A single-nucleus transcriptomic atlas of primate liver aging uncovers the pro-senescence role of SREBP2 in hepatocytes

Aging increases the risk ofliver diseases and systemic susceptibility to aging-related diseases.However,cell type-specific changes and the underlying mechanism of liver aging in higher vertebrates remain incompletely characterized.Here,we constructed the first single-nucleus transcriptomic landscape of primate liver aging,in which we resolved cell type-specific gene expression fluctuation in hepatocytes across three liver zonations and detected aberrant cell-cell interactions between hepatocytes and niche cells.Upon in-depth dissection of this rich dataset,we identifed impaired lipid metabolism and upregulation of chronic inflammation-related genes prominently associated with declined liver functions during aging.In particular,hyperactivated sterol regulatory element-binding protein(SREBP)signaling was a hallmark of the aged liver,and consequently,forced activation of SREBP2 in human primary hepatocytes recapitulated in vivo aging phenotypes,manifesting as impaired detoxification and accelerated cellular senescence.This study expands our knowledge of primate liver aging and informs the development of diagnostics and therapeutic interventions for liver aging and associated diseases.

Human ESC-derived vascular cells promote vascular regeneration in a HIF-1α dependent manner

Hypoxia-inducible factor(HIF-1α),a core transcription factor responding to changes in cellular oxygen levels,is closely associated with a wide range of physiological and pathological conditions.However,its differential impacts on vascular cell types and molecular programs modulating human vascular homeostasis and regeneration remain largely elusive.Here,we applied CRISPR/Cas9-mediated gene editing of human embryonic stem cells and directed differentiation to generate HIF-ia-deficient human vascular cells including vascular endothelial cells,vascular smooth muscle cells,and mesenchymal stem cells(MsCs),as a platform for discovering cell type-specific hypox-ia-induced response mechanisms.Through comparative molecular profiling across cell types under normoxic and hypoxic conditions,we provide insight into the indispensable role of HIF-1αin the promotion of ischemic vascular regeneration.We found human MSCs to be the vascular cell type most susceptible to HIF-1a deficiency,and that transcriptional inactivation of ANKZF1,an effector of HIF-1a,impaired pro-angiogenic processes.Altogether,our findings deepen the understanding of HIF-ia in human angiogenesis and support further explorations of novel therapeutic strategies of vascular regeneration against ischemic damage.

Single-nucleus profiling unveils a geroprotective role of the Foxo3 in primate skeletal muscle aging

Age-dependent loss of skeletal muscle mass and function is a feature of sarcopenia,and increases the risk of many aging-related metabolic diseases.Here,we report phenotypic and single-nucleus transcriptomic analyses of non-human primate skeletal muscle aging.A higher transcriptional fluctuation was observed in myonuclei relative to other interstitial cell types,indicating a higher susceptibility of skeletal muscle fiber to aging.We found a downregulation of Foxo3 in aged primate skeletal muscle,and identi-fied FOxo3 as a hub transcription factor maintaining skeletal muscle homeostasis.Through the establishment of a complementary experimental pipeline based on a human pluripotent stem cell-derived myotube model,we revealed that silence of Foxo3 accelerates human myotube senescence,whereas genetic activation of endogenous FOxO3 alleviates human myotube aging.Altogether,based on a combination of monkey skeletal muscle and human myotube aging research models,we unraveled the pivotal role of the FOxO3 in safeguarding primate skeletal muscle from aging,providing a comprehensive resource for the development of clinical diagnosis and targeted therapeutic interventions against human skeletal muscle aging and the onset of sarcopenia along with aging-relateddisorders.

A single-nucleus transcriptomic atlas of primate testicular aging reveals exhaustion of the spermatogonial stem cell reservoir and loss of Sertoli cell homeostasis

The testis is pivotal for male reproduction,and its progressive functional decline in aging is associated with infertility.However,the regulatory mechanism underlying primate testicular aging remains largely elusive.Here,we resolve the aging-related cellular and molecular alterations of primate testicular aging by establishing a single-nucleus transcriptomic atlas.Gene-expression patterns along the spermatogenesis trajectory revealed molecular programs associated with attrition of spermatogonial stem cell reservoir,disturbed meiosis and impaired spermiogenesis along the sequential continuum.Remarkably,Sertoli cell was identified as the cell type most susceptible to aging,given its deeply perturbed age-associated transcriptional profiles.Concomitantly,downregulation of the transcription factor Wilms'Tumor 1(WTi),essential for Sertoli cell homeostasis,was associated with accelerated cellular senescence,disrupted tight junctions,and a compromised cell identity signature,which altogether may help create a hostile microenvironment for spermatogenesis.Collectively,our study depicts in-depth transcriptomic traits of non-human primate(NHP)testicular aging at single-cell resolution,providing potential diagnostic biomarkers and targets for therapeutic interventions against testicular aging and age-related male reproductive diseases.

