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Endothelium dysfunction is a central problem for early rejection due to the host alloimmune response and the late status of arteriosclerosis in heart transplantation. However, reliable pieces of evidence are still limited concerning the source of the regenerated endothelium within the transplanted heart. We analyzed single-cell RNA sequencing data and constructed an inducible lineage tracing mouse, combined heart transplantation with bone marrow transplantation and a parabiosis model, cellular components, and endothelial cell populations in cardiac graft lesions. Our single-cell RNA sequencing analysis of a transplanted heart allowed for the establishment of an endothelial cell atlas with a heterogeneous population, including arterial, venous, capillary, and lymphatic endothelial cells. Along with genetic cell lineage tracing, we demonstrated that the donor cells were mostly replaced by recipient cells in the cardiac allograft, up to 83.29% 2 weeks after transplantation. Furthermore, recipient nonbone marrow CD34+ endothelial progenitors contributed significantly to extracellular matrix organization and immune regulation, with higher apoptotic ability in the transplanted hearts. Mechanistically, peripheral blood-derived human endothelial progenitor cells differentiate into endocardial cells via Vascular endothelial growth factor receptor-mediated pathways. Host circulating CD34+ endothelial progenitors could repair the damaged donor endothelium presumably through CCL3-CCR5 chemotaxis. Partial depletion of host CD34+ cells resulted in delayed endothelial regeneration. We created an annotated fate map of endothelial cells in cardiac allografts, indicating how recipient CD34+ cells could replace the donor endothelium via chemokine CCL3-CCR5 interactions. The mechanisms we discovered could have a potential therapeutic effect on the long-term outcomes of heart transplantation. [Display omitted] [ABSTRACT FROM AUTHOR]

Mouse models of lineage tracing have helped to describe the important subpopulations of hepatocytes responsible for liver regeneration. However, conflicting results have been obtained from different models. Herein, we aimed to reconcile these conflicting reports by repeating a key lineage-tracing study from pericentral hepatocytes and characterising this Axin2CreERT2 model in detail. We performed detailed characterisation of the labelled population in the Axin2CreERT2 model. We lineage traced this cell population, quantifying the labelled population over 1 year and performed in-depth phenotypic comparisons, including transcriptomics, metabolomics and analysis of proteins through immunohistochemistry, of Axin2CreERT2 mice to WT counterparts. We found that after careful definition of a baseline population, there are marked differences in labelling between male and female mice. Upon induced lineage tracing there was no expansion of the labelled hepatocyte population in Axin2CreERT2 mice. We found substantial evidence of disrupted homeostasis in Axin2CreERT2 mice. Offspring are born with sub-Mendelian ratios and adult mice have perturbations of hepatic Wnt/β-catenin signalling and related metabolomic disturbance. We find no evidence of predominant expansion of the pericentral hepatocyte population during liver homeostatic regeneration. Our data highlight the importance of detailed preclinical model characterisation and the pitfalls which may occur when comparing across sexes and backgrounds of mice and the effects of genetic insertion into native loci. Understanding the source of cells which regenerate the liver is crucial to harness their potential to regrow injured livers. Herein, we show that cells which were previously thought to repopulate the liver play only a limited role in physiological regeneration. Our data helps to reconcile differing conclusions drawn from results from a number of prior studies and highlights methodological challenges which are relevant to preclinical models more generally. [Display omitted] • Pericentral hepatocytes do not play a dominant role in physiological hepatocyte regeneration. • The Wnt pathway is suppressed by Axin2 haploinsufficiency. • Mouse genetics and environmental conditions should be rigorously controlled in preclinical models. [ABSTRACT FROM AUTHOR]

The Drosophila hindgut is a classical model to study organogenesis. The adult hindgut originates from the precursor cells in the larval hindgut. However, the territory of these cells has still not been well determined. A ring of wingless (wg)-expressing cells lies at the anterior zone of both the larval and adult hindgut. The larval Wg ring was thought as a portion of precursor of the adult hindgut. By applying a cell lineage tracing tool (G-TRACE), we demonstrate that larval wg -expressing cells have no cell lineage contribution to the adult hindgut. Additionally, adult Wg ring cells do not divide and move posteriorly to replenish the hindgut tissue. Instead, we determine that the precursors of the adult pylorus and ileum are situated in the cubitus interruptus (ci)-expressing cells in the anterior zone, and deduce that the precursor stem cells of the adult rectum locate in the trunk region of the larval pylorus including hedgehog (hh)-expressing cells. Together, this research advances our understanding of cell lineage origins and the development of the Drosophila hindgut. [Display omitted] • Adult Wg ring cells do not divide and move posteriorly to replenish the hindgut tissue. • Larval Wg ring in the anterior zone of hindgut was not the precursors of adult hindgut. • The progenitors of the adult pylorus and ileum were situated in the ci expressing cells in the larval anterior zone. • The progenitors of the adult rectum were located in the trunk region of pylorus including hh expressing cells. [ABSTRACT FROM AUTHOR]

Genetic technology using site-specific recombinases, such as the Cre-loxP system, has been widely employed for labeling specific cell populations and for studying their functions in vivo. To enhance the precision of cell lineage tracing and functional study, a similar site-specific recombinase system termed Drerox has been recently used in combination with Cre-loxP. To enable more specific cell lineage tracing and ablation through dual recombinase activity, we generated two mouse lines that render Dre- or Dre+Cre-mediated recombination to excise a stop codon sequence that prevents the expression of diphtheria toxin receptor (DTR) knocked into the ubiquitously expressed and safe Rosa26 locus. Using different Dre- and Cre-expressing mouse lines, we showed that the surrogate gene reporters tdTomato and DTR were simultaneously expressed in target cells and in their descendants, and we observed efficient ablation of tdTomato+ cells after diphtheria toxin administration. These mouse lines were used to simultaneously trace and deplete the target cells of interest through the inducible expression of a reporter and DTR using dual Cre and Dre recombinases, allowing a more precise and efficient study of the role of specific cell subsets within a heterogeneous population in pathophysiological conditions in vivo. [ABSTRACT FROM AUTHOR]

