Laminin-Inspired Cell-Instructive Microenvironments for Neural Stem Cells.

Laminin is a heterotrimeric glycoprotein with a key role in the formation and maintenance of the basement membrane architecture and properties, as well as on the modulation of several biological functions, including cell adhesion, migration, differentiation and matrix-mediated signaling. In the central nervous system (CNS), laminin is differentially expressed during development and homeostasis, with an impact on the modulation of cell function and fate. Within neurogenic niches, laminin is one of the most important and well described extracellular matrix (ECM) proteins. Specifically, efforts have been made to understand laminin assembly, domain architecture, and interaction of its different bioactive domains with cell surface receptors, soluble signaling molecules, and ECM proteins, to gain insight into the role of this ECM protein and its receptors on the modulation of neurogenesis, both in homeostasis and during repair. This is also expected to provide a rational basis for the design of biomaterial-based matrices mirroring the biological properties of the basement membrane of neural stem cell niches, for application in neural tissue repair and cell transplantation. This review provides a general overview of laminin structure and domain architecture, as well as the main biological functions mediated by this heterotrimeric glycoprotein. The expression and distribution of laminin in the CNS and, more specifically, its role within adult neural stem cell niches is summarized. Additionally, a detailed overview on the use of full-length laminin and laminin derived peptide/recombinant laminin fragments for the development of hydrogels for mimicking the neurogenic niche microenvironment is given. Finally, the main challenges associated with the development of laminin-inspired hydrogels and the hurdles to overcome for these to progress from bench to bedside are discussed.

Engineering hydrogels with affinity-bound laminin as 3D neural stem cell culture systems.

Laminin incorporation into biological or synthetic hydrogels has been explored to recapitulate the dynamic nature and biological complexity of neural stem cell (NSC) niches. However, the strategies currently explored for laminin immobilization within three-dimensional (3D) matrices do not address a critical aspect influencing cell-matrix interactions, which is the control over laminin conformation and orientation upon immobilization. This is a key feature for the preservation of the protein bioactivity. In this work, we explored an affinity-based approach to mediate the site-selective immobilization of laminin into a degradable synthetic hydrogel. Specifically, a four-arm maleimide terminated poly(ethylene glycol) (PEG-4MAL) macromer was functionalized with a mono-PEGylated recombinant human N-terminal agrin (NtA) domain, to promote high affinity binding of laminin. Different NtA concentrations (10, 50 and 100 µM) were used to investigate the impact of NtA density on laminin incorporation, hydrogel biophysical properties, and biological outcome. Laminin was efficiently incorporated for all the conditions tested (laminin incorporation >95%), and the developed hydrogels revealed mechanical properties (average storage modulus (G') ranging from 187 to 256 Pa) within the values preferred for NSC proliferation and neurite branching and extension. Affinity-bound laminin PEG-4MAL hydrogels better preserve laminin bioactivity, compared to unmodified hydrogels and hydrogels containing physically entrapped laminin, this effect being dependent on NtA concentration. This was evidenced by the 10 µM NtA-functionalized PEG-4MAL gels incorporating laminin that support enhanced human NSC proliferation and neurite extension, compared to the latter. Overall, this work highlights the potential of the proposed engineered matrices to be used as defined 3D platforms for the establishment of artificial NSC niches and as extracellular matrix-mimetic microenvironments to support human NSC transplantation.

Conjugation of the T1 sequence from CCN1 to fibrin hydrogels for therapeutic vascularization.

Hydrogel matrices with angiogenic properties are much desirable for therapeutic vascularization strategies, namely to provide vascular supply to ischemic areas, transplanted cells, or bioengineered tissues. Here we report the pro-angiogenic effect of fibrin (Fb) functionalization with the T1 sequence from the angiogenic inducer CCN1, forseeing its use in the injured brain and spinal cord. Fibrin functionalization with 40 µM of T1 peptide effectively improved cellular sprouting of human brain microvascular endothelial cells (hCMEC/D3) in the absence of vascular endothelial growth factor (VEGF), without impacting the viscoelastic properties of Fb, cell viability, or proliferation. The pro-angiogenic effect of immobilized T1 was potentiated in the presence of VEGF and partially mediated through α6ß1 integrin. The tethering of T1 also enhanced sprouting of human cord blood-derived outgrowth endothelial cells (OEC). Still, to elicit such effect, a higher input T1 concentration was required (60 µM), in line with the lower protein levels of α6 and ß1 integrin subunits found in OEC comparing to hCMEC/D3, prior to embedment in Fb gel. Finally, the ability of T1-functionalized Fb in inducing cappilary invasion in vivo was assessed using the CAM assay, which evidenced a significant increase in the number of newly formed vessels at sites of implantation of T1-functionalized Fb, in the absence of soluble angiogenic factors. Overall these results demonstrate the potential of T1 peptide-presenting gels for use in therapeutic vascularization approaches. Considering T1 neurite-extension promoting capability and pro-angiogenic properties, T1-functionalized Fb hydrogels are particularly promising for application in the injured central nervous system.

