Neural tube defects(NTDs)are severe congenital neurodevelopmental disorders arising from incomplete neural tube closure.Although folate supplementation has been shown to mitigate the incidence of NTDs,some cases,often...Neural tube defects(NTDs)are severe congenital neurodevelopmental disorders arising from incomplete neural tube closure.Although folate supplementation has been shown to mitigate the incidence of NTDs,some cases,often attributable to genetic factors,remain unpreventable.The SHROOM3 gene has been implicated in NTD cases that are unresponsive to folate supplementation;at present,however,the underlying mechanism remains unclear.Neural tube morphogenesis is a complex process involving the folding of the planar epithelium of the neural plate.To determine the role of SHROOM3 in early developmental morphogenesis,we established a neuroepithelial organoid culture system derived from cynomolgus monkeys to closely mimic the in vivo neural plate phase.Loss of SHROOM3 resulted in shorter neuroepithelial cells and smaller nuclei.These morphological changes were attributed to the insufficient recruitment of cytoskeletal proteins,namely fibrous actin(F-actin),myosin II,and phospho-myosin light chain(PMLC),to the apical side of the neuroepithelial cells.Notably,these defects were not rescued by folate supplementation.RNA sequencing revealed that differentially expressed genes were enriched in biological processes associated with cellular and organ morphogenesis.In summary,we established an authentic in vitro system to study NTDs and identified a novel mechanism for NTDs that are unresponsive to folate supplementation.展开更多
Video-based action recognition is becoming a vital tool in clinical research and neuroscientific study for disorder detection and prediction.However,action recognition currently used in non-human primate(NHP)research ...Video-based action recognition is becoming a vital tool in clinical research and neuroscientific study for disorder detection and prediction.However,action recognition currently used in non-human primate(NHP)research relies heavily on intense manual labor and lacks standardized assessment.In this work,we established two standard benchmark datasets of NHPs in the laboratory:Monkeyin Lab(Mi L),which includes 13 categories of actions and postures,and MiL2D,which includes sequences of two-dimensional(2D)skeleton features.Furthermore,based on recent methodological advances in deep learning and skeleton visualization,we introduced the Monkey Monitor Kit(Mon Kit)toolbox for automatic action recognition,posture estimation,and identification of fine motor activity in monkeys.Using the datasets and Mon Kit,we evaluated the daily behaviors of wild-type cynomolgus monkeys within their home cages and experimental environments and compared these observations with the behaviors exhibited by cynomolgus monkeys possessing mutations in the MECP2 gene as a disease model of Rett syndrome(RTT).Mon Kit was used to assess motor function,stereotyped behaviors,and depressive phenotypes,with the outcomes compared with human manual detection.Mon Kit established consistent criteria for identifying behavior in NHPs with high accuracy and efficiency,thus providing a novel and comprehensive tool for assessing phenotypic behavior in monkeys.展开更多
Parkinson’s disease is typically characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta.Many studies have been performed based on the supplementation of lost dopaminergic ...Parkinson’s disease is typically characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta.Many studies have been performed based on the supplementation of lost dopaminergic neurons to treat Parkinson’s disease.The initial strategy for cell replacement therapy used human fetal ventral midbrain and human embryonic stem cells to treat Parkinson’s disease,which could substantially alleviate the symptoms of Parkinson’s disease in clinical practice.However,ethical issues and tumor formation were limitations of its clinical application.Induced pluripotent stem cells can be acquired without sacrificing human embryos,which eliminates the huge ethical barriers of human stem cell therapy.Another widely considered neuronal regeneration strategy is to directly reprogram fibroblasts and astrocytes into neurons,without the need for intermediate proliferation states,thus avoiding issues of immune rejection and tumor formation.Both induced pluripotent stem cells and direct reprogramming of lineage cells have shown