Precisely deciphering the cellular plasticity in vivo is essential in understanding many key biological processes. Site-specific recombinases are genetic tools used for in vivo lineage tracing and gene manipulation. Conventional Cre- loxP , Dre- rox , and Flp- frt technologies form the orthogonal recombination systems that can also be used in combination to increase the precision. As such, more than one marker gene can be targeted for lineage tracing, studying cellular heterogeneity, recording cellular activities, or even genome editing. Their combinatory use has recently resolved some controversies in defining cellular fate plasticity. Focusing on cell fate studies, we introduce the design principles of orthogonal recombinases-based strategies, describe some working examples in resolving cell fate-related controversies, and discuss some of their technical strengths and limits. Orthogonal recombinase systems are superior to conventional single-recombinase for studying in vivo cell fate, as they employ multiple markers to increase the resolution and precision of cell fate mapping results. Orthogonal recombinase systems enable genetic labeling of the intersect or subtractive parts of multiple cell populations, which helps clarify some controversial issues in cell origins and fate plasticity. Transitional transcriptional activation could be genetically recorded by orthogonal recombinase systems that could continuously capture some brief yet biologically important cellular events or states. Orthogonal recombinase systems have broad applications in cell lineage tracing and gene manipulation in fields of developmental biology, neuroscience, cardiovascular biology, oncology, and regenerative medicine. [ABSTRACT FROM AUTHOR]

During adaptive immunity, B cells differentiate either into memory B cells or plasma cells and produce antibodies against foreign antigens to fight infection. Additionally, they behave as antigen-presenting cells and participate in T cell activation during cellular immune responses. However, their functional dysregulation can result in various autoimmune diseases and cancers. With significant breakthroughs in single cell technologies, assessing individual B cell genomics, transcriptomics, and proteomics can give deeper insights into mammalian B cell development, differentiation, antibody repertoire, and responses under conditions of homeostasis, infection, and aberrations during disease. In this review, we discuss the adoption of single cell approaches to identify different B cell gene signatures and biomarkers in normal and diseased tissues, and subsequent benefits for future therapeutic discoveries. Single cell techniques are used to study mammalian B cell signatures required for the differentiation of B cell subtypes within lymphoid organs. Single cell methods allow the capture of minute changes in B cell gene signatures during health and disease; these allow the translation of newly identified genes into putative biomarkers and/or therapeutic targets. B cell lineage tracing and signaling mechanisms have been refined due to high-resolution data from single cell methods. These high-throughput methods, which enable the discovery of new B cell subpopulations, pave the way for studies aiming to understand the functional diversity, plasticity, and heterogeneity of B cells. The integration of single cell methods and spatial transcriptomics can provide deeper insights into the cell-intrinsic and -extrinsic roles of B cell in healthy and diseased microenvironments. [ABSTRACT FROM AUTHOR]

Monocytes are progenitors to macrophages and a subclass of dendritic cells (monocyte-derived dendritic cells, MoDCs), but they also act as circulating sensors that respond to environmental changes and disease. Technological advances have defined the production of classical monocytes in the bone marrow through the identification of lineage-determining transcription factors (LDTFs) and have proposed alternative routes of differentiation. Monocytes released into the circulation can be recruited to tissues by specific chemoattractants where they respond to sequential niche-specific signals that determine their differentiation into terminal effector cells. New aspects of monocyte biology in the circulation are being revealed, exemplified by the influence of cancer on the systemic alteration of monocyte subset abundance and transcriptional profiles. These changes can act to enhance the metastatic spread of primary cancers and may offer therapeutic opportunities. Mammalian monocyte development in the bone marrow from hematopoietic stem cells is driven by evolving and dynamic genomic enhancer landscapes and sequential binding of LDTFs. Monocyte fates are highly diverse; their plasticity and niche-specific regulation are beginning to be revealed in the contexts of homeostasis, inflammation, and cancer. Murine models of liver injury reveal that the interactions of monocytes with liver sinusoidal cells can lead to sequential alterations of genomic enhancer landscapes that ultimately drive Kupffer cell-specific expression of enhancers and genes. Murine models of genetic fate mapping, combined with studies of human cancers using single-cell transcriptome and proteome analysis, as well as bioinformatic-inferred algorithms for mapping trajectories of cellular differentiation, demonstrate that the tumor-associated macrophage phenotype is driven by tumor-specific features, regardless of their origin from monocytes or from yolk sac-derived tissue-resident monocytes. [ABSTRACT FROM AUTHOR]

• Evidence that supports that DCs are a separate lineage to myeloid and lymphoid. • Summary of DC development at a clonal level. • Description of single cell lineage tracing technologies and their findings. • Placement of DC development within the emerging continuous model of haematopoiesis. • Reconciliation of controversies surrounding DC development. Understanding development of the dendritic cell (DC) subtypes continues to evolve. The origin and relationship of conventional DC type 1 (cDC1), cDC type 2 (cDC2) and plasmacytoid DCs (pDCs) to each other, and in relation to classic myeloid and lymphoid cells, has had a long and controversial history and is still not fully resolved. This review summarises the technological developments and findings that have been achieved at a clonal level, and how that has enhanced our knowledge of the process. It summarises the single cell lineage tracing technologies that have emerged, their application in in vitro and in vivo studies, in both mouse and human settings, and places the findings in a wider context of understanding haematopoiesis at a single cell or clonal level. In particular, it addresses the fate heterogeneity observed in many phenotypically defined progenitor subsets and how these findings have led to a departure from the classic ball-and-stick models of haematopoiesis to the emerging continuous model. Prior contradictions in DC development may be reconciled if they are framed within this revised model, where commitment to a lineage or cell type does not occur in an all-or-nothing process in defined progenitors but rather can occur at many stages of haematopoiesis in a dynamic process. [ABSTRACT FROM AUTHOR]

• Cellular lineage commitment within hematopoiesis is reviewed. • Insights from novel lineage tracing and single-cell transcriptomics studies are discussed. • How single cell proteomics has resolved cell state transitions in hematopoiesis is described. Blood production is essential to maintain human health, and even small perturbations in hematopoiesis can cause disease. Hematopoiesis has therefore been the focus of much research for many years. Experiments determining the lineage potentials of hematopoietic stem and progenitor cells (HSPCs) in vitro and after transplantation revealed a hierarchy of progenitor cell states, where differentiating cells undergo lineage commitment—a series of irreversible changes that progressively restrict their potential. New technologies have recently been developed that allow for a more detailed analysis of the molecular states and fates of differentiating HSPCs. Proteomic and lineage-tracing approaches, alongside single-cell transcriptomic analyses, have recently helped to reveal the biological complexity underlying lineage commitment during hematopoiesis. Recent insights from these new technologies were presented by Dr. Marjorie Brand and Dr. Allon Klein in the Summer 2019 ISEH Webinar, and are discussed in this Perspective. [ABSTRACT FROM AUTHOR]