Delivery of Antisense Oligonucleotides Mediated by a Hydrogel System: In Vitro and In Vivo Application in the Context of Spinal Cord Injury.

Biomaterials-based hydrogels are attractive drug-eluting vehicles in the context of RNA therapeutics, such as those utilizing antisense oligonucleotide or RNA interference based drugs, as they can potentially reduce systemic toxicity and enhance in vivo efficacy by increasing in situ concentrations. Here we describe the preparation of antisense oligonucleotide-loaded fibrin hydrogels exploring their applications in the context of the nervous system utilizing an organotypic dorsal root ganglion explant in vitro system and an in vivo model of spinal cord injury.

Opinions and main sources of information about menopause among middle-aged Brazilian women.

OBJECTIVES: The aim of this study was to evaluate women's opinions about menopause and the sources of information they use to deepen their understanding of the topic. METHODS: Population-based study with 749 Brazilian women aged 45 to 60 years. The answers to the question "What is menopause?" were typed and coded, and categories that emerged from the interviewees' own speech were created. The answers to the question: "Where or from whom did you get information about menopause?" were also analyzed. RESULTS: The mean age of women was 52.5 (± 4.4) years. Of them, 68% were postmenopausal. According to 67.5% of the interviewees, the concept of menopause encompassed changes in the menstrual cycle and hormones. For 48%, menopause meant physical changes such as "hot flushes and vaginal dryness." For 22.7%, menopause represented psychological changes. The concept of menopause was associated with some change in sexuality for 7.6% of the interviewees. Approximately 18% could not explain what menopause meant. Regarding the sources of information, 44.5% of the women attributed this knowledge to friends and relatives. Doctors or health services were mentioned by 44.3% of women. Television or radio was cited by 22.0%; magazines, newspapers, or books were cited by 14.0%; and the Internet was cited by 6.8% of women. Fifty-two women (6.9%) reported having no source of information about menopause. CONCLUSIONS: Most of the interviewees relate the term "menopause" to physiological events. There seems to be a suppressed demand for information on the various aspects of the menopausal transition among middle-aged Brazilian women.

An affinity-based approach to engineer laminin-presenting cell instructive microenvironments.

Laminin immobilization into diverse biological and synthetic matrices has been explored to replicate the microenvironment of stem cell niches and gain insight into the role of extracellular matrix (ECM) on stem cell behavior. However, the site-specific immobilization of this heterotrimeric glycoprotein and, consequently, control over its orientation and bioactivity has been a challenge that has limited many of the explored strategies to date. In this work, we established an affinity-based approach that takes advantage of the native high affinity interaction between laminin and the human N-terminal agrin (hNtA) domain. This interaction is expected to promote the site-selective immobilization of laminin to a specific substrate, while preserving the exposure of its key bioactive epitopes. Recombinant hNtA (rhNtA) domain was produced with high purity (>90%) and successfully conjugated at its N-terminal with a thiol-terminated poly(ethylene glycol) (PEG) without affecting its affinity to laminin. Self-assembled monolayers (SAMs) of mono-PEGylated rhNtA on gold (mPEG rhNtA-SAMs) were then prepared to evaluate the effectiveness of this strategy. The site-specific immobilization of laminin onto mPEG rhNtA-SAMs was shown to better preserve protein bioactivity in comparison to laminin immobilized on SAMs of thiol-PEG-succinimidyl glutaramide (HS-PEG-SGA), used for the non-selective covalent immobilization of laminin, as evidenced by its enhanced ability to efficiently self-polymerize and mediate cell adhesion and spreading of human neural stem cells. These results highlight the potential of this novel strategy to be used as an alternative to the conventional immobilization approaches in a wide range of applications, including engineered coatings for neuroelectrodes and cell culture, as well as biofunctionalization of 3D matrices.

