de la matriz extracelular (MEC) y del hueso subcondral. Afecta a más de 250 millones de individuos mundialmente y es la principal causa de dolor crónico y limitaciones de movilidad, sobre todo, en personas mayores. Sin embargo, a pesar de todos los avances en investigación, hasta la fecha no existe ningún tratamiento efectivo capaz de curar la artrosis, debido a la falta de conocimiento de la patogénesis de la enfermedad. Existen diferentes tratamientos paliativos e incluso técnicas de terapia celular, pero todavía son incapaces de restaurar el cartílago dañado y devolver la función normal a la articulación. Por tanto, los retos en investigación se centran en estudiar la fisiopatología de la artrosis para encontrar, en última instancia, un tratamiento definitivo. El objetivo de este estudio consistió en optimizar las condiciones de cultivo de una línea inmortalizada de condrocitos artrósicos que se pudiese utilizar como modelo in vitro para estudiar la fisiopatología de la artrosis. Los condrocitos fueron previamente inmortalizados en el laboratorio mediante la transducción del antígeno T grande del virus simio 40 (SV40) y la transcriptasa reversa de la telomerasa humana (hTERT). Estas células se cultivaron y expandieron en monocapa para después cultivarlas en tres dimensiones (3D) formando micromasas en diversos medios de cultivo. Se realizó un análisis histológico, un análisis de la concentración de glucosaminoglicanos (GAGs) y la medición de la expresión de los marcadores de condrocitos primarios [región determinante del sexo del cromosoma Y – caja 9 (SOX9), colágeno tipo II (COL II) y agrecano (ACAN)] mediante la reacción en cadena de la polimerasa cuantitativa (qPCR). Estos resultados confirmaron que no se detectó expresión de COL II y ACAN en las micromasas formadas en los medios de diferenciación condrogénica, mientras, [Resumo] A artrose é unha enfermidade heteroxénea que afecta ás articulacións móbiles e caracterízase por estrés celular e degradación da matriz extracelular (MEC) e do óso subcondral. Afecta a máis de 250 millóns de individuos en todo o mundo e é a principal causa de dor crónica e limitacións de mobilidade, especialmente en persoas maiores. Non obstante, a pesar de todos os avances na investigación, ata a data non existe un tratamento eficaz capaz de curar a artrose, debido ao descoñecemento da patoxénese da enfermidade. Existen diferentes tratamentos paliativos e incluso técnicas de terapia celular, pero aínda así son incapaces de restaurar a cartilaxe danada e devolver a función normal á articulación. Polo tanto, os retos da investigación céntranse en estudar a fisiopatoloxía da artrose para atopar, en definitiva, un tratamento definitivo. O obxectivo deste estudo foi optimizar as condicións de cultivo dunha liña inmortalizada de condrócitos artrósicos que se poderían empregar como modelo in vitro para estudar a fisiopatoloxía da artrose. Os condrócitos foron inmortalizados previamente no laboratorio polo antíxeno T grande 40 do virus simiano (SV40) e a transcriptase inversa da telomerasa humana (hTERT). Estas células cultiváronse e expandíronse nunha monocapa e despois cultiváronse en tres dimensións (3D) formando micromasa en varios medios. Realizouse unha análise histolóxica, unha análise da concentración de glicosaminoglicanos (GAGs) e a medición da expresión dos marcadores condrocitarios primarios [rexión determinante do sexo do cromosoma Y - caixa 9 (SOX9), coláxeno tipo II (COL II) e aggrecano (ACAN)] por reacción en cadea cuantitativa a tempo real da polimerasa (qPCR). Estes resultados confirmaron que non se detectou ningunha expresión de COL II e ACAN nas micromasas formadas nos medios de diferenciación condroxénicos, mentres que se detectou a expresión de SOX9 , un marcador inicial da condroxénese, en todas as mostras. A principal conclusión deste traba, [Abstract] Osteoarthritis is a heterogeneous illness that affects mobile joints and is characterised by cellular stress and degradation of the extracellular matrix (ECM) and subchondral bone. It affects more than 250 million people worldwide and is the main cause of chronic pain and mobility limitations, especially in older people. Nevertheless, despite the advances in research, nowadays there are not effective treatments capable of treating osteoarthritis, due to the lack of knowledge of its pathogenesis. There are different palliative treatments and even cell therapy techniques, but they are still unable to regenerate damaged cartilage and restore normal function to the joint. Therefore, research challenges focus on studying the pathophysiology of osteoarthritis to find a definitive treatment. The objective of this study was to optimize the culture conditions of an immortalized osteoarthritic chondrocytes line that could be used as an in vitro model to study the pathophysiology of osteoarthritis. Chondrocytes were previously immortalized in the laboratory by the transduction of simian virus 40 large T antigen (SV40) and human telomerase reverse transcriptase (hTERT). We cultured and expanded these cells in a monolayer culture and then cultured them in three dimensions (3D) by forming micromasses in different media. We performed a histological analysis, an analysis of the glycosaminoglycans (GAGs) concentration and the measurement of the expression of primary chondrocyte markers [SRYbox transcription factor 9 (SOX9), type II collagen (COL II) and aggrecan (ACAN)] by real time quantitative polymerase chain reaction (qPCR). These results confirmed that COL II and ACAN were not expressed in the micromasses formed in the chondrogenic differentiation media. On the other hand, we detected the expression of SOX9, an early marker of chondrogenesis, in all samples. The main conclusion of this study is that it has not been possible to correctly determine the ideal culture co