Unravelling stem cell fate plasticity is significant for understanding pathophysiology and exploring new therapeutic targets. The Cre-loxP genetic system is commonly used for cell lineage tracing. However, this conventional approach has limitations that have led to controversies in multiple fields. In recent years, there has been growing interest in a dual genetic approach based on Cre-loxP and another orthogonal recombinase. The dual genetic approach permits cell tracking at a significantly higher resolution and enables a more precise method for gene manipulation and cell fate control. Here, we review the recent progress in dual genetic approaches and highlight emerging work to demonstrate how the application of these approaches advances our understanding of stem cell fate plasticity for tissue repair and regeneration. [ABSTRACT FROM AUTHOR]

• Sox2 knockout in HCs increased the number of Type I HCs and decreased the number of Type II HCs. • The effect of Sox2 -knockout persisted into adulthood, resulting in an increased number of Type I HCs. • The results demonstrated that SOX2 plays a critical role in Type I and Type II HC fate determination. The vestibular system of the inner ear contains Type I and Type II hair cells (HCs) generated from sensory progenitor cells; however, little is known about how the HC subtypes are formed. Sox2 (encoding SRY-box 2) is expressed in Type II, but not in Type I, HCs. The present study aimed to investigate the role of SOX2 in cell fate determination in Type I vs. Type II HCs. First, we confirmed that Type I HCs developed from Sox2- expressing cells through lineage tracing of Sox2- positive cells using a CAG-tdTomato reporter mouse crossed with a Sox2-CreER mouse. Then, Sox2 loss of function was induced in HCs, using Sox2 flox transgenic mice crossed with a Gfi1-Cre driver mouse. Knockout of Sox2 in HCs increased the number of Type I HCs and decreased the number of Type II HCs, while the total number of HCs and Sox2- positive supporting cells did not change. In addition, the effect of Sox2 -knockout persisted into adulthood, resulting in an increased number of Type I HCs. These results demonstrate that SOX2 plays a critical role in the determination of Type II vs. Type I HC fate. The results suggested that Sox2 is a potential target for generating Type I HCs, which may be important for regenerative strategies for balance disorders. [ABSTRACT FROM AUTHOR]

Mammalian development involves continuous dynamic processes in which cells propagate, differentiate, orchestrate, and decease to produce high-order functions. Although accurate cell lineage information can provide a strong foundation to understand such complex processes, the cell lineages involved in development of the whole mammalian body remain largely unclear, except for in early embryogenesis, which is observable under a microscope. With CRISPR genome editing, the concept of 'evolving DNA barcodes' has rapidly emerged for large-scale, high-resolution cell lineage tracing, where cell-embedded DNA barcodes continuously accumulate random mutations that are inherited from mother to daughter cells. Similar to evolutionary tree reconstruction using species' DNA sequences, cell lineages can be reconstructed using shared mutation patterns in the DNA barcodes identified using massively parallel sequencing. The dramatic developments of single-cell and imaging technologies have enabled analyses of the molecular and spatial architecture of heterogeneous cells. The evolving DNA barcodes can also consolidate this information on a reconstructed cell lineage tree and accelerate our understanding of multicellular organisms. [ABSTRACT FROM AUTHOR]

Cardiac resident macrophages play vital roles in heart development, homeostasis, repair, and regeneration. Recent studies documented the hematopoietic potential of cardiac endothelium that supports the generation of cardiac macrophages and peripheral blood cells in mice. However, the conclusion was not strongly supported by previous genetic tracing studies, given the non-specific nature of conventional Cre- loxP tracing tools. Here, we develop an intercellular genetic labeling system that can permanently trace heart-specific endothelial cells based on cell-cell interaction in mice. Results from cell-cell contact-mediated genetic fate mapping demonstrate that cardiac endothelial cells do not exhibit hemogenic potential and do not contribute to cardiac macrophages or other circulating blood cells. This Matters Arising paper is in response to Shigeta et al. (2019), published in Developmental Cell. See also the response by Liu and Nakano (2023), published in this issue. [Display omitted] • Generation of intercellular genetic recording system to label neighboring cells • Genetic tracing of endocardium and coronary endothelial cells by cell-cell contact • Cardiac endothelium does not have hematopoietic potential in development • Cell-cell contact-mediated labeling enhances the specificity of genetic tracing In this Matters Arising article, Liu and Jin et al. re-evaluate the model that the developing endocardium is hemogenic and generates cardiac macrophages and circulating blood cells, which was proposed by Shigeta et al. By using an intercellular genetic tracing system that enables specific labeling of cardiac endothelium, they find that the cardiac endothelium does not have hematopoietic activity during heart development. [ABSTRACT FROM AUTHOR]

Diverse glutamatergic projection neurons (PNs) mediate myriad processing streams and output channels of the cerebral cortex. Yet, how different types of neural progenitors, such as radial glia (RGs) and intermediate progenitors (IPs), produce PN diversity, and hierarchical organization remains unclear. A fundamental issue is whether RGs constitute a homogeneous, multipotent lineage capable of generating all major PN types through a temporally regulated developmental program, or whether RGs comprise multiple transcriptionally heterogenous pools, each fated to generate a subset of PNs. Beyond RGs, the role of IPs in PN diversification remains underexplored. Addressing these questions requires tracking PN developmental trajectories with cell-type resolution – from transcription factor-defined RGs and IPs to their PN progeny, which are defined not only by laminar location but also by projection patterns and gene expression. Advances in cell-type resolution genetic fate mapping, axon tracing, and spatial transcriptomics may provide the technical capability for answering these fundamental questions. • The hierarchical organization of PN types is jointly defined by their transcriptome, anatomy, and physiology. • Lineage tracing needs to distinguish TF-defined RGs as well as projection and transcriptionally defined PNs. • Emerging evidence of fate-restricted RGs is not mutually exclusive with models of multipotency and temporal competence. • Direct and indirect neurogenesis differentially contribute to PN diversity. [ABSTRACT FROM AUTHOR]