Synthetic matrix enhances transplanted satellite cell engraftment in dystrophic and aged skeletal muscle with comorbid trauma.

Muscle satellite cells (MuSCs) play a central role in muscle regeneration, but their quantity and function decline with comorbidity of trauma, aging, and muscle diseases. Although transplantation of MuSCs in traumatically injured muscle in the comorbid context of aging or pathology is a strategy to boost muscle regeneration, an effective cell delivery strategy in these contexts has not been developed. We engineered a synthetic hydrogel-based matrix with optimal mechanical, cell-adhesive, and protease-degradable properties that promotes MuSC survival, proliferation, and differentiation. Furthermore, we establish a biomaterial-mediated cell delivery strategy for treating muscle trauma, where intramuscular injections may not be applicable. Delivery of MuSCs in the engineered matrix significantly improved in vivo cell survival, proliferation, and engraftment in nonirradiated and immunocompetent muscles of aged and dystrophic mice compared to collagen gels and cell-only controls. This platform may be suitable for treating craniofacial and limb muscle trauma, as well as postoperative wounds of elderly and dystrophic patients.

Factors associated with knowledge about menopause and hormone therapy in middle-aged Brazilian women: a population-based household survey.

OBJECTIVES: The aim of the study was to evaluate knowledge about menopause and hormone therapy (HT) among middle-aged Brazilian women and identify the factors associated with the level of knowledge. METHODS: A cross-sectional study was conducted with 749 women aged 45 to 60 years, residing in the Metropolitan Region of Campinas, Brazil. The dependent variable was knowledge about menopause and HT evaluated using a numerical score based on the answers given to three questions that had several response options. The independent variables were sociodemographic characteristics, personal habits, and health conditions. Statistical analysis was carried out by Mann-Whitney, Kruskal-Wallis, and a generalized linear model. RESULTS: The mean age was 52.5 (±4.4) years. Most of the women were postmenopausal (68%) and 19.5% had a history of current or previous use of HT. The final score for knowledge about menopause ranged from +1 to +11, with a mean of 4.59 (±1.86) and a median of 4. Women with higher levels of schooling (coefficient = 0.64, P < 0.001); with higher socioeconomic status (coefficient = 0.47, P < 0.002); with vaginal dryness (coefficient = 0.47, P = 0.003); who consumed alcohol (coefficient = 0.61, P = 0.006); who use or had ever used any type of treatment to counteract menopausal symptoms (coefficient = 0.41, P = 0.008); with anxiety (coefficient = 0.35, P = 0.037); and with depression (coefficient = 0.31, P = 0.043) possessed greater knowledge about menopause and HT. Women with a higher number (≥3) of vaginal deliveries (coefficient = -0.61, P < 0.001) and a higher body mass index (coefficient = -0.02, P = 0.046) possessed less knowledge about menopause and HT. CONCLUSIONS: The knowledge about menopause and HT demonstrated by the interviewees was low and possibly related to the time of exposure to qualified information.

Hydrogel-Assisted Antisense LNA Gapmer Delivery for In Situ Gene Silencing in Spinal Cord Injury.

After spinal cord injury (SCI), nerve regeneration is severely hampered due to the establishment of a highly inhibitory microenvironment at the injury site, through the contribution of multiple factors. The potential of antisense oligonucleotides (AONs) to modify gene expression at different levels, allowing the regulation of cell survival and cell function, together with the availability of chemically modified nucleic acids with favorable biopharmaceutical properties, make AONs an attractive tool for novel SCI therapy developments. In this work, we explored the potential of locked nucleic acid (LNA)-modified AON gapmers in combination with a fibrin hydrogel bridging material to induce gene silencing in situ at a SCI lesion site. LNA gapmers were effectively developed against two promising gene targets aiming at enhancing axonal regeneration-RhoA and GSK3ß. The fibrin-matrix-assisted AON delivery system mediated potent RNA knockdown in vitro in a dorsal root ganglion explant culture system and in vivo at a SCI lesion site, achieving around 75% downregulation 5 days after hydrogel injection. Our results show that local implantation of a AON-gapmer-loaded hydrogel matrix mediated efficient gene silencing in the lesioned spinal cord and is an innovative platform that can potentially combine gene regulation with regenerative permissive substrates aiming at SCI therapeutics and nerve regeneration.

