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One of the main challenges of tissue engineering in dentistry is to replace bone and dental tissues with strategies or techniques that simulate physiological tissue repair conditions. This systematic review of in vitro studies aimed to evaluate the influence of the addition of nanohydroxyapatite (NHap) to scaffolds on cell proliferation and osteogenic and odontogenic differentiation of human mesenchymal stem cells. In vitro studies on human stem cells that proliferated and differentiated into odontogenic and osteogenic cells in scaffolds containing NHap were included in this study. Searches in PubMed/MEDLINE, Scopus, Web of Science, OpenGrey, ProQuest, and Cochrane Library electronic databases were performed. The total of 333 articles was found across all databases. After reading and analyzing titles and abstracts, 8 articles were selected for full reading and extraction of qualitative data. Results showed that despite the large variability in scaffold composition, NHap-containing scaffolds promoted high rates of cell proliferation, increased alkaline phosphatase (ALP) activity during short culture periods, and induced differentiation, as evidenced by the high expression of genes involved in osteogenesis and odontogenesis. However, further studies with greater standardization regarding NHap concentration, type of scaffolds, and evaluation period are needed to observe possible interference of these criteria in the action of NHap on the proliferation and differentiation of human stem cells.

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ABSTRACT Objective: To assess the effects of cobalt chloride (CoCl2) as a hypoxia mimicking agent on human umbilical cord mesenchymal stem cells (hUCMSCs) expression of HIF-1α and mTOR for use in regenerative dentistry. Material and Methods: Human umbilical cord mesenchymal stem cells were isolated and then cultured. The characteristics of stemness were screened and confirmed by flow cytometry. The experiment was conducted on hypoxia (H) and normoxia (N) groups. Each group was divided and incubated into 24-, 48-, and 72-hours observations. Hypoxic treatment was performed using 100 µM CoCl2 on 5th passage cells in a conventional incubator (37°C; 5CO2). Then, immunofluorescence of HIF-1α and mTOR was done. Data was analyzed statistically using One-way ANOVA and Tukey's HSD. Results: Significant differences were found between normoxic and hypoxic groups on HIF-1α (p=0.015) and mTOR (p=0.000) expressions. The highest HIF-1α expression was found at 48 hours in the hypoxia group, while for mTOR at 24 hours in the hypoxia group. Conclusion: Hypoxia using cobalt chloride was able to increase human umbilical cord mesenchymal stem cells expression of HIF-1α and mTOR.

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Abstract Objective: To examine the cytotoxicity of calcium hydroxide on human umbilical cord mesenchymal stem cells (HUCMSC) to understand the characteristics for use in regenerative dentistry procedures especially regenerative endodontics. Material and Methods: HUCMSC was isolated, cultured, and confirmed by flow cytometry. The biological characteristics, such as cell morphology, proliferation, and protein expression, were screened. To check the cytotoxicity, HUCMSC was cultured and divided into two groups, the control group (cultured in minimum essential medium (MEM) alpha) and calcium hydroxide group (cultured in MEM alpha and calcium hydroxide). Methyl-thiazole-tetrazolium (MTT) assay was done on different concentrations of calcium hydroxide (0.39 to 25 µg/mL) and the cells were observed and counted. One-way ANOVA test was used with a significance level set at 5%. Results: Flow cytometric analysis confirmed positive of CD73, CD90, CD105, negative of CD45 and CD34. A significant difference was found between the concentration of 6.25 and 3.125 µg/mL (p=0.004). There was no significant difference among 6.25, 12.5 and 25 µg/mL concentrations. There was also no significant difference among 0.39, 0.78, 1.56, and 3.125 µg/mL concentrations. Conclusion: Even though calcium hydroxide is a medicament of choice in clinical endodontics, it decreases the viability of HUCMSC. The lower the concentration of calcium hydroxide, the higher the viability of HUCMSC.

