Author
David T. Breault
Other affiliations: University of Connecticut Health Center, Harvard University
Bio: David T. Breault is an academic researcher from Boston Children's Hospital. The author has contributed to research in topics: Stem cell & Adrenal cortex. The author has an hindex of 30, co-authored 92 publications receiving 4040 citations. Previous affiliations of David T. Breault include University of Connecticut Health Center & Harvard University.
Papers published on a yearly basis
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TL;DR: Using doxycycline-inducible vectors, it is shown that exogenous factors are required for about 10 days, after which cells enter a self-sustaining pluripotent state and markers are identified that define cell populations prior to and during this transition period.
908 citations
TL;DR: It is demonstrated that a slowly cycling stem cell population exists within the intestine and that mTert+ cells give rise to all differentiated intestinal cell types, persist long term, and contribute to the regenerative response following injury.
Abstract: The intestinal epithelium is maintained by a population of rapidly cycling (Lgr5+) intestinal stem cells (ISCs). It has been postulated, however, that slowly cycling ISCs must also be present in the intestine to protect the genome from accumulating deleterious mutations and to allow for a response to tissue injury. Here, we identify a subpopulation of slowly cycling ISCs marked by mouse telomerase reverse transcriptase (mTert) expression that can give rise to Lgr5+ cells. mTert-expressing cells distribute in a pattern along the crypt–villus axis similar to long-term label-retaining cells (LRCs) and are resistant to tissue injury. Lineage-tracing studies demonstrate that mTert+ cells give rise to all differentiated intestinal cell types, persist long term, and contribute to the regenerative response following injury. Consistent with other highly regenerative tissues, our results demonstrate that a slowly cycling stem cell population exists within the intestine.
491 citations
TL;DR: The Intestine Chip may be useful as a research tool for applications where normal intestinal function is crucial, including studies of metabolism, nutrition, infection, and drug pharmacokinetics, as well as personalized medicine.
Abstract: Here we describe a method for fabricating a primary human Small Intestine-on-a-Chip (Intestine Chip) containing epithelial cells isolated from healthy regions of intestinal biopsies. The primary epithelial cells are expanded as 3D organoids, dissociated, and cultured on a porous membrane within a microfluidic device with human intestinal microvascular endothelium cultured in a parallel microchannel under flow and cyclic deformation. In the Intestine Chip, the epithelium forms villi-like projections lined by polarized epithelial cells that undergo multi-lineage differentiation similar to that of intestinal organoids, however, these cells expose their apical surfaces to an open lumen and interface with endothelium. Transcriptomic analysis also indicates that the Intestine Chip more closely mimics whole human duodenum in vivo when compared to the duodenal organoids used to create the chips. Because fluids flowing through the lumen of the Intestine Chip can be collected continuously, sequential analysis of fluid samples can be used to quantify nutrient digestion, mucus secretion and establishment of intestinal barrier function over a period of multiple days in vitro. The Intestine Chip therefore may be useful as a research tool for applications where normal intestinal function is crucial, including studies of metabolism, nutrition, infection, and drug pharmacokinetics, as well as personalized medicine.
479 citations
TL;DR: The extended coculture of living human intestinal epithelium with stable communities of aerobic and anaerobic human gut microbiota is shown, using a microfluidic intestine-on-a-chip that permits the control and real-time assessment of physiologically relevant oxygen gradients.
Abstract: The diverse bacterial populations that comprise the commensal microbiome of the human intestine play a central role in health and disease. A method that sustains complex microbial communities in direct contact with living human intestinal cells and their overlying mucus layer in vitro would thus enable the investigation of host-microbiome interactions. Here, we show the extended coculture of living human intestinal epithelium with stable communities of aerobic and anaerobic human gut microbiota, using a microfluidic intestine-on-a-chip that permits the control and real-time assessment of physiologically relevant oxygen gradients. When compared to aerobic coculture conditions, the establishment of a transluminal hypoxia gradient in the chip increased intestinal barrier function and sustained a physiologically relevant level of microbial diversity, consisting of over 200 unique operational taxonomic units from 11 different genera and an abundance of obligate anaerobic bacteria, with ratios of Firmicutes and Bacteroidetes similar to those observed in human faeces. The intestine-on-a-chip may serve as a discovery tool for the development of microbiome-related therapeutics, probiotics and nutraceuticals.
428 citations
Columbia University Medical Center1, Stanford University2, Oregon Health & Science University3, City of Hope National Medical Center4, Stowers Institute for Medical Research5, University of North Carolina at Chapel Hill6, University of Oklahoma Health Sciences Center7, University of Pennsylvania8, University of Southern California9, National Institutes of Health10, Boston Children's Hospital11, University of California, Los Angeles12
TL;DR: The data suggest that the EE lineage, including mature EE cells, comprises a reservoir of homeostatic and injury-inducible ISCs, extending the understanding of cellular plasticity and stemness.