Conhecimento sobre a menopausa de acordo com mulheres brasileiras de meia-idade: um estudo de base populacional / Menopausal knowledge among Brazilian middle-aged women: a population-based household survey

Introdução: a síndrome do climatério é definida pelo conjunto de sintomas decorrentes da interação entre fatores endócrinos, socioculturais e psicológicos que ocorrem na mulher que envelhece. A procura por ajuda especializada e a utilização de algum método terapêutico neste período podem estar diretamente relacionadas ao nível de conhecimento apresentado pela mulher. Objetivos: avaliar a opinião sobre o significado da palavra menopausa, o conhecimento sobre o tema, os fatores associados ao nível de conhecimento e saber quais são as fontes de informação para mulheres de meia-idade residentes na Região Metropolitana de Campinas (RMC). Métodos: estudo transversal de base populacional com 749 mulheres brasileiras com idade entre 45 e 60 anos, residentes nos municípios que compõem a RMC. As respostas fornecidas à questão aberta "o que é a menopausa?" foram digitadas e codificadas, criando-se categorias que emergiram da fala das entrevistadas. Também foram analisadas as respostas à pergunta: "Onde ou de quem a senhora obteve informações sobre a menopausa?". Os conhecimentos sobre menopausa e Terapia Hormonal (TH) foram avaliados através de um escore calculado com base nas respostas fornecidas para três perguntas sobre o tema. As variáveis independentes foram as características sociodemográficas, hábitos de saúde e antecedentes ginecológicos. A análise estatística foi realizada através dos testes de Mann-Whitney, Kruskal-Wallis e Modelo Linear Generalizado. Resultados: a média etária das entrevistadas foi de 52,5 (+4,4) anos. Dessas, 16% estavam na pré-menopausa, 16% na perimenopausa e 68% estavam na pós-menopausa. Cento e quarenta e seis mulheres (19,5%) tinham antecedente de uso atual ou prévio de TH. De acordo com 67,5% das entrevistadas, o conceito de menopausa englobava alterações no ciclo menstrual e nos hormônios. Para 48% das entrevistadas, menopausa significava a ocorrência de alterações físicas, como "calores e ressecamento vaginal", e "envelhecimento". Para 22,7%, menopausa representava alterações psicológicas. O conceito de menopausa esteve associado a alguma alteração da sexualidade para 7,6% das entrevistadas. Aproximadamente 18% não tinham ideia do que significava menopausa. Quanto às fontes de informação, amigos, parentes e conhecidos foram os mais citados (44,5%). Médicos ou serviços de saúde vieram em seguida, mencionados por 44,3% das mulheres. Televisão ou rádio foram citados por 22,0%, revistas, jornais ou livros foram citados por 14,0% e a internet foi citada como fonte de informação sobre a menopausa por 6,8% das mulheres. O escore final de conhecimento variou entre +1 e +11, com uma média de 4,59 (+1,86) e uma mediana de 4. Mulheres com maior escolaridade (ß = 0,64; p 3 partos normais (ß= -0,61; p < 0,001) e com maior Índice de Massa Corporal (IMC) (ß = -0,02; p= 0,046) apresentaram menor conhecimento. Conclusões: a maioria das mulheres de meia-idade residentes na RMC relacionava o termo "menopausa" a eventos fisiológicos, porém uma parcela significativa delas os vivenciava de maneira negativa. O conhecimento sobre menopausa e TH demonstrado pelas entrevistadas foi baixo. Fatores pessoais relacionados a uma maior exposição à informação qualificada parecem se associar a um maior conhecimento sobre menopausa e TH. Aparentemente, há uma demanda reprimida por informação sobre os diversos aspectos da transição menopausal entre mulheres brasileiras de meia-idade.(AU)

