Progenitor Cell Types Research Articles

Dynamics of the Bone Marrow Microenvironment during Leukemic Progression Revealed By Codex Hyper-Parameter Tissue Imaging

IntroductionThe bone marrow (BM) microenvironment consists of various cell types such as mesenchymal stromal cells, endothelial cells, osteoblastic cells and multiple immune cell types including mature myeloid cells and lymphocytes. Recent studies have shown that leukemias can create and maintain a leukemia-supporting BM microenvironment, and vice versa, a dysfunctional BM microenvironment can contribute to leukemia development and progression. Moreover, in tumors the microenvironment is often immunosuppressive and restrains effective anti-tumoral immune responses by adaptive and innate immunity. A better understanding of the precise localization of microenvironmental and immune cell types in intact tissue, and how they physically interact with each other and with tumor cells, will improve our understanding of the mechanisms by which cancer reprograms its microenvironment and may form the basis for novel immunotherapies.MethodsCO-Detection by antibody indEXing (CODEX) is a multiplex fluorescence microscopy platform based on DNA-conjugated antibodies that allows the analysis of 50+ markers in a single tissue section. After staining with an antibody cocktail, tissues are imaged in a multi-cycle reaction using a microfluidics system and a fluorescence microscope with a computer automated X/Y/Z stage. DNA-conjugated antibodies are rendered visible using complementary fluorescent DNA probes, followed by imaging, probe stripping, washing and re-rendering. This process is repeated until all the antibodies present in the initial cocktail have been rendered and imaged. Here, we used CODEX to analyze intact BM at the single-cell level (~200nm resolution) during leukemic progression. Chronic myeloid leukemia (CML)-like disease was induced in non-irradiated mice using BCR-ABL1-GFP retrovirus. Tissue sections of femoral bones harvested at different time points after leukemia onset were stained using a 50+ marker CODEX antibody panel to simultaneously identify hematopoietic and leukemic stem and progenitor cells, multiple BM microenvironmental cell types, myeloid and lymphoid cells as well as functional markers.ResultsWe have built an integrated computational pipeline for the analysis of high-dimensional CODEX data that enables the identification and characterization of BM cell types as well as their spatial organization in situ. Raw images were concatenated and aligned using Hoechst nuclear stain as a reference, followed by deconvolution, segmentation, marker expression quantification and spatial compensation. Exported data were clustered in an unsupervised manner using VorteX algorithm, which identified 28 distinct cellular clusters based on marker expression values. All major BM compartments including stromal (vascular, pericytes, osteoblastic), lymphoid (T and B cell subsets), myeloid (megakaryocytes, macrophages, dendritic cells, granulocytes) and progenitor cell types, as well as leukemic cells, were represented. During leukemic progression, the BM microenvironment was dramatically rearranged. Besides the expected growth of the leukemic clone, we observed a massive increase in vascular and osteoblastic cell types, whereas immune cell clusters were significantly reduced. Interestingly, CD71, the transferrin receptor, was strongly up-regulated on tumor cells in advanced leukemia, indicating towards a role for iron metabolism in malignant progression. Furthermore, hierarchical clustering of tissue regions based on cellular composition using X/Y/Z positional information pointed towards the emergence of specific cell-cell interaction modules that developed during leukemic progression, including mutual attraction between B cells and central arterioles.ConclusionsHigh-dimensional imaging of the BM microenvironment by CODEX allows studying the abundance and distribution of cellular elements that are often underestimated or missed by traditional flow cytometry, such as stromal cells, vasculature and megakaryocytes. Importantly, CODEX identifies single cells in their tissue context during leukemic progression and facilitates the discovery of novel cell-cell interactions and cell types as well as unexpected marker constellations. DisclosuresSamusik:Akoya Biosciences: Consultancy, Equity Ownership, Honoraria, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Nolan:Akoya Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Goltsev:Akoya Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.

