Impact of hydrodynamics on iPSC-derived cardiomyocyte differentiation processes

Cardiomyocytes (CMs), derived from pluripotent stem cells (PSCs), have the potential to be used in cardiac repair. Addition of physical cues, such as electrical and mechanical stimulations, have proven to significantly effect morphology, density, cardiogenesis, maturity and functionality of differentiated CMs. This work combines rigorous fluid dynamics investigation and flow frequency analysis with iPSC differentiation experiments to identify and quantify the flow characteristics leading to a significant increase of differentiation yield. This is towards a better understanding of the physical relationship between frequency modulation and embryoid bodies suspension, and the development of dimensionless correlations applicable at larger scales. Laser Doppler Anemometry and Fast Fourier Transform analysis were used to identify characteristic flow frequencies under different agitation modes. Intermittent agitation resulted in a pattern of low intensity frequencies at reactor scale that could be controlled by varying three identified time components: rotational speed, interval and dwell times. A proof of concept biological study was undertaken, tuning the hydrodynamic environment through variation of dwell time based on the engineering study findings and a significant improvement in CM yield was obtained. This work introduces the concept of fine-tuning the physical hydrodynamic cues within a three-dimensional flow system to improve cardiomyocyte differentiation of iPSC

MonoMAC Syndrome Caused by a Novel GATA2 Mutation Successfully Treated by Allogeneic Hematopoietic Stem Cell Transplantation

info:eu-repo/semantics/publishedVersio

Macrofauna assemblages in a XVIIth century shipwreck: comparison with those on natural reefs and sandy bottoms

The macrofauna assemblages of a XVIIth century shipwreck off southern Portugal were studied and compared with those of nearby natural reefs and sandy bottoms, by underwater visual census. A total of 11 173 specimens of 224 different fauna taxa and 12 phyla were recorded. Natural reefs had the highest density of specimens (35 122 / 1000 m2) followed by the shipwreck (21 392 / 1000 m2) and the sandy bottoms (3771 / 1000 m2). Three biodiversity indices were estimated (Margalef, Shannon- Wiener and Pielou), with the natural reefs showing the highest values. However, the shipwreck presented values relatively similar to those of the natural reefs for the Shannon-Wiener and Pielou indices. The three habitats were clearly distinguishable by multivariate statistical analysis, with the average dissimilarity between sand and shipwreck, and between sand and natural reefs being much higher than that between the shipwreck and the natural reefs. The shipwreck had higher abundances of some commercially important species, such as the pouting Trisopterus luscus, European conger Conger conger, and common spider crab Maja squinado, as well as some vulnerable and threatened species such as the pink seafan Eunicella verrucosa. The results presented emphasize the importance of this habitat on the southern Portuguese coast.info:eu-repo/semantics/publishedVersio

Macrofauna assemblages in a XVIIth century shipwreck: comparison with those on natural reefs and sandy bottoms

The macrofauna assemblages of a XVIIth century shipwreck off southern Portugal were studied and compared with those of nearby natural reefs and sandy bottoms, by underwater visual census. A total of 11 173 specimens of 224 different fauna taxa and 12 phyla were recorded. Natural reefs had the highest density of specimens (35 122 / 1000 m2) followed by the shipwreck (21 392 / 1000 m2) and the sandy bottoms (3771 / 1000 m2). Three biodiversity indices were estimated (Margalef, Shannon- Wiener and Pielou), with the natural reefs showing the highest values. However, the shipwreck presented values relatively similar to those of the natural reefs for the Shannon-Wiener and Pielou indices. The three habitats were clearly distinguishable by multivariate statistical analysis, with the average dissimilarity between sand and shipwreck, and between sand and natural reefs being much higher than that between the shipwreck and the natural reefs. The shipwreck had higher abundances of some commercially important species, such as the pouting Trisopterus luscus, European conger Conger conger, and common spider crab Maja squinado, as well as some vulnerable and threatened species such as the pink seafan Eunicella verrucosa. The results presented emphasize the importance of this habitat on the southern Portuguese coast.info:eu-repo/semantics/publishedVersio

Macrofauna assemblages in a XVIIth century shipwreck: comparison with those on natural reefs and sandy bottoms