promising results in the treatment of Parkinson’s disease.However,there are also ethical concerns and the risk of tumor formation that need to be addressed.This review highlights the current application status of cell reprogramming in the treatment of Parkinson’s disease,focusing on the use of induced pluripotent stem cells in cell replacement therapy,including preclinical animal models and progress in clinical research.The review also discusses the advancements in direct reprogramming of lineage cells in the treatment of Parkinson’s disease,as well as the controversy surrounding in vivo reprogramming.These findings suggest that cell reprogramming may hold great promise as a potential strategy for treating Parkinson’s disease.展开更多
Mutations of PTEN-induced kinase I(PINK1)cause early-onset Parkinson’s disease(PD)with selective neurodegeneration in humans.However,current PINK1 knockout mouse and pig models are unable to recapitulate the typical ...Mutations of PTEN-induced kinase I(PINK1)cause early-onset Parkinson’s disease(PD)with selective neurodegeneration in humans.However,current PINK1 knockout mouse and pig models are unable to recapitulate the typical neurodegenerative phenotypes observed in PD patients.This suggests that generating PINK1 disease models in non-human primates(NHPs)that are close to humans is essential to investigate the unique function of PINK1 in primate brains.Paired single guide RNA(sgRNA)/Cas9-D10A nickases and truncated sgRNA/Cas9,both of which can reduce off-target effects without compromising on-target editing,are two optimized strategies in the CRISPR/Cas9 system for establishing disease animal models.Here,we combined the two strategies and injected Cas9-D10A mRNA and two truncated sgRNAs into one-cell-stage cynomolgus zygotes to target the PINK1 gene.We achieved precise and efficient gene editing of the target site in three newborn cynomolgus monkeys.The frame shift mutations of PINK1 in mutant fibroblasts led to a reduction in mRNA.However,western blotting and immunofluorescence staining confirmed the PINK1 protein levels were comparable to that in wild-type fibroblasts.We further reprogramed mutant fibroblasts into induced pluripotent stem cells(iPSCs),which showed similar ability to differentiate into dopamine(DA)neurons.Taken together,our results showed that co-injection of Cas9-D10A nickase mRNA and sgRNA into one-cell-stage cynomolgus embryos enabled the generation of human disease models in NHPs and target editing by pair truncated sgRNA/Cas9-D10A in PINK1 gene exon 2 did not impact protein expression.展开更多
Rett syndrome(RTT)is a progressive neurodevelopmental disorder that occurs mainly in girls with a range of typical symptoms of autism spectrum disorders.MeCP2 protein loss-of-function in neural lineage cells is the ma...Rett syndrome(RTT)is a progressive neurodevelopmental disorder that occurs mainly in girls with a range of typical symptoms of autism spectrum disorders.MeCP2 protein loss-of-function in neural lineage cells is the main cause of RTT pathogenicity.As it is still hard to understand the mechanism of RTT on the basis of only clinical patients or animal models,cell models cultured in vitro play indispensable roles.Here we reviewed the research progress in the pathogenesis of RTT at the cellular level,summarized the preclinical-research-related applications,and prospected potential future development.展开更多
Efforts have been made to establish various human pluripotent stem cell lines.However,such methods have not yet been duplicated in non-human primate cells.Here,we introduce a multiplexed single-cell sequencing techniq...Efforts have been made to establish various human pluripotent stem cell lines.However,such methods have not yet been duplicated in non-human primate cells.Here,we introduce a multiplexed single-cell sequencing technique to profile the molecular features of monkey pluripotent stem cells in published culture conditions.The results demonstrate suboptimized maintenance of pluripotency and show that the selected signaling pathways for resetting human stem cells can also be interpreted for establishing monkey cell lines.Overall,this work legitimates the translation of novel human cell line culture conditions to monkey cells and provides guidance for exploring chemical cocktails for monkey stem cell line derivation.展开更多