Single-cell profiling reveals a potent role of quercetin in promoting hair regeneration

Hair loss affects millions of people at some time in their life,and safe and efficient treatments for hair loss are a significant unmet medical need.We report that topical delivery of quercetin(Que)stimulates resting hair follicles to grow with rapid follicular keratinocyte proliferation and replenishes perifollicular microvasculature in mice.We construct dynamic single-cell transcriptome landscape over the course of hair regrowth and find that Que treatment stimulates the differentiation trajectory in the hair follicles and induces an angiogenic signature in dermal endothelial cells by activating HIF-1αin endothelial cells.Skin administration of a HIF-1αagonist partially recapitulates the pro-angiogenesis and hair-growing effects of Que.Together,these findings provide a molecular understanding for the efficacy of Que in hair regrowth,which underscores the translational potential of targeting the hair follicle niche as a strategy for regenerative medicine,and suggest a route of pharmacological intervention that may promote hair regrowth.

Differential stem cell aging kinetics in Hutchinson-Gilford progeria syndrome and Werner syndrome

progeria syndrome （HGPS） and Wemer syndrome （WS） are two of the best characterized human progeroid syndromes. HGPS is caused by a point mutation in lamin A （LMNA） gene, resulting in the production of a truncated protein product-progerin. WS is caused by mutations in 14/RN gem）, encoding a loss-of-function RecQ DNA helicase. Here, by gene editing we created isogenic human embryonic stem cells （ESCs） with heterozygous （G608G/＋） or homozygous （G608G/G608G） LMNA mutation and biallelic WRN knockout, for modeling HGPS and WS pathogenesis, respectively. While ESCs and endothelial cells （ECs） did not present any features of premature senescence, HGPS- and WS-mesenchymal stem cells （MSCs） showed aging-associated phenotypes with different kinetics. WS-MSCs had early-onset mild premature aging phenotypes while HGPS-MSCs exhibited iate-onset acute premature aging characterisitcs. Taken together, our study compares and contrasts the distinct pathologies underpinning the two premature aging disorders, and provides reliable stem-cell based models to identify new therapeutic strategies for pathological and physiological aging.

Modeling CADASIL vascular pathologies with patient-derived induced pluripotent stem cells

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy(CADASIL)is a rare hereditary cerebrovascular disease caused by a NOTCH3 mutation.However,the underlying cellular and molecular mechanisms remain unidentified.Here,we generated non-integrative induced pluripotent stem cells(iPSCs)from fibroblasts of a CADASIL patient harboring a heterozygous NOTCH3 mutation(c.3226C>T,p.R1076C).Vascular smooth muscle cells(VSMCs)differentiated from CADASIL-specific iPSCs showed gene expression changes associated with disease phenotypes,including activation of the NOTCH and NF-kB signaling pathway,cytoskeleton disorganization,and excessive cell proliferation.In comparison,these abnormalities were not observed in vascular endothelial cells(VECs)derived from the patients iPSCs.Importantly,the abnormal upregulation of NF-kB target genes in CADASIL VSMCs was diminished by a NOTCH pathway inhibitor,providing a potential therapeutic strategy for CADASIL.Overall,using this iPSCbased disease model,our study identified clues for studying the pathogenic mechanisms of CADASIL and developing treatment strategies for this disease.

Modeling xeroderma pigmentosum associated neurological pathologies with patients-derived iPSCs

Xeroderma pigmentosum （XP） is a group of genetic disorders caused by mutations of XP-associated genes, resulting in impairment of DNA repair. XP patients frequently exhibit neurological degeneration, but the underlying mechanism is unknown, in part due to lack of proper disease models. Here, we generated patientspecific induced pluripotent stem cells （iPSCs） harboring mutations in five different XP genes including XPA, XPB, XPC, XPG, and XPV. These iPSCs were further differentiated to neural cells, and their susceptibility to DNA damage stress was investigated. Mutation of XPA in either neural stem cells （NSCs） or neurons resulted in severe DNA damage repair defects, and these neural cells with mutant XPA were hyper-sensitive to DNA damage-induced apoptosis. Thus, XP-mutant neural cells represent valuable tools to clari the molecular mechanisms of neurological abnormalities in the XP patients.