[Abstract] Reprogramming somatic cells toward pluripotency became possible over a decade ago. Since then, induced pluripotent stem cells (iPSCs) have served as a versatile and powerful tool not only for basic research but also with the long-term goal of using them in human cell transplantation after differentiation. Nonetheless, downstream applications are frequently blurred by the difficulties that researchers have to face when working with iPSCs, such as trouble with clonal selection, in vitro culture and cryopreservation, adaptation to feeder-free conditions, or expansion of the cells. Therefore, in this article we aim to provide other researchers with practical and detailed information to successfully culture and adapt iPSCs. Specifically, we (1) describe the most common problems when in-vitro culturing iPSCs onto feeder cells as well as its possible troubleshooting, and (2) compare different matrices and culture media for adapting the iPSCs to feeder-free conditions. We believe that the troubleshooting and recommendations provided in this article can be of use to other researchers working with iPSCs and who may be experiencing similar issues, hopefully enhancing the appeal of this promising cell source to be used for biomedical investigations, such as tissue engineering or regenerative medicine applications.

[Abstract] Background: Mesenchymal stromal cells (MSCs) have the capacity for self-renewal and multi-differentiation, and for this reason they are considered a potential cellular source in regenerative medicine of cartilage and bone. However, research on this field is impaired by the predisposition of primary MSCs to senescence during culture expansion. Therefore, the aim of this study was to generate and characterize immortalized MSC (iMSC) lines from aged donors. Methods: Primary MSCs were immortalized by transduction of simian virus 40 large T antigen (SV40LT) and human telomerase reverse transcriptase (hTERT). Proliferation, senescence, phenotype and multi-differentiation potential of the resulting iMSC lines were analyzed. Results: MSCs proliferate faster than primary MSCs, overcome senescence and are phenotypically similar to primary MSCs. Nevertheless, their multi-differentiation potential is unbalanced towards the osteogenic lineage. There are no clear differences between osteoarthritis (OA) and non-OA iMSCs in terms of proliferation, senescence, phenotype or differentiation potential. Conclusions: Primary MSCs obtained from elderly patients can be immortalized by transduction of SV40LT and hTERT. The high osteogenic potential of iMSCs converts them into an excellent cellular source to take part in in vitro models to study bone tissue engineering.

[Abstract] Corneal cryopreservation can partially solve the worldwide concern regarding donor cornea shortage for keratoplasties. In this study, human corneas were cryopreserved using two standard cryopreservation protocols that are employed in the Tissue Bank of the Teresa Herrera Hospital (Spain) to store corneas for tectonic keratoplasties (TK protocol) and aortic valves (AV protocol), and two vitrification protocols, VS55 and DP6. Endothelial viability and general corneal state were evaluated to determine the protocol that provides the best results. The potential corneal cryopreservation protocol was studied in detail taking into consideration some cryopreservation-related variables and the endothelial integrity and stroma arrangement of the resulting cryopreserved corneas. TK corneas showed mostly viable endothelial cells, while the others showed few (AV) or none (DP6 and VS55). The corneal structure was well maintained in TK and AV corneas. TK corneas showed endothelial acellular areas surrounded by injured cells and a normal-like stromal fiber arrangement. Cryoprotectant solutions of the TK protocol presented an increasing osmolality and a physiological pH value. Cooling temperature rate of TK protocol was of 1 °C/min to −40 °C and 3 °C/min to −120 °C, and almost all of dimethyl sulfoxide left the tissue after washing. Future studies should be done changing cryopreservation-related variables of the TK protocol to store corneas of optical grade.

