Biology subjects

Rando, T. A.

Publications and source records attributed to Rando, T. A..

11 recordsLinked to original sources

bioRxiv2024

Spatiotemporal transcriptomic profiling and modeling of mouse brain at single-cell resolution reveals cell proximity effects of aging and rejuvenation

Old age is associated with a decline in cognitive function and an increase in neurodegenerative disease risk1. Brain aging is complex and accompanied by many cellular changes2-20. However, the influence that aged cells have on neighboring cells and how this contributes to tissue decline is unknown. More generally, the tools to systematically address this question in aging tissues have not yet been developed. Here, we generate spatiotemporal data at single-cell resolution for the mouse brain across lifespan, and we develop the first machine learning models based on spatial transcriptomics ( spatial aging clocks) to reveal cell proximity effects during brain aging and rejuvenation. We collect a single-cell spatial transcriptomics brain atlas of 4.2 million cells from 20 distinct ages and across two rejuvenating interventions--exercise and partial reprogramming. We identify spatial and cell type-specific transcriptomic fingerprints of aging, rejuvenation, and disease, including for rare cell types. Using spatial aging clocks and deep learning models, we find that T cells, which infiltrate the brain with age, have a striking pro-aging proximity effect on neighboring cells. Surprisingly, neural stem cells have a strong pro-rejuvenating effect on neighboring cells. By developing computational tools to identify mediators of these proximity effects, we find that pro-aging T cells trigger a local inflammatory response likely via interferon-{gamma} whereas pro-rejuvenating neural stem cells impact the metabolism of neighboring cells possibly via growth factors (e.g. vascular endothelial growth factor) and extracellular vesicles, and we experimentally validate some of these predictions. These results suggest that rare cells can have a drastic influence on their neighbors and could be targeted to counter tissue aging. We anticipate that these spatial aging clocks will not only allow scalable assessment of the efficacy of interventions for aging and disease but also represent a new tool for studying cell-cell interactions in many spatial contexts.

neuroscience↗

bioRxiv2023

A deep-learning-based screening platform in aging-relevant human motor neurons to identify therapeutic compounds for amyotrophic lateral sclerosis

Age is the primary risk factor for most neurodegenerative diseases including amyo-trophic lateral sclerosis (ALS). Despite the clear importance of age on the development and progression of ALS, age is rarely considered a factor in cellular or animal models of ALS. The recent advent of direct reprogramming methodologies, whereby somatic cells such as fibroblasts can be converted into neurons while retaining the age signature of the host, has facilitated the study of age-relevant human cells in vitro for the first time. Despite the promise of this technology, age is a complex, multidimensional, and dy-namic phenotype that makes the interpretation of the interplay between age and dis-ease a great challenge. To circumvent these challenges, we developed a screening platform for directly reprogrammed neurons in an age-relevant human cellular model to better model and identify disease-modifying targets and pathways for ALS. This plat-form uses deep learning image analysis to screen compounds for efficacy against ALS-associated cellular phenotypes and efficiently classifies ALS-like phenotypic signatures as well as predicts the age of the original fibroblast donor. Notably, our work identified NCB-0846, a TNIK/MAP4K7 inhibitor, as a novel compound with the ability to revert ALS-like phenotypes. Moreover, we show that chronic dosing of NCB-0846 in the SOD1G93A mouse model of ALS significantly reduced serum neurofilament-L levels. Our findings establish the power of a platform that combines direct reprogramming of human cells and deep learning as a powerful tool for combatting aging and age-related diseases.

neuroscience↗

bioRxiv2023

Dynamical modeling reveals RNA decay mediates the effect of matrix stiffness on aged muscle stem cell fate

