Identification of new human mesenchymal stem cell biomarker by aptamers / Identificação de novos biomarcadores de células tronco mesenquimais de origem humana por meio de aptâmeros

Stem cells are undifferentiated cells that can be distinguished from others by their ability to self-renew and to differentiate into new specific cell types. Mesenchymal stem cells (MSC) are adult stem cells that can be obtained from different sources, such as adipose tissue, bone marrow, dental pulp, and umbilical cord. They can either replicate, originating new identical cells, or differentiate into cells of mesodermal origin and from other germ layers. MSC have been studied as new tools for regenerative therapy. Although encouraging results have been demonstrated, MSC-based therapies still face a great barrier: the difficulty of isolating these cells from heterogeneous environments. MSC are currently characterized by immunolabelling through a set of multiple surface membrane markers, including CD29, CD73, CD90 and CD105, which are also expressed by other cell types. Hence, the present work aimed to identify new specific biomarkers for the characterization of human MSC using DNA aptamers produced by the SELEX (Systematic Evolution of Ligands by EXponential Enrichment) technique. Our results showed that MSC from different origins bound to DNA candidate aptamers, that is, DNA or RNA oligonucleotides selected from random libraries that bind specifically to biological targets. Aptamer-bound MSC could be isolated by fluorescenceactivated cell sorting (FACS) procedures, enhancing the induction of differentiation into specific phenotypes (chondrocytes, osteocytes and adipocytes) when compared to the whole MSC population. Flow cytometry analyses revealed that candidate aptamers bound to 50% of the MSC population from dental pulp and did not present significant binding rates to human fibroblasts or lymphocytes, both used as negative control. Moreover, immunofluorescence images and confocal analyses revealed staining of MSC by aptamers localized in the surfacemembrane of these cells. The results also showed internal staining of human monocytes by our investigated aptamers. A non-specific control aptamer (CNTR APT) obtained from the random pool was then utilized to compare the specificity of the aptamers bound to the analyzed non-apoptotic cells, showing no staining for MSC. However, 40% of the monocytes bound to the CNTR APT. Normalized data based on the cells bound to candidate aptamers compared to those bound to the CNTR APT, revealed a 10 to 16-fold higher binding rate for MSC against 2-fold for monocytes. Despite its low specificity, monocyte-aptamer binding occurs probably due to the expression of shared markers with MSC, since monocytes are derived from hematopoietic stem cells and are important for the immune system ability to internalize/phagocyte external molecules. Given that, we performed a pull-down assay followed by mass spectrometry analysis to detect which MSC-specific protein or other target epitope not coexpressed by monocytes or the CNTR APT would bind to the candidate aptamer. Distinguishing between MSC and monocyte epitopes is important, as both cells are involved in immunomodulatory effects after MSC transplantations. ADAM17 was found to be a target of the APT10, emerging as a possible biomarker of MSC, since its involvement in the inhibition of the TGF signaling cascade, which is responsible for the differentiation of MSC. Thus, MSC with a higher stemness profile should overexpress the protein ADAM17, which presents a catalytic site with affinity to APT10. Another target of Apt 10 is VAMP3, belonging to a transmembrane protein complex that is involved in endocytosis and exocytosis processes during immune and inflammatory responses. Overall, proteins identified as targets of APT10 may be cell surface MSC biomarkers, with importance for MSC-based cell and immune therapies

