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Osteosarcoma is a rare disease that mainly affects children, adolescents, and young people. It can develop in any bone, but the most common ones affected are the proximal humerus and the knees. Nowadays, treatment consists of chemotherapy and surgery. Despite the improvement in the patient’s survival, the 5-year event-free survival remains less than 20% in those with aggressive osteosarcoma. The lack of response in metastatic tumours is the reason why new strategies are needed. In this context, the use of T cells expressing chimeric antigen receptors (CAR T) has shown positive results in the treatment of haematological cancers, such as leukaemia or lymphomas. Nonetheless, the application of this therapy in the treatment of solid tumours, like osteosarcoma, must face some challenges due to the nature of this tumours: the homing of CAR T cells to the tumour, the antigen heterogeneity of solid tumours and the tumour microenvironment. In order to improve the outcome of patients, we need to solve the problems that these obstacles pose to us. The tumour microenvironment is mainly composed of tumour associated macrophages (TAMs), which are involved in numerous immunosuppressive pathways. This tumoral microenvironment (TME) decreases the efficacy of CAR T cell therapy. It was, therefore, the main aim of this work to study the role that macrophages have in the functionality of the CAR T cell therapy by developing an in vitro model. To achieve this goal, peripheral blood-derived human monocytes were isolated and differentiated to M1 and M2 macrophages. They were then cocultured with osteosarcoma cells and CAR T to study their interactions. A successful polarization of the macrophages was achieved. It has been found that M2 macrophages inhibit CAR T’s cytotoxic activity in the short term and M1 macrophages improve their functionality. However, as coculture time progresses, the expression of M1 markers increases in M2 macrophages losing their immunosuppressive potential. This i

Los sarcomas de hueso (SH) representan un grupo heterogéneo de enfermedades con diferente patología y malignidad. Históricamente, los SH han sido clasificados según el tejido o célula que asemejan; sin embargo todos ellos se originan en tejidos conectivos donde las MSCs juegan un papel fundamental. Las MSCs han sido propuestas como células iniciadoras de distintos sarcomas murinos y células con características similares han sido identificadas también en los cánceres humanos. Las CSCs han sido identificadas como las responsables de la heterogeneidad fenotípica y funcional típicas de los tumores. Sin embargo, el modelo de CSCs no es el único propuesto por explicar las dinámicas de crecimiento tumoral y la generación de la heterogeneidad tumoral. Los datos genómicos sugieren que existen distintos modelos de crecimiento tumoral, hecho que está relacionado con el pronóstico de los pacientes, con la progresión tumoral y con la respuesta a la terapia. Sin embargo, en este marco evolutivo las variaciones genéticas y fenotípicas no son los únicos factores que determinan el crecimiento tumoral. De hecho, las interacciones con el microambiente y en especial con el sistema inmune juegan un papel fundamental en la oncogénesis. Aunque se han descrito distintas estrategias de escape inmunológico, la caracterización del microambiente inmunitario no es completa..., Bone sarcomas (BS) comprise a heterogeneous group of diseases, including neoplasms with different pathological appearance and malignancy. BS have been historically classified according to the cell or tissue type which they most resemble, nevertheless they all originate in connective tissues where mesenchymal stem cells (MSCs) play a fundamental role. MSCs have been proposed as tumor initiating cells of different types of murine sarcomas, and cells with a stem-like status have been detected in human cancers. Cancer stem cells (CSCs) and the existence of tumor hierarchies have been suggested as mechanisms to explain tumor phenotypic and functional heterogeneity. However, CSCs model is not the only hypothesis to explain both the dynamics of cancer growth and heterogeneity generation in tumors. Genomic data suggest different models of cancer development, which are also related to patients’prognosis, tumor progression and therapeutic response. Not withstanding the former, in this evolutiveframe genetic and phenotypic variations are not the only factors determining cancer development. Indeed, microenvironmental interactions, with special mention to host immune system, play a fundamental role in oncogenesis. Different strategies of immune escape have been described, however, tumor-immune- microenvironment interplay have not been fully delineated...