Cancer refers to a diverse collection of diseases characterized by several well-established hallmarks,including the abilities to sustain proliferative signaling,evade growth suppressors,activate invasion and metastasi...Cancer refers to a diverse collection of diseases characterized by several well-established hallmarks,including the abilities to sustain proliferative signaling,evade growth suppressors,activate invasion and metastasis,enable replicative immortality,induce angiogenesis,and resist cell death1.Historically,genetic alterations(deletions,point mutations,and translocations)were thought to be the basis for tumor formation via the inactivation of tumor suppressors and activation of oncogenes.展开更多
Non-human primates(NHPs)are increasingly used in preclinical trials to test the safety and efficacy of biotech-nology therapies.Nonetheless,given the ethical issues and costs associated with this model,it would be hig...Non-human primates(NHPs)are increasingly used in preclinical trials to test the safety and efficacy of biotech-nology therapies.Nonetheless,given the ethical issues and costs associated with this model,it would be highly advantageous to use NHP cellular models in clinical studies.However,developing and maintaining the naive state of primate pluripotent stem cells(PSCs)remains difficult as does in vivo detection of PSCs,thus limiting biotech-nology application in the cynomolgus monkey.Here,we report a chemically defined,xeno-free culture system for culturing and deriving monkey PSCs in vitro.The cells display global gene expression and genome-wide hypometh-ylation patterns distinct from monkey-primed cells.We also found expression of signaling pathways components that may increase the potential for chimera formation.Crucially for biomedical applications,we were also able to integrate bioluminescent reporter genes into monkey PsCs and track them in chimeric embryos in vivo and in vitro.The engineered cells retained embryonic and extra-embryonic developmental potential.Meanwhile,we generated a chimeric monkey carrying bioluminescent cells,which were able to track chimeric cells for more than 2 years in living animals.Our study could have broad utility in primate stem cell engineering and in utilizing chimeric monkey models forclinical studies.展开更多
Pig and monkey are widely used models for exploration of human diseases and evaluation of drug efficiency and toxicity,but high cost limits their uses.Organoids have been shown to be promising models for drug test as ...Pig and monkey are widely used models for exploration of human diseases and evaluation of drug efficiency and toxicity,but high cost limits their uses.Organoids have been shown to be promising models for drug test as they reasonably preserve tissue structure and functions.However,colonic organoids of pig and monkey are not yet established.Here,we report a culture medium to support the growth of porcine and monkey colonic organoids.Wnt signaling and PGE2 are important for long-term expansion of the organoids,and their withdrawal results in lineage differentiation to mature cells.Furthermore,we observe that porcine colonic organoids are closer to human colonic organoids in terms of drug toxicity response.Successful establishment of porcine and monkey colonic organoids would facilitate the mechanistic investigation of the homeostatic regulation of the intestine of these animals and is useful for drug development and toxicity studies.展开更多
Splice-switching antisense oligonucleotides(ASOs)and engineered U7 small nuclear ribonucleoprotein(U7 Sm OPT)are the most commonly used methods for exon skipping.However,challenges remain,such as limited organ deliver...Splice-switching antisense oligonucleotides(ASOs)and engineered U7 small nuclear ribonucleoprotein(U7 Sm OPT)are the most commonly used methods for exon skipping.However,challenges remain,such as limited organ delivery and repeated dosing for ASOs and unknown risks of by-products produced by U7 Sm OPT.Here,we showed that antisense circular RNAs(AS-circRNAs)can effectively mediate exon skipping in both minigene and endogenous transcripts.We also showed a relatively higher exon skipping efficiency at the tested Dmd minigene than U7 Sm OPT.AS-circRNA specifically targets the precursor mRNA splicing without off-target effects.Moreover,AS-circRNAs with adeno-associated virus(AAV)delivery corrected the open reading frame and restored the dystrophin expression in a mouse model of Duchenne muscular dystrophy.In conclusion,we develop an alternative method for regulating RNA splicing,which might be served as a novel tool for genetic disease treatment.展开更多