Single-nucleus transcriptomic landscape of primate hippocampal aging

The hippocampus plays a crucial role in learning and memory,and its progressive deterioration with age is functionally linked to a variety of human neurodegenerative diseases.Yet a systematic profiling of the aging effects on various hippocampal cell types in primates is still missing.Here,we reported a variety of new aging-associated phenotypic changes of the primate hippocampus.These include,in particular,increased DNA damage and heterochromatin erosion with time,alongside loss of proteostasis and elevated inflammation.To understand their cellular and molecular causes,we established the first single-nucleus transcriptomic atlas of primate hippocampal aging.Among the 12 identified cell types,neural transiently amplifying progenitor cell(TAPC)and microglia were most affected by aging.In-depth dissection of gene-expression dynamics revealed impaired TAPC division and compromised neuronal function along the neurogenesis trajectory;additionally elevated pro-inflammatory responses in the aged microglia and oligodendrocyte,as well as dysregulated coagulation pathways in the aged endothelial cells may contribute to a hostile microenvironment for neurogenesis.This rich resource for understanding primate hippocampal aging may provide potential diagnostic biomarkers and therapeutic interventions against age-related neurodegenerative diseases.

CRISPR/Cas9-mediated targeted gene correction in amyotrophic lateral sclerosis patient iPSCs

Amyotrophic lateral sclerosis （ALS） is a complex neu- rodegenerative disease with cellular and molecular mechanisms yet to be fully described. Mutations in a number of genes including SOD1 and FUS are associated with familial ALS. Here we report the generation of induced pluripotent stem cells （iPSCs） from fibroblasts of familial ALS patients bearing SOD1＋1A27~c and FUS＋/GISe6A mutations, respectively. We further gener- ated gene corrected ALS iPSCs using CRISPR/Cas9 system. Genome-wide RNA sequencing （RNA-seq） analysis of motor neurons derived from SOD1＋~A272c and corrected iPSCs revealed 899 aberrant transcripts. Our work may shed light on discovery of early biomarkers and pathways dysregulated in ALS, as well as provide a basis for novel therapeutic strategies to treat ALS.

FOXO3-engineered human mesenchymal progenitor cells efficiently promote cardiac repair after myocardial infarction

Dear Editor,Myocardial infarction(MI)is the irreversible cardiomyocyte death resulting from prolonged oxygen deprivation due to obstructed blood supply(ischemia),leading to contractile dysfunction and cardiac remodeling.In recent decades,stem cell transplantation has been extensively investigated for the repair of injured heart in animal studies and clinical trials(Kanelidis et al.,2017;Gyongyosi et al.,2018).

Establishment of hepatic and neural differentiation platforms of Wilson’s disease specific induced pluripotent stem cells

The combination of disease-specific human induced pluripotent stem cells(iPSC)and directed cell differentiation offers an ideal platform for modeling and studying many inherited human diseases.Wilson’s disease(WD)is a monogenic disorder of toxic copper accumulation caused by pathologic mutations of the ATP7B gene.WD affects multiple organs with primary manifestations in the liver and central nervous system(CNS).In order to better investigate the cellular pathogenesis of WD and to develop novel therapies against various WD syndromes,we sought to establish a comprehensive platform to differentiate WD patient iPSC into both hepatic and neural lineages.Here we report the generation of patient iPSC bearing a Caucasian population hotspot mutation of ATP7B.Combining with directed cell differentiation strategies,we successfully differentiated WD iPSC into hepatocyte-like cells,neural stem cells and neurons.Gene expression analysis and cDNA sequencing confirmed the expression of the mutant ATP7B gene in all differentiated cells.Hence we established a platform for studying both hepatic and neural abnormalities of WD,which may provide a new tool for tissue-specific disease modeling and drug screening in the future.

Human induced pluripotent stem cells derived hepatocytes:rising promise for disease modeling,drug development and cell therapy

Recent advances in the study of human hepatocytes derived from induced pluripotent stem cells(iPSC)represent new promises for liver disease study and drug discovery.Human hepatocytes or hepatocyte-like cells differentiated from iPSC recapitulate many func-tional properties of primary human hepatocytes and have been demonstrated as a powerful and efficient tool to model human liver metabolic diseases and fa-cilitate drug development process.In this review,we summarize the recent progress in this field and discuss the future perspective of the application of human iPSC derived hepatocytes.

Exosomes from antler stem cells alleviate mesenchymal stem cell senescence and osteoarthritis

Dear Editor,Stem cell therapy holds enormous and revolutionary promise to treat various age-related diseases,such as diabetes,heart failure,and Parkinson’s disease.However,low retention and survival rate of delivered stem cells,partially due to immunological rejection,constitute major hurdles for the clinical implementation of stem cell therapy(Lei et al.,2021a).Since mounting evidence showed that several types of stem cells mainly exert their therapeutic effects through the secretion of paracrine effects,exosomes,which are released by stem cells and execute most paracrine functions,have begun to draw attention in the field(Tran and Damaser,2015).Exosomes are membrane-enclosed vesicles with an average diameter of∼100 nanometers secreted by the cells,containing cytokines.