Human-specific ARHGAP11B induces hallmarks of neocortical expansion in developing ferret neocortex.

The evolutionary increase in size and complexity of the primate neocortex is thought to underlie the higher cognitive abilities of humans. ARHGAP11B is a human-specific gene that, based on its expression pattern in fetal human neocortex and progenitor effects in embryonic mouse neocortex, has been proposed to have a key function in the evolutionary expansion of the neocortex. Here, we study the effects of ARHGAP11B expression in the developing neocortex of the gyrencephalic ferret. In contrast to its effects in mouse, ARHGAP11B markedly increases proliferative basal radial glia, a progenitor cell type thought to be instrumental for neocortical expansion, and results in extension of the neurogenic period and an increase in upper-layer neurons. Consequently, the postnatal ferret neocortex exhibits increased neuron density in the upper cortical layers and expands in both the radial and tangential dimensions. Thus, human-specific ARHGAP11B can elicit hallmarks of neocortical expansion in the developing ferret neocortex.

Metabolomic Analyses Reveal Extensive Progenitor Cell Deficiencies in a Mouse Model of Duchenne Muscular Dystrophy.

Duchenne muscular dystrophy (DMD) is a musculoskeletal disorder that causes severe morbidity and reduced lifespan. Individuals with DMD have an X-linked mutation that impairs their ability to produce functional dystrophin protein in muscle. No cure exists for this disease and the few therapies that are available do not dramatically delay disease progression. Thus, there is a need to better understand the mechanisms underlying DMD which may ultimately lead to improved treatment options. The muscular dystrophy (MDX) mouse model is frequently used to explore DMD disease traits. Though some studies of metabolism in dystrophic mice exist, few have characterized metabolic profiles of supporting cells in the diseased environment. Using nontargeted metabolomics we characterized metabolic alterations in muscle satellite cells (SCs) and serum of MDX mice. Additionally, live-cell imaging revealed MDX-derived adipose progenitor cell (APC) defects. Finally, metabolomic studies revealed a striking elevation of acylcarnitines in MDX APCs, which we show can inhibit APC proliferation. Together, these studies highlight widespread metabolic alterations in multiple progenitor cell types and serum from MDX mice and implicate dystrophy-associated metabolite imbalances in APCs as a potential contributor to adipose tissue disequilibrium in DMD.

Isolation and Characterization of Multipotent CD24+ Cells From the Renal Papilla of Swine.

Over 100,000 patients in the United States are currently waiting for a kidney transplant. With just over 10,000 cadaveric kidneys transplanted annually, it is of the utmost importance to optimize kidney viability upon transplantation. One exciting avenue may be xenotransplantation, which has rejuvenated interest after advanced gene editing techniques have been successfully used in swine. Simultaneously, acute kidney injury (AKI) is associated with high morbidity and mortality and currently lacks effective treatment. Animal models have been used extensively to address both of these issues, with recent emphasis on renal progenitor cells (RPCs). Due to anatomical similarities to humans we aimed to examine progenitor cells from the renal papillae of swine kidneys. To do this, RPCs were dissected from the renal papillae of healthy swine. Cell surface marker expression, proliferation, and differentiation of the RPCs were tested in vitro. Additionally, a mixed lymphocyte reaction was performed to examine immunomodulatory properties. RPCs displayed spindle shaped morphology with limited self-renewing capacity. Isolated RPCs were positive for CD24 and CD133 at early passages, but lost expression with subsequent passaging. Similarly, RPCs displayed myogenic, osteogenic, and adipogenic differentiation capacities at passage 2, but largely lost this by passage 6. Lastly, direct contact of RPCs with human lymphocytes increased release of IL6 and IL8. Taken together, RPCs from the papilla of porcine kidneys display transient stem cell properties that are lost with passaging, and either represent multiple types of progenitor cells, or a multipotent progenitor population. In instances of ischemic insult, augmentation of/with RPCs may potentiate regenerative properties of the kidney. While the use of swine for transplantation and ischemia studies confers obvious advantages, the populations of different progenitor cell populations within pig kidneys warrants further investigation. Ultimately, while gene editing techniques enhance the potential for xenotransplantation of organs or cells, the ultimate success of this strategy may be determined by the (dis)similarities of RPCs from different species.