Background:Aggression is a commonly hostile behavior linked to the hippocampal activity.Childhood trauma(CT)exposure has been associated with altered sensitization of the hypothalamic-pituitary-adrenal(HPA)axis and hi...Background:Aggression is a commonly hostile behavior linked to the hippocampal activity.Childhood trauma(CT)exposure has been associated with altered sensitization of the hypothalamic-pituitary-adrenal(HPA)axis and hippocampal volumewhich could increase violent aggressive behaviors.Additionally,Catechol-O-methyltransferase(COMT),the major dopamine metabolism enzyme,is impli-cated in stress responsivity,including aggression.Hence,CT exposure may affect aggression through the effect on the hippocampal function,which might also be modulated by the COMT variations.Objectives:This study examined whether both CT and haplotypes of COMT moderate hippocampal function and thus affect human aggressive behavior.Methods:We obtained bilateral hippocampal functional connectivity maps using resting state functional magnetic resonance imag-ing(MRI)data.COMT haplotype estimation was performed using Haploview 4.2 and PHASE 2.1.Then we constructed a moderated mediation model to study the effect of the CTQ×COMT on aggressive behavior.Results:Three major haplotypes were generated from thirteen single nucleotide polymorphisms(SNPs)within the COMT gene and formed three haplotypes corresponding to high,medium,and low enzymatic activity of COMT.The results showed interactive re-lationships between the Childhood Trauma Questionnaire(CTQ)and COMT with respect to the functional connectivity(FC)of the bilateral hippocampus(HIP)-orbital frontal cortex(OFC).Specifically,CT experience predicted lower negative HIP-OFC coupling in the APS and HPS haplotypes corresponding to the medium and high enzymatic activity of COMT,but greater FC in the LPS haplotypes corresponding to the low enzymatic activity.We also observed a conditional mediation effect of the right HIP-OFC coupling in the link between COMT and aggressive behavior that was moderated by CT experience.Conclusions:These results suggest that CT and COMT have a combined effect on aggressive behavior through hippocampal function.This mediation analysis sheds light on the influence of childhood experience on aggressive behavior in different genetic backgrounds.展开更多
Animal models are widely used for biomedical studies and drug evaluation.The small intestine plays key roles in nutrient absorption,hormone secretion,microbiota defense and drug absorption and metabolism.Although the ...Animal models are widely used for biomedical studies and drug evaluation.The small intestine plays key roles in nutrient absorption,hormone secretion,microbiota defense and drug absorption and metabolism.Although the intestinal structure of mammals is conserved,the differences on epithelial cell composition,functional assignments and drug absorption among mammals are largely unknown.Here,cross-species analysis of single-cell transcriptomic atlas of the ileum epithelium from mouse,rat,pig,macaque and human reveals the conserved and differential cell types and functions among species,identifies a new CA7+cell type in pig,macaque and human ileum,uncovers the distinct expression pattern in enterocytes,enteroendocrine cells and Paneth cells,and defines the conserved and species-specific intestinal stem cell signature genes.The examination of drug absorption across species suggests that drug metabolism in mouse ileum is closer to human while drug transport in macaque ileum is more similar to human.Together,our data provide the comprehensive information about cell composition and functional assignments in five species,and offer the valuable guidance for animal model selection and drug testing.展开更多
Animal models of human diseases are vital in better understanding the mechanism of pathogenesis and essential for evaluating and validating potential therapeutic interventions.As close relatives of humans,nonhuman pri...Animal models of human diseases are vital in better understanding the mechanism of pathogenesis and essential for evaluating and validating potential therapeutic interventions.As close relatives of humans,nonhuman primates(NHPs)play an increasingly indispensable role in advancing translational medicine research.In this review,we summarized the progress of NHP models generated by embryo engineering,analyzed their unique advantages in mimicking clinical patients,and discussed the remaining gap between basic research of NHP models to translational medicine.展开更多
