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Gut physiology is the epicenter of a web of internal communication systems (i.e., neural, immune, hormonal) mediated by cell-cell contacts, soluble factors, and external influences, such as the microbiome, diet, and the physical environment. Together these provide the signals that shape enteric homeostasis and, when they go awry, lead to disease. Faced with the seemingly paradoxical tasks of nutrient uptake (digestion) and retarding pathogen invasion (host defense), the gut integrates interactions between a variety of cells and signaling molecules to keep the host nourished and protected from pathogens. When the system fails, the outcome can be acute or chronic disease, often labeled as "idiopathic" in nature (e.g., irritable bowel syndrome, inflammatory bowel disease). Here we underscore the importance of a holistic approach to gut physiology, placing an emphasis on intercellular connectedness, using enteric neuroimmunophysiology as the paradigm. The goal of this opinion piece is to acknowledge the pace of change brought to our field via single-cell and -omic methodologies and other techniques such as cell lineage tracing, transgenic animal models, methods for culturing patient tissue, and advanced imaging. We identify gaps in the field and hope to inspire and challenge colleagues to take up the mantle and advance awareness of the subtleties, intricacies, and nuances of intestinal physiology in health and disease by defining communication pathways between gut resident cells, those recruited from the circulation, and "external" influences such as the central nervous system and the gut microbiota. [ABSTRACT FROM AUTHOR]

This article provides a summary of recent advances in kidney research published in Acta Physiologica. The research covers various aspects of renal physiology, including glomerular filtration dynamics, tubular transport mechanisms, hormonal regulation, and genetic and environmental determinants of renal health. The studies discussed in the article explore topics such as the effects of fasting on fibroblast growth factor 23 levels, the role of exercise in protecting against acute kidney injury, the potential benefits of the drug tempol in septic acute kidney injury, the importance of renin-producing cells in kidney regeneration, the impact of immunosuppressive drugs on kidney function, the involvement of Two-pore channels protein 1 in phosphate excretion, the use of MRI markers to assess renal oxygenation, the relationship between kidney dysfunction and atrial fibrillation, and the role of endothelial to mesenchymal transition in cardiovascular and kidney pathophysiology. The article also mentions review articles on glomerular crosstalk mechanisms and circadian rhythm in renal physiology and pathophysiology. These recent advancements have the potential to contribute to the development of targeted therapies and diagnostic tools for renal disorders. [Extracted from the article]

Loss of refreshment in nucleus pulposus (NP) cellularity leads to intervertebral disc (IVD) degeneration. Nevertheless, the cellular sequence of NP cell differentiation remains unclear, although an increasing body of literature has identified markers of NP progenitor cells (NPPCs). Notably, due to their fragility, the physical enrichment of NP‐derived cells has limited conventional transcriptomic approaches in multiple studies. To overcome this limitation, a spatially resolved transcriptional atlas of the mouse IVD is generated via the 10x Genomics Visium platform dividing NP spots into two clusters. Based on this, most reported NPPC‐markers, including Cathepsin K (Ctsk), are rare and predominantly located within the NP‐outer subset. Cell lineage tracing further evidence that a small number of Ctsk‐expressing cells generate the entire adult NP tissue. In contrast, Tie2, which has long suggested labeling NPPCs, is actually neither expressed in NP subsets nor labels NPPCs and their descendants in mouse models; consistent with this, an in situ sequencing (ISS) analysis validated the absence of Tie2 in NP tissue. Similarly, no Tie2‐cre‐mediated labeling of NPPCs is observed in an IVD degenerative mouse model. Altogether, in this study, the first spatial transcriptomic map of the IVD is established, thereby providing a public resource for bone biology. [ABSTRACT FROM AUTHOR]

Background: Most primary Triple Negative Breast Cancers (TNBCs) show amplification of the Epidermal Growth Factor Receptor (EGFR) gene, leading to increased protein expression. However, unlike other EGFR-driven cancers, targeting this receptor in TNBC yields inconsistent therapeutic responses. Methods: To elucidate the underlying mechanisms of this variability, we employ cellular barcoding and single-cell transcriptomics to reconstruct the subclonal dynamics of EGFR-amplified TNBC cells in response to afatinib, a tyrosine kinase inhibitor (TKI) that irreversibly inhibits EGFR. Results: Integrated lineage tracing analysis revealed a rare pre-existing subpopulation of cells with distinct biological signature, including elevated expression levels of Insulin-Like Growth Factor Binding Protein 2 (IGFBP2). We show that IGFBP2 overexpression is sufficient to render TNBC cells tolerant to afatinib treatment by activating the compensatory insulin-like growth factor I receptor (IGF1-R) signalling pathway. Finally, based on reconstructed mechanisms of resistance, we employ deep learning techniques to predict the afatinib sensitivity of TNBC cells. Conclusions: Our strategy proved effective in reconstructing the complex signalling network driving EGFR-targeted therapy resistance, offering new insights for the development of individualized treatment strategies in TNBC. [ABSTRACT FROM AUTHOR]

Background: Unlike in lower vertebrates, Müller glia (MG) in adult mammalian retinas lack the ability to reprogram into neurons after retinal injury or degeneration and exhibit reactive gliosis instead. Whether a transition in MG cell fate from gliosis to reprogramming would help preserve photoreceptors is still under exploration. Methods: A mouse model of retinitis pigmentosa (RP) was established using MG cell lineage tracing mice by intraperitoneal injection of sodium iodate (SI). The critical time point for the fate determination of MG gliosis was determined through immunohistochemical staining methods. Then, bulk-RNA and single-cell RNA seq techniques were used to elucidate the changes in RNA transcription of the retina and MG at that time point, and new genes that may determine the fate transition of MG were screened. Finally, the selected gene was specifically overexpressed in MG cells through adeno-associated viruses (AAV) in the mouse RP model. Bulk-RNA seq technique, immunohistochemical staining methods, and visual function testing were used to elucidate and validate the mechanism of new genes function on MG cell fate transition and retinal function. Results: Here, we found the critical time point for MG gliosis fate determination was 3 days post SI injection. Hmga2 was screened out as a candidate regulator for the cell fate transition of MG. After retinal injury caused by SI, the Hmga2 protein is temporarily and lowly expressed in MG cells. Overexpression of Hmga2 in MG down-regulated glial cell related genes and up-regulated photoreceptor related genes. Besides, overexpressing Hmga2 exclusively to MG reduced MG gliosis, made MG obtain cone's marker, and retained visual function in mice with acute retinal injury. Conclusion: Our results suggested the unique reprogramming properties of Hmga2 in regulating the fate transition of MG and neuroprotective effects on the retina with acute injury. This work uncovers the reprogramming ability of epigenetic factors in MG. [ABSTRACT FROM AUTHOR]