Abstract: Introduction: The climacteric syndrome is defined as a set of symptoms resulting from the interaction between endocrine, sociocultural, and psychological factors that occur in the aging woman. The search for specialized help and the use of some therapeutic method may be directly related to the level of knowledge that an individual acquires about a certain topic. Objectives: to evaluate the opinion about the meaning of the word menopause, the knowledge about the theme, the factors associated with the level of knowledge and to know the sources of information for middle-aged women living in the Metropolitan Region of Campinas (MRC). Methods: a population-based cross-sectional study was conducted with 749 women aged 45-60 years, residing in the Metropolitan Region of Campinas ¿ Brazil. The answers to the question "What is menopause?" were typed and coded, and categories that emerged from the interviewees¿ own speech were created. The answers to the question: "Where or from whom did you get information about menopause?" were also analyzed. Knowledge about menopause and hormone therapy (HT) was evaluated using a numerical score based on the answers given to 3 questions that had several response options. The independent variables were sociodemographic characteristics, personal habits, and gynecological background. Statistical analysis was carried out by Mann-Whitney, Kruskal-Wallis, and a generalized linear model. Results: the mean age was 52.5 (±4.4) years. Of them, 16% were premenopausal, 16% were perimenopausal and 68% were postmenopausal. One hundred and forty-six women (19.5%) had a history of current or previous use of HT. According to 67.5% of the interviewees, the concept of menopause encompassed changes in the menstrual cycle and hormones. For 48% of the interviewees, menopause meant physical changes such as "hot flushes and vaginal dryness" (42.5%), and "aging and getting old" (5.5%). For 22.7%, menopause represented psychological changes. The concept of menopause was associated with some change in sexuality for 7.6% of the interviewees. Approximately 18% could not explain and had no idea about what menopause meant. Regarding the sources of information, 44.5% of the women attributed this knowledge to friends and relatives. Doctors or health services were mentioned by 44.3% of women. Television or radio was cited by 22.0%; magazines, newspapers, or books was cited by 14.0%; and the internet was cited by 6.8% of women. The final score for knowledge about menopause ranged from +1 to +11, with a mean of 4.59 (±1.86) and a median of 4. Women with higher levels of schooling (?=0.64, p<0.001); with higher socioeconomic status (?=0.47, p<0.002); with vaginal dryness (?=0.47, p=0.003); who consumed alcohol (?=0.61, p=0.006); who use or had ever used any type of treatment to counteract menopausal symptoms (?=0.41, p=0.008); with anxiety (?=0.35, p=0.037); and with depression (?=0.31, p=0.043) possessed greater knowledge about menopause and HT. Women with a higher number (?3) of vaginal deliveries (?=-0.61, p<0.001) and a higher BMI (?=-0.02, p=0.046) possessed less knowledge about menopause and HT. Conclusions: most middle-aged women living in the Metropolitan Region of Campinas relate the term "menopause" to physiological events, but a significant portion of them experience them in a negative way. The knowledge about menopause and HT demonstrated by the interviewees was low. Personal factors related to a greater exposure to qualified information seem to be associated with greater knowledge about menopause and HT. There appears to be a suppressed demand for information on the various aspects of the menopausal transition among middle-aged Brazilian women(AU)

Fibrin functionalization with synthetic adhesive ligands interacting with α6ß1 integrin receptor enhance neurite outgrowth of embryonic stem cell-derived neural stem/progenitors.

To enhance fibrin hydrogel affinity towards pluripotent stem cell-derived neural stem/progenitor cells (NSPCs) and its capacity to support NSPC migration and neurite extension, we explored the tethering of synthetic peptides engaging integrin α6ß1, a cell receptor enriched in NSPCs. Six α6ß1 integrin ligands were tested for their ability to support integrin α6ß1-mediated adhesion of embryonic stem cell-derived NSPCs (ES-NSPs) and sustain ES-NSPC viability, migration, and neuronal differentiation. Due to their better performance, peptides T1, HYD1, and A5G81 were immobilized into fibrin and functionalized gels characterized in terms of peptide binding efficiency, structure and viscoelastic properties. Tethering of T1 or HYD1 successfully enhanced cell outgrowth from ES-NSPC neurospheres (up to 2.4-fold increase), which exhibited a biphasic response to peptide concentration. Inhibition assays evidenced the involvement of α6ß1 and α3ß1 integrins in mediating radial outgrowth on T1-/HYD1-functionalized gels. Fibrin functionalization also promoted neurite extension of single ES-NSPCs in fibrin, without affecting cell proliferation and neuronal differentiation. Finally, HYD1-functionalized gels were found to provide a permissive environment for axonal regeneration, leading up to a 2.0-fold increase in neurite extension from rat dorsal root ganglia explants as compared to unmodified fibrin, and to significant improved locomotor function after spinal cord injury (complete transection), along with a trend toward a higher area positive for growth associated protein 43 (marker for axonal growth cone formation). Our results suggest that conjugation of α6ß1 integrin-binding motifs is of interest to increase the biofunctionality of hydrogels used in 3D platforms for ES-NSPC culture and potentially, in matrix-assisted ES-NSPC transplantation. STATEMENT OF SIGNIFICANCE: Impact statement: The transplantation of NSPCs derived from pluripotent stem cells holds much promise for the treatment of central nervous system disorders. Moreover, the combinatorial use of biodegradable hydrogels with NSPCs was shown to contribute to the establishment of a more permissive environment for survival and integration of transplanted cells. In this study, fibrin hydrogels functionalized with a synthetic peptide engaging integrin α6ß1 (HYD1) were shown to promote neurite extension of ES-NSPCs, which is fundamental for the formation of functional neuronal relay circuits after NSPC transplantation. Notably, HYD1-functionalized fibrin per se led to enhanced axonal growth ex vivo and to an improvement in locomotor function after implantation in a rat model of spinal cord injury. Conjugation of α6ß1 integrin-binding motifs may therefore be of interest to confer bioactivity to NSPC hydrogel vehicles.