Cynomolgus macaques(Macaca fascicularis)belong to the genus Macaca.They are the most widespread nonhuman primate(NHP)and share a common ancestor with humans from about 25 million years ago(Kumar and Hedges,1998).Becau...Cynomolgus macaques(Macaca fascicularis)belong to the genus Macaca.They are the most widespread nonhuman primate(NHP)and share a common ancestor with humans from about 25 million years ago(Kumar and Hedges,1998).Because of their phylogenetic proximity to humans,macaques are an attractive NHP research model for a wide range of biomedical research(Yan et al.,2011).展开更多
Animal models of human diseases are vital in better understanding the mechanism of pathogenesis and essential for evaluating and validating potential therapeutic interventions.As close relatives of humans,nonhuman pri...Animal models of human diseases are vital in better understanding the mechanism of pathogenesis and essential for evaluating and validating potential therapeutic interventions.As close relatives of humans,nonhuman primates(NHPs)play an increasingly indispensable role in advancing translational medicine research.In this review,we summarized the progress of NHP models generated by embryo engineering,analyzed their unique advantages in mimicking clinical patients,and discussed the remaining gap between basic research of NHP models to translational medicine.展开更多
Direct Ink Writing(DIW)has demonstrated great potential as a versatile method to 3D print multifunctional structures.In this work,we report the implementation of hydrogel meta-structures using DIW at room temperature,...Direct Ink Writing(DIW)has demonstrated great potential as a versatile method to 3D print multifunctional structures.In this work,we report the implementation of hydrogel meta-structures using DIW at room temperature,which seamlessly integrate large specific surface areas,interconnected porous characteristics,mechanical toughness,biocompatibility,and water absorption and retention capabilities.Robust but hydrophobic polymers and weakly crosslinked nature-origin hydrogels form a balance in the self-supporting ink,allowing us to directly print complex meta-structures without sacrificial materials and heating extrusion.Mechanically,the mixed bending or stretching of symmetrical re-entrant cellular lattices and the unique curvature patterns are combined to provide little lateral expansion and large compressive energy absorbance when external forces are applied on the printed meta-structures.In addition,we have successfully demonstrated ear,aortic valve conduits and hierarchical architectures.We anticipate that the reported 3D meta-structured hydrogel would offer a new strategy to develop functional biomaterials for tissue engineering applications in the future.展开更多
Background Although postpartum depression(PPD)and non-peripartum major depressive disorder(MDD)occurring within and outside the postpartum period share many clinical characteristics,whether PPD and MDD are the same or...Background Although postpartum depression(PPD)and non-peripartum major depressive disorder(MDD)occurring within and outside the postpartum period share many clinical characteristics,whether PPD and MDD are the same or not remains controversial.Methods The current study was devoted to identify the shared and different neural circuits between PPD and MDD by resting-state functionalmagnetic resonance imaging data from 77 participants(22 first-episodic drug-naleMDD,26 drug-nale PPD,and 29 healthy controls(HC)).Results Both the PPD andMDD groups exhibited higher fractional amplitude of low-frequency fluctuation(fALFF)in left temporal pole relative to the HC group;the MDD group showed specifically increased degree centrality in the right cerebellum while PPD showed specifically decreased fALFF in the left supplementary motor area and posterior middle temporal gyrus(pMTG_L),and specifically decreased functional connectivities between pMTG and precuneus and between left subgeneual anterior cingulate cortex(sgACC_L)and right sgACC.Moreover,sgACC and left thalamus showed abnormal regional homogeneity of functional activities between any pair of HC,MDD,and PPD.Conclusions These results provide initial evidence that PPD and MDD have common and distinct neural circuits,which may facilitate understanding the neurophysiological basis and precision treatment for PPD.展开更多
