Time-resolved fate mapping identifies the intestinal upper crypt zone as an origin of Lgr5+ crypt base columnar cells.

In the prevailing model, Lgr5+ cells are the only intestinal stem cells (ISCs) that sustain homeostatic epithelial regeneration by upward migration of progeny through elusive upper crypt transit-amplifying (TA) intermediates. Here, we identify a proliferative upper crypt population marked by Fgfbp1, in the location of putative TA cells, that is transcriptionally distinct from Lgr5+ cells. Using a kinetic reporter for time-resolved fate mapping and Fgfbp1-CreERT2 lineage tracing, we establish that Fgfbp1+ cells are multi-potent and give rise to Lgr5+ cells, consistent with their ISC function. Fgfbp1+ cells also sustain epithelial regeneration following Lgr5+ cell depletion. We demonstrate that FGFBP1, produced by the upper crypt cells, is an essential factor for crypt proliferation and epithelial homeostasis. Our findings support a model in which tissue regeneration originates from upper crypt Fgfbp1+ cells that generate progeny propagating bi-directionally along the crypt-villus axis and serve as a source of Lgr5+ cells in the crypt base.

Cellular origins and lineage relationships of the intestinal epithelium.

Knowledge of the development and hierarchical organization of tissues is key to understanding how they are perturbed in injury and disease, as well as how they may be therapeutically manipulated to restore homeostasis. The rapidly regenerating intestinal epithelium harbors diverse cell types and their lineage relationships have been studied using numerous approaches, from classical label-retaining and genetic lineage tracing methods to novel transcriptome-based annotations. Here, we describe the developmental trajectories that dictate differentiation and lineage specification in the intestinal epithelium. We focus on the most recent single-cell RNA-sequencing (scRNA-seq)-based strategies for understanding intestinal epithelial cell lineage relationships, underscoring how they have refined our view of the development of this tissue and highlighting their advantages and limitations. We emphasize how these technologies have been applied to understand the dynamics of intestinal epithelial cells in homeostatic and injury-induced regeneration models.

Keratin Profiling by Single-Cell RNA-Sequencing Identifies Human Prostate Stem Cell Lineage Hierarchy and Cancer Stem-Like Cells.

Single prostate stem cells can generate stem and progenitor cells to form prostaspheres in 3D culture. Using a prostasphere-based label retention assay, we recently identified keratin 13 (KRT13)-enriched prostate stem cells at single-cell resolution, distinguishing them from daughter progenitors. Herein, we characterized the epithelial cell lineage hierarchy in prostaspheres using single-cell RNA-seq analysis. Keratin profiling revealed three clusters of label-retaining prostate stem cells; cluster I represents quiescent stem cells (PSCA, CD36, SPINK1, and KRT13/23/80/78/4 enriched), while clusters II and III represent active stem and bipotent progenitor cells (KRT16/17/6 enriched). Gene set enrichment analysis revealed enrichment of stem and cancer-related pathways in cluster I. In non-label-retaining daughter progenitor cells, three clusters were identified; cluster IV represents basal progenitors (KRT5/14/6/16 enriched), while clusters V and VI represent early and late-stage luminal progenitors, respectively (KRT8/18/10 enriched). Furthermore, MetaCore analysis showed enrichment of the "cytoskeleton remodeling-keratin filaments" pathway in cancer stem-like cells from human prostate cancer specimens. Along with common keratins (KRT13/23/80/78/4) in normal stem cells, unique keratins (KRT10/19/6C/16) were enriched in cancer stem-like cells. Clarification of these keratin profiles in human prostate stem cell lineage hierarchy and cancer stem-like cells can facilitate the identification and therapeutic targeting of prostate cancer stem-like cells.

Hematopoiesis and T-cell specification as a model developmental system.

The pathway to generate T cells from hematopoietic stem cells guides progenitors through a succession of fate choices while balancing differentiation progression against proliferation, stage to stage. Many elements of the regulatory system that controls this process are known, but the requirement for multiple, functionally distinct transcription factors needs clarification in terms of gene network architecture. Here, we compare the features of the T-cell specification system with the rule sets underlying two other influential types of gene network models: first, the combinatorial, hierarchical regulatory systems that generate the orderly, synchronized increases in complexity in most invertebrate embryos; second, the dueling 'master regulator' systems that are commonly used to explain bistability in microbial systems and in many fate choices in terminal differentiation. The T-cell specification process shares certain features with each of these prevalent models but differs from both of them in central respects. The T-cell system is highly combinatorial but also highly dose-sensitive in its use of crucial regulatory factors. The roles of these factors are not always T-lineage-specific, but they balance and modulate each other's activities long before any mutually exclusive silencing occurs. T-cell specification may provide a new hybrid model for gene networks in vertebrate developmental systems.

Autocrine amphiregulin signaling sustains castration-resistant Ly6d+ prostate cancer cells.

Mechanisms underlying the development of castration-resistant prostate cancer (CRPC) remain incompletely understood. A recent study by Steiner et al. shows that Ly6d+ prostate tumor cells survive androgen deprivation through an autocrine amphiregulin signaling pathway.