Generation of Cardiomyocytes From Vascular Adventitia-Resident Stem Cells.

Regeneration of lost cardiomyocytes is a fundamental unresolved problem leading to heart failure. Despite several strategies developed from intensive studies performed in the past decades, endogenous regeneration of heart tissue is still limited and presents a big challenge that needs to be overcome to serve as a successful therapeutic option for myocardial infarction. One of the essential prerequisites for cardiac regeneration is the identification of endogenous cardiomyocyte progenitors and their niche that can be targeted by new therapeutic approaches. In this context, we hypothesized that the vascular wall, which was shown to harbor different types of stem and progenitor cells, might serve as a source for cardiac progenitors. We describe generation of spontaneously beating mouse aortic wall-derived cardiomyocytes without any genetic manipulation. Using aortic wall-derived cells (AoCs) of WT (wild type), αMHC (α-myosin heavy chain), and Flk1 (fetal liver kinase 1)-reporter mice and magnetic bead-associated cell sorting sorting of Flk1+ AoCs from GFP (green fluorescent protein) mice, we identified Flk1+CD (cluster of differentiation) 34+Sca-1 (stem cell antigen-1)-CD44- AoCs as the population that gives rise to aortic wall-derived cardiomyocytes. This AoC subpopulation delivered also endothelial cells and macrophages with a particular accumulation within the aortic wall-derived cardiomyocyte containing colonies. In vivo, cardiomyocyte differentiation capacity was studied by implantation of fluorescently labeled AoCs into chick embryonic heart. These cells acquired cardiomyocyte-like phenotype as shown by αSRA (α-sarcomeric actinin) expression. Furthermore, coronary adventitial Flk1+ and CD34+ cells proliferated, migrated into the myocardium after mouse myocardial infarction, and expressed Isl-1+ (insulin gene enhancer protein-1) indicative of cardiovascular progenitor potential. Our data suggest Flk1+CD34+ vascular adventitia-resident stem cells, including those of coronary adventitia, as a novel endogenous source for generating cardiomyocytes. This process is essentially supported by endothelial cells and macrophages. In summary, the therapeutic manipulation of coronary adventitia-resident cardiac stem and their supportive cells may open new avenues for promoting cardiac regeneration and repair after myocardial infarction and for preventing heart failure.

Cortical progenitor biology: key features mediating proliferation versus differentiation.

The cerebral cortex is a highly organized structure whose development depends on diverse progenitor cell types, namely apical radial glia, intermediate progenitors, and basal radial glia cells, which are responsible for the production of the correct neuronal output. In recent years, these progenitor cell types have been deeply studied, particularly basal radial glia and their role in cortical expansion and gyrification. We review here a broad series of factors that regulate progenitor behavior and daughter cell fate. We first describe the different neuronal progenitor types, emphasizing the differences between lissencephalic and gyrencephalic species. We then review key factors shown to influence progenitor proliferation versus differentiation, discussing their roles in progenitor dynamics, neuronal production, and potentially brain size and complexity. Although spindle orientation has been considered a critical factor for mode of division and daughter cell output, we discuss other features that are emerging as crucial for these processes such as organelle and cell cycle dynamics. Additionally, we highlight the importance of adhesion molecules and the polarity complex for correct cortical development. Finally, we briefly discuss studies assessing progenitor multipotency and its possible contribution to the production of specific neuronal populations. This review hence summarizes recent aspects of cortical progenitor cell biology, and pinpoints emerging features critical for their behavior.