Loss of muscle stem cell (MuSC) self-renewal with aging reflects a combination of influences from the intracellular (e.g., post-transcriptional modifications) and extracellular (e.g., matrix stiffness) environment. Whereas conventional single cell analyses have revealed valuable insights into factors contributing to impaired self-renewal with age, most are limited by static measurements that fail to capture nonlinear dynamics. Using bioengineered matrices mimicking the stiffness of young and old muscle, we showed that while young MuSCs were unaffected by aged matrices, old MuSCs were phenotypically rejuvenated by young matrices. Dynamical modeling of RNA velocity vector fields in silico revealed that soft matrices promoted a self-renewing state in old MuSCs by attenuating RNA decay. Vector field perturbations demonstrated that the effects of matrix stiffness on MuSC self-renewal could be circumvented by fine-tuning the expression of the RNA decay machinery. These results demonstrate that post-transcriptional dynamics dictate the negative effect of aged matrices on MuSC self-renewal. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC="FIGDIR/small/529950v2_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@1a840corg.highwire.dtl.DTLVardef@d354cforg.highwire.dtl.DTLVardef@650556org.highwire.dtl.DTLVardef@41587c_HPS_FORMAT_FIGEXP M_FIG Graphical abstract description: The balance of self-renewal and differentiation in young muscle stem cells (MuSCs) is robust to perturbations of the biophysical microenvironment. In contrast, aged MuSCs are highly sensitive to extrinsic perturbations, and exposure to a youthful microenvironment rejuvenates the self-renewing potential of aged MuSCs by modulating post-transcriptional RNA dynamics. C_FIG

cell biology↗

bioRxiv2022

Death-seq identifies regulators of cell death and senolytic therapies

Selectively ablating senescent cells ("senolysis") is an evolving therapeutic approach for age-related diseases. Current senolytics are limited to local administration by potency and side effects. While genetic screens could identify senolytics, current screens are underpowered for identifying genes that regulate cell death due to limitations in screen methodology. Here, we establish Death-seq, a positive selection CRISPR screen optimized to identify enhancers and mechanisms of cell death. Our screens identified synergistic enhancers of cell death induced by the known senolytic ABT-263, a BH3 mimetic. SMAC mimetics, enhancers in our screens, synergize with ABT-199, another BH3 mimetic that is not senolytic alone, clearing senescent cells in models of age-related disease while sparing human platelets, avoiding the thrombocytopenia associated with ABT-263. Death-seq enables the systematic screening of cell death pathways to uncover molecular mechanisms of regulated cell death subroutines and identify drug targets for diverse pathological states such as senescence, cancer, and neurodegeneration.

cell biology↗

bioRxiv2022

Cell type-specific aging clocks to quantify aging and rejuvenation in regenerative regions of the brain

Aging manifests as progressive dysfunction culminating in death. The diversity of cell types is a challenge to the precise quantification of aging and its reversal. Here we develop a suite of aging clocks based on single cell transcriptomic data to characterize cell type-specific aging and rejuvenation strategies. The subventricular zone (SVZ) neurogenic region contains many cell types and provides an excellent system to study cell-level tissue aging and regeneration. We generated 21,458 single-cell transcriptomes from the neurogenic regions of 28 mice, tiling ages from young to old. With these data, we trained a suite of single cell-based regression models (aging clocks) to predict both chronological age (passage of time) and biological age (fitness, in this case the proliferative capacity of the neurogenic region). Both types of clocks perform well on independent cohorts of mice. Genes underlying the single cell-based aging clocks are mostly cell-type specific, but also include a few shared genes in the interferon and lipid metabolism pathways. We used these single cell-based aging clocks to measure transcriptomic rejuvenation, by generating single cell RNA-seq datasets of SVZ neurogenic regions for two interventions - heterochronic parabiosis (young blood) and exercise. Interestingly, the use of aging clocks reveals that both heterochronic parabiosis and exercise reverse transcriptomic aging in the niche, but in different ways across cell types and genes. This study represents the first development of high-resolution aging clocks from single cell transcriptomic data and demonstrates their application to quantify transcriptomic rejuvenation.

genomics↗

bioRxiv2022

Exercise reprograms the inflammatory landscape of multiple stem cell compartments during mammalian aging