Cochlear melanocytes are intermediate cells in the stria vascularis that generate endocochlear potentials required for auditory function. Human PAX3 mutations cause Waardenburg syndrome and abnormalities of skin and retinal melanocytes, manifested as congenital hearing loss (~ 70%) and hypopigmentation of skin, hair and eyes. However, the underlying mechanism of hearing loss remains unclear. Cochlear melanocytes in the stria vascularis originated from Pax3-traced melanoblasts and Plp1-traced Schwann cell precursors, both of which derive from neural crest cells. Here, using a Pax3-Cre knock-in mouse that allows lineage tracing of Pax3-expressing cells and disruption of Pax3, we found that Pax3 deficiency causes foreshortened cochlea, malformed vestibular apparatus, and neural tube defects. Lineage tracing and in situ hybridization show that Pax3+ derivatives contribute to S100+, Kir4.1+ and Dct+ melanocytes (intermediate cells) in the developing stria vascularis, all of which are significantly diminished in Pax3 mutant animals. Taken together, these results suggest that Pax3 is required for the development of neural crest cell-derived cochlear melanocytes, whose absence may contribute to congenital hearing loss of Waardenburg syndrome in humans. [ABSTRACT FROM AUTHOR]

In various biological contexts, cells receive signals and stimuli that prompt them to change their current state, leading to transitions into a future state. This change underlies the processes of development, tissue maintenance, immune response, and the pathogenesis of various diseases. Following the path of cells from their initial identity to their current state reveals how cells adapt to their surroundings and undergo transformations to attain adjusted cellular states. DNA-based molecular barcoding technology enables the documentation of a phylogenetic tree and the deterministic events of cell lineages, providing the mechanisms and timing of cell lineage commitment that can either promote homeostasis or lead to cellular dysregulation. This review comprehensively presents recently emerging molecular recording technologies that utilize CRISPR/Cas systems, base editing, recombination, and innate variable sequences in the genome. Detailing their underlying principles, applications, and constraints paves the way for the lineage tracing of every cell within complex biological systems, encompassing the hidden steps and intermediate states of organism development and disease progression. [ABSTRACT FROM AUTHOR]

This article discusses the limitations of using rodent models in heart transplantation research and highlights the shift towards using human specimens for discovery research. The authors focus on a study that uses single-nucleus RNA sequencing to investigate the cellular landscape of transplanted pediatric hearts. The study identifies important candidate mediators of heart transplant failure, such as graft-infiltrating macrophages and natural killer cells. The authors also explore the dynamics of immune cells within the failing transplanted heart and highlight the potential therapeutic targets that could be investigated further. However, the study has limitations due to the small sample size and the difficulty in obtaining healthy heart transplant tissue. Overall, the article emphasizes the potential of single-cell sequencing and other molecular pathology techniques in advancing our understanding of heart transplantation. [Extracted from the article]

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]

Insect halteres, as specialised hind wings, play an important role during aerial manoeuvres. In Drosophila, halteres and wings are homologous appendages with different morphology. Previous studies have focused on the metamorphosis of halteres, while current knowledge about its cell lineage and regional compartmentalization is still limited. In this study, we performed cell‐lineage tracing of canonical landmark signals in halteres and present a simple model for haltere development. Cell lineage tracing in wings was used as a reference. The nub showed wing‐like expressions in halteres, whereas hth and pnr exhibited different expressions in adult wings and halteres. The lineage tracing revealed that the pouch region gives rise to end‐bulb, and hinge cells contribute to proximal haltere formation. Moreover, we demonstrated that twi‐expressing cells participate in the cell population of the distal end‐bulb. Haematoxylin and eosin staining indicated that muscle cells were present at the distal end‐bulb. These results indicated that adult halteres displayed unique cell lineage patterns and the muscle cells are important components of end‐bulbs. [ABSTRACT FROM AUTHOR]

Research in neuroscience has greatly benefited from the development of genetic approaches that enable lineage tracing, cell type targeting, and conditional gene regulation. Recent advances in combinatorial strategies, which integrate multiple cellular features, have significantly enhanced the spatiotemporal precision and flexibility of these manipulations. In this minireview, we introduce the concept and design of these strategies and provide a few examples of their application in genetic fate mapping, cell type targeting, and reversible conditional gene regulation. These advancements have facilitated in‐depth investigation into the developmental principles underlying the assembly of brain circuits, granting experimental access to highly specific cell lineages and subtypes, as well as offering valuable new tools for modeling and studying neurological diseases. Additionally, we discuss future directions aimed at expanding and improving the existing genetic toolkit for a better understanding of the development, structure, and function of healthy and diseased brains. [ABSTRACT FROM AUTHOR]

Glioma-associated oncogene homolog 1 (Gli1) is a transcriptional activator of hedgehog (Hh) signaling that regulates target gene expression and several cellular biological processes. Cell lineage tracing techniques have highlighted Gli1 as an ideal marker for mesenchymal stem cells (MSCs) in vivo. Gli1+ MSCs are critical for the osteogenesis of the craniofacial bone; however, the regulatory mechanism by which Gli1+ MSCs mediate the bone development and tissue regeneration of craniofacial bone has not been systematically outlined. This review comprehensively elucidates the specific roles of Gli1+ MSCs in craniofacial bone osteogenesis. In addition to governing craniofacial bone development, Gli1+ MSCs are associated with the tissue repair of craniofacial bone under pathological conditions. Gli1+ MSCs promote intramembranous and endochondral ossification of the craniofacial bones, and assist the osteogenesis of the craniofacial bone by improving angiopoiesis. This review summarizes the novel role of Gli1+ MSCs in bone development and tissue repair in craniofacial bones, which offers new insights into bone regeneration therapy. [ABSTRACT FROM AUTHOR]

