The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Todd C Mcdevitt - One of the best experts on this subject based on the ideXlab platform.
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osteogenic Embryoid Body derived material induces bone formation in vivo
Scientific Reports, 2015Co-Authors: Ken Sutha, Zvi Schwartz, Yun Wang, Sharon L Hyzy, Barbara D Boyan, Todd C McdevittAbstract:The progressive loss of endogenous regenerative capacity that accompanies mammalian aging has been attributed at least in part to alterations in the extracellular matrix (ECM) composition of adult tissues. Thus, creation of a more regenerative microenvironment, analogous to embryonic morphogenesis, may be achieved via pluripotent embryonic stem cell (ESC) differentiation and derivation of devitalized materials as an alternative to decellularized adult tissues, such as demineralized bone matrix (DBM). Transplantation of devitalized ESC materials represents a novel approach to promote functional tissue regeneration and reduce the inherent batch-to-batch variability of allograft-derived materials. In this study, the osteoinductivity of Embryoid Body-derived material (EBM) was compared to DBM in a standard in vivo ectopic osteoinduction assay in nude mice. EBM derived from EBs differentiated for 10 days with osteogenic media (+β-glycerophosphate) exhibited similar osteoinductivity to active DBM (osteoinduction score = 2.50 ± 0.27 vs. 2.75 ± 0.16) based on histological scoring and exceeded inactive DBM (1.13 ± 0.13, p < 0.005). Moreover, EBM stimulated formation of new bone, ossicles and marrow spaces, similar to active DBM. The potent osteoinductivity of EBM demonstrates that morphogenic factors expressed by ESCs undergoing osteogenic differentiation yield a novel devitalized material capable of stimulating de novo bone formation in vivo.
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single cell analysis of Embryoid Body heterogeneity using microfluidic trapping array
Biomedical Microdevices, 2014Co-Authors: Jenna L Wilson, Shalu Suri, Ankur Singh, Catherine A Rivet, Todd C McdevittAbstract:The differentiation of pluripotent stem cells as Embryoid bodies (EBs) remains a common method for inducing differentiation toward many lineages. However, differentiation via EBs typically yields a significant amount of heterogeneity in the cell population, as most cells differentiate simultaneously toward different lineages, while others remain undifferentiated. Moreover, physical parameters, such as the size of EBs, can modulate the heterogeneity of differentiated phenotypes due to the establishment of nutrient and oxygen gradients. One of the challenges in examining the cellular composition of EBs is the lack of analytical methods that are capable of determining the phenotype of all of the individual cells that comprise a single EB. Therefore, the objective of this work was to examine the ability of a microfluidic cell trapping array to analyze the heterogeneity of cells comprising EBs during the course of early differentiation. The heterogeneity of single cell phenotype on the basis of protein expression of the pluripotent transcription factor OCT-4 was examined for populations of EBs and single EBs of different sizes at distinct stages of differentiation. Results from the cell trap device were compared with flow cytometry and whole mount immunostaining. Additionally, single cells from dissociated pooled EBs or individual EBs were examined separately to discern potential differences in the value or variance of expression between the different methods of analysis. Overall, the analytical method described represents a novel approach for evaluating how heterogeneity is manifested in EB cultures and may be used in the future to assess the kinetics and patterns of differentiation in addition to the loss of pluripotency.
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systematic analysis of embryonic stem cell differentiation in hydrodynamic environments with controlled Embryoid Body size
Integrative Biology, 2012Co-Authors: Melissa A Kinney, Rabbia Saeed, Todd C McdevittAbstract:The sensitivity of stem cells to environmental perturbations has prompted many studies which aim to characterize the influence of mechanical factors on stem cell morphogenesis and differentiation. Hydrodynamic cultures, often employed for large scale bioprocessing applications, impart complex fluid shear and transport profiles, and influence cell fate as a result of changes in media mixing conditions. However, previous studies of hydrodynamic cultures have been limited in their ability to distinguish confounding factors that may affect differentiation, including modulation of Embryoid Body size in response to changes in the hydrodynamic environment. In this study, we demonstrate the ability to control and maintain Embryoid Body (EB) size using a combination of forced aggregation formation and rotary orbital suspension culture, in order to assess the impact of hydrodynamic cultures on ESC differentiation, independent of EB size. Size-controlled EBs maintained at different rotary orbital speeds exhibited similar morphological features and gene expression profiles, consistent with ESC differentiation. The similar differentiation of ESCs across a range of hydrodynamic conditions suggests that controlling EB formation and resultant size may be important for scalable bioprocessing applications, in order to standardize EB morphogenesis. However, perturbations in the hydrodynamic environment also led to subtle changes in differentiation toward certain lineages, including temporal modulation of gene expression, as well changes in the relative efficiencies of differentiated phenotypes, thereby highlighting important tissue engineering principles that should be considered for implementation in bioreactor design, as well as for directed ESC differentiation.