Los sarcomas de hueso (SH) representan un grupo heterogéneo de enfermedades con diferente patología y malignidad. Históricamente, los SH han sido clasificados según el tejido o célula que asemejan; sin embargo todos ellos se originan en tejidos conectivos donde las MSCs juegan un papel fundamental. Las MSCs han sido propuestas como células iniciadoras de distintos sarcomas murinos y células con características similares han sido identificadas también en los cánceres humanos. Las CSCs han sido identificadas como las responsables de la heterogeneidad fenotípica y funcional típicas de los tumores. Sin embargo, el modelo de CSCs no es el único propuesto por explicar las dinámicas de crecimiento tumoral y la generación de la heterogeneidad tumoral. Los datos genómicos sugieren que existen distintos modelos de crecimiento tumoral, hecho que está relacionado con el pronóstico de los pacientes, con la progresión tumoral y con la respuesta a la terapia. Sin embargo, en este marco evolutivo las variaciones genéticas y fenotípicas no son los únicos factores que determinan el crecimiento tumoral. De hecho, las interacciones con el microambiente y en especial con el sistema inmune juegan un papel fundamental en la oncogénesis. Aunque se han descrito distintas estrategias de escape inmunológico, la caracterización del microambiente inmunitario no es completa...

On-site sanitation is generally advocated as a means to eradicate the health hazards associated with open defecation. While this has provided a welcome upgrade to the livelihoods of millions of people in low-income countries, improved sanitation facilities are increasingly becoming a threat to domestic groundwater-based supplies. Within this context, a survey of pit latrines, domestic wells and improved water sources was carried out in a large rural village of southern Mali. All households were surveyed for water, sanitation and hygiene habits. Domestic wells and improved water sources were georeferenced and sampled for water quality (pH, electric conductivity, temperature, turbidity, total dissolved solids, thermotolerant coliforms, chloride and nitrate) and groundwater level, while all latrines were inspected and georeferenced. A GIS database was then used to evaluate the proportion of water points within the influence area of latrines, as well as to underpin multiple regression models to establish the determinants for fecal contamination in drinking supplies. Moreover, an appraisal of domestic water treatment practices was carried out. This revealed that nearly two-thirds of the population uses bleach to purify drinking supplies, but also that domestic-scale treatment as currently implemented by the population is far from effective. It is thus concluded that existing habits could be enhanced as a means to make water supplies safer. Furthermore, population, well and latrine density were all identified as statistically significant predictors for fecal pollution at different spatial scales. These findings are policy-relevant in the context of groundwater-dependent human settlements, since many countries in the developing world currently pursue the objective of eliminating open defecation., Agencia Española de Cooperación Internacional al Desarrollo (AECID), Depto. de Geodinámica, Estratigrafía y Paleontología, Fac. de Ciencias Geológicas, TRUE, pub

[Background]: Age-associated changes in genomic DNA methylation have been primarily attributed to 5-methylcytosine (5mC). However, the recent discovery of 5-hydroxymethylcytosine (5hmC) suggests that this epigenetic mark might also play a role in the process. [Methods]: Here, we analyzed the genome-wide profile of 5hmc in mesenchymal stem cells (MSCs) obtained from bone-marrow donors, aged 2-89 years. [Results]: We identified 10,685 frequently hydroxymethylated CpG sites in MSCs that were, as in other cell types, significantly associated with low density CpG regions, introns, the histone posttranslational modification H3k4me1 and enhancers. Study of the age-associated changes to 5hmC identified 785 hyper- and 846 hypo-hydroxymethylated CpG sites in the MSCs obtained from older individuals. [Conclusions]: DNA hyper-hydroxymethylation in the advanced-age group was associated with loss of 5mC, which suggests that, at specific CpG sites, this epigenetic modification might play a role in DNA methylation changes during lifetime. Since bone-marrow MSCs have many clinical applications, and the fact that the epigenomic alterations in this cell type associated with aging identified in this study could have associated functional effects, the age of donors should be taken into account in clinical settings.