The macrofauna assemblages of a XVIIth century shipwreck off southern Portugal were studied and compared with those of nearby natural reefs and sandy bottoms, by underwater visual census. A total of 11 173 specimens of 224 different fauna taxa and 12 phyla were recorded. Natural reefs had the highest density of specimens (35 122 / 1000 m²) followed by the shipwreck (21 392 / 1000 m²) and the sandy bottoms (3771 / 1000 m²). Three biodiversity indices were estimated (Margalef, Shannon-Wiener and Pielou), with the natural reefs showing the highest values. However, the shipwreck presented values relatively similar to those of the natural reefs for the Shannon-Wiener and Pielou indices. The three habitats were clearly distinguishable by multivariate statistical analysis, with the average dissimilarity between sand and shipwreck, and between sand and natural reefs being much higher than that between the shipwreck and the natural reefs. The shipwreck had higher abundances of some commercially important species, such as the pouting Trisopterus luscus, European conger Conger conger, and common spider crab Maja squinado, as well as some vulnerable and threatened species such as the pink seafan Eunicella verrucosa. The results presented emphasize the importance of this habitat on the southern Portuguese coast.As comunidades de macrofauna de um naufrágio do século XVII ocorrido ao largo da costa Sul de Portugal, foram estudadas e comparadas com recifes naturais e fundos de areia através de census visuais subaquáticos. Foram registados 11 173 espécimes pertencentes a 224 taxa faunísticos e 12 phyla. Os recifes naturais apresentaram a maior densidade de espécimes (35 122 / 1000 m²) seguidos do naufrágio (21 392 / 1000 m²) e dos fundos de areia (3771 / 1000 m²). Foram calculados três índices de biodiversidade (Margalef, Shannon-Wiener e Pielou), com os recifes naturais que apresentarem os valores mais elevados. No entanto, o naufrágio mostrou valores relativamente semelhantes aos fundos rochosos nos índices de Shannon-Wiener e Pielou. Os três habitats foram separados por estatística multivariada, com a dissimilaridade média entre areia e naufrágio, e entre areia e recifes naturais sendo muito superior à dissimilaridade entre os recifes naturais e o naufrágio. O naufrágio apresentou elevada abundância de algumas espécies comercialmente importantes, como a faneca Trisopterus luscus, o safio Conger conger e a santola Maja squinado, assim como de espécies vulneráveis e ameaçadas como a gorgonia rosa Eunicella verrucosa. Os resultados apresentados realçam a importância desse habitat para a costa Sul de Portugal

Compreender a Recorrência dos Mixomas Cardíacos. Um Caso Clínico

Recurrence of cardiac myxoma is a rare condition, observed in about 3% of patients in sporadic cases, although it is more frequent in familial ones. Several mechanisms have been proposed to explain such recurrence, and the importance of increased vascularization as a facilitating feature is the subject of debate. The authors report the case of a non-familial right atrial myxoma, unusual for both its histopathology and recurrence

Decreasing population selection rates of resistance mutation K65R over time in HIV-1 patients receiving combination therapy including tenofovir

Objectives The use of tenofovir is highly associated with the emergence of mutation K65R, which confers broad resistance to nucleoside/nucleotide analogue reverse transcriptase inhibitors (NRTIs), especially when tenofovir is combined with other NRTIs also selecting for K65R. Although recent HIV-1 treatment guidelines discouraging these combinations resulted in reduced K65R selection with tenofovir, updated information on the impact of currently recommended regimens on the population selection rate of K65R is presently lacking. Methods In this study, we evaluated changes over time in the selection rate of resistance mutation K65R in a large population of 2736 HIV-1-infected patients failing combination antiretroviral treatment between 2002 and 2010. Results The K65R resistance mutation was detected in 144 patients, a prevalence of 5.3%. A large majority of observed K65R cases were explained by the use of tenofovir, reflecting its wide use in clinical practice. However, changing patterns over time in NRTIs accompanying tenofovir resulted in a persistent decreasing probability of K65R selection by tenofovir-based therapy. The currently recommended NRTI combination tenofovir/emtricitabine was associated with a low probability of K65R emergence. For any given dual NRTI combination including tenofovir, higher selection rates of K65R were consistently observed with a non-nucleoside reverse transcriptase inhibitor than with a protease inhibitor as the third agent. Discussion Our finding of a stable time trend of K65R despite elevated use of tenofovir illustrates increased potency of current HIV-1 therapy including tenofovi

A Genomic Signature and the Identification of New Sporulation Genes

Bacterial endospores are the most resistant cell type known to humans, as they are able to withstand extremes of temperature, pressure, chemical injury, and time. They are also of interest because the endospore is the infective particle in a variety of human and livestock diseases. Endosporulation is characterized by the morphogenesis of an endospore within a mother cell. Based on the genes known to be involved in endosporulation in the model organism Bacillus subtilis, a conserved core of about 100 genes was derived, representing the minimal machinery for endosporulation. The core was used to define a genomic signature of about 50 genes that are able to distinguish endospore-forming organisms, based on complete genome sequences, and we show this 50-gene signature is robust against phylogenetic proximity and other artifacts. This signature includes previously uncharacterized genes that we can now show are important for sporulation in B. subtilis and/or are under developmental control, thus further validating this genomic signature. We also predict that a series of polyextremophylic organisms, as well as several gut bacteria, are able to form endospores, and we identified 3 new loci essential for sporulation in B. subtilis: ytaF, ylmC, and ylzA. In all, the results support the view that endosporulation likely evolved once, at the base of the Firmicutes phylum, and is unrelated to other bacterial cell differentiation programs and that this involved the evolution of new genes and functions, as well as the cooption of ancestral, housekeeping functions.FCT grant: (PEst-OE/EQB/LA0004/2011), FCT Ph.D. fellowship: (SFRH/BPD/36328/2007), FCT postdoc fellowship: (SFRH/BPD/65605/2009), Instituto Gulbenkian de Ciência research fellowship

inTB - a data integration platform for molecular and clinical epidemiological analysis of tuberculosis