Células tronco são células indiferenciadas que podem ser distinguidas de outros tipos celulares por meio da habilidade de se auto renovarem e de se diferenciarem em novos tipos celulares. Células tronco mesenquimais (MSC) são células tronco adultas encontradas em diferentes tecidos como tecido adiposo, polpa de dente e cordão umbilical. Estas células podem se autodividir em células idênticas ou se diferenciarem em células de origem mesodermal. Estas células têm sido estudadas em novas aplicações que envolvem terapia regenerativas. Embora resultados encorajadores tenham sido demonstrados, terapias que utilizam MSC ainda encontram uma grande barreira: a dificuldade no isolamento destas células a partir de um ambiente heterogêneo. MSC são caracterizadas por populações positivas em ensaios de imunomarcação para os epítopos membranares CD29, CD73, CD90 e CD105, presentes também em outros tipos celulares. Assim, o presente trabalho tem o objetivo de identificar novos biomarcadores de MSC de origem humana, utilizando aptâmeros de DNA produzidos pela técnica SELEX (Systematic Evolution of Ligands by EXponential Enrichment) como ferramenta. Nossos resultados mostraram que MSC de diferentes origens ligam-se a aptâmeros (oligonucleotídeos de DNA ou RNA que atuam como ligantes específicos de alvos moleculares) de DNA candidatos que atuam no isolamento de MSC por meio da técnica FACS de separação celular, promovendo uma maior indução de diferenciação em células específicas (condrócitos, osteócitos e adipócitos) comparada com a população total de MSC. Análises de citometria de fluxo mostraram que os aptâmeros candidatos se ligam a 50% das MSC de polpa de dente e não apresentam taxa de ligação significante para fibroblastos e linfócitos de origem humana - utilizados como controles negativo. Além domais, imagens de imunofluorescência e confocal mostraram ligação na superfície da membrana de MSC e a marcação interna de monócitos a estes aptâmeros. Portanto, um aptâmero controle (CNTR APT) foi utilizado para comparar a especificidade dos aptâmeros ligados a células viáveis, mostrando a não ligação deste aptâmero a MSC. Porém, 40% da população de monócitos ligou-se ao CNTR APT. Uma normalização baseada na comparação entre as taxas de ligação entre células ligadas com aptâmeros candidatos e o aptâmero controle gerou uma taxa de especificidade entre 10-16 vezes maior para MSC contra 2,5 vezes para os monócitos. Deste modo, embora os resultados tenham mostrado uma taxa de ligação entre monócitos e aptâmeros, as MSC ligadas aos aptâmeros candidatos possuem uma maior taxa de especificidade devido a uma maior presença de antígenos que são expressos em ambas as células. Um ensaio de Pull Down seguido de espectrometria de massas foi utilizado para a identificação de biomarcadores que se ligariam aos aptâmeros candidatos, e que não seriam co-expressos por monócitos e por antígenos ligados ao aptâmero controle. Deste modo, a proteína ADAM17 foi identificada nas amostras de APT10 ligadas às MSC. Tal proteína está relacionada à inibição de uma cascata de sinalização da família de proteínas TGF, responsável pela diferenciação de MSC. Assim, MSC com maior potencial tronco deveriam expressar ADAM17 em maior quantidade. Tal proteína apresenta um sítio catalítico que demonstra interagir com o APT10, de acordo com predição Docking entre proteína e DNA. Foi identificada também, a proteína VAMP3, que pertence a um complexo proteico transmembranar responsável pelos processos de endocitose e exocitose, e que podem ter um papel importante na liberação de citocinas e outras moléculas relacionadas às respostas imune e inflamatórias. Deste modo, o APT10 identificou proteínas importantes que devem estar relacionas com a melhora de imunoterapias que utilizam MSC

Obtención y caracterización de células madre mesenquimales adultas de la pulpa dental humana / Obtaining and characterizing adult mesenchymal stem cells from human dental pulp

El presente trabajo de investigación tiene como objetivo principal el aislar, expandir y caracterizar inmunofenotípicamente células madre mesenquimales de la pulpa dental humana, según los criterios mínimos propuestos por The International Society for Cellular Therapy (ISCT), como así también establecer la puesta a punto de las técnicas y protocolos de procedimientos para tal fin. Los cultivos fueron permanentemente monitoreados mediante microscopio invertido con contraste de fase y la inmunotipificación fue realizada por citometría de flujo (AU)

Papila Apical Dentaria como fuente de Células Madre: Relato del Caso / Papila Apical Dentária como Fonte de Células Tronco: Relato de Caso / Dental Apical Papila Stem Cell: Case Report

Las células madre mesenquimales (MSCs) son células madre adultas que tienen la capacidad de diferenciarse en varios tipos de células. Una rica fuente de células madre mesenquimales puede ser obtenida de tejidos dentales como la papila apical. El objetivo de este trabajo fue extraer las células madre de la papila apical de terceros molares humano para mirar su viabilidad de usarse en la práctica clínica. La de separación celular con Stro-1 y estudio de cinéticas fueron realizados. Como resultado, esas células, presentaron alta tasa de proliferación, formación de colonias y fácil acceso. Concluimos que el uso de células madre obtenidas de diente puede ser una buena alternativa por ser de fácil acceso, alta viabilidad celular y expresión positiva para marcadores celulares mesenquimales. La regeneración tisular o la formación de estructuras craneofaciales constituyen el futuro de la medicina regenerativa, ofreciendo una posibilidad para el tratamiento de malformaciones congénitas, traumas y otras enfermedades.

As células tronco mesenquimais (CTMs) são células-tronco adultas que têm a capacidade de diferenciar em vários tipos de células. Uma rica fonte de células tronco mesenquimais pode ser obtida de tecidos dentais como a papila apical. O objetivo deste trabalho foi extrair as células tronco da papila apical de terceiros molares humano para ver sua viabilidade de ser usado na prática clinica. Teste separação celular com Stro-1 e estudo de cinéticas foram realizados. Como resultado essas células apresentaram alta taxa de proliferação, formação de colônias e de fácil acesso. Concluímos que o uso de células tronco obtidas de dente pode ser uma boa alternativa por ser fácil acesso, alta viabilidade celular e expressão positiva para marcadores celulares mesenquimais. A regeneração tecidual ou a formação de novo de estruturas craniofaciais é o futuro da medicina regenerativa, oferecendo uma possibilidade para tratamento de malformações congênitas, traumas e outras doenças.