Defining the Emerging Blood System During Development at Single-Cell Resolution.

Developmental hematopoiesis differs from adult and is far less described. In the developing embryo, waves of lineage-restricted blood precede the ultimate emergence of definitive hematopoietic stem cells (dHSCs) capable of maintaining hematopoiesis throughout life. During the last two decades, the advent of single-cell genomics has provided tools to circumvent previously impeding characteristics of embryonic hematopoiesis, such as cell heterogeneity and rare cell states, allowing for definition of lineage trajectories, cellular hierarchies, and cell-type specification. The field has rapidly advanced from microfluidic platforms and targeted gene expression analysis, to high throughput unbiased single-cell transcriptomic profiling, single-cell chromatin analysis, and cell tracing-offering a plethora of tools to resolve important questions within hematopoietic development. Here, we describe how these technologies have been implemented to address a wide range of aspects of embryonic hematopoiesis ranging from the gene regulatory network of dHSC formation via endothelial to hematopoietic transition (EHT) and how EHT can be recapitulated in vitro, to hematopoietic trajectories and cell fate decisions. Together, these studies have important relevance for regenerative medicine and for our understanding of genetic blood disorders and childhood leukemias.

Endogenous lung stem cells for lung regeneration.

INTRODUCTION: Lifelong maintenance of a healthy lung requires resident stem cells to proliferate according to tissue requirements. Once thought to be a quiescent tissue, evolving views of the complex differentiation landscape of lung stem and progenitor cells have broad implications for our understanding of how the lung is maintained, as well as the development of new therapies for promoting endogenous regeneration in lung disease. AREAS COVERED: This review collates a large body of research relating to the hierarchical organization of epithelial stem cells in the adult lung and their role in tissue homeostasis and regeneration after injury. To identify relevant studies, PubMed was queried using one or a combination of the terms 'lung', 'airway', 'alveoli', 'stem cells', 'progenitor', 'repair' and 'regeneration'. EXPERT OPINION: This review discusses how new technologies and injury models have challenged the demarcations between stem and progenitor cell populations.

Studying hematopoiesis using single-cell technologies.

Hematopoiesis is probably the best-understood stem cell differentiation system; hematopoietic stem cell (HSC) transplantation represents the most widely used regenerative therapy. The classical view of lineage hierarchy in hematopoiesis is built on cell type definition system by a group of cell surface markers. However, the traditional model is facing increasing challenges, as many classical cell types are proved to be heterogeneous. Recently, the developments of new technologies allow genome, transcriptome, proteome, and epigenome analysis at the single-cell level. For the first time, we can study hematopoietic system at single-cell resolution on a multi-omic scale. Here, we review recent technical advances in single-cell analysis technology, as well as their current applications. We will also discuss the impact of single-cell technologies on both basic research and clinical application in hematology.

Reconstructing Lineage Hierarchies of Mouse Uterus Epithelial Development Using Single-Cell Analysis.

The endometrial layer comprises luminal and glandular epithelia that both develop from the same simple layer of fetal uterine epithelium. Mechanisms of uterine epithelial progenitor self-renewal and differentiation are unclear. This study aims to systematically analyze the molecular and cellular mechanisms of uterine epithelial development by single-cell analysis. An integrated set of single-cell transcriptomic data of uterine epithelial progenitors and their differentiated progenies is provided. Additionally the unique molecular signatures of these cells, characterized by sequential upregulation of specific epigenetic and metabolic activities, and activation of unique signaling pathways and transcription factors, were also investigated. Finally a unique subpopulation of early progenitor, as well as differentiated luminal and glandular lineages, were identified. A complex cellular hierarchy of uterine epithelial development was thus delineated. Our study therefore systematically decoded molecular markers and a cellular program of uterine epithelial development that sheds light on uterine developmental biology.

Prostate epithelial stem and progenitor cells.

The classic androgen ablation and replacement experiment demonstrates that prostate epithelia possess extensive regenerative capacities and implies the existence of the prostate stem/progenitor cells. These cells may serve as the cells of origin for prostate cancer and their intrinsic property may dictate the clinical behaviors of the resulting diseases. Therefore, detailed characterization of these cells will potentially benefit disease prevention, diagnosis and prognosis. In this review, we describe several major in vitro and in vivo approaches that have been employed in the studies of the prostate stem cell activities, summarize the major progress that has been made during the last two decades regarding the identity of prostate stem/progenitor cells and their niches, and discuss some remaining outstanding questions in the field.

Cancer stem cells in brain tumors and their lineage hierarchy.

Despite recent advances in the development of novel targeted chemotherapies, the prognosis of malignant glioma remains dismal. The chemo-resistance of this tumor is attributed to tumor heterogeneity. To explain this unique chemo- resistance, the concept of cancer stem cells has been evoked. Cancer stem cells, a subpopulation of whole tumor cells, are now regarded as candidate therapeutic targets. Here, the author reviews and discusses the cancer stem cell concept.