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Abstract The aim of this study was to investigate the viability of human dental pulp cells from extracted teeth kept at standard room temperature and atmospheric pressure for different periods of time. Twenty-one healthy permanent teeth were used. They were divided into five groups according to the expected time from extraction to processing. One group was tested immediately after extraction; the other groups were each tested at one of the following time points: 30 minutes, 1 hour, 2 hours, and 5 hours post-extraction. Cell morphology was analysed by light microscopy; cell proliferation was analysed using MTT assay and by counting the viable cells in a haemocytometer. Similar results were observed in all groups (p < 0.05). A delay of up to five hours for tooth processing and tissue collection does not preclude the establishment of dental pulp cell cultures, affect the morphology of these cells, or reduce their proliferative potential.

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MatrigelBD is a hydrogel scaffold with three-dimensional intercrossed networks of hydrophilic polymers with high water content. Human gingival tissue might represent a better source of MSCs, allowing these cells to be easily obtained in a relatively non-invasive way. The objective of this study was to evaluate the biocompatibility of MatrigelBD with GMSCs in vitro. Gingival connective tissue samples were obtained from healthy donors. Fresh tissue was minced and cultured during two weeks, after which cells at passage fourth were analyzed for their immune phenotype by flow cytometry. Differentiation into osteogenic, chondrogenic, and adipogenic lineages was induced and evaluated by culture staining. The "construct" was made of MatrigelBD with GMSC. To assess the biocompatibility, an MTT cellular proliferation assay was performed. The differentiation potential of the cells toward the osteogenic, adipogenic, and chondrogenic lineages was analyzed after 21 days of growth in MatrigelBD with induction differentiation media. The MTT analysis showed that MatrigelBD stimulated cell proliferation; the GMSCs maintained the expression of MSC markers. Importantly, the growth of GMSCs within the MatrigelBD did not interfere with the cell differentiation potential. These findings indicate that MatrigelBD is biocompatible with GMSCs, and this matrix improves cell proliferation in vitro.

MatrigelBD es un andamiaje de hidrogel con redes tridimensionales entrecruzadas de polímeros hidrófilos con un alto contenido de agua. El tejido gingival humano podría representar una mejor fuente de MSCs, estas células pueden obtenerse fácilmente de una manera relativamente no invasiva. El objetivo de este estudio fue evaluar la biocompatibilidad de MatrigelBD con GMSCs in vitro. Muestras gingivales de tejido conectivo se obtuvieron de donantes sanos. El tejido se trituró y se cultivó durante dos semanas, y cuando las células se encontraban en el cuarto pasaje se les analizó su fenotipo inmunológico utilizando citometría de flujo. Se indujo la diferenciación hacia los linajes osteogénico, condrogénico y adipogénico, evaluandose con tinciones. El "constructo" se hizo de MatrigelBD con GMSC. Para evaluar la biocompatibilidad, se realizó un ensayo de proliferación celular: MTT. Se analizó el potencial de diferenciación de las células hacia los linajes osteogénico, adipogénico y condrogénico después de 21 días de cultivo en MatrigelBD con medio de diferenciación de inducción. El análisis de MTT mostró que MatrigelBD estimula la proliferación celular; GMSCs mantiene la expresión de marcadores de MSC. Es importante destacar que el crecimiento de GMSCs en MatrigelBD no interfirió con el potencial de diferenciación celular. Estos hallazgos indican que MatrigelBD es biocompatible con GMSCs, y esta matriz mejora la proliferación celular in vitro.

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Aim. The purpose of this article is to summarize the way young medical professionals view these modern biomedical procedures and their moral acceptability. Materials and methods. A survey, filled in online, analyzing items in four main areas: genetic techniques, cloning, stem cell research, and assisted reproduction. Results. Genetics related items. Most subjects agreed that the right to the genetic material should be a fundamental human right and that genetic engineering should be used if it could lead to the elimination os severe genetic diseases like cystic fibrosis and thalassemia. The least acceptance rate was obtained for techniques that would either change physical traits (like eye or hair color) or augment them. Assisted reproductive techniques. Most subjects agreed that the prenatal screening should be mandatory, and if the screening detects a severe congenital malformation the physician should recommend therapeutic abortion. Cloning. Most subjects disagreed that cloning of any type, either therapeutic or reproductive, using human, animal, or vegetal genetic material. Stem cell research. Most subjects agreed with the collection and storage of cord blood stem cells and the use of adult stem cells, and most of them disagreed with the creation of embryos specifically for obtaining stem cells. Conclusions. Even if the national legislation in this area is very scarce, the responses have usually identified the highly controversial techniques. If however the national legislation has elements similar to the items from the survey, they tended to take the respective items as morally acceptable without trying to analyze them critically.