Mesenchymal stem cells (MSCs) are adult stem cells that have the ability to diffe-rentiate into various cell types. A rich source of mesenchymal stem cells can be obtained from dental tissues such as the apical papilla. The objective of this work was to extract stem cells from the apical papilla of third human molars to see its feasibility to be used in clinical practice. Testing of cell separation with Stro-1 and study of kinetics were performed. As a result, these cells had a high rate of pro-liferation, formation of colonies and easy access. We conclude that the use of stem cells obtained from teeth can be a good alternative because they are easy to ac-cess, high cell viability and positive ex-pression for mesenchymal cell markers. Tissue regeneration or new formation of craniofacial structures is the future of re-generative medicine, offering a possibili-ty for treatment of congenital malforma-tions, traumas and other diseases.

Awareness, attitude and ethical concern of dental practitioners about stem cells: a cross-sectional study / Consciência, atitude e preocupação ética dos dentistas sobre as células-tronco: um estudo transversal

Objective: To evaluate the level of awareness and attitude among dental practitioners regarding the use of stem cells in dentistry and to determine their knowledge of ethical concerns related to the recent therapy. Material and Methods: A cross-sectional survey-based study was conducted at Taibah University Dental College and Hospital. Medina and at governmental and private dental clinics at the western region of Saudi Arabia. Responses of dental practitioners who completed the survey were recorded between March 2019 and July 2019 without containing any personal identifiers. Level of awareness and attitude and knowledge about ethical issues in relation to stem cell therapy was established. Results: A total of 214 male and female dental practitioners participated in this study and the majority were registered at the Saudi Commission for Health Specialists 128 (59.8%). Dental consultants reported the highest percentage of awareness about dental stem cells (96%, p= 0.005), whereas general dental practitioners (56.2%, p= 0.005) and specialists (52%, p= 0.005), respectively had a lower percentage. When ethical concerns were determined, dental consultants had the highest percentage of knowledge (56%, p= 0.005), whereas dental practitioners (71.2%, p= 0.005) with < 5 years of experience (69.1, p= 0.002) lacked information about related ethical issues. Conclusion: Ways to increase stem cell awareness among dental practitioners in this study recommended including stem cell topics in the dental curriculum and organizing frequent seminars and conferences on this subject. (AU)

Objetivo: Avaliar o nível de consciência e a atitude dos dentistas em relação ao uso de células-tronco na odontologia, e determinar o conhecimento desses profissionais sobre as questões éticas relacionadas à terapia recente. Material e Métodos: Um estudo transversal baseado em pesquisa foi conduzido na Faculdade e Hospital de Odontologia da Universidade Taibah de Medina e em clínicas odontológicas governamentais e privadas na região oeste da Arábia Saudita. As respostas dos dentistas que responderam à pesquisa foram registradas entre março de 2019 e julho de 2019, sem conter nenhum identificador pessoal. Foi estabelecido o nível de consciência, atitude e conhecimento sobre questões éticas em relação à terapia com células-tronco. Resultados: Um total de 214 dentistas do sexo masculino e feminino participaram deste estudo, sendo 128 (59,8%) desses cadastrada na Comissão Saudita de Especialistas em Saúde. Os consultores em odontologia relataram o maior percentual de conhecimento sobre as células-tronco dentárias (96%, p = 0,005), enquanto os dentistas gerais (56,2%, p = 0,005) e especialistas (52%, p = 0,005) tiveram um percentual menor. Quando as questões éticas foram determinadas, os consultores em odontologia tiveram o maior percentual de conhecimento (56%, p = 0,005), enquanto os dentistas (71,2%, p = 0,005) com menos de 5 anos de experiência (69,1%, p = 0,002) tinham menos informações sobre questões éticas relacionadas. Conclusão: Formas de aumentar a conscientização sobre as células-tronco entre os dentistas deste estudo incluem tópicos sobre células-tronco no currículo de odontologia, além de frequentemente organizar seminários e conferências sobre o assunto. (AU)

Establecimiento e implementación de un protocolo simplificado de expansión y cultivo de Células Madre de Pulpa Dental Humana (DPSCh) / Estabelecimento e implementação de um protocolo simplificado para expansão e cultura de células-tronco da polpa dentária humana (DPSCh) / Establishing and implementing a simplified protocol for the expansion and culture of human dental pulp stem cells (hDPSCs)

Resumen Objetivos: Establecer e implementar un protocolo simplificado de extracción, aislamiento primario y cultivo de células madre derivadas de la pulpa dental humana (DPSCh). Analizar cuantitativamente y cualitativamente las células aisladas. Metodología: 10 terceros molares sanos donados por pacientes que concurrieron a la Facultad de Odontología, UdelaR y otorgaron su consentimiento escrito fueron procesados antes de las 48 hs. Se realizó la fractura de la pieza para la obtención del tejido pulpar y se procesó por el método explante. Se analizó viabilidad celular y expresión de marcadores por citometría de flujo en pasajes 4 y 12 y se corroboró mediante inmunocitoquímica. Resultados: Las células obtenidas presentaron una vitalidad mayor al 90% en todos los pasajes, observándose una morfología característica y expresión de marcadores de células madre mesenquimales CD90, C105, CD73, CD29 y 166 mediante citometría de flujo en ambos pasajes. Conclusiones: Se logró establecer un protocolo de aislamiento y expansión celular, con alta tasa de éxito de una población de DPSCh.