Entre neurônios e sinapses: as contribuições de Cajal e Athias para a medicina ibérica entre os séculos XIX e XX / Between neurons and synapses: the contributions of Cajal and Athias to Iberian medicine between the nineteenth and twentieth centuries

Resumo O presente estudo reflete sobre o trajeto da histologia na transição do século XIX para o XX, em Portugal e em Espanha, para estabelecer um paralelo entre as contribuições de Santiago Ramón y Cajal e Marck Athias, ambos determinantes para o desenvolvimento da medicina experimental na Península Ibérica, reconhecidos como pilares de uma nova mentalidade científica europeia, na alvorada do século XX. Neste estudo de caso, refletimos sobre as vicissitudes da construção da ciência na “periferia” europeia, no contexto da categoria historiográfica centro-periferia desenvolvida pelo grupo Science and Technology in the European Periphery (Step), confrontando a realidade ibérica com o modelo de ciência alemã, no período em estudo.

Abstract The trajectory of histology at the cusp of the twentieth century in Portugal and Spain is investigated to draw a parallel between the contributions of Santiago Ramón y Cajal and Marck Athias, both of whom were instrumental in the development of experimental medicine in the Iberian Peninsula and recognized as pillars of a new European scientific mindset at the dawn of the twentieth century. In this case study we reflect on the vicissitudes of the construction of science in the “periphery” of Europe, in the context of the historiographical category of center-periphery developed by STEP (Science and Technology in the European Periphery), contrasting the reality in Iberia with the model of German science in the period under study.

Rotary orbital suspension culture of embryonic stem cell-derived neural stem/progenitor cells: impact of hydrodynamic culture on aggregate yield, morphology and cell phenotype.

Embryonic stem (ES)-derived neural stem/progenitor cells (ES-NSPCs) constitute a promising cell source for application in cell therapies for the treatment of central nervous system disorders. In this study, a rotary orbital hydrodynamic culture system was applied to single-cell suspensions of ES-NSPCs, to obtain homogeneously-sized ES-NSPC cellular aggregates (neurospheres). Hydrodynamic culture allowed the formation of ES-NSPC neurospheres with a narrower size distribution than statically cultured neurospheres, increasing orbital speeds leading to smaller-sized neurospheres and higher neurosphere yield. Neurospheres formed under hydrodynamic conditions (72 h at 55 rpm) showed higher cell compaction and comparable percentages of viable, dead, apoptotic and proliferative cells. Further characterization of cellular aggregates provided new insights into the effect of hydrodynamic shear on ES-NSPC behaviour. Rotary neurospheres exhibited reduced protein levels of N-cadherin and ß-catenin, and higher deposition of laminin (without impacting fibronectin deposition), matrix metalloproteinase-2 (MMP-2) activity and percentage of neuronal cells. In line with the increased MMP-2 activity levels found, hydrodynamically-cultured neurospheres showed higher outward migration on laminin. Moreover, when cultured in a 3D fibrin hydrogel, rotary neurospheres generated an increased percentage of neuronal cells. In conclusion, the application of a constant orbital speed to single-cell suspensions of ES-NSPCs, besides allowing the formation of homogeneously-sized neurospheres, promoted ES-NSPC differentiation and outward migration, possibly by influencing the expression of cell-cell adhesion molecules and the secretion of proteases/extracellular matrix proteins. These findings are important when establishing the culture conditions needed to obtain uniformly-sized ES-NSPC aggregates, either for use in regenerative therapies or in in vitro platforms for biomaterial development or pharmacological screening. Copyright © 2016 John Wiley & Sons, Ltd.