Objetivo. El propósito de este artículo es recoger la forma en que jóvenes profesionales médicos ven los procedimientos biomédicos modernos y su aceptabilidad moral. Materiales y métodos: Una encuesta, rellenada online, que analiza elementos en cuatro áreas principales: técnicas genéticas, clonación, investigación con células madre y reproducción asistida. Resultados: Elementos relacionados con la genética: La mayoría de los sujetos acepta que el derecho a material genético debería ser un derecho humano fundamental y que la ingeniería genética debería usarse si pudiese eliminar enfermedades genéticas severas como la fibrosis quística y la talasemia. Se obtuvo una frecuencia de aceptación menor para técnicas que pudieran o cambiar características físicas (como el color de los ojos o del pelo) o aumentarlas. Técnicas de reproducción asistida: La mayoría de los sujetos acepta que el examen de detección prenatal debiera ser mandatorio y si se detecta una deformación congénita severa, el médico debería recomendar aborto terapéutico. Clonación: La mayoría de los sujetos no acepta clonación de ningún tipo, terapéutica o reproductiva, usando material genético humano, animal o vegetal. Investigación con células madre: La mayoría de los sujetos acepta recoger y almacenar células madre del cordón umbilical y el uso de células madre adultas y está en desacuerdo con la creación de embriones específicamente para obtener células madre. Conclusiones: aunque la legislación nacional en esta área es muy escasa, las respuestas por lo general han identificado las técnicas altamente controversiales. Sin embargo, si la legislación nacional tiene elementos similares a los temas de la encuesta, se tiende a tomarlos respectivamente como moralmente aceptables sin tratar de analizarlos críticamente.

Objetivo. A proposta deste artigo é sumarizar o modo de ver dos jovens profissionais médicos sobre procedimentos biomédicos modernos e sua aceitação moral. Materiais e métodos. Uma pesquisa de opinião realizada online, analisou ítens de quatro principais áreas: técnicas genéticas, clonagem, pesquisa com células-tronco, e reprodução assistida. Resultados. Itens relacionados à Genética. A maioria dos sujeitos concordaram que o direito ao material genético deveria ser um direito humano fundamental e que a engenharia genética poderia ser usada se puder levar à eliminação de doenças genéticas severas como a fibrose cística e a talassemia. A menor taxa de aceitação foi obtida para técnicas que pudessem modificar o aspecto físico individual (como olho e cor do cabelo) ou aumentá-los. Técnicas de reprodução assistida. A maioria dos sujeitos concordaram que a seleção pré-natal (screening) deverá ser impositiva, e que se o "screening" detetar uma severa malformação congênita o médico deveria recomendar o aborto terapêutico. Clonagem. A maioria dos sujeitos discordaram da clonagem de qualquer tipo, terapêutica ou reprodutiva, com material genético de uso humano, animal, ou vegetal. Pesquisa com células-tronco. A maioria dos sujeitos concordaram com a obtenção e estocagem de células-tronco de sangue do cordão umbilical e a utilização de células-tronco adultas , e a maioria deles discordaram da criação de embriões especificamente para a obtenção de células-tronco. Conclusões. Mesmo que a legislação nacional na área seja muito escassa, as respostas usualmente identificaram as técnicas como altamente controversas. Quando a legislação nacional oferece elementos semelhantes aos ítens obtidos pela pesquisa de opinião, eles tenderiam a tomar os respectivos ítens como moralmente aceitáveis sem tentar analisá-los criticamente.

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As células-tronco mesenquimais (CTM) constituem uma ferramenta promissora para o campo de terapia celular. Além de seu potencial de diferenciação em diferentes tipos celulares, as CTM apresentam a habilidade de secretar moléculas bioativas e, assim, exercer múltiplos efeitos biológicos, tais como indução da regeneração de tecidos lesionados, redução de fibrose e modulação do sistema imune. A superexpressão dos fatores de crescimento G-CSF e IGF-1, conhecidos por seus efeitos sobre os processos de imunomodulação, sobrevivência celular e reparo tecidual, pode ampliar as ações terapêuticas das CTM. O objetivo deste trabalho consiste em gerar e caracterizar linhagens de CTM de camundongo superexpressando hGCSF ou hIGF-1. Um sistema lentiviral de segunda geração foi utilizado para modificação de CTM para expressão ectópica dos genes de interesse. As sequências codificantes de hG-CSF e hIGF-1 foram amplificadas por PCR e subclonadas em um vetor lentiviral de transferência, contendo um promotor constitutivo. As partículas lentivirais foram produzidas a partir da cotransfecção de células da linhagem HEK293FT...