Exercise has the ability to rejuvenate stem cells and improve tissue homeostasis and regeneration in aging animals. However, the cellular and molecular changes elicited by exercise have not been systematically studied across a broad range of cell types in stem cell compartments. To gain better insight into the mechanisms by which exercise affects niche and stem cell function, we subjected young and old mice to aerobic exercise and generated a single cell transcriptomic atlas of muscle, neural and hematopoietic stem cells with their niche cells and progeny. Complementarily, we also performed whole transcriptome analysis of single myofibers from these animals. We identified common and unique pathways that are compromised across these tissues and cell types in aged animals. We found that exercise has a rejuvenating effect on subsets of stem cells, and a profound impact in the composition and transcriptomic landscape of both circulating and tissue resident immune cells. Exercise ameliorated the upregulation of a number of inflammatory pathways as well as restored aspects of cell-cell communication within these stem cell compartments. Our study provides a comprehensive view of the coordinated responses of multiple aged stem cells and niche cells to exercise at the transcriptomic level.

cell biology↗

bioRxiv2022

Fasting Induces a Highly Resilient Deep Quiescent State in Muscle Stem Cells via Ketone Body Signaling

Short-term fasting is beneficial for the regeneration of multiple tissue types. However, the effects of fasting on muscle regeneration are largely unknown. Here we report that fasting slows muscle repair both immediately after the conclusion of fasting as well as after multiple days of refeeding. We show that ketosis, either endogenously produced during fasting or a ketogenic diet, or exogenously administered, promotes a deep quiescent state in muscle stem cells (MuSCs). Although deep quiescent MuSCs are less poised to activate, slowing muscle regeneration, they have markedly improved survival when facing sources of cellular stress. Further, we show that ketone bodies, specifically {beta}-hydroxybutyrate, directly promote MuSC deep quiescence via a non-metabolic mechanism. We show that {beta}-hydroxybutyrate functions as an HDAC inhibitor within MuSCs leading to acetylation and activation of an HDAC1 target protein p53. Finally, we demonstrate that p53 activation contributes to the deep quiescence and enhanced resilience observed during fasting.

cell biology↗

bioRxiv2021

Tabula Microcebus: A transcriptomic cell atlas of mouse lemur, an emerging primate model organism

Mouse lemurs are the smallest, fastest reproducing, and among the most abundant primates, and an emerging model organism for primate biology, behavior, health and conservation. Although much has been learned about their physiology and their Madagascar ecology and phylogeny, little is known about their cellular and molecular biology. Here we used droplet- and plate-based single cell RNA-sequencing to profile 226,000 cells from 27 mouse lemur organs and tissues opportunistically procured from four donors clinically and histologically characterized. Using computational cell clustering, integration, and expert cell annotation, we defined and biologically organized over 750 mouse lemur molecular cell types and their full gene expression profiles. These include cognates of most classical human cell types, including stem and progenitor cells, and the developmental programs for spermatogenesis, hematopoiesis, and other adult tissues. We also described dozens of previously unidentified or sparsely characterized cell types and subtypes. We globally compared cell type expression profiles to define the molecular relationships of cell types across the body, and explored primate cell and gene expression evolution by comparing mouse lemur cell transcriptomes to those of human, mouse, and macaque. This revealed cell type specific patterns of primate specialization, as well as many cell types and genes for which lemur provides a better human model than mouse. The atlas provides a cellular and molecular foundation for studying this primate model organism, and establishes a general approach for other emerging model organisms.

genetics↗

bioRxiv2021

RNA-BINDING PROTEINS DIRECT MYOGENIC CELL FATE DECISIONS

RNA-binding proteins (RBPs) are essential for skeletal muscle regeneration and RBP dysfunction causes muscle degeneration and neuromuscular disease. How ubiquitously expressed RBPs orchestrate complex tissue regeneration and direct cell fate decisions in skeletal muscle remains poorly understood. Single cell RNA-sequencing of regenerating skeletal muscle reveals that RBP expression, including numerous neuromuscular disease-associated RBPs, is temporally regulated in skeletal muscle stem cells and correlates to stages of myogenic differentiation. By combining machine learning with RBP engagement scoring, we discover that the neuromuscular disease associated RBP Hnrnpa2b1 is a differentiation-specifying regulator of myogenesis controlling myogenic cell fate transitions during terminal differentiation. The timing of RBP expression specifies cell fate transitions by providing a layer of post-transcriptional regulation needed to coordinate stem cell fate decisions during complex tissue regeneration.