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microsphere size effects on Embryoid Body incorporation and embryonic stem cell differentiation
Journal of Biomedical Materials Research Part A, 2010Co-Authors: Richard L Carpenedo, Todd C Mcdevitt, Scott A SeamanAbstract:Received 14 July 2009; revised 28 September 2009; accepted 22 October 2009Published online 8 March 2010 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.a.32710Abstract: Differentiation of pluripotent embryonic stemcells (ESCs) in vitro via multicellular spheroids calledEmbryoid bodies (EBs) is commonly performed to modelaspects of early mammalian development and initiate dif-ferentiation of cells for regenerative medicine technolo-gies. However, the three-dimensional nature of EBs posesunique challenges for directed ESC differentiation, includ-ing limited diffusion into EBs of morphogenic moleculescapable of specifying cell fate. Degradable polymer micro-spheres incorporated within EBs can present morphogenicmolecules to ESCs in a spatiotemporally controlled man-ner to more efficiently direct differentiation. In this study,the effect of microsphere size on incorporation into EBsand ESC differentiation in response to microsphere-mediated morphogen delivery were assessed. PLGAmicrospheres with mean diameters of 1, 3, or 11 lm werefabricated and mixed with ESCs during EB formation.Smaller microspheres were incorporated more efficientlythroughout EBs than larger microspheres, and regardlessof size, retained for at least 10 days of differentiation.Retinoic acid release from incorporated microspheresinduced EB cavitation in a size-dependent manner, withsmaller microspheres triggering accelerated and morecomplete cavitation than larger particles. These resultsdemonstrate that engineering the size of microspheredelivery vehicles incorporated within stem cell environ-ments can be used to modulate the course of differentia-tion. 2010 Wiley Periodicals, Inc. J Biomed Mater Res94A: 466–475, 2010Key words: Embryoid Body; embryonic stem cell; micro-sphere; differentiation; retinoic acid
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engineering the Embryoid Body microenvironment to direct embryonic stem cell differentiation
Biotechnology Progress, 2009Co-Authors: Andres M Brattleal, Todd C Mcdevitt, Richard L CarpenedoAbstract:Embryonic stem cells (ESCs) are pluripotent cells capable of differentiating into all somatic and germ cell types. The intrinsic ability of pluripotent cells to generate a vast array of different cells makes ESCs a robust resource for a variety of cell transplantation and tissue engineering applications, however, efficient and controlled means of directing ESC differentiation is essential for the development of regenerative therapies. ESCs are commonly differentiated in vitro by spontaneously self-assembling in suspension culture into 3D cell aggregates called Embryoid bodies (EBs), which mimic many of the hallmarks of early embryonic development, yet the 3D organization and structure of EBs also presents unique challenges to effectively direct the differentiation of the cells. ESC differentiation is strongly influenced by physical and chemical signals comprising the local extracellular microenvironment, thus current methods to engineer EB differentiation have focused primarily on spatially controlling EB size, adding soluble factors to the media, or culturing EBs on or within natural or synthetic extracellular matrices. Although most such strategies aim to influence differentiation from the exterior of EBs, engineering the microenvironment directly within EBs enables new opportunities to efficiently direct the fate of the cells by locally controlling the presentation of morphogenic cues.
Richard L Carpenedo - One of the best experts on this subject based on the ideXlab platform.