GABAergic interneurons are key regulators of cortical circuit function. Among the dozens of reported transcriptionally distinct subtypes of cortical interneurons, neurogliaform cells (NGCs) are unique: they are recruited by long-range excitatory inputs, are a source of slow cortical inhibition and are able to modulate the activity of large neuronal populations. Despite their functional relevance, the developmental emergence and diversity of NGCs remains unclear. Here, by combining single-cell transcriptomics, genetic fate mapping, and electrophysiological and morphological characterization, we reveal that discrete molecular subtypes of NGCs, with distinctive anatomical and molecular profiles, populate the mouse neocortex. Furthermore, we show that NGC subtypes emerge gradually through development, as incipient discriminant molecular signatures are apparent in preoptic area (POA)-born NGC precursors. By identifying NGC developmentally conserved transcriptional programs, we report that the transcription factor Tox2 constitutes an identity hallmark across NGC subtypes. Using CRISPR-Cas9-mediated genetic loss of function, we show that Tox2 is essential for NGC development: POA-born cells lacking Tox2 fail to differentiate into NGCs. Together, these results reveal that NGCs are born from a spatially restricted pool of Tox2+ POA precursors, after which intra-type diverging molecular programs are gradually acquired post-mitotically and result in functionally and molecularly discrete NGC cortical subtypes. [ABSTRACT FROM AUTHOR]

Outflow tract (OFT) develops from cardiac progenitor cells in the second heart field (SHF) domain. APJ, a G-Protein Coupled Receptor, is expressed by cardiac progenitors in the SHF. By lineage tracing APJ+SHF cells, we show that these cardiac progenitors contribute to the cells of OFT, which eventually give rise to aorta and pulmonary trunk/artery upon its morphogenesis. Furthermore, we show that early APJ ​+ ​cells give rise to both aorta and pulmonary cells but late APJ ​+ ​cells predominantly give rise to pulmonary cells. APJ is expressed by the outflow tract progenitors in the SHF but its role is unclear. We performed knockout studies to determine the role of APJ in SHF cell proliferation and survival. Our data suggested that APJ knockout in the SHF reduced the proliferation of SHF progenitors, while there was no significant impact on survival. In addition, we show that ectopic overexpression of WNT in these cells disrupted aorta and pulmonary morphogenesis from OFT. Overall, our study has identified APJ ​+ ​progenitor population within the SHF that give rise to aorta and pulmonary trunk/artery cells. Furthermore, we show that APJ signaling stimulates proliferation of these cells in the SHF. [Display omitted] • APJ is expressed by cells within the second heart field (SHF) region. • APJ+ cells in the SHF give rise to the cells of the outflow tract (OFT). • Early APJ+ cells give rise to the cells of both the aorta and pulmonary trunk. • Late APJ+ cells predominantly give rise to the cells of pulmonary trunk. • APJ stimulates the proliferation of APJ+ cardiac progenitor cells in the SHF. [ABSTRACT FROM AUTHOR]

Clonal expansion and development of immunological memory are two hallmarks of adaptive immune responses. Resolving the intricate pathways that regulate cell cycle activity and lead to the generation of diverse effector and memory T cell subsets is essential for improving our understanding of protective T cell immunity. A deeper knowledge of cell cycle regulation in T cells also has translational implications for adoptive cell therapies and vaccinations against infectious diseases. Here, we summarize recent evidence for an early diversification of effector and memory CD8+ T cell fates and discuss how this process is coupled to discrete changes in division speed. We further review technical advances in lineage tracing and cell cycle analysis and outline how these techniques have shed new light on the population dynamics of CD8+ T cell responses, thereby refining our current understanding of the developmental organization of the memory T cell pool. [ABSTRACT FROM AUTHOR]

The thick ascending limb (TAL) is critical for renal control of fluid and ion homeostasis. The function of the TAL depends on the activity of the bumetanide-sensitive Na þ-Kþ-2Cl-cotransporter (NKCC2), which is highly abundant in the luminal membrane of TAL cells. TAL function is regulated by various hormonal and nonhormonal factors. However, many of the underlying signal transduction pathways remain elusive. Here, we describe and characterize a novel gene-modified mouse model for an inducible and specific Cre/Lox-mediated gene modification in the TAL. In these mice, tamoxifen-dependent Cre (CreERT2) was inserted into the 30-untranslated region of the Slc12a1 gene, which encodes NKCC2 (Slc12a1-CreERT2). Although this gene modification strategy slightly reduced endogenous NKCC2 expression at the mRNA and protein levels, the lowered NKCC2 abundance was not associated with altered urinary fluid and ion excretion, urinary concentration, and the renal response to loop diuretics. Immunohistochemistry on kidneys from Slc12a1-CreERT2 mice revealed strong Cre expression exclusively in TAL cells but not in any other nephron portion. Cross-breeding of these mice with the mT/mG reporter mouse line showed a very low recombination rate (-0% in male mice and <3% in female mice) at baseline but complete (-100%) recombination after repeated tamoxifen administration in male and female mice. The achieved recombination encompassed the entire TAL and also included the macula densa. Thus, the new Slc12a1-CreERT2 mouse line allows inducible and very efficient gene targeting in the TAL and hence promises to be a powerful tool to advance our understanding of the regulation of TAL function. [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]