Abstract SHP2 is a ubiquitously expressed protein tyrosine phosphatase, which is involved in many signaling pathways to regulate the skeletal development. In endochondral ossification, SHP2 is known to modify the osteogenic fate of osteochondroprogenitors and to impair the osteoblastic transdifferentiation of hypertrophic chondrocytes. However, how SHP2 regulates osteoblast differentiation in intramembranous ossification remains incompletely understood. To address this question, we generated a mouse model to ablate SHP2 in the Prrx1-expressing mesenchymal progenitors by using "Cre-loxP"-mediated gene excision and examined the development of calvarial bone, in which the main process of bone formation is intramembranous ossification. Phenotypic characterization showed that SHP2 mutants have severe defects in calvarial bone formation. Cell lineage tracing and in situ hybridization data showed less osteoblast differentiation of mesenchymal cells and reduced osteogenic genes expression, respectively. Further mechanistic studies revealed enhanced TGFβ and suppressed BMP2 signaling in SHP2 ablated mesenchymal progenitors and their derivatives. Our study uncovered the critical role of SHP2 in osteoblast differentiation through intramembranous ossification and might provide a potential target to treat craniofacial skeleton disorders. Highlights • SHP2 is required for intramembranous ossification. • SHP2 regulates the osteogenic differentiation of mesenchymal progenitors. • SHP2 in Prrx1+ mesenchymal progenitors modifies BMP2 and TGFβ signaling and osteogenic transcription factor expression. [ABSTRACT FROM AUTHOR]

Abstract Phenotypes of myeloid-lineage cells remain poorly understood in the rainbow trout, and were the focus of this study, including effects of in vivo challenge to Flavobacterium psychrophilum (Fp), the cause of Bacterial Cold Water Disease (BCWD). A genetic line was used that is highly resistant to BCWD (R-line) as well as a susceptible control line (S-line). Using flow cytometry, we describe two Pax5-negative, myeloid-lineage populations: Population 1 consisted of small cells with high SSC and strong staining for Q4E, MPO, Pu1, EBF, and IL- 1β, which we named "neutrophil-like" cell. Population 2 had high Q4E, but weaker MPO, Pu1, EBF, and IL-1β staining. Five days after Fp-challenge, both genetic lines had a reduced abundance of neutrophil-like cells in anterior kidney, PBL, and spleen. Pop. 2 abundance was reduced in anterior kidney, and increased in spleen. S-line fish responded more strongly to Fp-challenge compared to R-line fish. Challenged fish with a higher abundance of neutrophil-like cells had significantly lower Fp-loads after challenge, suggesting that these cells aid in the resistance to BCWD. Highlights • A line of Rainbow trout resistant to BCWD differed from a susceptible line in abundance of myeloid-lineage populations. • After in vivo challenge with Flavobacterium psychrophilum , changes in abundance of neutrophil-like cells was more strongly seen in susceptible fish. • Fp-challenged fish with higher abundance of neutrophil-like cells in their anterior kidney or PBLs had significantly lower Fp-loads. [ABSTRACT FROM AUTHOR]

Highlights • Cell reprogramming requires genome-wide changes to endogenous gene expression. • Synthetic transcription factors (Syn-TFs) can regulate endogenous gene expression in vitro and in vivo. • Syn-TFs are capable of initiating cell reprogramming in vitro and in vivo. • Syn-TFs can deterministically remodel chromatin and establish autonomous gene expression networks. • High throughput screens with libraries of Syn-TFs can discover novel regulators of reprogramming. The ability to reprogram cell lineage specification through the activity of master regulatory transcription factors has transformed disease modeling, drug screening, and cell therapy for regenerative medicine. Recent advances in the engineering of synthetic transcription factors to modulate endogenous gene expression networks and chromatin states have generated a new set of tools with unique advantages to study and enhance cell reprogramming methods. Several studies have applied synthetic transcription factors in various cell reprogramming paradigms in human and murine cells. Moreover, the adaption of CRISPR-based transcription factors for high-throughput screening will enable the systematic identification of optimal factors and gene network perturbations to improve current reprogramming protocols and enable conversion to more diverse, highly specified, and mature cell types. The rapid development of next-generation technologies with more robust and versatile functionality will continue to expand the application of synthetic transcription factors for cell reprogramming. [ABSTRACT FROM AUTHOR]

Highlights • iNKT subsets are defined by expression of lineage specifying transcription factors. • Epigenetic mechanisms are critical for regulating gene expression. • DNA methyltransferases and TET proteins control the dynamics of DNA methylation. • TET proteins exert multifaceted roles in regulating gene expression. • iNKT cell lineage specification is controlled by TET proteins. Abstract During the last years, intensive research has shed light in the transcriptional networks that shape the invariant NKT (iNKT) cell lineage and guide the choices towards functionally distinct iNKT cell subsets (Constantinides and Bendelac, 2013; Engel and Kronenberg, 2014; Gapin, 2016; Kim et al., 2015). However, the epigenetic players that regulate gene expression and orchestrate the iNKT cell lineage choices remain poorly understood. Here, we summarize recent advances in our understanding of epigenetic regulation of iNKT cell development and lineage choice. Particular emphasis is placed on DNA modifications and the Ten Eleven Translocation (TET) family of DNA demethylases. [ABSTRACT FROM AUTHOR]

In vitro experiments with cultured cells are essential for studying their growth and migration pattern and thus, for gaining a better understanding of cancer progression and its treatment. Recent progress in lens-free microscopy (LFM) has rendered it an inexpensive tool for label-free, continuous live cell imaging, yet there is only little work on analysing such time-lapse image sequences. We propose (1) a cell detector for LFM images based on fully convolutional networks and residual learning, and (2) a probabilistic model based on moral lineage tracing that explicitly handles multiple detections and temporal successor hypotheses by clustering and tracking simultaneously. (3) We benchmark our method in terms of detection and tracking scores on a dataset of three annotated sequences of several hours of LFM, where we demonstrate our method to produce high quality lineages. (4) We evaluate its performance on a somewhat more challenging problem: estimating cell lineages from the LFM sequence as would be possible from a corresponding fluorescence microscopy sequence. We present experiments on 16 LFM sequences for which we acquired fluorescence microscopy in parallel and generated annotations from them. Finally, (5) we showcase our methods effectiveness for quantifying cell dynamics in an experiment with skin cancer cells. [ABSTRACT FROM AUTHOR]

Although osteoblasts and osteocytes are descended from the same lineage, they each have unique and essential roles in bone. Targeting gene deletion to osteoblasts and osteocytes using the Cre/loxP system has greatly increased our current understanding of how these cells function. Additionally, the use of the Cre/loxP system in conjunction with cell-specific reporters has enabled lineage tracing of these bone cells both in vivo and ex vivo. However, concerns have been raised regarding the specificity of the promoters used and the resulting off-target effects on cells within and outside of the bone. In this review, we have summarized the main mouse models that have been used to determine the functions of specific genes in osteoblasts and osteocytes. We discuss the expression patterns and specificity of the different promoter fragments during osteoblast to osteocyte differentiation in vivo. We also highlight how their expression in non-skeletal tissues may complicate the interpretation of study results. A thorough understanding of when and where these promoters are activated will enable improved study design and greater confidence in data interpretation. • Specific targeting of osteocytes or osteoblasts in vivo is challenging. • Relevant osteoblast and osteocyte-targeted promoter models are described. • Limitations due to off-target promoter expression are discussed. • Future strategies to improve specific targeting at different stages of osteoblast-osteocyte differentiation are proposed. [ABSTRACT FROM AUTHOR]