Resumo Objetivos: Estabelecer e implementar um protocolo simplificado para a extração, isolamento primário e cultura de células-tronco da polpa dentária humana (DPSCh). Analise as células isoladas quantitativa e qualitativamente. Metodologia: 10 terceiros molares saudáveis ​​doados por pacientes que frequentaram a Faculdade de Odontologia UdelaR e deram consentimento por escrito foram processados ​​antes de 48 horas. A fratura da peça foi realizada para obtenção do tecido pulpar e processada pelo método do explante. A viabilidade celular e a expressão do marcador foram analisadas por citometría de fluxo nas passagens 4 e 12 e confirmadas por inmunocitoquímica. Resultados: As células obtidas apresentaram viabilidade superior a 90% em todas as passagens, observando uma morfologia característica e expressão dos marcadores de células-tronco mesenquimais CD90, C105, CD73, CD29 e 166 por citometría de fluxo em ambas as passagens. Conclusões: Foi possível estabelecer um protocolo de isolamento celular, com alta taxa de sucesso e segurança para isolar o DPSCh.

Abstract Objectives: To establish and implement a simplified protocol for the extraction, primary isolation, and culture of human dental pulp stem cells (hDPSCs). To analyze the isolated cells quantitatively and qualitatively. Methodology: Ten healthy third molars were donated by patients who attended the School of Dentistry, UdelaR, and gave their written consent. The teeth were processed within 48 hours. The teeth were sectioned to obtain the pulp tissue and processed with the explant method. Cell viability and marker expression were analyzed by flow cytometry at passages 4 and 12 and verified by immunocytochemistry. Results: The cells obtained had a vitality greater than 90% in all passages. We found the characteristic morphology and the expression of CD90, C105, CD73, CD29 and 166 mesenchymal stem cell markers by flow cytometry in both passages. Conclusion: It was possible to establish a cell isolation protocol that is highly successful and safe to isolate hDPSC.

Identification and characterization of murine adipose tissue-derived somatic stem cells of Shenque (CV8) acupoint / 中华医学杂志(英文版)

BACKGROUND@#Shenque (CV8) acupoint is located on the navel and has been therapeutically used for more than 2000 years in Traditional Chinese Medicine (TCM). However, clinical research on the underlying therapeutic molecular mechanisms of the CV8 acupoint lags far behind. This study aimed to study the mechanisms of umbilical acupoint therapy by using stem cells.@*METHODS@#The morphological characteristics of CV8 acupoint were detected under a stereomicroscope using hematoxylin and eosin (H&E) staining. Oil Red, Masson, and immunohistochemical staining on multi-layered slices were used to identify the type of cells at the CV8 acupoint. Cell proliferation was measured by a cell counting kit-8 (CCK-8) method. Flow cytometry and immunohistochemistry were used for cell identification. Induced differentiation was used to compare the differentiation of cells derived from CV8 acupoint and non-acupoint somatic stem cells into other cell types, such as osteogenic, adipogenic, and neural stem cell-like cells.@*RESULTS@#Morphological observations showed that adipose tissues at the linea alba of the CV8 acupoint in mice had a mass-like distribution. Immunohistochemical staining confirmed the distribution of stem cell antigen-1 (Sca-1) positive cells in the multi-layered slices of CV8 acupoint tissues. Cells isolated from adipose tissues at the CV8 acupoint exhibited high expression of Sca-1 and CD44 and low expression of CD31 and CD34, and these cells possessed osteogenic, adipogenic, and neurogenic stem cell-like cell differentiation ability. The cell proliferation (day 4: 0.5138 ± 0.0111 vs. 0.4107 ± 0.0180, t = 8.447, P = 0.0011; day 5: 0.6890 ± 0.0070 vs. 0.5520 ± 0.0118, t = 17.310, P 100 μm: 2.6000 ± 0.5477 vs. 0.8000 ± 0.8367, t = 4.025, P = 0.0038) were significantly enhanced in somatic stem cells derived from the CV8 acupoint compared to somatic stem cells from the groin non-acupoint. However, cells possessed significantly weaker osteogenicity ([2.697 ± 0.627]% vs. [7.254 ± 0.958]%, t = 6.893, P = 0.0023) in the CV8 acupoint group.@*CONCLUSIONS@#Our study showed that CV8 acupoint was rich with adipose tissues that contained abundant somatic stem cells. The biological examination of somatic stem cells derived from the CV8 acupoint provided novel insights for future research on the mechanisms of umbilical therapy.