Down syndrome (DS) is a genetic disorder caused by an extra copy of chromosome 21 that presents developmental dysfunction and intellectual disability. To better understand the cellular changes associated with DS, we investigated the cell composition in blood, brain, and buccal swab samples from DS patients and controls using DNA methylation-based cell-type deconvolution. We used genome-scale DNA methylation data from Illumina HumanMethylation450k and HumanMethylationEPIC arrays to profile cell composition and trace fetal lineage cells in blood samples (DS N = 46; control N = 1469), brain samples from various regions (DS N = 71; control N = 101), and buccal swab samples (DS N = 10; control N = 10). In early development, the number of cells from the fetal lineage in the blood is drastically lower in DS patients (Δ = 17.5%), indicating an epigenetically dysregulated maturation process for DS patients. Across sample types, we observed significant alterations in relative cell-type proportions for DS subjects compared with the controls. Cell-type proportion alterations were present in samples from early development and adulthood. Our findings provide insight into DS cellular biology and suggest potential cellular interventional targets for DS. [ABSTRACT FROM AUTHOR]

Agonists of trained immunity induce epigenetic changes in hematopoietic stem and progenitor cells (HSPCs) to generate long-lasting immune protection. Although trained HSPCs generate myeloid cells with increased responsiveness to secondary challenges, whether their differentiation kinetics is affected by prior exposure to inducers of trained immunity remains elusive. Here, we used lineage tracing to examine the cell fates of endothelial protein C receptor-positive hematopoietic stem cells (EPCR+ HSCs) and fms-like tyrosine kinase 3-positive multipotent progenitor cells (Flt3+ MPPs) in β-glucan-induced trained immunity. We found that although β-glucan triggered the expected expansion of myeloid progenitors, the differentiation behaviors of EPCR+ HSCs and Flt3+ MPPs in multiple cycles of hematopoietic regeneration were hardly affected. Thus, our results rule out changed kinetics in cell differentiation by EPCR+ HSC and Flt3+ MPP as the cause of enhanced myelopoiesis upon secondary immune challenges. [ABSTRACT FROM AUTHOR]

Endothelial–mesenchymal transition (EndMT) drives the endothelium to contribute to vascular calcification in diabetes mellitus. In our previous study, we showed that glycogen synthase kinase-3β (GSK3β) inhibition induces β-catenin and reduces mothers against DPP homolog 1 (SMAD1) to direct osteoblast-like cells toward endothelial lineage, thereby reducing vascular calcification in Matrix Gla Protein (Mgp) deficiency. Here, we report that GSK3β inhibition reduces vascular calcification in diabetic Ins2Akita/wt mice. Cell lineage tracing reveals that GSK3β inhibition redirects endothelial cell (EC)-derived osteoblast-like cells back to endothelial lineage in the diabetic endothelium of Ins2Akita/wt mice. We also find that the alterations in β-catenin and SMAD1 by GSK3β inhibition in the aortic endothelium of diabetic Ins2Akita/wt mice are similar to Mgp−/− mice. Together, our results suggest that GSK3β inhibition reduces vascular calcification in diabetic arteries through a similar mechanism to that in Mgp−/− mice. [ABSTRACT FROM AUTHOR]

BACKGROUND: Spinal cord ependymal cells exhibit neural stem/progenitor cell properties after injury in adult mammalian. OBJECTIVE: Ependymal cells were labeled with Nestin and Foxj1 transgenic mice to track the proliferation and differentiation fate of ependymal cells and their progeny after spinal cord injury in adult mice. METHODS: A 1-mm section was completely removed from the T8 segment of the spinal cord in transgenic mice. Ependymal cells were dynamically observed by continuous intraperitoneal injection of BrdU at 1-7 days after spinal cord injury. At different time points (3, 7, 14, 28, 56 days) after spinal cord injury, the genetic fate mapping of ependymal cells in the lesion edge was observed by immunofluorescence staining of BrdU, GFAP, Tuj1, and NeuN. RESULTS AND CONCLUSION: (1) In the uninjured spinal cord, Nestin-positive ependymal cells were quiescent. Ependymal cells were activated and proliferated around the lesion edge at 3 days after spinal cord injury. (2) Approximately 3.3% of Nestin-positive ependymal cells expressed neuronal marker Tuj1 at 28 days after spinal cord injury. (3) At 56 days after injury, approximately 25.7% of Nestin-positive ependymal cells differentiated into astrocytes and formed the core of glial scar, participating in the formation of glial scar. (4) This article can provide theoretical basis for understanding the pathological process and new ideas for spinal cord injury repair by detecting the spatiotemporal changes, proliferation and differentiation characteristics of ependymal cells after spinal cord injury. [ABSTRACT FROM AUTHOR]

Background: CD34+ cells have been used to treat the patients with heart failure, but the outcome is variable. It is of great significance to scrutinize the fate and the mechanism of CD34+ cell differentiation in vivo during heart failure and explore its intervention strategy. Methods: We performed single-cell RNA sequencing (scRNA-seq) of the total non-cardiomyocytes and enriched Cd34-tdTomato+ lineage cells in the murine (male Cd34-CreERT2; Rosa26-tdTomato mice) pressure overload model (transverse aortic constriction, TAC), and total non-cardiomyocytes from human adult hearts. Then, in order to determine the origin of CD34+ cell that plays a role in myocardial fibrosis, bone marrow transplantation model was performed. Furthermore, to further clarify the role of CD34 + cells in myocardial remodeling in response to TAC injury, we generated Cd34-CreERT2; Rosa26-eGFP-DTA (Cre/DTA) mice. Results: By analyzing the transcriptomes of 59,505 single cells from the mouse heart and 22,537 single cells from the human heart, we illustrated the dynamics of cell landscape during the progression of heart hypertrophy, including CD34+ cells, fibroblasts, endothelial and immune cells. By combining genetic lineage tracing and bone marrow transplantation models, we demonstrated that non-bone-marrow-derived CD34+ cells give rise to fibroblasts and endothelial cells, while bone-marrow-derived CD34+ cell turned into immune cells only in response to pressure overload. Interestingly, partial depletion of CD34+ cells alleviated the severity of myocardial fibrosis with a significant improvement of cardiac function in Cd34-CreERT2; Rosa26-eGFP-DTA model. Similar changes of non-cardiomyocyte composition and cellular heterogeneity of heart failure were also observed in human patient with heart failure. Furthermore, immunostaining showed a double labeling of CD34 and fibroblast markers in human heart tissue. Mechanistically, our single-cell pseudotime analysis of scRNA-seq data and in vitro cell culture study revealed that Wnt-β-catenin and TGFβ1/Smad pathways are critical in regulating CD34+ cell differentiation toward fibroblasts. Conclusions: Our study provides a cellular landscape of CD34+ cell-derived cells in the hypertrophy heart of human and animal models, indicating that non-bone-marrow-derived CD34+ cells differentiating into fibroblasts largely account for cardiac fibrosis. These findings may provide novel insights for the pathogenesis of cardiac fibrosis and have further potential therapeutic implications for the heart failure. [ABSTRACT FROM AUTHOR]