We present a novel computational framework for the analysis of high-throughput microscopy videos of living cells. The proposed framework is generally useful and can be applied to different datasets acquired in a variety of laboratory settings. This is accomplished by tying together two fundamental aspects of cell lineage construction, namely cell segmentation and tracking, via a Bayesian inference of dynamic models. In contrast to most existing approaches, which aim to be general, no assumption of cell shape is made. Spatial, temporal, and cross-sectional variation of the analysed data are accommodated by two key contributions. First, time series analysis is exploited to estimate the temporal cell shape uncertainty in addition to cell trajectory. Second, a fast marching (FM) algorithm is used to integrate the inferred cell properties with the observed image measurements in order to obtain image likelihood for cell segmentation, and association. The proposed approach has been tested on eight different time-lapse microscopy data sets, some of which are high-throughput, demonstrating promising results for the detection, segmentation and association of planar cells. Our results surpass the state of the art for the Fluo-C2DL-MSC data set of the Cell Tracking Challenge (Maška et al., 2014). [ABSTRACT FROM AUTHOR]

Ocular infection with herpes simplex virus (HSV) can result in a chronic immune inflammatory lesion that is a significant cause of human blindness. A key to controlling stromal keratitis (SK) lesion severity is to identify cellular and molecular events responsible for tissue damage and to counteract them. One potentially useful approach to achieve such therapy is Retinoic Acid (RA). Here we show that RA therapy reduces the severity of SK by having inhibitory effects on the T effector subtypes responsible for orchestrating SK. RA also served to stabilize the function of regulatory T cell (Treg) which counteract inflammatory cell activity. The Treg stabilizing effect was demonstrated by in vitro studies where RA was shown to retain Foxp3 expression when exposed to proinflammatory conditions such as IL-12 and IL-6+TGF-β. in vivo studies revealed that RA exerted its stabilizing effects by downregulating IL-6R expression on Treg after HSV-1 infection and this helped to control the progression of SK. Since the therapy was effective when used both early and after the initiation of lesions, it may represent a valuable means of therapy when used alone or along with additional therapies. [ABSTRACT FROM AUTHOR]

The anaphase of mitosis is one of the most critical stages of the cell division cycle in that it can reveal precious information on the fate of a cell lineage. Indeed, most types of nuclear DNA segregation defects visualized during anaphase are manifestations of genomic instability and augur dramatic outcomes, such as cell death or chromosomal aberrations characteristic of cancer cells. Although chromatin bridges and lagging chromatin are always pathological (generating aneuploidy or complex genomic rearrangements), the main subject of this article, the ultrafine anaphase bridges, might, in addition to potentially driving genomic instability, play critical roles for the maintenance of chromosome structure in rapidly proliferating cells. [ABSTRACT FROM AUTHOR]

Background aims We have previously reported the generation of a current Good Manufacture Practice (cGMP)-compliant induced pluripotent stem cell (iPSC) line for clinical applications. Here we show that multiple cellular products currently being considered for therapy can be generated from a single master cell bank of this or any other clinically compliant iPSC line Methods Using a stock at passage 20 prepared from the cGMP-compliant working cell bank (WCB), we tested differentiation into therapeutically relevant cell types of the three germ layers using standardized but generic protocols. Cells that we generated include (i) neural stem cells, dopaminergic neurons and astrocytes; (ii) retinal cells (retinal pigment epithelium and photoreceptors); and (iii) hepatocyte, endothelial and mesenchymal cells. To confirm that these generic protocols can also be used for other iPSC lines, we tested the reproducibility of our methodology with a second clinically compliant line Results Our results confirmed that well-characterized iPSC lines have broad potency, and, despite allelic variability, the same protocols could be used with minimal modifications with multiple qualified lines. In addition, we introduced a constitutively expressed GFP cassette in Chr13 safe harbor site using a standardized previously described method and observed no significant difference in growth and differentiation between the engineered line and the control line indicating that engineered products can be made using a standardized methodology Conclusions We believe that our demonstration that multiple products can be made from the same WCB and that the same protocols can be used with multiple lines offers a path to a cost-effective strategy for developing cellular products from iPSC lines. [ABSTRACT FROM AUTHOR]

Delineating the lineage of neural cells that captures the progressive steps in their specification is fundamental to understanding brain development, organization, and function. Since the earliest days of embryology, lineage questions have been addressed with methods of increasing specificity, capacity, and resolution. Yet, a full realization of individual cell lineages remains challenging for complex systems. A recent explosion of technical advances in genome-editing and single-cell sequencing has enabled lineage analysis in an unprecedented scale, speed, and depth across different species. In this review, we discuss the application of available as well as future genetic labeling techniques for tracking neural lineages in vivo in the mammalian nervous system. [ABSTRACT FROM AUTHOR]

Advances in single-cell RNA-sequencing have helped reveal the previously underappreciated level of cellular heterogeneity present during cellular differentiation. A static snapshot of single-cell transcriptomes provides a good representation of the various stages of differentiation as differentiation is rarely synchronized between cells. Data from numerous single-cell analyses has suggested that cellular differentiation and development can be conceptualized as continuous processes. Consequently, computational algorithms have been developed to infer lineage relationships between cell types and construct developmental trajectories along which cells are re-ordered such that similarity between successive cell pairs is maximized. Here, we compare and contrast the existing computational methods, and illustrate how they may be applied to build mouse myeloid progenitor lineages from massively parallel RNA single-cell sequencing data. [ABSTRACT FROM AUTHOR]

Most single-cell approaches to date are based on destructive snapshot measurements which do not permit to correlate a current molecular state with future fate. However, to understand how cell fate choices are established by transcription factor networks (TFNs) regulating cell fates, TFN dynamics must be continuously monitored in single cells. Here we review how quantitative time-lapse imaging can contribute to understanding TFN dependent cell fate regulation at the single-cell level. We outline potentials of the technology and highlight challenges for interpreting the dynamics of fluorescent protein reporters that may interfere with endogenous TF function. We provide an outlook on how continuous observation of TF dynamics and single-cell fates may be complemented by perturbation studies and be linked to multidimensional molecular profiling in the future. [ABSTRACT FROM AUTHOR]