Molecular signature of progenitor cells isolated from young and adult human hearts

The loss of endogenous cardiac regenerative capacity within the first week of postnatal life has intensified clinical trials to induce cardiac regeneration in the adult mammalian heart using different progenitor cell types. We hypothesized that donor age-related phenotypic and functional characteristics of cardiac progenitor cells (CPC) account for mixed results of cell-based cardiac repair. We compared expression profiles and cell turnover rates of human heart-derived c-kitpos progenitors (c-kitpos CPC) and cardiosphere-derived cells (CDC) from young and adult donor origin and studied their in vitro angiogenic and cardiac differentiation potential, which can be relevant for cardiac repair. We report that 3-dimensional CDC expansion recapitulates a conducive environment for growth factor and cytokine release from adult donor cells (aCDC) that optimally supports vascular tube formation and vessel sprouting. Transdifferentiation capacity of c-kitpos CPCs and CDCs towards cardiomyocyte-like cells was modest, however, most notable in young c-kitpos cells and adult CDCs. Progenitors isolated with different methods thus show cell- and donor-specific characteristics that may account for variable contributions in functional myocardial recovery.

A Method for Isolating and Characterizing Mesenchymal Stromal Cell-derived Extracellular Vesicles.

The unit describes protocols for isolating and characterizing extracellular vesicles (EVs) derived from human adipose tissue-derived mesenchymal stromal cells (MSCs). EVs are a mixed population of membrane-surrounded structures with overlapping composition and size. Advances made in recent years have led to a better understanding of the biological role of EVs. In particular, they can be considered key factors responsible for MSC-paracrine activity, mediating their anti-inflammatory effects towards innate immune cells, such as macrophages. The topics comprise description of the MSC-conditioned medium containing vesicles preparation, EV isolation, and characterization mainly by specifically set up flow cytometry and electron microscopy approaches, and in vitro methodologies involved in testing the EV anti-inflammatory capacity. The procedures described here can be easily reproduced and can be employed regardless of the type of progenitor cells used to secrete EVs. © 2018 by John Wiley & Sons, Inc.

PD33-12 IN VIVO EVALUATION OF FUNCTIONALIZED MUSCLE SCAFFOLDS FOR RECONSTRUCTION

INTRODUCTION: Every tissue in the body contains some type of stem or progenitor cells. These cells are believed to be part of underlying regenerative machinery that is responsible for daily maintenance and repair of injured tissue. In this study, we aimed to regenerate muscle functionality following volumetric muscle loss through the use of a target-specific scaffolding system. The objectives of this study were to evaluate the functionalized decellularized tissue scaffolds on the host muscle cell migration and muscle function recovery in vivo.

The transcription factor Dach1 is essential for podocyte function.

Dedifferentiation and loss of podocytes are the major cause of chronic kidney disease. Dach1, a transcription factor that is essential for cell fate, was found in genome‐wide association studies to be associated with the glomerular filtration rate. We found that podocytes express high levels of Dach1 in vivo and to a much lower extent in vitro. Parietal epithelial cells (PECs) that are still under debate to be a type of progenitor cell for podocytes expressed Dach1 only at low levels. The transfection of PECs with a plasmid encoding for Dach1 induced the expression of synaptopodin, a podocyte‐specific protein, demonstrated by immunocytochemistry and Western blot. Furthermore, synaptopodin was located along actin fibres in a punctate pattern in Dach1‐expressing PECs comparable with differentiated podocytes. Moreover, dedifferentiating podocytes of isolated glomeruli showed a significant reduction in the expression of Dach1 together with synaptopodin after 9 days in cell culture. To study the role of Dach1 in vivo, we used the zebrafish larva as an animal model. Knockdown of the zebrafish ortholog Dachd by morpholino injection into fertilized eggs resulted in a severe renal phenotype. The glomeruli of the zebrafish larvae showed morphological changes of the glomerulus accompanied by down‐regulation of nephrin and leakage of the filtration barrier. Interestingly, glomeruli of biopsies from patients suffering from diabetic nephropathy showed also a significant reduction of Dach1 and synaptopodin in contrast to control biopsies. Taken together, Dach1 is a transcription factor that is important for podocyte differentiation and proper kidney function.