Background: We previously identified Leucine-rich repeats and immunoglobulin-like domains 1 (Lrig1) as a marker of long-term neurogenic stem cells in the lateral wall of the adult mouse brain. The morphology of the stem cells thus identified differed from the canonical B1 type stem cells, raising a question about their cellular origin. Thus, we investigated the development of these stem cells in the postnatal and juvenile brain. Furthermore, because Lrig1 is a known regulator of quiescence, we also investigated the effect(s) of its deletion on the cellular proliferation in the lateral wall. Methods: To observe the development of the Lrig1-lineage stem cells, genetic inducible fate mapping studies in combination with thymidine analog administration were conducted using a previously published Lrig1T2A-iCreERT2 mouse line. To identify the long-term consequence(s) of Lrig1 germline deletion, old Lrig1 knock-out mice were generated using two different Lrig1 null alleles in the C57BL/6J background. The lateral walls from these mice were analyzed using an optimized whole mount immunofluorescence protocol and confocal microscopy. Results: We observed the Lrig1-lineage labeled cells with morphologies consistent with neurogenic stem cell identity in postnatal, juvenile, and adult mouse brains. Interestingly, when induced at postnatal or juvenile ages, morphologically distinct cells were revealed, including cells with the canonical B1 type stem cell morphology. Almost all of the presumptive stem cells labeled were non-proliferative at these ages. In the old Lrig1 germline knock-out mice, increased proliferation was observed compared to wildtype littermates without concomitant increase in apoptosis. Conclusions: Once set aside during embryogenesis, the Lrig1-lineage stem cells remain largely quiescent during postnatal and juvenile development until activation in adult age. The absence of premature proliferative exhaustion in the Lrig1 knock-out stem cell niche during aging is likely due to a complex cascade of effects on the adult stem cell pool. Thus, we suggest that the adult stem cell pool size may be genetically constrained via Lrig1. [ABSTRACT FROM AUTHOR]

Tissue-resident macrophages are essential to protect from pathogen invasion and maintain organ homeostasis. The ability of thymic macrophages to engulf apoptotic thymocytes is well appreciated, but little is known about their ontogeny, maintenance, and diversity. Here, we characterized the surface phenotype and transcriptional profile of these cells and defined their expression signature. Thymic macrophages were most closely related to spleen red pulp macrophages and Kupffer cells and shared the expression of the transcription factor (TF) SpiC with these cells. Single-cell RNA sequencing (scRNA-Seq) showed that the macrophages in the adult thymus are composed of two populations distinguished by the expression of Timd4 and Cx3cr1. Remarkably, Timd4+ cells were located in the cortex, while Cx3cr1+ macrophages were restricted to the medulla and the cortico-medullary junction. Using shield chimeras, transplantation of embryonic thymuses, and genetic fate mapping, we found that the two populations have distinct origins. Timd4+ thymic macrophages are of embryonic origin, while Cx3cr1+ macrophages are derived from adult hematopoietic stem cells. Aging has a profound effect on the macrophages in the thymus. Timd4+ cells underwent gradual attrition, while Cx3cr1+ cells slowly accumulated with age and, in older mice, were the dominant macrophage population in the thymus. Altogether, our work defines the phenotype, origin, and diversity of thymic macrophages. [ABSTRACT FROM AUTHOR]

The article presents a study on T-box transcription factor 3 (Tbx3)-derived neuronal lineage and Tbx3-dependent neuronal specification which found that Tbx3 hierarchically established and maintained the identity of KNDy neurons for triggering puberty. Topics include effect of Tbx3 deficiency on the identity establishment of KNDy, specific deletion of Tbx3 in KNDy neurons which undermines their identity maintenance, and significance of identity maintenance of KNDy neurons for puberty onset.

The article discusses a 2022 study on a minority population of LMO2+ breast cancer cells that integrate into the vasculature and initiate metastasis. Topics covered include the reduction of lung metastasis by LMO2 knockdown in human breast tumors, and the binding of LMO2 to STAT3 that is required for the latter's activation. Also noted is the identification of cells with angiogenic features that establishes the LMO2-STAT3 signaling axis as a therapeutic target in breast cancer metastasis.

The oligodendrocyte (OL) lineage transcription factor Olig2 is expressed throughout oligodendroglial development and is essential for oligodendroglial progenitor specification and differentiation. It was previously reported that deletion of Olig2 enhanced the maturation and myelination of immature OLs and accelerated the remyelination process. However, by analyzing multiple Olig2 conditional KO mouse lines (male and female), we conclude that Olig2 has the opposite effect and is required for OL maturation and remyelination. We found that deletion of Olig2 in immature OLs driven by an immature OL-expressing Plp1 promoter resulted in defects in OL maturation and myelination, and did not enhance remyelination after demyelination. Similarly, Olig2 deletion during premyelinating stages in immature OLs using Mobp or Mog promoter-driven Cre lines also did not enhance OL maturation in the CNS. Further, we found that Olig2 was not required for myelin maintenance in mature OLs but was critical for remyelination after lysolecithin-induced demyelinating injury. Analysis of genomic occupancy in immature and mature OLs revealed that Olig2 targets the enhancers of key myelination-related genes for OL maturation from immature OLs. Together, by leveraging multiple immature OL-expressing Cre lines, these studies indicate that Olig2 is essential for differentiation and myelination of immature OLs and myelin repair. Our findings raise fundamental questions about the previously proposed role of Olig2 in opposing OL myelination and highlight the importance of using Cre-dependent reporter(s) for lineage tracing in studying cell state progression. [ABSTRACT FROM AUTHOR]

The article presents a developmental neuroscience research on how tissue-wide genetic and cellular landscape shapes the execution of sequential PRC2 functions in neural stem cell lineage progression. Topics include critical role for the genetic and cellular niche environment in neural stem cell behavior; and , cortical astrocyte production and maturation is cell-autonomously controlled by PRC2-dependent transcriptional regulation.