Determining the lineage origin of cell types is a major goal in developmental biology. Furthermore, lineage tracing is a powerful approach for understanding the origin of developmental defects as well as the origin of diseases such as cancer. There is now a variety of complementary approaches for identifying lineage relationships, ranging from direct observation of cell divisions by light microscopy to genetic labeling of cells using inducible recombinases and fluorescent reporters. A recent development, and the main topic of this review article, is the use of high-throughput sequencing data for lineage analysis. This emerging approach holds the promise of increased multiplexing capacity, allowing lineage analysis of large cell numbers up to the organism-wide level combined with simultaneous transcription profiling by single cell RNA sequencing. [ABSTRACT FROM AUTHOR]

Purpose of review The hierarchical nature of the hematopoietic system provides an ideal model system to illustrate the features of lineage tracing. We have outlined the utility of lineage tracing methods in establishing the origin and development of hematopoietic cells. Recent findings Methods such as CRISPR/Cas9, Polylox barcoding, and single-cell RNA-sequencing have improved our understanding of hematopoiesis. Summary This review chronicles the fate of the hematopoietic cells emerging from the mesoderm that subsequently develops into the adult blood lineages. Specifically, we explain classic techniques utilized in lineage tracing for the hematopoietic system, as well as novel state-of-the-art methods to elucidate clonal hematopoiesis and cell fate mapping at a single-cell level. [ABSTRACT FROM AUTHOR]

Oligodendrocytes (OLs) are glial cells in the central nervous system (CNS), which produce myelin, a lipid-rich membrane that insulates neuronal axons. The main function ascribed to OLs is to regulate the speed of electric pulse transmission, and as such OLs have been widely considered as a single and discrete population. Nevertheless, OLs and their precursor cells (OPCs) throughout the CNS have different morphologies and regional functional differences have been observed. Moreover, OLs have recently been involved in other functional processes such as metabolic coupling with axons. In this review, we focus on recent advances in single-cell transcriptomics suggesting that OLs are more heterogeneous than previously thought, with defined subpopulations and cell states that are associated with different stages of lineage progression and might also represent distinct functional states. [ABSTRACT FROM AUTHOR]

Peripheral glia includes predominantly myelinating and non-myelinating Schwann cells in addition to satellite, terminal and enteric glia as well as other unresolved subtypes with localized functions. Of these subtypes, all of them originate from neural crest-derived embryonic Schwann cell precursors (SCPs). Specific gene regulatory networks control neural crest specification and downstream events, including SCP differentiation and myelination. Embryonic SCPs are multipotent and generate neuroendocrine cells, parasympathetic and enteric neurons, melanocytes and other cell types. The evolutionary origin of peripheral Schwann cell lineage is not widely discussed in the literature, despite numerous similarities between central and peripheral glia. Here, we review the major features of the Schwann cell lineage and proceed to an evolutionary discussion around possible relations between central and peripheral glial cells. [ABSTRACT FROM AUTHOR]

Stem cells are at the basis of organismal development, characterized by their potential to differentiate towards specific lineages upon receiving proper signals. To understand the molecular principles underlying gain and loss of pluripotency, proteomics plays an increasingly important role owing to technical developments in mass spectrometry and implementation of innovative biochemical approaches. Here we review how quantitative proteomics has been used to investigate protein expression, localization, interaction and modification in stem cells both in vitro and in vivo , thereby complementing other omics approaches to study fundamental properties of stem cell plasticity. [ABSTRACT FROM AUTHOR]

Over the last decade significant advances have been made toward reprogramming the fate of somatic cells, typically by overexpression of cell lineage-determinant transcription factors. As key regulators of cell fate, the SOX family of transcription factors has emerged as potent drivers of direct somatic cell reprogramming into multiple lineages, in some cases as the sole overexpressed factor. The vast capacity of SOX factors, especially those of the SOXB1, E and F subclasses, to reprogram cell fate is enlightening our understanding of organismal development, cancer and disease, and offers tremendous potential for regenerative medicine and cell-based therapies. Understanding the molecular mechanisms through which SOX factors reprogram cell fate is essential to optimize the development of novel somatic cell transdifferentiation strategies. [ABSTRACT FROM AUTHOR]

To understand organ morphogenetic mechanisms, it is essential to clarify how spatiotemporally-regulated molecular/cellular dynamics causes physical tissue deformation. In the case of vertebrate limb development, while some of the genes and oriented cell behaviors underlying morphogenesis have been revealed, tissue deformation dynamics remains incompletely understood. We here introduce our recent work on the reconstruction of tissue deformation dynamics in chick limb development from cell lineage tracing data. This analysis has revealed globally-aligned anisotropic tissue deformation along the proximo-distal axis not only in the distal region but also in the whole limb bud. This result points to a need, as a future challenge, to find oriented molecular/cellular behaviors for realizing the observed anisotropic tissue deformation in both proximal and distal regions, which will lead to systems understanding of limb morphogenesis. [ABSTRACT FROM AUTHOR]

T cells are continually generated in the thymus in a highly dynamic process comprising discrete steps of lineage commitment, T cell receptor (TCR) gene rearrangement, and selection. These steps are linked to distinct rates of proliferation, survival, and cell death, but a quantitative picture of T cell development is only beginning to emerge. Here we summarize recent technical advances, including genetic fate mapping, barcoding, and molecular timers, that have allowed the implementation of computational models to quantify developmental dynamics in the thymus. Coupling new techniques with mathematical models has recently resulted in the emergence of new paradigms in early hematopoiesis and might similarly open new perspectives on T cell development. [ABSTRACT FROM AUTHOR]

An ideal model for HIV-1 research is still unavailable. However, infection of non-human primates (NHP), such as macaques, with Simian Immunodeficiency Virus (SIV) recapitulates most virological, immunological and clinical hallmarks of HIV infection in humans. It has become the most suitable model to study the mechanisms of transmission and physiopathology of HIV/AIDS. On the other hand, natural hosts of SIV, such as African green monkeys and sooty mangabeys that when infected do not progress to AIDS, represent an excellent model to elucidate the mechanisms involved in the capacity of controlling inflammation and disease progression. The use of NHP-SIV models has indeed enriched our knowledge in the fields of: i) viral transmission and viral reservoirs, ii) early immune responses, iii) host cell-virus interactions in tissues, iv) AIDS pathogenesis, v) virulence factors, vi) prevention and vii) drug development. The possibility to control many variables during experimental SIV infection, together with the resemblance between SIV and HIV infections, make the NHP model the most appropriate, so far, for HIV/AIDS research. Nonetheless, some limitations in using these models have to be considered. Alternative models for HIV/AIDS research, such as humanized mice and recombinant forms of HIV-SIV viruses (SHIV) for NHP infection, have been developed. The improvement of SHIV viruses that mimic even better the natural history of HIV infection and of humanized mice that develop a greater variety of human immune cell lineages, is ongoing. None of these models is perfect, but they allow contributing to the progress in managing or preventing HIV infection. [ABSTRACT FROM AUTHOR]