Isolation and characterization of stem cells derived by human dental pulp from harvest based in rotary and manual techniques used in endodontic therapy / Isolamento e caracterização de células tronco da polpa dentária humana obtidas em instrumentação mecanizada e manual durante tratamento endodôntico

Objective: The aim of this study was to evaluate the impact on the isolation and characterization of stem cells from pulp tissues obtained through rotary instrumentation techniques compared to the manual technique. Material and Methods: Thirty permanent teeth were included, 15 of which were instrumented with rotational technique (Protaper SX) and other 15 with manual technique. Cells obtained were characterized by flow cytometry and proliferation was evaluated by the MTT assay. The plasticity was evaluated for adipogenic, osteogenic and odontogenic differentiations. Results: Cells isolated from the pulp of permanent teeth, by manual techniques, presented fibroblast morphology and were able to differentiate successfully. All lineages expressed CD29, CD73, CD90, CD105, CD146, CD166 and were negative for CD31, CD34 and CD45. MTT assay showing significantly increased proliferation of hDPSCs in 5 and 7 days of the culture. Conclusions: The present study demonstrated that manual instrumentation technique is one of the best candidates to harvest dental pulp tissue as the dental stem cell source due to ability effective expanded with less tissue invasion. The technique of rotational instrumentation proved to be very harmful to the tissues of the dental pulp, and we can't obtain cells using this technique. (AU)

Objetivo: O objetivo deste estudo foi avaliar o impacto no isolamento e caracterização de células-tronco de tecidos pulpares obtidos por meio de técnicas de instrumentação rotatória em comparação à técnica manual. Material e Métodos: Trinta dentes permanentes foram incluídos, 15 dos quais foram instrumentados com técnica mecanizada (Protaper SX) e outros 15 com técnica manual. As células obtidas foram caracterizadas por citometria de fluxo e a proliferação foi avaliada pelo ensaio MTT. A plasticidade foi avaliada quanto às diferenciações adipogênica, osteogênica e odontogênica. Resultados: células isoladas da polpa de dentes permanentes, por técnicas manuais, apresentaram morfologia de fibroblastos e foram capazes de se diferenciar com sucesso. Todas as linhagens expressaram CD29, CD73, CD90, CD105, CD146, CD166 e foram negativas para CD31, CD34 e CD45. O teste de MTT mostrou proliferação significativamente aumentada de hDPSCs em 5 e 7 dias da cultura. Conclusões: O presente estudo demonstrou que a técnica de instrumentação manual é um dos melhores candidatos para a colheita de tecido pulpar como fonte de células tronco dentárias devido à boa capacidade de proliferação celular com menor invasão tecidual. A técnica de instrumentação rotatória provou ser muito prejudicial para os tecidos da polpa dentária, e não possibilitou obter células. (AU)

Autophagy Mediates Astrogenesis in Adult Hippocampal Neural Stem Cells

Neural stem cells (NSCs) have the ability to self-renew and differentiate into neurons, oligodendrocytes, and astrocytes. Highly dynamic nature of NSC differentiation requires the intimate involvement of catabolic processes such as autophagy. Autophagy is a major intracellular degradation pathway necessary for cellular homeostasis and remodeling. Autophagy is important for mammalian development and its role in neurogenesis has recently drawn much attention. However, little is known about how autophagy is associated with differentiation of NSCs into other neural lineages. Here, we report that autophagy plays a critical role in differentiation of adult rat hippocampal neural stem (HCN) cells into astrocytes. During differentiation, autophagy flux peaked at early time points, and remained high. Pharmacological or genetic suppression of autophagy by stable knockdown of Atg7, LC3 or CRISPR-Cas9-mediated knockout (KO) of p62 impaired astrogenesis, while reintroduction of p62 recovered astrogenesis in p62 KO HCN cells. Taken together, our findings suggest that autophagy plays a key role in astrogenesis in adult NSCs.

Periodic Methylation Patterns in the Background Mucosa of Gastric Cancer

BACKGROUND/AIMS: Gastrointestinal glandular stem cells renew every 8 years. New stem cells with impeded housekeeping gene methylation have unstable phenotypes and are prone to transform into malignant cells. Age-related changes in methylation in the gastric mucosa were evaluated to define the period of cancer-prone stem cell replacement. MATERIALS AND METHODS: Endoscopic biopsy specimens of normal-appearing gastric mucosa were obtained from 148 Helicobacter pylori-negative controls, 124 H. pylori-positive controls, and 69 gastric cancer patients with closed-type mucosal atrophy. Methylation-variable sites of two stomach-specific genes (TFF2 and TFF3) and four housekeeping genes (CDH1, ARRDC4, MMP2, and CDKN2A) were analyzed using radioisotope-labeled methylation-specific polymerase chain reaction. Age-related methylation was evaluated depending on the gastric mucosal atrophy at 2-year intervals. RESULTS: TFF2 methylation peaked periodically at 40 to 41, 48 to 49, 56 to 57, and 64 to 65 years of age in H. pylori-negative controls. Periodic peaks of TFF2 methylation were also found in H. pylori-positive controls. Housekeeping-gene methylation troughed at 48 to 49, 56 to 57, and 68 to 69 years of age in cancer patients. Trough methylation of CDH1 and ARRDC4 was lower in cancer patients than in H. pylori-positive controls. CONCLUSIONS: Methylation peaks of stomach-specific TFF2 in controls and methylation troughs of housekeeping genes in cancer patients were found every 8 years. Periodic methylation patterns may be used to identify individuals at high risk for gastric cancer.