[Resumo] A rexeneración do óso e da cartilaxe tras sufrir un traumatismo ou unha enfermidade dexenerativa segue sendo un gran desafío clínico. Debido á súa capacidade de auto-renovación e multi-diferenciación, as células mesenquimais estromais (MSC) son unha fonte celular moi prometedora para a rexeneración destes tecidos, pero a investigación neste campo está limitada pola tendencia das MSC á senescencia ao seren expandidas en cultivo. A inmortalización das MSC permítelles superar a senescencia, o que supón un impulso para os avances na investigación. Neste estudo desenvolveuse un método para inmortalizar MSC derivadas de doantes de idade avanzada mediante inoculación centrífuga de dous xenes de inmortalización: o antíxeno T grande do virus de simio 40 (SV40LT) e a transcriptase reversa da telomerase humana (hTERT). As MSC inmortalizadas son fenotipicamente similares ás MSC primarias e son capaces de diferenciarse cara ás tres liñaxes esqueléticas, aínda que se inclinan cara á ruta de diferenciación osteoxénica. Os condrocitos articulares e os sinoviocitos pódense inmortalizar empregando o mesmo método, pero os condrocitos inmortalizados son metabolicamente diferentes dos condrocitos articulares primarios. Estas células poden ser útiles como parte de modelos in vitro de rexeneración dos tecidos articulares ou de enfermidade osteocondral., [Resumen] La regeneración del hueso y el cartílago tras sufrir un traumatismo o una enfermedad degenerativa continúa siendo un gran desafío clínico. Debido a su capacidad de auto-renovación y multi-diferenciación, las células mesenquimales estromales (MSC) son una fuente celular prometedora para la regeneración de estos tejidos, pero la investigación en este campo se ve limitada por la tendencia de las MSC a la senescencia en cultivo. La inmortalización de las MSC les permite superar la senescencia, impulsando así los avances en la investigación. En este estudio, se ha desarrollado un método para inmortalizar MSC derivadas de donantes de edad avanzada mediante inoculación centrífuga de dos genes de inmortalización: el antígeno T grande del virus de simio 40 (SV40LT) y la transcriptasa reversa de la telomerasa humana (hTERT). Las MSC inmortalizadas son fenotípicamente similares a las MSC primarias y son capaces de diferenciarse hacia los tres linajes esqueléticos, aunque tienen tendencia a seguir la ruta de diferenciación osteogénica. Los condrocitos articulares y los sinoviocitos se pueden inmortalizar utilizando el mismo método, pero los condrocitos inmortalizados son metabólicamente diferentes de los condrocitos articulares primarios. Estas células pueden ser útiles como parte de modelos in vitro de regeneración de los tejidos articulares o de enfermedad osteocondral., [Abstract] Regeneration of bone and cartilage after trauma or age-related degenerative diseases remains a major clinical challenge. Due to their self-renewal and multi-differentiation potential, mesenchymal stromal cells (MSCs) are a promising cell source for bone and cartilage regeneration, but research on this field is impaired by MSCs’ predisposition to senescence when culture-expanded. Immortalization of MSCs allows them to bypass senescence, thus boosting the advances in MSC research. In this study, a method has been developed to immortalize MSCs derived from elderly donors by spinoculation of two immortalization genes: simian virus 40 large T antigen (SV40LT) and human telomerase reverse transcriptase (hTERT). Immortalized MSCs are phenotypically similar to primary MSCs and are able to differentiate to the three skeletal lineages, although their multi-differentiation potential is unbalanced towards the osteogenic pathway. Articular chondrocytes and synoviocytes can also be immortalized by the same method, but immortalized chondrocytes are metabolically different from primary articular chondrocytes. These immortalized cells can be useful as part of in vitro models of osteochondral regeneration and disease.