The twenty-first century has already recorded more than ten major epidemics or pandemics of viral disease,including the devastating COVID-19.Novel effective antivirals with broad-spectrum coverage are urgently needed....The twenty-first century has already recorded more than ten major epidemics or pandemics of viral disease,including the devastating COVID-19.Novel effective antivirals with broad-spectrum coverage are urgently needed.Herein,we reported a novel broad-spectrum antiviral compound PAC5.Oral administration of PAC5 eliminated HBV cccDNA and reduced the large antigen load in distinct mouse models of HBV infection.Strikingly,oral administration of PAC5 in a hamster model of SARS-CoV-2 omicron(BA.1)infection significantly decreases viral loads and attenuates lung inflammation.Mechanistically,PAC5 binds to a pocket near Asp49 in the RNA recognition motif of hnRNPA2B1.PAC5-bound hnRNPA2B1 is extensively activated and translocated to the cytoplasm where it initiates the TBK1-IRF3 pathway,leading to the production of type I IFNs with antiviral activity.Our results indicate that PAC5 is a novel small-molecule agonist of hnRNPA2B1,which may have a role in dealing with emerging infectious diseases now and in the future.展开更多
Rapid detection and response to visual threats are critical for survival in animals.The amygdala(AMY)is hypothesized to be involved in this process,but how it interacts with the visual system to do this remains unclea...Rapid detection and response to visual threats are critical for survival in animals.The amygdala(AMY)is hypothesized to be involved in this process,but how it interacts with the visual system to do this remains unclear.By recording flash-evoked potentials simultaneously from the superior colliculus(SC),lateral posterior nucleus of the thalamus,AMY,lateral geniculate nucleus(LGN)and visual cortex,which belong to the cortical and subcortical pathways for visual fear processing,we investigated the temporal relationship between these regions in visual processing in rats.A quick flash-evoked potential(FEP)component was identified in the AMY.This emerged as early as in the LGN and was approximately 25 ms prior to the earliest component recorded in the SC,which was assumed to be an important area in visual fear.This quick P1 component in the AMY was not affected by restraint stress or corticosterone injection,but was diminished by RU38486,a glucocorticoid receptor blocker.By injecting a monosynaptic retrograde AAV tracer into the AMY,we found that it received a direct projection from the retina.These results confirm the existence of a direct connection from the retina to the AMY,that the latency in the AMY to flashes is equivalent to that in the sensory thalamus,and that the response is modulated by glucocorticoids.展开更多
基金supported by the National Natural Science Foundation of China (81930121,82125008 to Y.C.C.)National Key Research and Development Program of China (2018YFA0107902 to Y.C.C.and 2018YFA0801403 to Z.B.W.)+1 种基金Major Basic Research Project of Science and Technology of Yunnan (202001BC070001 to Y.C.C.)Natural Science Foundation of Yunnan Province (202102AA100053 to Y.C.C.)。
文摘Neural tube defects(NTDs)are severe congenital neurodevelopmental disorders arising from incomplete neural tube closure.Although folate supplementation has been shown to mitigate the incidence of NTDs,some cases,often attributable to genetic factors,remain unpreventable.The SHROOM3 gene has been implicated in NTD cases that are unresponsive to folate supplementation;at present,however,the underlying mechanism remains unclear.Neural tube morphogenesis is a complex process involving the folding of the planar epithelium of the neural plate.To determine the role of SHROOM3 in early developmental morphogenesis,we established a neuroepithelial organoid culture system derived from cynomolgus monkeys to closely mimic the in vivo neural plate phase.Loss of SHROOM3 resulted in shorter neuroepithelial cells and smaller nuclei.These morphological changes were attributed to the insufficient recruitment of cytoskeletal proteins,namely fibrous actin(F-actin),myosin II,and phospho-myosin light chain(PMLC),to the apical side of the neuroepithelial cells.Notably,these defects were not rescued by folate supplementation.RNA sequencing revealed that differentially expressed genes were enriched in biological processes associated with cellular and organ morphogenesis.In summary,we established an authentic in vitro system to study NTDs and identified a novel mechanism for NTDs that are unresponsive to folate supplementation.