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microsphere size effects on Embryoid Body incorporation and embryonic stem cell differentiation
Journal of Biomedical Materials Research Part A, 2010Co-Authors: Richard L Carpenedo, Todd C Mcdevitt, Scott A SeamanAbstract:Received 14 July 2009; revised 28 September 2009; accepted 22 October 2009Published online 8 March 2010 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.a.32710Abstract: Differentiation of pluripotent embryonic stemcells (ESCs) in vitro via multicellular spheroids calledEmbryoid bodies (EBs) is commonly performed to modelaspects of early mammalian development and initiate dif-ferentiation of cells for regenerative medicine technolo-gies. However, the three-dimensional nature of EBs posesunique challenges for directed ESC differentiation, includ-ing limited diffusion into EBs of morphogenic moleculescapable of specifying cell fate. Degradable polymer micro-spheres incorporated within EBs can present morphogenicmolecules to ESCs in a spatiotemporally controlled man-ner to more efficiently direct differentiation. In this study,the effect of microsphere size on incorporation into EBsand ESC differentiation in response to microsphere-mediated morphogen delivery were assessed. PLGAmicrospheres with mean diameters of 1, 3, or 11 lm werefabricated and mixed with ESCs during EB formation.Smaller microspheres were incorporated more efficientlythroughout EBs than larger microspheres, and regardlessof size, retained for at least 10 days of differentiation.Retinoic acid release from incorporated microspheresinduced EB cavitation in a size-dependent manner, withsmaller microspheres triggering accelerated and morecomplete cavitation than larger particles. These resultsdemonstrate that engineering the size of microspheredelivery vehicles incorporated within stem cell environ-ments can be used to modulate the course of differentia-tion. 2010 Wiley Periodicals, Inc. J Biomed Mater Res94A: 466–475, 2010Key words: Embryoid Body; embryonic stem cell; micro-sphere; differentiation; retinoic acid
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engineering the Embryoid Body microenvironment to direct embryonic stem cell differentiation
Biotechnology Progress, 2009Co-Authors: Andres M Brattleal, Todd C Mcdevitt, Richard L CarpenedoAbstract:Embryonic stem cells (ESCs) are pluripotent cells capable of differentiating into all somatic and germ cell types. The intrinsic ability of pluripotent cells to generate a vast array of different cells makes ESCs a robust resource for a variety of cell transplantation and tissue engineering applications, however, efficient and controlled means of directing ESC differentiation is essential for the development of regenerative therapies. ESCs are commonly differentiated in vitro by spontaneously self-assembling in suspension culture into 3D cell aggregates called Embryoid bodies (EBs), which mimic many of the hallmarks of early embryonic development, yet the 3D organization and structure of EBs also presents unique challenges to effectively direct the differentiation of the cells. ESC differentiation is strongly influenced by physical and chemical signals comprising the local extracellular microenvironment, thus current methods to engineer EB differentiation have focused primarily on spatially controlling EB size, adding soluble factors to the media, or culturing EBs on or within natural or synthetic extracellular matrices. Although most such strategies aim to influence differentiation from the exterior of EBs, engineering the microenvironment directly within EBs enables new opportunities to efficiently direct the fate of the cells by locally controlling the presentation of morphogenic cues.
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rotary suspension culture enhances the efficiency yield and homogeneity of Embryoid Body differentiation
Stem Cells, 2007Co-Authors: Richard L Carpenedo, Todd C Mcdevitt, Carolyn Y SargentAbstract:Embryonic stem (ES) cells hold great promise as a robust cell source for cell-based therapies and as a model of early embryonic development. Current experimental methods for differentiation of ES cells via Embryoid Body (EB) formation are either inherently incapable of larger-scale production or exhibit limited control over cell aggregation during EB formation and subsequent EB agglomeration. This report describes and characterizes a novel method for formation of EBs using rotary orbital motion that simultaneously addresses both concerns. EBs formed under rotary suspension conditions were compared with hanging-drop and static EBs for efficiency of EB formation, cell and EB yield, homogeneity of EB size and shape, and gene expression. A 20-fold enhancement in the number of cells incorporated into primitive EBs in rotary versus static conditions was detected after the first 12 hours, and a fourfold increase in total cell yield was achieved by rotary culture after 7 days. Morphometric analysis of EBs demonstrated formation and maintenance of a more uniform EB population under rotary conditions compared with hanging-drop and static conditions. Quantitative gene expression analysis indicated that rotary EBs differentiated normally, on the basis of expression of ectoderm, endoderm, and mesoderm markers. Increased levels of endoderm gene expression, along with cystic EB formation, indicated by histological examination, suggested that differentiation was accelerated in rotary EBs. Thus, the rotary suspension culture method can produce a highly uniform population of efficiently differentiating EBs in large quantities in a manner that can be easily implemented by basic research laboratories conducting ES cell differentiation studies.
Utkan Demirci - One of the best experts on this subject based on the ideXlab platform.
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embryonic stem cell bioprinting for uniform and controlled size Embryoid Body formation
Biomicrofluidics, 2011Co-Authors: Banupriya Sridharan, Shuqi Wang, Umut A Gurkan, Brian C Syverud, Utkan DemirciAbstract:Embryonic stem cells (ESCs) are pluripotent with multilineage potential to differentiate into virtually all cell types in the organism and thus hold a great promise for cell therapy and regenerative medicine. In vitro differentiation of ESCs starts with a phase known as Embryoid Body (EB) formation. EB mimics the early stages of embryogenesis and plays an essential role in ESC differentiation in vitro. EB uniformity and size are critical parameters that directly influence the phenotype expression of ESCs. Various methods have been developed to form EBs, which involve natural aggregation of cells. However, challenges persist to form EBs with controlled size, shape, and uniformity in a reproducible manner. The current hanging-drop methods are labor intensive and time consuming. In this study, we report an approach to form controllable, uniform-sized EBs by integrating bioprinting technologies with the existing hanging-drop method. The approach presented here is simple, robust, and rapid. We present signific...
George T J Huang - One of the best experts on this subject based on the ideXlab platform.