[Abstract] Osteoarthritis (OA) is the most common musculoskeletal disorder. Although joint replacement remains the standard of care for knee OA patients, knee joint distraction (KJD), which works by temporarily off-loading the joint for 6–8 weeks, is becoming a novel joint-sparing alternative for younger OA sufferers. The biological mechanisms behind KJD structural improvements remain poorly understood but likely involve jointresident regenerative cells including multipotent stromal cells (MSCs). In this study, we hypothesized that KJD leads to beneficial cartilage-anabolic and anti-catabolic changes in joint-resident MSCs and investigated gene expression profiles of synovial fluid (SF) MSCs following KJD as compared with baseline. To obtain further insights into the effects of local biomechanics on MSCs present in late OA joints, SF MSC gene expression was studied in a separate OA arthroplasty cohort and compared with subchondral bone (SB) MSCs from medial (more loaded) and lateral (less loaded) femoral condyles from the same joints. In OA arthroplasty cohort (n = 12 patients), SF MSCs expressed lower levels of ossification- and hypotrophy-related genes [bone sialoprotein (IBSP), parathyroid hormone 1 receptor (PTH1R), and runt-related transcription factor 2 (RUNX2)] than did SB MSCs. Interestingly, SF MSCs expressed 5- to 50-fold higher levels of transcripts for classical extracellular matrix turnover molecules matrix metalloproteinase 1 (MMP1), a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5), and tissue inhibitor of metalloproteinase-3 (TIMP3), all (p < 0.05) potentially indicating greater cartilage remodeling ability of OA SF MSCs, compared with SB MSCs. In KJD cohort (n = 9 patients), joint off-loading resulted in sustained, significant increase in SF MSC colonies’ sizes and densities and a notable transcript upregulation of key cartilage core protein aggrecan (ACAN) (weeks 3 and 6), as well as reduction in pro-inflammatory C–C motif

[Abstract] Induced pluripotent stem cells (iPSCs) represent an unlimited source of pluripotent cells capable of di erentiating into any cell type of the body. Several studies have demonstrated the valuable use of iPSCs as a tool for studying the molecular and cellular mechanisms underlying disorders a ecting bone, cartilage and muscle, as well as their potential for tissue repair. Musculoskeletal diseases are one of the major causes of disability worldwide and impose an important socio-economic burden. To date there is neither cure nor proven approach for e ectively treating most of these conditions and therefore new strategies involving the use of cells have been increasingly investigated in the recent years. Nevertheless, some limitations related to the safety and di erentiation protocols among others remain, which humpers the translational application of these strategies. Nonetheless, the potential is indisputable and iPSCs are likely to be a source of di erent types of cells useful in the musculoskeletal field, for either disease modeling or regenerative medicine. In this review, we aim to illustrate the great potential of iPSCs by summarizing and discussing the in vitro tissue regeneration preclinical studies that have been carried out in the musculoskeletal field by using iPSCs.

[Abstract] Background: The biological mechanisms that contribute to atrophic long bone non-union are poorly understood. Multipotential mesenchymal stromal cells (MSCs) are key contributors to bone formation and are recognised as important mediators of blood vessel formation. This study examines the role of MSCs in tissue formation at the site of atrophic non-union. Materials and Methods: Tissue and MSCs from non-union sites (n = 20) and induced periosteal (IP) membrane formed following the Masquelet bone reconstruction technique (n = 15) or bone marrow (n = 8) were compared. MSC content, differentiation, and influence on angiogenesis were measured in vitro. Cell content and vasculature measurements were performed by flow cytometry and histology, and gene expression was measured by quantitative polymerase chain reaction (qPCR). Results: MSCs from non-union sites had comparable differentiation potential to bone marrow MSCs. Compared with induced periosteum, non-union tissue contained similar proportion of colony-forming cells, but a greater proportion of pericytes (p = 0.036), and endothelial cells (p = 0.016) and blood vessels were more numerous (p = 0.001) with smaller luminal diameter (p = 0.046). MSCs showed marked differences in angiogenic transcripts depending on the source, and those from induced periosteum, but not non-union tissue, inhibited early stages of in vitro angiogenesis. Conclusions: In vitro, non-union site derived MSCs have no impairment of differentiation capacity, but they differ from IP-derived MSCs in mediating angiogenesis. Local MSCs may thus be strongly implicated in the formation of the immature vascular network at the non-union site. Attention should be given to their angiogenic support profile when selecting MSCs for regenerative therapy.