Hyperthermia differentially affects specific human stem cells and their differentiated derivatives

Dear Editor,The human body operates optimally at a core temperature of 37 degrees Celsius.Homeostasis at this temperature is essential for cellular and physiological functions(Cheshire,2016).However,infectious diseases,inflammation,injury,neoplasia,and elevated climate temperature can cause a regulated rise in body core temperature,i.e.,fever(Pasi-khova et al,2017).Indeed,an acute or chronic increase in temperature leads to detrimental effects on vasculature by altering a number of indices of vascular structure and function(DuBose et al.,1998).

Compound screening platform using human induced pluripotent stem cells to identify small molecules that promote chondrogenesis

Articular cartilage,which is mainly composed of collagen Ⅱ,enables smooth skeletal movement.Degeneration of collagen Ⅱ can be caused by various events,such as injury,but degeneration especially increases over the course of normal aging.Unfortunately,the body does not fully repair itself from this type of degeneration,resulting in impaired movement.Microfracture,an articular cartilage repair surgical technique,has been commonly used in the clinic to induce the repair of tissue at damage sites.Mesenchymal stem cells(MSC)have also been used as cell therapy to repair degenerated cartilage.However,the therapeutic outcomes of all these techniques vary in different patients depending on their age,health,lesion size and the extent of damage to the cartilage.The repairing tissues either form fibrocartilage or go into a hypertrophic stage,both of which do not reproduce the equivalent functionality of endogenous hyaline cartilage.One of the reasons for this is inefficient chondrogenesis by endogenous and exogenous MSC.Drugs that promote chondrogenesis could be used to induce self-repair of damaged cartilage as a non-invasive approach alone,or combined with other techniques to greatly assist the therapeutic outcomes.The recent development of human induced pluripotent stem cell(iPSCs),which are able to self-renew and differentiate into multiple cell types,provides a potentially valuable cell resource for drug screening in a“more relevant”cell type.Here we report a screening platform using human iPSCs in a multi-well plate format to identify compounds that could promote chondrogenesis.

αKLOTHO and sTGFβR2 treatment counteract the osteoarthritic phenotype developed in a rat model

Dear Editor,Homeostasis and repair are critical biological processes that allow for tissue and organ preservation and function in multicellular organisms.Their regulation and extension vary drastically across the animal kingdom,and mammals show limited tissue-specific regenerative capacity that declines with age.During aging,articular cartilage is one of the tissues that undergo substantial changes in the matrix structure,molecular composition,metabolic activity,and mechanical properties(Loeser et al.2016).

Mutations in foregut SOX2^+ cells induce efficient proliferation via CXCR2 pathway

Identification of the precise molecular pathways involved in oncogene-induced transformation may help us gain a better understanding of tumor initiation and promotion. Here, we demonstrate that SOX2^+ foregut epithelial cells are prone to oncogenic transformation upon mutagenic insults, such as Kras^G12D and p53 deletion. GFP-based lineage-tracing experiments indicate that SOX2^+ cells are the cells-of-origin of esophagus and stomach hyperplasia. Our observations indicate distinct roles for oncogenic KRAS mutation and P53 deletion. p53 homozygous deletion is required for the acquisition of an invasive potential, and Kras^G12D expression, but not p53 deletion, suffices for tumor formation. Global gene expression analysis reveals secreting factors upregulated in the hyperplasia induced by oncogenic KRAS and highlights a crucial role for the CXCR2 pathway in driving hyperplasia. Collectively, the array of genetic models presented here demonstrate that stratified epithelial cells are susceptible to oncogenic insults, which may lead to a better understanding of tumor initiation and aid in the design of new cancer therapeutics.

First progeria monkey model generated using base editor

In this issue,Wang et al.report on the generation of a nonhuman primate model of Hutchinson-Gilford progeria syndrome(HGPS)using a base editor.Base editing is an emerging novel genome editing technique for modifying a single base pair at specific sites in the genome.Base editors(BEs)have two prin cipal comp orients,a catalytically in active or single strand cleaving Cas-variant,which binds to the guide RNA and a nucleobase deaminase domain to convert specific base pairs at the target loci(Komor et al.,2016;Nishida et al.,2016;Gaudelli et al.,2017).Cytosine base editor(CBE)and adenine base editor(ABE)are two baseeditors,which con vert Cytosi ne-Gua nine(C-G)to Thymine-Adenine(T-A)and A-T to G-C,respectively.

A widely adaptable approach to generate integration-free iPSCs from non-invasively acquired human somatic cells

Dear Editor Human pluripotent stem cells including human embryonic stem cells （ESCs） and induced pluripotent stem cells （iPSCs） are cells displaying abilities of unlimited self-renewal and differentiation into any somatic cell type. These unique properties make them increasingly attractive for novel applications in disease modeling, drug discovery, and cell therapy （Buganim et al., 2014; Liu et al., 2011; Liu et al., 2012; Sanchez Alvarado and Yamanaka, 2014）. Moreover, iPSCs hold great potential for personalized cell therapy as they avoid some of the ethical concerns as well as the immunological rejection issues ascribed to ESCs.