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human dental stem cell derived transgene free ipscs generate functional neurons via Embryoid Body mediated and direct induction methods
Journal of Tissue Engineering and Regenerative Medicine, 2018Co-Authors: Ikbale El Ayachi, Jun Zhang, Xiaoying Zou, Wei Wei, Kristen M S Oconnell, George T J HuangAbstract:Induced pluripotent stem cells (iPSCs) give rise to neural stem/progenitor cells, serving as a good source for neural regeneration. Here, we established transgene-free (TF) iPSCs from dental stem cells (DSCs) and determined their capacity to differentiate into functional neurons in vitro. Generated TF iPSCs from stem cells of apical papilla and dental pulp stem cells underwent two methods-Embryoid Body-mediated and direct induction, to guide TF-DSC iPSCs along with H9 or H9 Syn-GFP (human embryonic stem cells) into functional neurons in vitro. Using the Embryoid Body-mediated method, early stage neural markers PAX6, SOX1, and nestin were detected by immunocytofluorescence or reverse transcription-real time polymerase chain reaction (RT-qPCR). At late stage of neural induction measured at Weeks 7 and 9, the expression levels of neuron-specific markers Nav1.6, Kv1.4, Kv4.2, synapsin, SNAP25, PSD95, GAD67, GAP43, and NSE varied between stem cells of apical papilla iPSCs and H9. For direct induction method, iPSCs were directly induced into neural stem/progenitor cells and guided to become neuron-like cells. The direct method, while simpler, showed cell detachment and death during the differentiation process. At early stage, PAX6, SOX1 and nestin were detected. At late stage of differentiation, all five genes tested, nestin, βIII-tubulin, neurofilament medium chain, GFAP, and Nav, were positive in many cells in cultures. Both differentiation methods led to neuron-like cells in cultures exhibiting sodium and potassium currents, action potential, or spontaneous excitatory postsynaptic potential. Thus, TF-DSC iPSCs are capable of undergoing guided neurogenic differentiation into functional neurons in vitro, thereby may serve as a cell source for neural regeneration.
Athanasios Mantalaris - One of the best experts on this subject based on the ideXlab platform.
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in vitro direct chondrogenesis of murine embryonic stem cells by bypassing Embryoid Body formation
Stem Cells and Development, 2008Co-Authors: Yushik Hwang, Julia M Polak, Athanasios MantalarisAbstract:Current approaches on the chondrogenic differentiation of embryonic stem cells (ESCs) involve Embryoid Body (EB) formation, resulting in a fragmented process where control of differentiation, integ...
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in vitro direct osteogenesis of murine embryonic stem cells without Embryoid Body formation
Stem Cells and Development, 2008Co-Authors: Yushik Hwang, Julia M Polak, Athanasios MantalarisAbstract:Embryonic stem cells (ESCs) posses the ability to self-renew and differentiate into a multitude of lineages, including the osteogenic lineage in vitro. Currently, most approaches have focused on embryonic Body (EB)-mediated osteogenic differentiation, which relies on formation of all three germ layers resulting in limited yields and labour-intensive culture processes. Our study aimed at developing an efficient culture strategy resulting in the upregulated in vitro osteogenic differentiation of murine ESCs (mESCs), which completely avoided EB formation. Specifically, mESCs were cultured in HepG2 conditioned medium for 3 days and then directed into osteogenic differentiation for 21 days without prior EB formation. The mineralised bone nodules generated were characterized by Alizarin red S-staining, phenotypic alkaline phosphatase expression, time-course analysis of ALPase activity, the presence of type I collagen and osteopontin, and osteocalcin, cbfa-1/runx-2, and osterix gene expression. Our method of dir...
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enhanced derivation of osteogenic cells from murine embryonic stem cells after treatment with hepg2 conditioned medium and modulation of the Embryoid Body formation period application to skeletal tissue engineering
Tissue Engineering, 2006Co-Authors: Yushik Hwang, Julia M Polak, Wesley L Randle, Robert C Bielby, Athanasios MantalarisAbstract:Despite the considerable progress made in directing embryonic stem cell (ESC) differentiation to therapeutically useful lineages, several issues remain to be resolved before ESCs can be used for cell therapy: 1) increasing the efficiency of specific lineage generation, and 2) developing time- and costeffective culture systems for controlling ESC differentiation. Our study aimed to develop efficient methods to enhance mesodermal differentiation and thereby upregulate osteogenic differentiation of ESCs. Specifically, murine ESCs (mESCs) were cultured in the presence of 50% conditioned medium (CM) from the human hepatocarcinoma cell line HepG2, which resulted in enhanced mesoderm formation during Embryoid Body (EB) formation in the CM-treated mESCs (CM-mESCs). By varying the length of EB culture time, we achieved the selective control and stimulation of osteogenic differentiation and suppression of cardiogenic differentiation. Hence, reducing the EB culture of the CM-mESCs to 1 day resulted in 5–10-fold enha...