The absence of a conventional association between the cell–cell cohabitation and its emergent dynamics into cliques during development has hindered our understanding of how cell populations proliferate, differentiate, and compete (i.e., the cell ecology). With the recent advancement of single-cell RNA sequencing (RNA-seq), we can potentially describe such a link by constructing network graphs that characterize the similarity of the gene expression profiles of the cell-specific transcriptional programs and analyze these graphs systematically using the summary statistics given by the algebraic topology. We propose single-cell topological simplicial analysis (scTSA). Applying this approach to the single-cell gene expression profiles from local networks of cells in different developmental stages with different outcomes reveals a previously unseen topology of cellular ecology. These networks contain an abundance of cliques of single-cell profiles bound into cavities that guide the emergence of more complicated habitation forms. We visualize these ecological patterns with topological simplicial architectures of these networks, compared with the null models. Benchmarked on the single-cell RNA-seq data of zebrafish embryogenesis spanning 38,731 cells, 25 cell types, and 12 time steps, our approach highlights gastrulation as the most critical stage, consistent with the consensus in developmental biology. As a nonlinear, model-independent, and unsupervised framework, our approach can also be applied to tracing multi-scale cell lineage, identifying critical stages, or creating pseudo-time series. [ABSTRACT FROM AUTHOR]

CELL DEATH Cytotoxic pores require the right splice Granzyme A (GzmA) is one of the cytotoxic granule proteins that killer lymphocytes deploy to kill targeted tumor cells. MARINE PRODUCTIVITY A dusting of nutrients Most of the nutrients that fuel primary production in the oceans come from water upwelled to the surface from deeper depths, but atmospheric aerosols have long been recognized as another potential source. Cyclical tissue strain from normal breathing movements was required to maintain cell fate in the lung alveolus, and loss of these movements led to AT1-AT2 cell reprogramming. [Extracted from the article]

We previously showed the importance of TGFβ signaling in development of the mouse axial skeleton. Here, we provide the first direct evidence that TGFβ signaling is required for resegmentation of the sclerotome using chick embryos. Lipophilic fluorescent tracers, DiO and DiD, were microinjected into adjacent somites of embryos treated with or without TGFβRI inhibitors, SB431542, SB525334 or SD208, at developmental day E2.5 (HH16). Lineage tracing of labeled cells was observed over the course of 4 days until the completion of resegmentation at E6.5 (HH32). Vertebrae were malformed and intervertebral discs were small and misshapen in inhibitor injected embryos. Hypaxial myofibers were also increased in thickness after treatment with the inhibitor. Inhibition of TGFβ signaling resulted in alterations in resegmentation that ranged between full, partial, and slanted shifts in distribution of DiO or DiD labeled cells within vertebrae. Patterning of rostro-caudal markers within sclerotome was disrupted at E3.5 after treatment with TGFβRI inhibitor with rostral domains expressing both rostral and caudal markers. We propose that TGFβ signaling regulates rostro-caudal polarity and subsequent resegmentation in sclerotome during spinal column development. [Display omitted] • Injection of TGFBR1 inhibitor in chick somites resulted in defects in axial skeleton. • TGFβ signaling was required for normal resegmentation of sclerotome. • Inhibition of TGFβ signaling resulted in altered sclerotome polarity. • Lineage tracer dyes and drug inhibitors can be used to study somite development in chick. [ABSTRACT FROM AUTHOR]

Although several observations suggest that the constitutive biological, genetic or physiological changes leading to autism spectrum disorders (ASD) start in utero, their early impact on the number and density of neurons in the brain remains unknown. Using genetic fate mapping associated with the immunollabeling‐enabled three‐dimensional imaging of solvent‐cleared organs (iDISCO) clearing method we identified and counted a selective population of neocortical and hippocampal pyramidal neurons in the in utero valproate (VPA) mouse model of autism. We report that 1 day before birth, the number of pyramidal neurons born at E14.5 in the neocortex and hippocampus of VPA mice is smaller than in age‐matched controls. VPA also induced a reduction of the neocortical—but not hippocampal—volume 1 day before birth. Interestingly, VPA mice present an increase in both neocortical and hippocampal volumes 2 days after birth compared with controls. These results suggest that the VPA‐exposed hippocampus and neocortex differ substantially from controls during the highly complex perinatal period, and specially 1 day before birth, reflecting the early pathogenesis of ASD. [ABSTRACT FROM AUTHOR]

Structural changes known as airway remodeling characterize chronic/severe asthma and contribute to lung dysfunction. We previously reported that neonatal SSEA-1+ pulmonary stem/progenitor cells (PSCs) ameliorated airway inflammation in asthmatic mice. However, the molecular mechanisms by which endogenous SSEA-1+ PSC of adult mice afford beneficial effects in alveolar homeostasis and lung repair after allergen challenge remain incompletely understood. To analyze the expression profile and clarify the biological significance of endogenous adult lung SSEA-1+ cells in asthmatic mice. Lung SSEA-1+ cells and circulating SSEA-1+ cells in peripheral blood were determined by confocal microscopy and cytometric analysis. GFP chimeric mice were used to trace cell lineage in vivo. The roles of circulating SSEA-1+ cells were verified in ovalbumin-induced and house dust mite-induced allergic asthmatic models. In asthmatic mice, endogenous lung SSEA-1+ cells almost disappeared; however, a unique population of circulating SSEA-1+ cells was enriched after the challenge phase. In asthmatic mice, adoptive transfer of circulating SSEA-1+ cells had a specific homing preference for the lung in response to inhaled antigen through upregulating CXCR7–CXCL11 chemokine axis. Circulating SSEA-1+ cells can transdifferentiate in the alveolar space and ameliorate lung inflammation and structural damage through inhibiting the infiltration of inflammatory cells into peribronchovascular and goblet cell hyperplasia areas, reducing the thickened smooth muscle layers and PAS-positive mucus-containing goblet cells. Reinforcing bone marrow-derived circulating SSEA-1+ cells from peripheral blood into lung tissue which create a rescue mechanism in maintaining alveolar homeostasis and tissue repair to mediate lung protection for emergency responses after allergen challenge in asthmatic conditions. [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]