A Pdgf-cCreERT2 knock-in mouse model for tracing PDGF-C cell lineages during development.

PDGF-C, a member of the platelet-derived growth factor (PDGF) family, plays important roles in the development of craniofacial structures, the neural system, the vascular system, and tumors. PDGF-C could also be required for the regulation of certain types of stem or progenitor cells as suggested by its expression in the regions where these cells are located. To further characterize the role of PDGF-C in development, we generated a Pdgf-cCreERT2 mouse strain, in which a tamoxifen-inducible Cre (CreERT2) cDNA was specifically targeted into the Pdgf-c genomic locus and controlled by the endogenous Pdgf-c regulatory elements. We also showed that Cre activity in this mouse strain could be specifically induced by tamoxifen, which allowed the fate of PDGF-C-expressing cells to be traced at various stages of development. Using this model system, we demonstrated for the first time that PDGF-C-expressing cells could be multipotent, generating multiple cell lineages required for the formation of the cerebellum. Therefore, the Pdgf-cCreERT2 mouse strain generated in this study will be a valuable transgenic tool for exploring the function of PDGF-C in development and stem cell biology.

How Cells Fold the Cerebral Cortex.

Folding of the cerebral cortex is as highly intriguing as poorly understood. At first sight, this may appear as simple tissue crumpling inside an excessively small cranium, but the process is clearly much more complex and developmentally predetermined. Whereas theoretical modeling supports a critical role for biomechanics, experimental evidence demonstrates the fundamental role of specific progenitor cell types, cellular processes, and genetic programs on cortical folding.Dual Perspectives Companion Paper: How Forces Fold the Cerebral Cortex, by Christopher D. Kroenke and Philip V. Bayly.

Conserved properties of dentate gyrus neurogenesis across postnatal development revealed by single-cell RNA sequencing.

The dentate gyrus of the hippocampus is a brain region in which neurogenesis persists into adulthood; however, the relationship between developmental and adult dentate gyrus neurogenesis has not been examined in detail. Here we used single-cell RNA sequencing to reveal the molecular dynamics and diversity of dentate gyrus cell types in perinatal, juvenile, and adult mice. We found distinct quiescent and proliferating progenitor cell types, linked by transient intermediate states to neuroblast stages and fully mature granule cells. We observed shifts in the molecular identity of quiescent and proliferating radial glia and granule cells during the postnatal period that were then maintained through adult stages. In contrast, intermediate progenitor cells, neuroblasts, and immature granule cells were nearly indistinguishable at all ages. These findings demonstrate the fundamental similarity of postnatal and adult neurogenesis in the hippocampus and pinpoint the early postnatal transformation of radial glia from embryonic progenitors to adult quiescent stem cells.

Engraftment of Reprogramed Human Bile Duct Cells Transiently Rescues Diabetes in Mice

Differentiation of adult stem/progenitor cells into functional beta cells to provide a successful autologous cell therapy for Type 1 diabetes patients has not yet been achieved. Progenitor cells in the adult pancreas or liver have been considered potential sources of endocrine beta cells based on their ability to repopulate the organ following injury. Here we describe the isolation and reprogramming of a lineage-committed progenitor population of cells in the human bile duct towards a beta cell fate. These cells, which possess SOX9 as a marker, were able to manifest beta cell characteristics upon ectopic expression of pancreatic transcription factors. The beta cells derived from SOX9+ progenitor cells were also able to ameliorate high blood glucose in diabetic mice. The insulin+ islet-like clusters which developed from this progenitor cell type demonstrate the potential of this approach to generate functional, autologous beta cells.