Dental stem cell and dental tissue regeneration / 医学前沿

The teeth are highly differentiated chewing organs formed by the development of tooth germ tissue located in the jaw and consist of the enamel, dentin, cementum, pulp, and periodontal tissue. Moreover, the teeth have a complicated regulatory mechanism, special histologic origin, diverse structure, and important function in mastication, articulation, and aesthetics. These characteristics, to a certain extent, greatly complicate the research in tooth regeneration. Recently, new ideas for tooth and tissue regeneration have begun to appear with rapid developments in the theories and technologies in tissue engineering. Numerous types of stem cells have been isolated from dental tissue, such as dental pulp stem cells (DPSCs), stem cells isolated from human pulp of exfoliated deciduous teeth (SHED), periodontal ligament stem cells (PDLSCs), stem cells from apical papilla (SCAPs), and dental follicle cells (DFCs). All these cells can regenerate the tissue of tooth. This review outlines the cell types and strategies of stem cell therapy applied in tooth regeneration, in order to provide theoretical basis for clinical treatments.

mA Regulates Neurogenesis and Neuronal Development by Modulating Histone Methyltransferase Ezh2 / 基因组蛋白质组与生物信息学报·英文版

N-methyladenosine (mA), catalyzed by the methyltransferase complex consisting of Mettl3 and Mettl14, is the most abundant RNA modification in mRNAs and participates in diverse biological processes. However, the roles and precise mechanisms of mA modification in regulating neuronal development and adult neurogenesis remain unclear. Here, we examined the function of Mettl3, the key component of the complex, in neuronal development and adult neurogenesis of mice. We found that the depletion of Mettl3 significantly reduced mA levels in adult neural stem cells (aNSCs) and inhibited the proliferation of aNSCs. Mettl3 depletion not only inhibited neuronal development and skewed the differentiation of aNSCs more toward glial lineage, but also affected the morphological maturation of newborn neurons in the adult brain. mA immunoprecipitation combined with deep sequencing (MeRIP-seq) revealed that mA was predominantly enriched in transcripts related to neurogenesis and neuronal development. Mechanistically, mA was present on the transcripts of histone methyltransferase Ezh2, and its reduction upon Mettl3 knockdown decreased both Ezh2 protein expression and consequent H3K27me3 levels. The defects of neurogenesis and neuronal development induced by Mettl3 depletion could be rescued by Ezh2 overexpression. Collectively, our results uncover a crosstalk between RNA and histone modifications and indicate that Mettl3-mediated mA modification plays an important role in regulating neurogenesis and neuronal development through modulating Ezh2.

Adult Stem Cells in Hibernation: Future Perspectives of Space Travel

Space traveling is imperative for mankind in the future. Expectedly, hibernation will become an option for space traveler to overcome the endless voyage. With regard to some of the studies pointed out that during hibernation, muscle will undergo atrophy and meantime neurogenesis will reduce, these obstacles were frequently related with stem cell regeneration. Thus, investigation on whether hibernation will lead to dysfunction of stem cell becomes an important issue. By going through four main systems in this article, such as, hematopoietic system, skeletal muscle system, central nervous system and orthopedic system, we are expecting that stem cells regeneration capacity will be affected by hibernation. To date, these researches are majorly the read-out from short term or seasonal hibernating mammals. Proposing and creating a simulated long-term hibernation animal model is turning essential for the further investigation on the effect of longer period of hibernation to human stem cells.

Adult Stem Cells: Beyond Regenerative Tool, More as a Bio-Marker in Obesity and Diabetes

Obesity, diabetes, and cardiovascular diseases are increasing rapidly worldwide and it is therefore important to know the effect of exercise and medications for diabetes and obesity on adult stem cells. Adult stem cells play a major role in remodeling and tissue regeneration. In this review we will focus mainly on two adult stem/progenitor cells such as endothelial progenitor cells and mesenchymal stromal cells in relation to aerobic exercise and diabetes medications, both of which can alter the course of regeneration and tissue remodelling. These two adult precursor and stem cells are easily obtained from peripheral blood or adipose tissue depots, as the case may be and are precursors to endothelium and mesenchymal tissue (fat, bone, muscle, and cartilage). They both are key players in maintenance of cardiovascular and metabolic homeostasis and can act also as useful biomarkers.

Current Stem Cell Therapy for Osteoarthritis / 대한내과학회지

Osteoarthritis is a musculoskeletal disease representative of an aging society. As medical conditions are usually complicated in an aging population, osteoarthritis becomes more frequently encountered in the physician's office. There is a growing need, therefore, for physicians to pay attention to this common orthopedic condition. Cartilage degeneration, arthritic pain, and joint dysfunction are major manifestations of osteoarthritis, and degenerated cartilage is difficult to repair with conventional treatment modalities. Scientists and physicians have developed various therapeutic strategies, including the use of stem cells. Here, we discuss previous and current progress in cartilage regenerative therapy against osteoarthritis.