This deposit is composed by the main article plus the supplementary materials of the publication.Tuberculosis is currently the second highest cause of death from infectious diseases worldwide. The emergence of multi and extensive drug resistance is threatening to make tuberculosis incurable. There is growing evidence that the genetic diversity of Mycobacterium tuberculosis may have important clinical consequences. Therefore, combining genetic, clinical and socio-demographic data is critical to understand the epidemiology of this infectious disease, and how virulence and other phenotypic traits evolve over time. This requires dedicated bioinformatics platforms, capable of integrating and enabling analyses of this heterogeneous data.Instituto Gulbenkian de Ciência, Programa Nacional de Luta contra a Tuberculose, Instituto Nacional de Saúde Ricardo Jorge, Administração Regional de Saúde de Lisboa e Vale do Tejo

Enginnering cardiac tissue using human induced pluripotent stem cell derivatives: Proteomic characterization of co-cultures of cardiomyocytes and endothelial cells

Prediction of cardiac toxicity effect is extremely relevant in the development of new drugs for different medical applications. In this way, it is important to develop more predictable human cell-based models which physiologically better mimic the human heart and allow the prediction of this toxic effect as well as establish the tools that enable the characterization of these complex cell models. To recreate engineered cardiac tissue, it is essential to reproduce the complexity of the heart by resorting to different cell types. Cardiomyocytes (CMs) are functional contractile units of the heart, and it is known that their communication with endothelial cells (ECs) is crucial for cardiac homeostasis. The aim of this study is to recreate a human pluripotent stem cells (hiPSC)-based cardiac tissue model and evaluate the impact of communication between both cell types on the phenotype of CMs. Co-cultures of hiPSC-CM and hiPSC-EC were established and maintained for 12 days as confirmed by immunofluorescence microscopy. Quantitative whole-proteome analysis was performed using SWATH Mass Spectrometry tools to compare the conditions of hiPSC-CM mono-culture and the co-culture of hiPSC-CM and hiPSC-EC. Our data showed relative increase of expression ratios of morphological maturation-related cardiac proteins in hiPSC-CM co-cultures. In particular, the expression ratios of MYH7/MYH6, MYL2/MYL7, TNNI3/TNNI1 increased 2.4-, 5.1-, and 5-fold, respectively, when compared to the mono-culture condition, indicating that in the presence of hiPSC-EC, hPSC-CM display a more adult- and ventricular- like phenotype. Changes in the extracellular matrix composition were also observed, especially related with the increased expression of ECM proteins in co-culture condition namely, collagens I and III (8.6-fold and 6-fold, respectively), fibronectin (3.5-fold) and thrombospondin-4 (2.5-fold). Other growth factors attributed to the extracellular space (e.g. CTGF, PAI1, CRTAP, IGFBP7, and NPPB) that may be responsible for the communication between both cell types have also shown to be up-regulated in the co-culture condition. The presence of a SMA+ (myofibroblast-like) population in the co-culture condition was observed by immunofluorescence microscopy images, which is in agreement with the more complex and fibrotic extracellular matrix found by whole proteome analysis. Ultrastructure characterization of CMs was carried out by transmission electron microscopy. In both conditions, hiPSC-CM displayed aligned myofibrils composed by sarcomeres with organized Z-disks, A- and I-bands, intercalated discs between adjacent cells as well as abundant mitochondria. Noteworthy, sarcomere length was higher in hiPSC-CM cultured with hiPSC-EC, suggesting structural changes associated with cardiomyocyte maturation. Calcium imaging is being performed to evaluate calcium handling of hiPSC-CMs and their response to drugs. All together our data revealed that promoting the communication of hiPSC-CM and hiPSC-EC induced structural changes in hiPSC-CM associated with maturation. This study provides important insights towards the development of more complex cardiac tissues and establishes potent analytical tools for the characterization of these models. This work was supported by Fundação para a Ciência e Tecnologia (FCT)-funded project CARDIOSTEM (MITP-TB/ECE/0013/2013); and iNOVA4Health UID/Multi/04462/2013, a program supported by FCT/Ministério da Educação e Ciência, through national funds and cofounded by FEDER under the PT2020 Partnership Agreement. BA. was supported by FCT Grant SFRH/BD/52475/201