[Abstract] Human bone marrow-derived mesenchymal stromal cells (MSCs) obtained from aged patients are prone to senesce and diminish their differentiation potential, therefore limiting their usefulness for osteochondral regenerative medicine approaches or to study age-related diseases, such as osteoarthiritis (OA). MSCs can be transduced with immortalizing genes to overcome this limitation, but transduction of primary slow-dividing cells has proven to be challenging. Methods for enhancing transduction efficiency (such as spinoculation, chemical adjuvants, or transgene expression inductors) can be used, but several parameters must be adapted for each transduction system. In order to develop a transduction method suitable for the immortalization of MSCs from aged donors, we used a spinoculation method. Incubation parameters of packaging cells, speed and time of centrifugation, and valproic acid concentration to induce transgene expression have been adjusted. In this way, four immortalized MSC lines (iMSC#6, iMSC#8, iMSC#9, and iMSC#10) were generated. These immortalized MSCs (iMSCs) were capable of bypassing senescence and proliferating at a higher rate than primary MSCs. Characterization of iMSCs showed that these cells kept the expression of mesenchymal surface markers and were able to differentiate towards osteoblasts, adipocytes, and chondrocytes. Nevertheless, alterations in the CD105 expression and a switch of cell fate-commitment towards the osteogenic lineage have been noticed. In conclusion, the developed transduction method is suitable for the immortalization of MSCs derived from aged donors. The generated iMSC lines maintain essential mesenchymal features and are expected to be useful tools for the bone and cartilage regenerative medicine research.

[Abstract] Here, we report the establishment of the human iPS cell line N1-FiPS4F#7 generated from skin cells of a patient with no rheumatic diseases, thus obtaining an appropriate control iPS cell line for researchers working in the field of rheumatic diseases. The reprogramming factors Oct4, Sox2, Klf4 and c-Myc were introduced using a non-integrating reprogramming strategy involving Sendai Virus.

[Abstract] Knowledge and research results about hand osteoarthritis (hOA) are limited due to the lack of samples and animal models of the disease. Here, we report the generation of two induced pluripotent stem cell (iPSC)-lines from patients with radiographic hOA. Furthermore, we wondered whether these iPSC-lines carried single nucleotide polymorphisms (SNPs) within genes that have been associated with hOA. Finally, we performed chondrogenic differentiation of the iPSCs in order to prove their usefulness as cellular models of the disease. We performed a non-integrative reprogramming of dermal fibroblasts obtained from two patients with radiographic rhizarthrosis and non-erosive hOA by introducing the transcriptional factors Oct4, Sox2, Klf4 and c-Myc using Sendai virus. After reprogramming, embryonic stem cell-like colonies emerged in culture, which fulfilled all the criteria to be considered iPSCs. Both iPSC-lines carried variants associated with hOA in the four studied genes and showed differences in their chondrogenic capacity when compared with a healthy control iPSC-line. To our knowledge this is the first time that the generation of iPSC-lines from patients with rhizarthrosis and non-erosive hOA is reported. The obtained iPSC-lines might enable us to model the disease in vitro, and to deeper study both the molecular and cellular mechanisms underlying hOA.

[Abstract] Culture expansion of MSCs has detrimental effects on various cell characteristics and attributes (e.g., phenotypic changes and senescence), which, in addition to inherent interdonor variability, negatively impact the standardization and reproducibility of their therapeutic potential. The identification of innate distinct functional MSC subpopulations, as well as the description of ex vivo protocols aimed at maintaining phenotypes and enhancing specific functions have the potential to overcome these limitations. The incorporation of those approaches into cell-based therapy would significantly impact the field, as more reproducible clinical outcomes may be achieved.