Mesenchymal stem cells (MSCs) are a promising tool for the cell therapy field. In addition to their potential for differentiation into different cell types, MSCs have the ability to secrete bioactive molecules and thus exert multiple biological effects such as induction of the injured tissue regeneration, fibrosis reduction and modulation of the immune system. The overexpression of the growth factors G-CSF and IGF-1, known for their effects on immune modulation processes, cell survival and tissue repair, can result in a magnification of MSCs' therapeutic actions. The objective of this work is to generate and characterize mouse MSCs lines overexpressing hG-CSF or hIGF-1. A second generation lentiviral system was used to modify MSCs derived from mice for the ectopic expression of the genes of interest. The coding sequences of hG-CSF and hIGF-1 were amplified by PCR and subcloned into a lentiviral transfer vector containing a constitutive promoter. The lentiviral particles were produced from the co-transfection of HEK293FT...

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O desenvolvimento de medicamentos e terapias baseados nos princípios científicos das biociências e das biotecnologias da saúde - com base em células vivas e difíceis de serem estandardizadas - tem sido um tema de amplos debates públicos em nível global. A área tem sido recentemente demarcada como medicina regenerativa, que inclui as pesquisas e terapias com células-tronco (PCT e TC), foco deste estudo. No presente artigo, apresentam-se os principais eventos históricos na área de pesquisa celular, descreve-se o estágio atual na evolução da medicina regenerativa e as características das principais políticas desenvolvidas, em especial pelos países de liderança global, e em relação à regulamentação dos direitos de propriedade intelectual. Desenvolve-se análise quantitativa e qualitativa com dados secundários coletados em nível internacional, revisão bibliográfica e de informações em arquivos das instituições de regulamentação globais, resenhas jornalísticas atualizadas, assim como de artigos especializados publicados em revistas internacionais. A revisão da informação é guiada pelas seguintes perguntas: quais são as trajetórias principais de inovação em ciência e saúde nessa área? Quais fatores incidem principalmente na sua evolução? Conclui-se com reflexões específicas sobre os impactos dos desenvolvimentos associados na Saúde Coletiva...

Medicine and therapy developments based upon scientific principles of biosciences and health biotechnologies - with the use of live cells which are difficult to standardize - have been subject of wide public debates at the global level. The area has been recently defined as one of regenerative medicine that includes stem cell research and therapy, the focus of our study. This paper presents the main historical research events in cellular research, describes regenerative medicine's present stage of evolution and the characteristics of the main public policies developed, most especially in the leading countries and in relation to the regulation of intellectual property rights. A quantitative and qualitative analysis is developed, drawing upon different sets of secondary data collected internationally, bibliographic and archival information from global regulatory institutions, updated journal reviews as well as of specialized articles published in international journals. This information is reviewed guided by the following questions: which are the main trajectories in health innovation in this area? Which factors have most highly influenced its evolution? The paper concludes with reflections on the specific impacts of associated developments on collective health...

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Noções de natureza desempenham um papel na cobertura midiática e nas percepções do público sobre as biotecnologias. Neste estudo, fez-se uma análise de discurso das ideias de natureza e natural usadas na cobertura jornalística brasileira da clonagem e das células-tronco. Nos discursos oponentes, as técnicas biotecnológicas foram postas fora da ordem natural e consideradas imorais. Nos discursos defensores, apareceu uma natureza indiferente ou cruel para com o destino humano, ou obstáculo a ser superado; por outro lado, a clonagem e as células-tronco embrionárias foram naturalizadas, e a ovelha Dolly foi antropomorfizada. As representações mitificantes ou transcendentes da natureza não incidem somente sobre as percepções públicas, têm implicações éticas e políticas.