Invited commentary

Invited commentary

Microparticles Carrying Peroxisome Proliferator-Activated Receptor Alpha Restore the Reduced Differentiation and Functionality of Bone Marrow-Derived Cells Induced by High-Fat Diet.

Metabolic pathologies such as diabetes and obesity are associated with decreased level of circulating and bone marrow (BM)‐derived endothelial progenitor cells (EPCs). It is known that activation of peroxisome proliferator‐activated receptor alpha (PPARα) may stimulate cell differentiation. In addition, microparticles (MPs), small membrane vesicles produced by activated and apoptotic cells, are able to reprogram EPCs. Here, we evaluated the role of MPs carrying PPARα on both phenotype and function of progenitor cells from mice fed with a high‐fat diet (HFD). HFD reduced circulating EPCs and, after 7 days of culture, BM‐derived EPCs and monocytic progenitor cells from HFD‐fed mice displayed impaired differentiation. At the same time, we show that MPs bearing PPARα, MPsPPARα+/+, increased the differentiation of EPCs and monocytic progenitors from HFD‐fed mice, whereas MPs taken from PPARα knockout mice (MPsPPARα–/–) had no effect on the differentiation of all types of progenitor cells. Furthermore, MPsPPARα+/+ increased the ability of progenitor cells to promote in vivo angiogenesis in mice fed with HFD. The in vitro and in vivo effects of MPsPPARα+/+ were abolished in presence of MK886, a specific inhibitor of PPARα. Collectively, these data highlight the ability of MPs carrying PPARα to restore the failed differentiation and functionality of BM‐derived cells induced by HFD. stem cells translational medicine 2018;7:135–145

Postimplantation Mga expression and embryonic lethality of two gene-trap alleles

BackgroundThe dual-specificity T-box/basic helix-loop-helix leucine zipper transcription factor MGA is part of the MAX-interacting network of proteins. In the mouse, MGA is necessary for the survival of the pluripotent epiblast cells of the peri-implantation embryo and a null, gene-trap allele MgaGt results in embryonic lethality shortly after implantation. We have used this allele to document expression of Mga in postimplantation embryos and also investigated a second, hypomorphic gene-trap allele, MgaInv. ResultsCompound heterozygotes, MgaGt/MgaInv, die prior to midgestation. The extraembryonic portion of the embryos appears to develop relatively normally while the embryonic portion, including the pluripotent cells of the epiblast, is severely retarded by E7.5. Mga expression is initially limited to the pluripotent inner cell mass of the blastocyst and epiblast, but during organogenesis it is widely expressed, notably in the central nervous system and sensory organs, reproductive and excretory systems, heart, somites and limbs. ConclusionsWidespread yet specific areas of expression of Mga during organogenesis raise the possibility that the transcription factor may play roles in controlling proliferation and potency in the progenitor cell populations of different organ systems. Documentation of these patterns sets the stage for the investigation of specific progenitor cell types.

Plasma stem cell factor levels are associated with risk of cardiovascular disease and death.

Stem cell factor (SCF) is a key growth factor for several types of stem and progenitor cells. There is experimental evidence that such cells are of importance for maintaining the integrity of the cardiovascular system. We investigated the association between circulating levels of SCF and risk for development of cardiovascular events and death. SCF was analysed by the proximity extension assay technique in plasma from 4742 subjects participating in the Malmö Diet and Cancer Study. Cardiovascular events and death were monitored through national registers with a mean follow-up time of 19.2 years. Subjects with high baseline levels of SCF had lower cardiovascular (n = 340) and all-cause mortality (n = 1159) as well as a lower risk of heart failure (n = 177), stroke (n = 318) and myocardial infarction (n = 452). Smoking, diabetes and high alcohol consumption were associated with lower levels of SCF. Single nucleotide polymorphisms in the gene region encoding PDX1 C-terminal inhibiting factor 1 (PCIF1) and matrix metalloproteinase-9 were associated with plasma SCF levels. The highest SCF quartile remained independently associated with a lower risk of a lower risk of cardiovascular [hazard ratio and 95% confidence interval 0.59 (0.43-0.81)] and all-cause mortality [0.68 (0.57-0.81)], heart failure [0.50 (0.31-0.80)] and stroke [0.66 (0.47-0.92)], but not with MI [0.96 (0.72-1.27)] as compared with the lowest quartile when adjusting for traditional cardiovascular risk factors in Cox proportional hazard regression models. This prospective population-based study demonstrates that subjects with high levels of SCF have a lower risk of cardiovascular events and death. The findings provide clinical support for a protective role of SCF in maintaining cardiovascular integrity.