264 citations
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TL;DR: The core Wnt/β-catenin signaling pathway is described, how it controls stem cells, and contributes to disease, and strategies for Wnt-based therapies are discussed.
2,663 citations
TL;DR: The generation of induced pluripotent stem cells from patients with a variety of genetic diseases with either Mendelian or complex inheritance are described, offering an unprecedented opportunity to recapitulate both normal and pathologic human tissue formation in vitro, thereby enabling disease investigation and drug development.
2,195 citations
TL;DR: The production of virus-free iPS cells, albeit from embryonic fibroblasts, addresses a critical safety concern for potential use of i PS cells in regenerative medicine.
Abstract: Induced pluripotent stem (iPS) cells have been generated from mouse and human somatic cells by introducing Oct3/4 and Sox2 with either Klf4 and c-Myc or Nanog and Lin28 using retroviruses or lentiviruses. Patient-specific iPS cells could be useful in drug discovery and regenerative medicine. However, viral integration into the host genome increases the risk of tumorigenicity. Here, we report the generation of mouse iPS cells without viral vectors. Repeated transfection of two expression plasmids, one containing the complementary DNAs (cDNAs) of Oct3/4, Sox2, and Klf4 and the other containing the c-Myc cDNA, into mouse embryonic fibroblasts resulted in iPS cells without evidence of plasmid integration, which produced teratomas when transplanted into mice and contributed to adult chimeras. The production of virus-free iPS cells, albeit from embryonic fibroblasts, addresses a critical safety concern for potential use of iPS cells in regenerative medicine.
2,089 citations
TL;DR: It is shown that the individual PB insertions can be removed from established iPS cell lines, providing an invaluable tool for discovery, and the traceless removal of reprogramming factors joined with viral 2A sequences delivered by a single transposon from murine iPS lines is demonstrated.
Abstract: Transgenic expression of just four defined transcription factors (c-Myc, Klf4, Oct4 and Sox2) is sufficient to reprogram somatic cells to a pluripotent state. The resulting induced pluripotent stem (iPS) cells resemble embryonic stem cells in their properties and potential to differentiate into a spectrum of adult cell types. Current reprogramming strategies involve retroviral, lentiviral, adenoviral and plasmid transfection to deliver reprogramming factor transgenes. Although the latter two methods are transient and minimize the potential for insertion mutagenesis, they are currently limited by diminished reprogramming efficiencies. piggyBac (PB) transposition is host-factor independent, and has recently been demonstrated to be functional in various human and mouse cell lines. The PB transposon/transposase system requires only the inverted terminal repeats flanking a transgene and transient expression of the transposase enzyme to catalyse insertion or excision events. Here we demonstrate successful and efficient reprogramming of murine and human embryonic fibroblasts using doxycycline-inducible transcription factors delivered by PB transposition. Stable iPS cells thus generated express characteristic pluripotency markers and succeed in a series of rigorous differentiation assays. By taking advantage of the natural propensity of the PB system for seamless excision, we show that the individual PB insertions can be removed from established iPS cell lines, providing an invaluable tool for discovery. In addition, we have demonstrated the traceless removal of reprogramming factors joined with viral 2A sequences delivered by a single transposon from murine iPS lines. We anticipate that the unique properties of this virus-independent simplification of iPS cell production will accelerate this field further towards full exploration of the reprogramming process and future cell-based therapies.
1,884 citations
TL;DR: This work generated mouse induced pluripotent stem cells from fibroblasts and liver cells by using nonintegrating adenoviruses transiently expressing Oct4, Sox2, Klf4, and c-Myc, providing strong evidence that insertional mutagenesis is not required for in vitro reprogramming.
Abstract: Pluripotent stem cells have been generated from mouse and human somatic cells by viral expression of the transcription factors Oct4, Sox2, Klf4, and c-Myc. A major limitation of this technology is the use of potentially harmful genome-integrating viruses. We generated mouse induced pluripotent stem (iPS) cells from fibroblasts and liver cells by using nonintegrating adenoviruses transiently expressing Oct4, Sox2, Klf4, and c-Myc. These adenoviral iPS (adeno-iPS) cells show DNA demethylation characteristic of reprogrammed cells, express endogenous pluripotency genes, form teratomas, and contribute to multiple tissues, including the germ line, in chimeric mice. Our results provide strong evidence that insertional mutagenesis is not required for in vitro reprogramming. Adenoviral reprogramming may provide an improved method for generating and studying patient-specific stem cells and for comparing embryonic stem cells and iPS cells.
1,714 citations