Different conceptions of nature influence media coverage and public opinion about biotechnology. This study reports on a discourse analysis of the ideas about nature and what is natural expressed in Brazilian media coverage of cloning and stem cell research. In the discourse against this research, the biotechnologies in question are placed outside the natural order of things and deemed immoral. In the discourse of those who defend it, nature is portrayed as indifferent to the fate of humans or even cruel, or else a barrier to be overcome, while cloning and embryonic stem cells are naturalized and Dolly the sheep is anthropomorphized. The mythifying or transcendental representations of nature do not just influence public opinion, but also have ethical and political implications.

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O artigo visa a apresentar uma revisão crítica da literatura internacional contemporânea sobre novas formas de governança e da participação pública, focando nas biociências da saúde de ponta, na área da saúde coletiva e pública. A nova genética confronta os indivíduos, as comunidades e os Estados nacionais com uma reformulação de supostos previamente aceitos, sobre a natureza da vida e das relações entre os seres vivos. Esses desenvolvimentos, nos países industrializados, têm rearticulado as relações entre cidadãos, cientistas e agentes de políticas públicas, e levado a adotar estratégias que envolvem simultaneamente o público em geral e excluem algumas das suas contribuições para o desenho de sistemas de regulação. O objetivo do artigo é apresentar elementos conceituais sobre esses temas que permitam, por um lado, refletir sobre a evolução das teorias e, por outro, difundir algumas das suas contribuições relevantes para a participação pública no contexto brasileiro. Exemplifica-se com o caso das pesquisas com células-tronco (PCTs) e das terapias celulares, segundo informações de uma pesquisa em andamento coordenada pela autora desse artigo. No final, o artigo analisa os tipos de consultas públicas elaboradas sobre a PCT nacional e a inclusão limitada das organizações da sociedade civil brasileira.

This paper presents a critical review of contemporary international literature on new forms of governance and public engagement, and focuses on cutting-edge health biosciences in the fields of social and public health. New genetics confronts individuals, communities and national States with a redefinition of conventional assumptions on the nature of life and of relationships between living beings. In industrialized countries, these developments have contributed to reorganize relationships between citizens, scientists and policymakers, and have led to the design of public policy strategies that, simultaneously, promote the engagement of the general public and exclude some of their contributions from regulatory frameworks. The main aim of this paper is to discuss selected theoretical concepts on these topics, as an incentive, on the one hand, for a reflection on the evolution of theories and, on the other, for the dissemination of contributions eventually relevant to public participation in the Brazilian context. Examples are provided on the case of stem cell research and therapy (SCR), based on data from an ongoing research project coordinated by the author. Towards the end, the text analyzes the public consultation processes on stem cell research developed in Brazil and their limited inclusion of voices from civil society organizations.

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This article analyzes the main rhetorical approaches, themes, and strategies in the contemporary public debate on stem cell research and therapies in Brazil. Based on previous studies, analysis of the transcripts from public hearings, newspaper clippings, and 15 semi-structured interviews with researchers, public policymakers, and representatives of organized civil society, the article discusses divergences and convergences between the proponents and opponents of such research. The results are compared to trends in the United Kingdom, a leading country in the field, concluding that the main difference between the two countries lies in the form of social inclusion and public participation.

Dans cet article, on examine quelques recours rhétoriques, thèmes et stratégies mis en oeuvre pendant les débats publics actuels au Brésil, concernant la recherche sur les cellules souches et les thérapies cellulaires. À partir de travaux universitaires antérieurs, de l'analyse de transcriptions d'audiences publiques et d'articles de presse ainsi que de 15 entretiens semi-structurés avec des chercheurs, des agents de politiques publiques et des représentants de la société civile au Brésil, on évalue les divergences et convergences entre les défenseurs et opposants de ces recherches. On termine sur une réflexion comparative de controverses choisies au Brésil et au Royaume Uni - ce dernier étant pionnier dans ce domaine et dans sa réglementation -, par rapport à des propositions de politiques publiques, une construction technique, scientifique et morale des discours, un rôle de participation de la société civile et l'insertion globale des pays en génétique et biotechnologie. On en conclut que les plus grandes divergences reposent sur les formes d'inclusion et l'engagement du public.