Mesenchymal stem cells (MSCs) are key components of the bone marrow microenvironment which contribute to the maintenance of the hematopoietic stem cell niche and exert immunoregulatory functions in innate and adaptive immunity. We analyze the immunobiology of MSCs derived from acute lymphoblastic leukemia (ALL) patients and their impact on NK cell function. In contrast to the inhibitory effects on the immune response exerted by MSCs from healthy donors (Healthy-MSCs), we demonstrate that MSCs derived from low/intermediate risk ALL patients at diagnosis (ALL-MSCs) promote an efficient NK cell response including cytokine production, phenotypic activation and most importantly, cytotoxicity. Longitudinal studies indicate that these immunostimulatory effects of ALL-MSCs are progressively attenuated. Healthy-MSCs adopt ALL-MSC-like immunomodulatory features when exposed to leukemia cells, acquiring the ability to stimulate NK cell antitumor function. The mechanisms underlying to these functional changes of ALL-MSCs include reduced production of soluble inhibitory factors, differential expression of costimulatory and coinhibitory molecules, increased expression of specific TLRs and Notch pathway activation. Collectively our findings indicate that, in response to leukemia cells, ALL-MSCs could mediate a host beneficial immunomodulatory effect by stimulating the antitumor innate immune response., Ministerio de Economía, Comercio y Empresa (España), Comunidad de Madrid, Asociación Española contra el Cáncer 2010, Instituto de Salud Carlos III, Depto. de Biología Celular, Fac. de Ciencias Biológicas, TRUE, pub

RESEARCHER ID M-3378-2014 (María Vallet Regí) ORCID 0000-0002-6104-4889 (María Vallet Regí) ORCID 0000-0002-9042-8865 (Alejandro Baeza) RESEARCHER ID: K-8193-2014(Alejandro Baeza), A novel phototriggered drug delivery nanocarrier, which exhibits very high tumor cytotoxicity against human tumoral cells, is presented. This device is based on mesoporous silica nanoparticles decorated with a biocompatible protein shell cleavable by light irradiation. The proteins that compose the protein shell (avidin, streptavidin and biotinylated transferrin) act as targeting and capping agents at the same time, avoiding the use of redundant systems. The light responsive behavior is provided by a biotinylated photocleavable cross-linker covalently grafted on the mesoporous surface, which suffers photocleavage by UV radiation (366 nm). Human tumoral cells incubated in the presence of a very low particle concentration enter into the apoptotic stage after a short irradiation time. Thus, the system described here could be applied to the treatment of exposed tumors that affect the skin, oesophagus, and stomach, among others, and are easily accessible for light irradiation., Ministerio de Economía y Competitividad (MINECO), Comunidad de Madrid, Instituto de Salud Carlos III/Fondos FEDER, RTICC, Depto. de Química en Ciencias Farmacéuticas, Fac. de Farmacia, TRUE, pub

Importance of the field: In the last decade, knowledge of mesenchymal stem cells (MSCs) has evolved rapidly; their immunomodulatory properties and paracrine interactions with specific cell types in damaged tissues and promising results in some clinical applications have made these cells an attractive option for the treatment of certain diseases. Areas covered in this review: We present some relevant methodological issues and biological properties of MSCs, as well as clinical applications of MSC therapies with particular emphasis in the treatment of graft versus host disease (GVHD), complex perianal fistula and refractory metastatic neuroblastoma. Other topical aspects relevant to the application of cellular therapies such as biosafety studies and cellular production of MSCs are also discussed in this review. What the reader will gain: The growing optimism regarding MSCs research is based on the promising results obtained in in vitro and in vivo studies. The rapid translational research with MSCs necessitated standardization of methodology and terminology and greater focus on other aspects such as biosafety and cellular production, especially for clinical use of MSCs. Take home message: Much has been learned about the biology and applications of MSCs and much remains to be learned

Background. We previously reported the in vitro spontaneous transformation of human mesenchymal stem cells (MSC) generating a population with tumorigenic potential, that we termed transformed mesenchymal cells (TMC)., Methodology/Principal Findings. Here we have characterized the molecular changes associated with TMC generation. Using microarrays techniques we identified a set of altered pathways and a greater number of downregulated than upregulated genes during MSC transformation, in part due to the expression of many untranslated RNAs in MSC. Microarray results were validated by qRT-PCR and protein detection., Conclusions/Significance. In our model, the transformation process takes place through two sequential steps; first MSC bypass senescence by upregulating c-myc and repressing p16 levels. The cells then bypass cell crisis with acquisition of telomerase activity, Ink4a/Arf locus deletion and Rb hyperphosphorylation. Other transformation-associated changes include modulation of mitochondrial metabolism, DNA damage-repair proteins and cell cycle regulators. In this work we have characterized the molecular mechanisms implicated in TMC generation and we propose a two-stage model by which a human MSC becomes a tumor cell.