[Abstract] This work proposes a method based on image analysis and machine and statistical learning to model and estimate osteocyte growth (in type I collagen scaffolds for bone regeneration systems) and the collagen degradation degree due to cellular growth. To achieve these aims, the mass of collagen -subjected to the action of osteocyte growth and differentiation from stem cells- was measured on 3 days during each of 2 months, under conditions simulating a tissue in the human body. In addition, optical microscopy was applied to obtain information about cellular growth, cellular differentiation, and collagen degradation. Our first contribution consists of the application of a supervised classification random forest algorithm to image texture features (the structure tensor and entropy) for estimating the different regions of interest in an image obtained by optical microscopy: the extracellular matrix, collagen, and image background, and nuclei. Then, extracellular-matrix and collagen regions of interest were determined by the extraction of features related to the progression of the cellular growth and collagen degradation (e.g., mean area of objects and the mode of an intensity histogram). Finally, these critical features were statistically modeled depending on time via nonparametric and parametric linear and nonlinear models such as those based on logistic functions. Namely, the parametric logistic mixture models provided a way to identify and model the degradation due to biological activity by estimating the corresponding proportion of mass loss. The relation between osteocyte growth and differentiation from stem cells, on the one hand, and collagen degradation, on the other hand, was determined too and modeled through analysis of image objects’ circularity and area, in addition to collagen mass loss. This set of imaging techniques, machine learning procedures, and statistical tools allowed us to characterize and parameterize type I collagen biodegradation when

[Abstract] The unavailability of sufficient numbers of human primary cells is a major roadblock for in vitro repair of bone and/or cartilage, and for performing disease modelling experiments. Immortalized mesenchymal stromal cells (iMSCs) may be employed as a research tool for avoiding these problems. The purpose of this review was to revise the available literature on the characteristics of the iMSC lines, paying special attention to the maintenance of the phenotype of the primary cells from which they were derived, and whether they are effectively useful for in vitro disease modeling and cell therapy purposes. This review was performed by searching on Web of Science, Scopus, and PubMed databases from 1 January 2015 to 30 September 2019. The keywords used were ALL = (mesenchymal AND (“cell line” OR immortal*) AND (cartilage OR chondrogenesis OR bone OR osteogenesis) AND human). Only original research studies in which a human iMSC line was employed for osteogenesis or chondrogenesis experiments were included. After describing the success of the immortalization protocol, we focused on the iMSCs maintenance of the parental phenotype and multipotency. According to the literature revised, it seems that the maintenance of these characteristics is not guaranteed by immortalization, and that careful selection and validation of clones with particular characteristics is necessary for taking advantage of the full potential of iMSC to be employed in bone and cartilage-related research.

[Abstract] Osteoarthritis (OA) is the most common degenerative joint disorder. Multipotential stromal cells (MSCs) have a crucial role in joint repair, but how OA severity affects their characteristics remains unknown. Knee OA provides a good model to study this, as osteochondral damage is commonly more severe in the medial weight-bearing compartment compared to lateral side of the joint. This study utilised in vitro functional assays, cell sorting, gene expression and immunohistochemistry to compare MSCs from medial and lateral OA femoral condyles. Despite greater cartilage loss and bone sclerosis in medial condyles, there was no significant differences in MSC numbers, growth rates or surface phenotype. Culture-expanded and freshly-purified medial-condyle MSCs expressed higher levels of several ossification-related genes. Using CD271-staining to identify MSCs, their presence and co-localisation with TRAP-positive chondroclasts was noted in the vascular channels breaching the osteochondral junction in lateral condyles. In medial condyles, MSCs were additionally found in small cavities within the sclerotic plate. These data indicate subchondral MSCs may be involved in OA progression by participating in cartilage destruction, calcification and sclerotic plate formation and that they remain abundant in severe disease. Biological or biomechanical modulation of these MSCs may be a new strategy towards cartilage and bone restoration in knee OA.