Advancing our understanding of osteoblast biology and differentiation is critical to elucidate the pathological mechanisms responsible for skeletal diseases such as osteoporosis. Histology and histomorphometry, the classical methods to study osteoblast biology, identify osteoblasts based on their location and morphology and ability to mineralize matrix, but do not clearly define their stage of differentiation. Introduction of visual transgenes into the cells of osteoblast lineage has revolutionized the field and resulted in a paradigm shift that allowed for specific identification and isolation of subpopulations within the osteoblast lineage. Knowledge acquired from the studies based on GFP transgenes has allowed for more precise interpretation of studies analyzing targeted overexpression or deletion of genes in the osteoblast lineage. Here, we provide a condensed overview of the currently available promoter-fluorescent reporter transgenic mice that have been generated and evaluated to varying extents. We cover different stages of the lineage as transgenes have been utilized to identify osteoprogenitors, pre-osteoblasts, osteoblasts, or osteocytes. We show that each of these promoters present with advantages and disadvantages. The studies based on the use of these reporter mice have improved our understanding of bone biology. They constitute attractive models to target osteoblasts and help to understand their cell biology. [ABSTRACT FROM AUTHOR]

Bones are an important component of vertebrates; they grow explosively in early life and maintain their strength throughout life. Bones also possess amazing capabilities to repair — the bone is like new without a scar after complete repair. In recent years, a substantial progress has been made in our understanding on mammalian bone stem cells. Mouse genetic models are powerful tools to understand the cell lineage, giving us better insights into stem cells that regulate bone growth, maintenance and repair. Recent findings about these stem cells raise new questions that require further investigations. [ABSTRACT FROM AUTHOR]

During mouse embryogenesis initial specification of the cell fates depends on the type of division during 8- to 16- and 16- to 32-cell stage transition. A conservative division of a blastomere creates two polar outer daughter cells, which are precursors of the trophectoderm (TE), whereas a differentiative division gives rise to a polar outer cell and an apolar inner (the presumptive inner cell mass – ICM) cell. We hypothesize that the type of division may depend on the interactions between blastomeres of the embryo. To investigate whether modification of these interactions influences divisions, we analyzed the pattern of blastomere division and cell lineage specification in chimeric embryos obtained by injection of a different number of mouse embryonic stem cells (ESCs) into 8-cell embryos. As the ESCs populate only the ICM of the resulting chimeric blastocysts, they emulated in our model additional inner cells. We found that introduction of ESCs decreased the number of inner, apolar blastomeres at the 8- to 16-cell stage transition and reduced the number of ICM cells of host embryo-origin during formation of the blastocyst. Moreover, we showed that the proportion of inner blastomeres and their fate (EPI or PE) in chimeric blastocysts was dependent on the number of ESCs injected. Our results suggest the existence of a regulative mechanism, which links number of inner cells with a proportion of conservative vs. differentiative blastomere divisions during the cleavage and thus dictates their developmental fate. [ABSTRACT FROM AUTHOR]

Gastropod snails in the genus Crepidula have emerged as model systems for studying a metazoan super clade, the Spiralia. Recent work on one species in particular, Crepidula fornicata , has produced high-resolution cell lineage fate maps, details of morphogenetic events during gastrulation, key insights into the molecular underpinnings of early development, and the first demonstration of CRISPR/Cas9 genome editing in the Spiralia. Furthermore, invasive species of Crepidula are a significant ecological threat, while one of these, C. fornicata , is also being harvested for food. This review highlights progress towards developing these animals as models for evolutionary, developmental, and ecological studies. Such studies have contributed greatly to our understanding of biology in a major clade of bilaterians. This information may also help us to control and cultivate these snails. [ABSTRACT FROM AUTHOR]

The combinatorial action of transcription factors drives cell-type-specific gene expression patterns. However, transcription factor binding and gene regulation occur in the context of chromatin, which modulates DNA accessibility. High-resolution chromatin interaction maps have defined units of chromatin that are in spatial proximity, called topologically associated domains (TADs). TADs can be further classified based on expression activity, replication timing, or the histone marks or non-histone proteins associated with them. Independently, other chromatin domains have been defined by their likelihood to interact with non-DNA structures, such as the nuclear lamina. Lamina-associated domains (LADs) correlate with low gene expression and late replication timing. TADs and LADs have recently been evaluated with respect to cell-type-specific gene expression. The results shed light on the relevance of these forms of chromatin organization for transcriptional regulation, and address specifically how chromatin sequestration influences cell fate decisions during organismal development. [ABSTRACT FROM AUTHOR]

Microglia are tissue resident macrophages of the central nervous system (CNS) that maintain homeostasis and respond to immune challenges. New genetic fate mapping tools have revealed a yolk sac origin of microglia. Once established in the CNS, microglia persist throughout the lifetime of the organism behind the blood–brain barrier and maintain themselves by self-renewal. Recent studies uncovered a broad spectrum of microglial functions that are influenced by the dynamism of brain formation and neuronal wiring. This review focuses on current findings concerning microglia origin and formation during development and discusses the factors important for microglia survival and function. [ABSTRACT FROM AUTHOR]

Endocardial development involves a complex orchestration of cell fate decisions that coordinate with endoderm formation and other mesodermal cell lineages. Historically, investigations into the contribution of endocardium in the developing embryo was constrained to the heart where these cells give rise to the inner lining of the myocardium and are a major contributor to valve formation. In recent years, studies have continued to elucidate the complexities of endocardial fate commitment revealing a much broader scope of lineage potential from developing endocardium. These studies cover a wide range of species and model systems and show direct contribution or fate potential of endocardium giving rise to cardiac vasculature, blood, fibroblast, and cardiomyocyte lineages. This review focuses on the marked expansion of knowledge in the area of endocardial fate potential. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel. [ABSTRACT FROM AUTHOR]