Stem cell therapy in pain medicine / 대한통증학회지

Stem cells are attracting attention as a key element in future medicine, satisfying the desire to live a healthier life with the possibility that they can regenerate tissue damaged or degenerated by disease or aging. Stem cells are defined as undifferentiated cells that have the ability to replicate and differentiate themselves into various tissues cells. Stem cells, commonly encountered in clinical or preclinical stages, are largely classified into embryonic, adult, and induced pluripotent stem cells. Recently, stem cell transplantation has been frequently applied to the treatment of pain as an alternative or promising approach for the treatment of severe osteoarthritis, neuropathic pain, and intractable musculoskeletal pain which do not respond to conventional medicine. The main idea of applying stem cells to neuropathic pain is based on the ability of stem cells to release neurotrophic factors, along with providing a cellular source for replacing the injured neural cells, making them ideal candidates for modulating and possibly reversing intractable neuropathic pain. Even though various differentiation capacities of stem cells are reported, there is not enough knowledge and technique to control the differentiation into desired tissues in vivo. Even though the use of stem cells is still in the very early stages of clinical use and raises complicated ethical problems, the future of stem cells therapies is very bright with the help of accumulating evidence and technology.

Adult Stem Cell Treatment for Rheumatoid Arthritis

Since methotrexate began to be used in the treatment of rheumatoid arthritis (RA) 30 years ago, RA treatments have advanced rapidly from only reducing joint pain and inflammation to suppressing disease progression and joint destruction. In particular, the development of biologics and targeted anti-rheumatic drugs has almost made it possible to induce remission in patients with RA. On the other hand, the current RA treatments are still limited by adverse effects and treatment failure. Stem cell therapy has been suggested as an alternative treatment of RA, and preclinical studies and clinical trials using representative adult stem cells (ASCs), hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs), are currently underway. HSC therapy in RA has mostly progressed based on the concept of ‘immune reset’, in which the existing immune cells are replaced with healthy ones. HSC transplantation was completed relatively safely, and the patients showed a positive treatment response. Nevertheless, the treatment response of HSCs in RA depends on the conditioning regimen, and the efficacy did not persist for a long time. The MSCs possessed a hypo-immunogenicity, immune modulation effect and tissue regeneration capability, making them another promising candidate for the RA treatment. MSC transplantation in RA was found to be safe with few adverse effects, such as immune rejection or embolism, but it showed a partial and transient response. This review addresses the characteristics of ASCs, focusing specifically on HSCs and MSCs, and summarizes the results of preclinical studies and clinical trials of ASC therapy in RA.

For Better Fat Graft Outcome in Soft Tissue Augmentation: Systematic Review and Meta-Analysis

BACKGROUND: Autologous fat grafts are widely used in plastic surgery, but they have the disadvantage of unpredictability due to variable resorption. This meta-analysis examined the literature on the survival rate of autologous fat grafts using objective markers, and investigated the factors that affected the survival rate. METHODS: The reviewers searched the PubMed, EMBASE, and Cochrane Library databases from January 2001 to December 2017. A meta-analysis was performed to estimate fat graft survival and to identify variables that influenced the survival rate. RESULTS: A total of 27 studies (1,066 cases) were included in the meta-analysis. The mean survival rate of grafted fat was 56.5%. The survival rate was significantly higher for cell-assisted lipotransfer (CAL) than for non-CAL (62% vs. 53.4%; P=0.015). The survival rate for procedures performed to correct lipoatrophy was higher than that of procedures performed for other purposes (64.6%; P=0.014), and was significantly higher in patients who underwent breast pre-expansion using the BRAVA device (66.2% vs. 50.35%; P=0.001). There were no significant differences in the survival rate according to the recipient site, harvesting method, or refinement method. CONCLUSIONS: Fat transplantation showed a varying survival rate, with an average of 56%. In patients who underwent CAL or breast pre-expansion with the BRAVA device, the survival rate of transplanted fat was higher than in their counterparts, supporting the use of these techniques in fat transplantation.

Rejuvenating Aged Hematopoietic Stem Cells Through Improvement of Mitochondrial Function

Mitochondria are the powerhouses of the cell as well as the primary site of hematopoiesis, which also occurs in the cytoplasm. Hematopoietic stem cells (HSCs) are characterized by a very high turnover rate, and are thus considered to be relatively free from the age-related insults generated by mitochondria. However, HSCs are also subject to these age-related insults, including the incidence of myeloid proliferative diseases, marrow failure, hematopoietic neoplasms, and deterioration of the adaptive human immune system. Recently, NAD⁺ dietary supplements, known as niacin or vitamin B₃, including tryptophan, nicotinic acid, nicotinamide, and the newly identified NAD⁺ precursor nicotinamide riboside, have been shown to play a role in restoring adult stem cell function through the amelioration of mitochondrial dysfunction. This insight motivated a study that focused on reversing aging-related cellular dysfunction in adult mouse muscle stem cells by supplementing their diet with nicotinamide riboside. The remedial effect of nicotinamide riboside enhanced mitochondrial function in these muscle stem cells in a SIRT1-dependent manner, affecting cellular respiration, membrane potential, and production of ATP. Accordingly, numerous studies have demonstrated that sirtuins, under nuclear/mitochondrial control, have age-specific effects in determining HSC phenotypes. Based on the evidence accumulated thus far, we propose a clinical intervention for the restoration of aged HSC function by improving mitochondrial function through NAD⁺ precursor supplementation.