molecular biology↗

bioRxiv2019

Exercise and angiotensin receptor blockade enhance recovery after orthopaedic trauma in mice by decreasing pain and improving muscle regeneration

Chronic pain and disability after limb injury are major public health problems. One key obstacle to addressing these adverse outcomes is that we do not know when exercise should be initiated or whether the beneficial effects of exercise can be reproduced using pharmacological tools. In these studies, we developed and used a murine model of orthopaedic trauma combining tibia fracture and pin fixation with tibialis anterior muscle damage. Behavioral measures included mechanical nociceptive thresholds and distances run on exercise wheels. Bone healing was quantified using microCT scanning, and muscle fiber size distribution as well as fibrosis were followed using immunohistochemistry. We found that the model provided robust mechanical allodynia, fibrosis and a shift to smaller average muscle fiber size lasting up to 5 weeks from injury. We also observed that allowing \"late\" (weeks 1-2) rather than \"early\" (weeks 0-1) wheel running after injury resulted in greater overall running activity and greater reversal of allodynia. In parallel, the late running paradigm was also associated with lower levels of muscle fibrosis and a return towards normal muscle fiber diameters. Providing the anti-fibrotic angiotensin receptor blocker losartan to mice in drinking water blocked TGFbeta production while reducing both allodynia and muscle fibrosis. Combining losartan and late exercise provided no additional benefit. We conclude that early healing after orthopaedic trauma must be allowed prior to the initiation of exercise to achieve optimal pain, functional and physiological outcomes. Losartan may provide many of the same pain, functional and physiological outcomes by its regulation of TGFbeta signaling and is a viable candidate for translational studies.\n\nKey points summaryO_LIOur tibial fracture orthopaedic injury model in mice recapitulates the major manifestations of complex trauma including nociceptive sensitization, bone fracture, muscle fibrosis and muscle fiber loss.\nC_LIO_LIDelayed exercise after complex orthopaedic trauma results in decreased muscle fibrosis and improved pain\nC_LIO_LILosartan, an angiotensin-receptor blocker with antifibrotic abilities, recapitulates the effect of exercise on post-injury recovery\nC_LI

neuroscience↗

bioRxiv2019

Transient non-integrative nuclear reprogramming promotes multifaceted reversal of aging in human cells

Aging is characterized by a gradual loss of function occurring at the molecular, cellular, tissue and organismal levels1-3. At the chromatin level, aging is associated with the progressive accumulation of epigenetic errors that eventually lead to aberrant gene regulation, stem cell exhaustion, senescence, and deregulated cell/tissue homeostasis3. The technology of nuclear reprogramming to pluripotency, through over-expression of a small number of transcription factors, can revert both the age and the identity of any cell to that of an embryonic cell by driving epigenetic reprogramming2,4,5. Recent evidence has shown that transient transgenic reprogramming can ameliorate age-associated hallmarks and extend lifespan in progeroid mice6. However, it is unknown how this form of epigenetic rejuvenation would apply to physiologically aged cells and, importantly, how it might translate to human cells. Here we show that transient reprogramming, mediated by transient expression of mRNAs, promotes a rapid reversal of both cellular aging and of epigenetic clock in human fibroblasts and endothelial cells, reduces the inflammatory profile in human chondrocytes, and restores youthful regenerative response to aged, human muscle stem cells, in each case without abolishing cellular identity. Our method, that we named Epigenetic Reprogramming of Aging (ERA), paves the way to a novel, potentially translatable strategy for ex vivo cell rejuvenation treatment. In addition, ERA holds promise for in vivo tissue rejuvenation therapies to reverse the physiological manifestations of aging and the risk for the development of age-related diseases.

cell biology↗