Uncovering the mechanisms that underlie central noradrenergic neuron heterogeneity is essential to understanding selective subtype vulnerability to disease and environmental insult. Using recombinase-based intersectional genetic fate mapping we have previously demonstrated that molecularly distinct progenitor populations give rise to mature noradrenergic neurons differing in their anatomical location, axon morphology and efferent projection pattern. Here we review the findings from our previous study and extend our analysis of the noradrenergic subpopulation defined by transient developmental expression of Hoxb1. Using a combination of intersectional genetic fate mapping and analysis of a targeted loss of function mutation in Hoxb1 , we have now uncovered additional heterogeneity based on the requirement of some noradrenergic neurons for Hoxb1 expression. By comparing the distribution of noradrenergic neurons derived from the Hoxb1 expression domain in wild-type and mutant mice, we demonstrate that Hoxb1 expression is required by a subset of neurons in the pons. Additional fate mapping, using a Hoxb1 enhancer element that drives Cre recombinase expression exclusively in rhombomere 4 of the hindbrain, reveals the existence of a subpopulation of noradrenergic neurons in the pons with more restricted axonal targets than the full Hoxb1 -derived subpopulation. The unique projection profile of this newly defined subpopulation suggests that it may be functionally distinct. These analyses shed new light on the molecular determinants of noradrenergic identity in the pons and the overall complexity of the central noradrenergic system. This article is part of a Special Issue entitled SI: Noradrenergic System . [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is pathologically characterized by extracellular deposition of insoluble amyloid-β (Aβ) plaques and intracellular tangles made up of phosphorylated tau in brain. Several therapeutic approaches are being carried out in animal AD models for testing their safety and efficacy in altering disease pathology and behavioral deficits. Very few studies have examined the effect of human umbilical cord blood (hUCB) derived stem cells in degenerative disease models despite growing number of cord blood banks worldwide. Here we have examined the therapeutic efficacy of hUCB derived lineage negative (Lin −ve) stem cells in alleviating behavioral and neuropathological deficits in a mouse model of cognitive impairment induced by bilateral intrahippocampal injection of Aβ-42. Lin −ve cells were transplanted at two doses (50,000 and 100,000) at the site of injury and examined at 10 and 60 days post transplantation for rescue of memory deficits. These cells were found to ameliorate cognitive impairment in 50,000—60 days and 100,000—10 days groups whereas, 50,000—10 days and 100,000—60 days groups could not exert any significant improvement. Further, mice showing spatial memory improvement were mediated by up-regulation of BDNF, CREB and also by concomitant down regulation of Fas-L in their brain. The transplanted cells were found in the host tissue and survived up to 60 days without expressing markers of neuronal differentiation or reducing Aβ burden in mouse brain. We suggest that these undifferentiated cells could exert neuroprotective effects either through inhibiting apoptosis and/or trophic effects in the brain. [ABSTRACT FROM AUTHOR]

A large proportion of the cellular transcriptome of higher vertebrates consists of non-protein coding transcripts, among them the long noncoding RNAs (lncRNAs). Although lncRNAs are functionally extremely divergent, many ncRNAs have been shown to interact with chromatin modifying complexes and/or with transcriptional regulators. Via such interactions, many lncRNAs are involved in controlling the activity and expression level of target genes, including important regulators of embryonic processes, and thereby fine-tune gene regulatory networks controlling cell fate, lineage balance, and organogenesis. Intriguingly, an increase in organ complexity during evolution parallels a rise in lncRNA abundance. The current data suggest that lncRNAs support the generation of cell diversity and organ complexity during embryogenesis, and thereby have promoted the evolution of more complex organisms. [ABSTRACT FROM AUTHOR]

Human neural stem cells (hNSCs) hold great promise for the treatment of neurological diseases. Considerable progress has been made to induce neural differentiation in the cell culture in vitro and upon transplantation in vivo [2] in order to explore restoration of damaged neuronal circuits. However, in vivo conventional strategies are limited to post mortem analysis. Here, we apply our developed first fate mapping platform to monitor neuronal differentiation in vivo by magnetic resonance imaging, bioluminescence imaging, and fluorescence imaging. Ferritin, Luciferase and GFP under neuronal-specific promoters for immature and mature neurons, respectively, were used to generate transgenic hNSCs. Differentiation-linked imaging reporter expression was validated in vitro . The time profile of spontaneous neuronal maturation after transplantation into mouse brain cortex demonstrated early neuronal differentiation within 6 weeks. Fully mature neurons expressing synaptogenesis were observed only after three months or longer. Our trimodal fate mapping strategy represents a unique non-invasive tool to monitor the time course of neuronal differentiation of transplanted stem cells in vivo . [ABSTRACT FROM AUTHOR]

Identifying the matrix properties that permit directing stem cell fate is crucial for expanding desired cell lineages ex vivo for disease treatment. Such efforts require knowledge of matrix surface chemistry and the cell responses they elicit. Recent progress in analyzing biomaterial composition and identifying cell phenotype with two label-free chemical imaging techniques, TOF-SIMS and Raman spectroscopy are presented. TOF-SIMS is becoming indispensable for the surface characterization of biomaterial scaffolds. Developments in TOF-SIMS data analysis enable correlating surface chemistry with biological response. Advances in the interpretation of Raman spectra permit identifying the fate decisions of individual, living cells with location specificity. Here we highlight this progress and discuss further improvements that would facilitate efforts to develop artificial scaffolds for tissue regeneration. [ABSTRACT FROM AUTHOR]

The article discusses the research on how the Cre-loxP system can circumvent the problem of label dilution through induction of reporter genes and how it can be used to visualize cell lineages. It states that it allows to follow cell fate decision of stem/progenitor cell populations. It mentions that the expression of reporter genes and Cre-inducing agents have direct effect on the behavior of cell due to lack of suitable controls.

The myogenic regulatory genes Myf5, Mrf4, Myod, and Myogenin likely arose by gene duplications during evolution, presumably to address the more demanding requirements of the vertebrate body plan. Two cell lineages were proposed to be regulated independently by Myf5 and Myod to safeguard against tissue failure. Here we report severe muscle loss following ablation of Myf5-expressing cells. Using both lineage-specific and ubiquitous reporter alleles, we show that the remaining muscles in Myf5Cre-DTA embryos arise mainly from Myf5+ escaper cells. Elimination of Myf5Cre-DTA cells on a Myod null background did not result in the total absence of skeletal muscles, as would be expected if a Myod+/Myf5-independent cell population played a major role in this scenario. Therefore, these observations are incompatible with a previously proposed functional two-lineage model. These findings will have an impact on the interpretation of phenotypes obtained using similar strategies in other tissues. [ABSTRACT FROM AUTHOR]