基金supported by the National Key R&D Program of China (2021ZD0202805,2019YFA0709504,2021ZD0200900)National Defense Science and Technology Innovation Special Zone Spark Project (20-163-00-TS-009-152-01)+4 种基金National Natural Science Foundation of China (31900719,U20A20227,82125008)Innovative Research Team of High-level Local Universities in Shanghai,Science and Technology Committee Rising-Star Program (19QA1401400)111 Project (B18015)Shanghai Municipal Science and Technology Major Project (2018SHZDZX01)Shanghai Center for Brain Science and Brain-Inspired Technology。
文摘Video-based action recognition is becoming a vital tool in clinical research and neuroscientific study for disorder detection and prediction.However,action recognition currently used in non-human primate(NHP)research relies heavily on intense manual labor and lacks standardized assessment.In this work,we established two standard benchmark datasets of NHPs in the laboratory:Monkeyin Lab(Mi L),which includes 13 categories of actions and postures,and MiL2D,which includes sequences of two-dimensional(2D)skeleton features.Furthermore,based on recent methodological advances in deep learning and skeleton visualization,we introduced the Monkey Monitor Kit(Mon Kit)toolbox for automatic action recognition,posture estimation,and identification of fine motor activity in monkeys.Using the datasets and Mon Kit,we evaluated the daily behaviors of wild-type cynomolgus monkeys within their home cages and experimental environments and compared these observations with the behaviors exhibited by cynomolgus monkeys possessing mutations in the MECP2 gene as a disease model of Rett syndrome(RTT).Mon Kit was used to assess motor function,stereotyped behaviors,and depressive phenotypes,with the outcomes compared with human manual detection.Mon Kit established consistent criteria for identifying behavior in NHPs with high accuracy and efficiency,thus providing a novel and comprehensive tool for assessing phenotypic behavior in monkeys.
基金supported by the National Natural Science Foundation of China,No.31960120Yunnan Science and Technology Talent and Platform Plan,No.202105AC160041(both to ZW).
文摘Parkinson’s disease is typically characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta.Many studies have been performed based on the supplementation of lost dopaminergic neurons to treat Parkinson’s disease.The initial strategy for cell replacement therapy used human fetal ventral midbrain and human embryonic stem cells to treat Parkinson’s disease,which could substantially alleviate the symptoms of Parkinson’s disease in clinical practice.However,ethical issues and tumor formation were limitations of its clinical application.Induced pluripotent stem cells can be acquired without sacrificing human embryos,which eliminates the huge ethical barriers of human stem cell therapy.Another widely considered neuronal regeneration strategy is to directly reprogram fibroblasts and astrocytes into neurons,without the need for intermediate proliferation states,thus avoiding issues of immune rejection and tumor formation.Both induced pluripotent stem cells and direct reprogramming of lineage cells have shown promising results in the treatment of Parkinson’s disease.However,there are also ethical concerns and the risk of tumor formation that need to be addressed.This review highlights the current application status of cell reprogramming in the treatment of Parkinson’s disease,focusing on the use of induced pluripotent stem cells in cell replacement therapy,including preclinical animal models and progress in clinical research.The review also discusses the advancements in direct reprogramming of lineage cells in the treatment of Parkinson’s disease,as well as the controversy surrounding in vivo reprogramming.These findings suggest that cell reprogramming may hold great promise as a potential strategy for treating Parkinson’s disease.