[Between neurons and synapses: the contributions of Cajal and Athias to Iberian medicine between the nineteenth and twentieth centuries]. / Entre neurônios e sinapses: as contribuições de Cajal e Athias para a medicina ibérica entre os séculos XIX e XX.

The trajectory of histology at the cusp of the twentieth century in Portugal and Spain is investigated to draw a parallel between the contributions of Santiago Ramón y Cajal and Marck Athias, both of whom were instrumental in the development of experimental medicine in the Iberian Peninsula and recognized as pillars of a new European scientific mindset at the dawn of the twentieth century. In this case study we reflect on the vicissitudes of the construction of science in the "periphery" of Europe, in the context of the historiographical category of center-periphery developed by STEP (Science and Technology in the European Periphery), contrasting the reality in Iberia with the model of German science in the period under study.

Three-dimensional culture of single embryonic stem-derived neural/stem progenitor cells in fibrin hydrogels: neuronal network formation and matrix remodelling.

In an attempt to improve the efficacy of neural stem/progenitor cell (NSPC) based therapies, fibrin hydrogels are being explored to provide a favourable microenvironment for cell survival and differentiation following transplantation. In the present work, the ability of fibrin to support the survival, proliferation, and neuronal differentiation of NSPCs derived from embryonic stem (ES) cells under monolayer culture was explored. Single mouse ES-NSPCs were cultured within fibrin (fibrinogen concentration: 6 mg/ml) under neuronal differentiation conditions up to 14 days. The ES-NSPCs retained high cell viability and proliferated within small-sized spheroids. Neuronal differentiation was confirmed by an increase in the levels of ßIII-tubulin and NF200 over time. At day 14, cell-matrix constructs mainly comprised NSPCs and neurons (46.5% ßIII-tubulin+ cells). Gamma-aminobutyric acid (GABA)ergic and dopaminergic/noradrenergic neurons were also observed, along with a network of synaptic proteins. The ES-NSPCs expressed matriptase and secreted MMP-2/9, with MMP-2 activity increasing along time. Fibronectin, laminin and collagen type IV deposition was also detected. Fibrin gels prepared with higher fibrinogen concentrations (8/10 mg/ml) were less permissive to neurite extension and neuronal differentiation, possibly owing to their smaller pore area and higher rigidity. Overall, it is shown that ES-NSPCs within fibrin are able to establish neuronal networks and to remodel fibrin through MMP secretion and extracellular matrix (ECM) deposition. This three-dimensional (3D) culture system was also shown to support cell viability, neuronal differentiation and ECM deposition of human ES-NSPCs. The settled 3D platform is expected to constitute a valuable tool to develop fibrin-based hydrogels for ES-NSPC delivery into the injured central nervous system. Copyright © 2016 John Wiley & Sons, Ltd.

Análise dos processos de atenção e interação em criança com deficiência múltipla sensorial / Analysis of interaction and attention processes in a child with multiple disabilities

RESUMO Crianças com deficiência múltipla sensorial podem não desenvolver a linguagem de forma efetiva, pois a combinação de deficiências tende a reduzir a participação em seu meio e prejudicar o desenvolvimento da linguagem. O objetivo desta pesquisa foi analisar os processos interacionais, ou seja, os comportamentos de atenção (atenção à pessoa, atenção ao objeto e atenção conjunta) e comunicativos, entre uma aluna com deficiência múltipla sensorial (4 anos e 6 meses de idade) e sua professora, especializada na área da surdocegueira e deficiência múltipla sensorial. Os dados foram coletados durante atividades em sala de aula, por meio de observação participante e de gravações audiovisuais da interação/díade, e analisados com auxilio do programa ATLAS.TI. Os resultados apontaram que a aluna apresentou atenção ao objeto, em atividades que envolveram música e ritmo. Como potencial forma de comunicação não verbal, observou-se olhar, movimentos corporais e vocalização. As formas de comunicação da professora foram verbal, toque, visual, auditiva (ritmo) e sinais de Língua Brasileira de Sinais. A aluna apresentou potenciais trocas de turnos apenas quando a ação foi iniciada pela professora. A qualidade das atividades, os materiais utilizados e a participação mostraram ter impacto sobre os níveis de atenção e comunicação. Mais pesquisas devem considerar esses resultados, como forma de definir quais as atividades que podem contribuir para apoiar o desenvolvimento e a qualidade de vida das crianças com deficiência múltipla sensorial. Assim, professor e fonoaudiólogo devem conhecer a forma como cada criança se comunica e manterem-se alertas para os comportamentos não verbais, a fim de estabelecerem uma comunicação efetiva.