Nestin-expressing cell types in the temporal lobe and hippocampus: Morphology, differentiation, and proliferative capacity.

Nestin is expressed in immature neuroepithelial and progenitor cell types and transiently upregulated in proliferative neuroglial cells responding to acute brain injury, including following seizures. In 36 temporal lobe (TLobe) specimens from patients with TLobe epilepsy (age range 8–60 years) we studied the number, distribution and morphology of nestin‐expressing cells (NEC) in the pes, hippocampus body, parahippocampal gyrus, amygdala, temporal cortex and pole compared with post mortem control tissues from 26 cases (age range 12 gestational weeks to 76 years). The proliferative fraction of NEC was evaluated in selected regions, including recognized niches, using MCM2. Their differentiation was explored with neuronal (DCX, mushashi, βIII tubulin, NeuN) and glial (GFAP, GFAPdelta, glutamine synthetase, aquaporin4, EAAT1) markers, both in sections or following culture. Findings were correlated with clinical parameters. A stereotypical pattern in the distribution and morphologies of NEC was observed, reminiscent of patterns in the developing brain, with increased densities in epilepsy than adult controls (p < .001). Findings included MCM2‐positive radial glial‐like cells in the periventricular white matter and rows of NEC in the hippocampal fimbria and sulcus. Nestin cells represented 29% of the hippocampal proliferative fraction in epilepsy cases; 20% co‐expressed βIII tubulin in culture compared with 28% with GFAP. Significant correlations were noted between age at surgery, memory deficits and nestin populations. TLobe NEC with ongoing proliferative capacity likely represent vestiges of developmental migratory streams and resident reactive cell populations of potential relevance to hippocampal epileptogenesis, TLobe pathology, and co‐morbidities, including memory decline.

Reconstructing blood stem cell regulatory network models from single-cell molecular profiles

Adult blood contains a mixture of mature cell types, each with specialized functions. Single hematopoietic stem cells (HSCs) have been functionally shown to generate all mature cell types for the lifetime of the organism. Differentiation of HSCs toward alternative lineages must be balanced at the population level by the fate decisions made by individual cells. Transcription factors play a key role in regulating these decisions and operate within organized regulatory programs that can be modeled as transcriptional regulatory networks. As dysregulation of single HSC fate decisions is linked to fatal malignancies such as leukemia, it is important to understand how these decisions are controlled on a cell-by-cell basis. Here we developed and applied a network inference method, exploiting the ability to infer dynamic information from single-cell snapshot expression data based on expression profiles of 48 genes in 2,167 blood stem and progenitor cells. This approach allowed us to infer transcriptional regulatory network models that recapitulated differentiation of HSCs into progenitor cell types, focusing on trajectories toward megakaryocyte–erythrocyte progenitors and lymphoid-primed multipotent progenitors. By comparing these two models, we identified and subsequently experimentally validated a difference in the regulation of nuclear factor, erythroid 2 (Nfe2) and core-binding factor, runt domain, alpha subunit 2, translocated to, 3 homolog (Cbfa2t3h) by the transcription factor Gata2. Our approach confirms known aspects of hematopoiesis, provides hypotheses about regulation of HSC differentiation, and is widely applicable to other hierarchical biological systems to uncover regulatory relationships.