Nanotechnology and multipotent adult progenitor cells in Reparative Medicine: therapeutic perspectives / Nanotecnologia e as células progenitoras adultas multipotentes na Medicina Reparativa: perspectivas terapêuticas

ABSTRACT The biology of stem cells is one of the most dynamic and promising fields of the biological sciences, since it is the basis for the development of organisms. Its biological complexity demands efforts from several lines of research aimed mainly at its therapeutic use. Nanotechnology has been emerging as a new field of study, which shows great potential in the treatment of various diseases. This new area of health has been called "Nanomedicine" or "Bionanotechnology", which can be applied in Medicine by transport and drug delivery systems, robotic tools to be used in diagnostic and surgical processes, nanobiomaterials, gene therapies, nanobiomedical devices, among others. Because stem cells and Nanotechnology are two areas of extremely promising science, a new field of study, called "stem cell Nanotechnology", has gradually emerged. In this, Nanotechnology is used to help the stem cells apply their therapeutic potential in the treatment, cure, and repair of the damaged tissues, in an effective and safe way. In this way, stem cell Nanotechnology has generated great interest, since it may result in significant contributions to Regenerative Medicine and tissue engineering. The present work aims to present the state-of-the-art regarding its therapeutic use in Human Medicine.

RESUMO A biologia das células-tronco é um dos campos mais dinâmicos e promissores das ciências biológicas, pois é a base do desenvolvimento dos organismos. Sua complexidade biológica demanda esforços de diversas linhas de pesquisa, visando principalmente à sua utilização terapêutica. A Nanotecnologia surge como um novo campo de estudo, o qual demonstra grande potencial no que tange ao tratamento de diversas doenças. Esta nova área da saúde vem sendo denominada "Nanomedicina" ou "Bionanotecnologia", a qual pode ser aplicada na Medicina por meio da utilização de sistemas de transporte e liberação de fármacos, ferramentas robóticas a serem utilizadas em processos de diagnóstico e cirurgia, nanobiomateriais, terapias gênicas, dispositivos nanobiomédicos, entre outros. Em razão das células-tronco e a Nanotecnologia serem duas áreas da ciência extremante promissoras, um novo campo de estudo, denominado "Nanotecnologia das células-tronco", surge gradativamente. Neste, a Nanotecnologia é utilizada de forma a auxiliar as células-tronco a exercerem seu potencial terapêutico no tratamento, na cura e na reparação dos tecidos lesionados, de forma eficaz e segura. A Nanotecnologia das células-tronco tem gerado grande interesse, podendo resultar em contribuições significativas na Medicina Regenerativa e na engenharia de tecidos. O presente trabalho teve por objetivo apresentar o estado da arte visando à sua utilização terapêutica na Medicina Humana.

Células madre mesenquimales para la regeneración de tejidos: ¿Por qué siguen siendo una promesa sin cumplir? / Mesenchymal stem cells for tissue regeneration: ¿Why are they still an unfulfilled promise?

Desde su descripción inicial, hace ya más de 40 años, las células madre mesenquimales (MSC) fueron reconocidas como una importante alternativa para el manejo de enfermedades caracterizadas por la pérdida aguda o crónica de tejido, gracias a su capacidad de proliferación y diferenciación, lo cual les permitiría sustituir las células perdidas y de esta forma recuperar la estructura y función. Cada vez es más abundante la evidencia que sugiere el potencial de estas células para el manejo de un amplio grupo de enfermedades, al menos en modelos experimentales preclínicos. No obstante, esta capacidad no ha podido refrendarse contundente y consistentemente en ensayos clínicos. Con la presente revisión, se pretende presentar una visión del estado actual del desarrollo conceptual en torno a las capacidades terapéuticas de las MSC y un análisis crítico de algunos de los factores, que han impedido que estas sean una opción terapéutica usable en la práctica clínica diaria

Since they were initially identified more than 40 years ago, mesenchymal stem cells (MSCs) were recognized as an important therapeutic alternative for diseases characterized by acute or chronic loss of tissue, thanks to their proliferation and differentiation ability, which would allow the replacement of lost cells and their structural and functional recuperation. Evidence suggesting the therapeutic potential of these cells for a vast group of diseases increases day by day, at least in preclinical experimental models. However, clinical trials have been unable to obtain consistent and categorical results to demonstrate this capacity. This review aims to provide, an overview on the current status of the conceptual development on the therapeutic properties of MSCs, and a critical analysis of some factors which have hindered the application of this therapeutic option in daily clinical practice.