ABSTRACT Children with multiple sensory disabilities may not develop the language effectively, given that the combination of disabilities tends to reduce their participation in the environment and harm language development. The objective of this research was to analyze the interaction processes, i.e., the attention (attention to the person, attention to the object and joint attention) and communicative behaviors between a student with multiple disabilities (age: 4 years and 6 months) and her teacher specialized in the field of sensory multiple disabilities. The data were collected during activities in the classroom, through participant observations and audiovisual recordings of teacher/child interactions and analyzed with the ATLAS.TI program. The results indicated that the student showed attention to the object in activities involving music and rhythm. As potential forms of nonverbal communication it was observed eye gaze body movement and vocalizations. The teacher forms of communication were verbal, touch, visual, auditory (rhythm) and Brazilian Sign Language. The student presented potential turn-taking only when the action was initiated by the teacher. The quality of the activities, the materials used and the participation showed to have impact on the levels of attention and communication. More researches should consider these results as a way to define which activities that can contribute to support the development and quality of life of children with sensory multiple disabilities. Thus, teacher and speech/language therapist should know the way each child communicates and to be alert to nonverbal behaviors as a way to establish effective communication.

Analysis of Interaction and Attention Processes in a Child With Congenital Deafblindness.

Children with deafblindness need support to be able to understand the world and to have access to information. The authors analyzed a dyad consisting of a child with congenital deafblindness and a specialized teacher. The study included participant observations and audiovisual recordings. It was found that the child showed attention to the teacher in activities involving music and rhythm. As potential forms of nonverbal communication, the child presented vocalization, touch, body contact, body movements, facial expressions, and tears. The teacher's forms of communication were verbal, touch, visual, rhythm, and sign language. It was concluded that a significant communication partner is essential to identify, interpret, and respond to attention and communicative behaviors. Use of other forms of communication must comply with individual characteristics so that the child with deafblindness can receive information from the environment through these senses and thus be guaranteed access to the world.

Propaganda and Philanthropy: The Institute Bento da Rocha Cabral, the Lisbon Site of Biochemistry (1925-1953).

This paper focuses on the internal organization and dynamics of the Institute Bento da Rocha Cabral (IRC) in Lisbon, a privately-funded institution devoted to biomedical research, from the particular vantage point of its laboratory of biochemistry; in particular, the process through which the institution turned from medically-related to chemically-related research in the period spanning from 1925 to 1953. The history of the IRC raises interesting questions regarding the social politics of science as it materialized the desire of leading physicians of the Faculty of Medicine of Lisbon to create proper physical facilities for medically-related scientific research. We argue that the process which led to the creation of the IRC coincided with the gradual professional and political ascendance of physicians in Portuguese society initiated in the late nineteenth century, and is closely associated with Portuguese republicanism and the process of Lisbon becoming the scientific capital.

Biomimetic synthetic self-assembled hydrogels for cell transplantation.

The development of three-dimensional matrices capable of recapitulating the main features of native extracellular matrix and contribute for the establishment of a favorable microenvironment for cell behavior and fate is expected to circumvent some of the main limitations of cell-based therapies. In this context, self-assembly has emerged as a promising strategy to engineer cell-compatible hydrogels. A wide number of synthetically-derived biopolymers, such as proteins, peptides and DNA/RNA, with intrinsic ability to self-assemble into well-defined nanofibrous structures, are being explored. The resulting hydrogels, in addition to closely resembling the architecture of native cellular microenvironments, present a versatile and dynamic behavior that allows them to be designed to undergo sol-to-gel transition in response to exogenous stimulus. This review presents an overview on the state-of-the-art of the different strategies being explored for the development of injectable synthetic self-assembled hydrogels for cell transplantation and/or recruitment of endogenous cells, with an emphasis on their biological performance, both in vitro and in vivo. Systems based on peptides are the most widely explored and have already generated promising results in pre-clinical in vivo studies involving different repair/regenerative scenarios, including cartilage, bone, nerve and heart. On the other hand, systems based on DNA and hybrid hydrogels are now emerging for application in the biomedical field with high potential. Finally, the main challenges hampering the translation of these systems to the clinic and the issues that need to be addressed for these to progress from bench-to-bedside are discussed.