Cells of most bacterial species are around 2 micrometers in length, with some of the largest specimens reaching 750 micrometers. Using fluorescence, x-ray, and electron microscopy in conjunction with genome sequencing, we characterized Candidatus (Ca.) Thiomargarita magnifica, a bacterium that has an average cell length greater than 9000 micrometers and is visible to the naked eye. These cells grow orders of magnitude over theoretical limits for bacterial cell size, display unprecedented polyploidy of more than half a million copies of a very large genome, and undergo a dimorphic life cycle with asymmetric segregation of chromosomes into daughter cells. These features, along with compartmentalization of genomic material and ribosomes in translationally active organelles bound by bioenergetic membranes, indicate gain of complexity in the Thiomargarita lineage and challenge traditional concepts of bacterial cells. [ABSTRACT FROM AUTHOR]

Progenitors of the developing human neocortex reside in the ventricular and outer subventricular zones (VZ and OSVZ, respectively). However, whether cells derived from these niches have similar developmental fates is unknown. By performing fate mapping in primary human tissue, we demonstrate that astrocytes derived from these niches populate anatomically distinct layers. Cortical plate astrocytes emerge from VZ progenitors and proliferate locally, while putative white matter astrocytes are morphologically heterogeneous and emerge from both VZ and OSVZ progenitors. Furthermore, via single-cell sequencing of morphologically defined astrocyte subtypes using Patch-seq, we identify molecular distinctions between VZ-derived cortical plate astrocytes and OSVZ-derived white matter astrocytes that persist into adulthood. Together, our study highlights a complex role for cell lineage in the diversification of human neocortical astrocytes. [ABSTRACT FROM AUTHOR]

Vascular establishment is one of the early events in embryogenesis. It is believed that vessel-initiating endothelial progenitors cluster to form the first primitive vessel. Understanding the molecular identity of these progenitors is crucial in order to elucidate lineage hierarchy. In this study, we identify protein C receptor (Procr) as an endothelial progenitor marker and investigate the role of Procr+ progenitors during embryonic vascular development. Using a ProcrmGFP-2A-lacZ reporter, we reveal a much earlier Procr expression (embryonic day 7.5) than previously acknowledged (embryonic day 13.5). Genetic fate-mapping experiments using ProcrCre and ProcrCreER demonstrate that Procr+ cells give rise to blood vessels throughout the entire embryo proper. Single-cell RNA-sequencing analyses place Procr+ cells at the start of endothelial commitment and maturation. Furthermore, targeted ablation of Procr+ cells results in failure of vessel formation and early embryonic lethality. Notably, genetic fate mapping and scRNA-seq pseudotime analysis support the view that Procr+ progenitors can give rise to hemogenic endothelium. In this study, we establish a Procr expression timeline and identify Procr+ vessel-initiating progenitors, and demonstrate their indispensable role in establishment of the vasculature during embryo development. [ABSTRACT FROM AUTHOR]

During bone development and repair, osteoblasts are recruited to bone deposition sites. To identify the origin of recruited osteoblasts, cell lineage tracing using Cre/loxP recombination is commonly used. However, a confounding factor is the use of transgenic Cre drivers that do not accurately recapitulate endogenous gene expression or the use of knock-in Cre drivers that alter endogenous protein activity or levels. Here, we describe a CRISPR/Cas9 homology-directed repair knock-in approach that allows efficient generation of Cre drivers controlled by the endogenous gene promoter. In addition, a self-cleaving peptide preserves the reading frame of the endogenous protein. Using this approach, we generated col10a1p2a-CreERT2 knock-in medaka and show that tamoxifen-inducible CreERT2 efficiently recombined loxP sites in col10a1 cells. Similar knock-in efficiencies were obtained when two unrelated loci (osr1 and col2a1a) were targeted. Using live imaging, we traced the fate of col10a1 osteoblast progenitors during bone lesion repair in the medaka vertebral column. We show that col10a1 cells at neural arches represent a mobilizable cellular source for bone repair. Together, our study describes a previously unreported strategy for precise cell lineage tracing via efficient and non-disruptive knock-in of Cre. [ABSTRACT FROM AUTHOR]

The vertebrate musculoskeletal system is known to be formed by mesenchymal stem cells condensing into tissue elements, which then differentiate into cartilage, bone, tendon/ligament, and muscle cells. These lineage-committed cells mature into end-stage differentiated cells, like hypertrophic chondrocytes and osteocytes, which are expected to expire and to be replaced by newly differentiated cells arising from the same lineage pathway. However, there is emerging evidence of the role of cell transdifferentiation in bone development and disease. Although the concept of cell transdifferentiation is not new, a breakthrough in cell lineage tracing allowed scientists to trace cell fates in vivo. Using this powerful tool, new theories have been established: (1) hypertrophic chondrocytes can transdifferentiate into bone cells during endochondral bone formation, fracture repair, and some bone diseases, and (2) tendon cells, beyond their conventional role in joint movement, directly participate in normal bone and cartilage formation, and ectopic ossification. The goal of this review is to obtain a better understanding of the key roles of cell transdifferentiation in skeletal development and diseases. We will first review the transdifferentiation of chondrocytes to bone cells during endochondral bone formation. Specifically, we will include the history of the debate on the fate of chondrocytes during bone formation, the key findings obtained in recent years on the critical factors and molecules that regulate this cell fate change, and the role of chondrocyte transdifferentiation in skeletal trauma and diseases. In addition, we will also summarize the latest discoveries on the novel roles of tendon cells and adipocytes on skeletal formation and diseases. [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]

The article discusses that Gonadal sex determination represents a unique model for studying cell fate decisions. Complete understanding of the different cell lineages forming the developing testis and ovary remains elusive. The research was conducted on the origin, specification, and subsequent sex-specific differentiation of a previously uncharacterized population of supporting-like cells (SLCs) in the developing mouse gonads.