[Abstract] The poly(DL-lactide-co-glycolide) (PDLGA) copolymers have been specifically designed and performed as biomaterials, taking into account their biodegradability and biocompatibility properties. One of the applications of statistical degradation models in material engineering is the estimation of the materials degradation level and reliability. In some reliability studies, as the present case, it is possible to measure physical degradation (mass loss, water absorbance, pH) depending on time. To this aim, we propose an expert system able to provide support in collagen degradation analysis through computer vision methods and statistical modelling techniques. On this base, the researchers can determine which statistical model describes in a better way the biomaterial behaviour. The expert system was trained and evaluated with a corpus of 63 images (2D photographs obtained by electron microscopy) of human mesenchymal stem cells (CMMh-3A6) cultivated in a laboratory experiment lasting 44 days. The collagen type-1 sponges were arranged in 3 groups of 21 samples (each image was obtained in intervals of 72 hours).

[Abstract] The Poly(DL-lactide-co-glycolide) copolymers (PDLGA) have designed and performed as biomaterials, taking into account their biodegradability and biocompatibility properties. These materials have a wide range of application in medicine such as orthopedic implants, general surgical implants (suture materials), osteosynthesis, bone cement, among many others. For these reasons, in this paper, we present an intelligent educational environment that can be used for both, researchers and students interested in the analysis of the biomaterial behavior under certain conditions. Our platform includes a Learning Objects (LOs) for MOODLE, and in the same way, implements several digital image processing techniques as well as a decision support module based on a random forest algorithm and statistical modeling. With the aim of determining the real feasibility of this proposal, we have presented the system to 34 Ecuadorian engineering students. After testing the platform, the students answered a survey aimed at determining their perception of the system. The results provide several guidelines to continue with the developing of the platform.

[Abstract] The establishment of cartilage regenerative medicine is an important clinical issue, but the search for cell sources able to restore cartilage integrity proves to be challenging. Human mesenchymal stromal cells (MSCs) are prone to form epiphyseal or hypertrophic cartilage and have an age-related limited proliferation. On the other hand, it is difficult to obtain functional chondrocytes from human embryonic stem cells (ESCs). Moreover, the ethical issues associated with human ESCs are an additional disadvantage of using such cells. Since their discovery in 2006, induced pluripotent stems cells (iPSCs) have opened many gateways to regenerative medicine research, especially in cartilage tissue engineering therapies. iPSCs have the capacity to overcome limitations associated with current cell sources since large numbers of autologous cells can be derived from small starting populations. Moreover, problems associated with epiphyseal or hypertrophic-cartilage formation can be overcome using iPSCs. iPSCs emerge as a promising cell source for treating cartilage defects and have the potential to be used in the clinical field. For this purpose, robust protocols to induce chondrogenesis, both in vitro an in vivo, are required. This review summarises the recent progress in iPSC technology and its applications for cartilage repair.

[Abstract] The integrity of the articular cartilage is necessary for the proper functioning of the diarthrodial joint. The self-repair capacity of this tissue is very limited and, currently, there is no effective treatment capable of restoring it. The degradation of the articular cartilage leads to osteoarthritis (OA), a leading cause of pain and disability mainly among older people. Different cell treatments have been developed with the aim of forming a repair tissue with the characteristics of native articular cartilage, including cellular therapy and tissue engineering. Cell therapy-based approaches include bone marrow-stimulating techniques, implants of periosteum and perichondrium, ostechondral grafting and implantation of chondrogenic cells as chondrocytes, mesenchymal stem cells or induced pluripotent stem cells. In tissue engineering-based approaches cell-free scaffolds capable of recruiting endogenous cells or chondrogenic cell-loaded scaffolds may be used. However, despite the numerous treatments available nowadays, no technique has been able to consistently regenerate native articular cartilage in clinical trials. Although many cell therapy and tissue engineering studies have shown promising results and clinical improvement, these treatments generate a fibrocartilaginous tissue different from native articular cartilage. More research is needed to improve cell-based approaches and prove its efficacy.

[Purpose] Bone-marrow mesenchymal stem cells (BM-MSCs) are multipotent self-renewal adult cells with potential to regenerate the damaged tissues in degenerative diseases such as osteoarthritis (OA). Nevertheless, research require in vitro expansion of BM-MSCs, a process which eventually causes cell senescence. To overcome this problem cell lines can be used but, currently, BM-MSC lines available are scarce and present limitations regarding their differentiation capacities. For this reason, the aim of this study was to generate and characterize human BM-MSCs lines, derived from an OA patient and a healthy donor, with high chondrogenic and osteogenic capacities for their use in research on Regenerative Medicine for OA.