Reconstructing the development of lineage relationships and cell fate mapping has been a fundamental problem in biology. Using advanced molecular biology and single-cell RNA sequencing, we have profiled transcriptomes at the single-cell level and mapped cell fates during development. Recently, CRISPR/Cas9 barcode editing for large-scale lineage tracing has been used to reconstruct the pseudotime trajectory of cells and improve lineage tracing accuracy. This review presents the progress of the latest CbLT (CRISPR-based Lineage Tracing) and discusses the current limitations and potential technical pitfalls in their application and other emerging concepts. [ABSTRACT FROM AUTHOR]

Background: Tamoxifen is widely used for Cre‐estrogen receptor‐mediated genomic recombination in transgenic mouse models to mark cells for lineage tracing and to study gene function. However, recent studies have highlighted off‐target effects of tamoxifen in various tissues and cell types when used for induction of Cre recombination. Despite the widespread use of these transgenic Cre models to assess gastrointestinal (GI) function, the effect of tamoxifen exposure on GI motility has not been described. Methods: We examined the effects of tamoxifen on GI motility by measuring total GI transit, gastric emptying, small intestinal transit, and colonic contractility in wild‐type adult mice. Key Results: We observed a significant delay in total GI transit in tamoxifen‐treated mice, with unaltered gastric emptying, accelerated small intestinal transit, and abnormal colonic motility. Conclusion: Our findings highlight the importance of considering GI motility alterations induced by tamoxifen when designing protocols that utilize tamoxifen as a Cre‐driver for studying GI function. [ABSTRACT FROM AUTHOR]

Accurate discovery of somatic mutations in a cell is a challenge that partially lays in immaturity of dedicated analytical approaches. Approaches comparing a cell's genome to a control bulk sample miss common mutations, while approaches to find such mutations from bulk suffer from low sensitivity. We developed a tool, All2, which enables accurate filtering of mutations in a cell without the need for data from bulk(s). It is based on pair-wise comparisons of all cells to each other where every call for base pair substitution and indel is classified as either a germline variant, mosaic mutation, or false positive. As All2 allows for considering dropped-out regions, it is applicable to whole genome and exome analysis of cloned and amplified cells. By applying the approach to a variety of available data, we showed that its application reduces false positives, enables sensitive discovery of high frequency mutations, and is indispensable for conducting high resolution cell lineage tracing. Author summary: DNA make up in cells of a human is slightly different from one another because of cell specific mutations called mosaic mutations. Mosaic mutations can be introduced during early development in a fetus, during normal cell division throughout life or during an aggressive onset of cell division such as cancer. Thus, the extent of accumulation, time of acquiring and specific location of mosaic mutations in the genome are vital for the understanding of the biology of normal development as well as diseases that are caused by it. The ultimate way of discovering and analyzing mosaic mutations is by studying the genome of single cells. Our method, All2, uses a novel approach to compare the genome of single cells to one another to accurately recognize true mosaic mutations from natural variations and from noise by implementing a unique scoring method. We have shown that our method performs better than discovery of mutations from a bulk or from comparing cells to a bulk. We have applied the method to high resolution cell lineage tracing and demonstrated it superb performance for reconstructing individualized cell ancestry trees starting from the zygote. [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]

Pancreatic ductal metaplasia (PDM) is the transformation of potentially various types of cells in the pancreas into ductal or ductal-like cells, which eventually replace the existing differentiated somatic cell type(s). PDM is usually triggered by and manifests its ability to adapt to environmental stimuli and genetic insults. The development of PDM to atypical hyperplasia or dysplasia is an important risk factor for pancreatic intraepithelial neoplasia (PanIN) and pancreatic ductal adenocarcinoma (PDA). Recent studies using genetically engineered mouse models, cell lineage tracing, single-cell sequencing and others have unraveled novel cellular and molecular insights in PDM formation and evolution. Those novel findings help better understand the cellular origins and functional significance of PDM and its regulation at cellular and molecular levels. Given that PDM represents the earliest pathological changes in PDA initiation and development, translational studies are beginning to define PDM-associated cell and molecular biomarkers that can be used to screen and detect early PDA and to enable its effective intervention, thereby truly and significantly reducing the dreadful mortality rate of PDA. This review will describe recent advances in the understanding of PDM biology with a focus on its underlying cellular and molecular mechanisms, and in biomarker discovery with clinical implications for the management of pancreatic regeneration and tumorigenesis. [ABSTRACT FROM AUTHOR]

Background: DCLEAR is an R package used for single cell lineage reconstruction. The advances of CRISPR-based gene editing technologies have enabled the prediction of cell lineage trees based on observed edited barcodes from each cell. However, the performance of existing reconstruction methods of cell lineage trees was not accessed until recently. In response to this problem, the Allen Institute hosted the Cell Lineage Reconstruction Dream Challenge in 2020 to crowdsource relevant knowledge from across the world. Our team won sub-challenges 2 and 3 in the challenge competition. Results: The DCLEAR package contained the R codes, which was submitted in response to sub-challenges 2 and 3. Our method consists of two steps: (1) distance matrix estimation and (2) the tree reconstruction from the distance matrix. We proposed two novel methods for distance matrix estimation as outlined in the DCLEAR package. Using our method, we find that two of the more sophisticated distance methods display a substantially improved level of performance compared to the traditional Hamming distance method. DCLEAR is open source and freely available from R CRAN and from under the GNU General Public License, version 3. Conclusions: DCLEAR is a powerful resource for single cell lineage reconstruction. [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]

The article presents the findings of the study which suggest that PDGFRA signaling is implicated in the pathogenesis, and inhibition of this pathway is a potential therapeutic strategy in FLNC-related cardiomyopathies. It mentions that FLNC truncating mutations (FLNCtv) are prevalent causes of inherited dilated cardiomyopathy (DCM), with a high risk of developing arrhythmogenic cardiomyopathy.