基金supported by the National Key Research and Development Program(2016YFA0101401 and 2018YFA0801400)Major Basic Research Project of Science and Technology of Yunnan(2019FY002 and 202001BC070001)。
文摘Mutations of PTEN-induced kinase I(PINK1)cause early-onset Parkinson’s disease(PD)with selective neurodegeneration in humans.However,current PINK1 knockout mouse and pig models are unable to recapitulate the typical neurodegenerative phenotypes observed in PD patients.This suggests that generating PINK1 disease models in non-human primates(NHPs)that are close to humans is essential to investigate the unique function of PINK1 in primate brains.Paired single guide RNA(sgRNA)/Cas9-D10A nickases and truncated sgRNA/Cas9,both of which can reduce off-target effects without compromising on-target editing,are two optimized strategies in the CRISPR/Cas9 system for establishing disease animal models.Here,we combined the two strategies and injected Cas9-D10A mRNA and two truncated sgRNAs into one-cell-stage cynomolgus zygotes to target the PINK1 gene.We achieved precise and efficient gene editing of the target site in three newborn cynomolgus monkeys.The frame shift mutations of PINK1 in mutant fibroblasts led to a reduction in mRNA.However,western blotting and immunofluorescence staining confirmed the PINK1 protein levels were comparable to that in wild-type fibroblasts.We further reprogramed mutant fibroblasts into induced pluripotent stem cells(iPSCs),which showed similar ability to differentiate into dopamine(DA)neurons.Taken together,our results showed that co-injection of Cas9-D10A nickase mRNA and sgRNA into one-cell-stage cynomolgus embryos enabled the generation of human disease models in NHPs and target editing by pair truncated sgRNA/Cas9-D10A in PINK1 gene exon 2 did not impact protein expression.
基金The Major Basic Research Project of Science and Technology of YunnanGrant/Award Number:202001BC070001 and 202105AC160041+3 种基金National Natural Science Foundation of ChinaGrant/Award Number:81930121 and 31960120The National Key Research and Development Program of ChinaGrant/Award Number:2018YFA0107902 and 2018YFA0801403。
文摘Rett syndrome(RTT)is a progressive neurodevelopmental disorder that occurs mainly in girls with a range of typical symptoms of autism spectrum disorders.MeCP2 protein loss-of-function in neural lineage cells is the main cause of RTT pathogenicity.As it is still hard to understand the mechanism of RTT on the basis of only clinical patients or animal models,cell models cultured in vitro play indispensable roles.Here we reviewed the research progress in the pathogenesis of RTT at the cellular level,summarized the preclinical-research-related applications,and prospected potential future development.
基金supported by the National Key R&D Program of China(Nos.2021YFA0805700 and 2021YFA1102000)the National Natural Science Foundation of China(No.U2102204)the Natural Science Foundation of Yunnan Province,China(Nos.202001BC070001 and 202102AA100053)。
文摘Efforts have been made to establish various human pluripotent stem cell lines.However,such methods have not yet been duplicated in non-human primate cells.Here,we introduce a multiplexed single-cell sequencing technique to profile the molecular features of monkey pluripotent stem cells in published culture conditions.The results demonstrate suboptimized maintenance of pluripotency and show that the selected signaling pathways for resetting human stem cells can also be interpreted for establishing monkey cell lines.Overall,this work legitimates the translation of novel human cell line culture conditions to monkey cells and provides guidance for exploring chemical cocktails for monkey stem cell line derivation.
基金supported by grants from the Natural Science Foundation of Yunnan Province(Grant Nos.202001BC070001 and 202102AA100053)the National Natural Science Foundation of China(Grant No.32270845)+4 种基金Yunnan Fundamental Research Project(Grant No.202301AT070304)“Xingdian Talent Support Program”of Yunnan Province(Grant No.KKRD202273103)the Key Project of“Double First-Class”Initiative Joint Plan between Science and Technology of Yunnan and KUST(Grant No.202201BE070001-006)supported by grants from the US National Institutes of Health(Grant Nos.1R01AI12140301A1 and 1R01HD112473-01)Department of Defense(Grant No.ME220215).
文摘Cancer refers to a diverse collection of diseases characterized by several well-established hallmarks,including the abilities to sustain proliferative signaling,evade growth suppressors,activate invasion and metastasis,enable replicative immortality,induce angiogenesis,and resist cell death1.Historically,genetic alterations(deletions,point mutations,and translocations)were thought to be the basis for tumor formation via the inactivation of tumor suppressors and activation of oncogenes.
基金supported by grants from the National Key R&D Program of China(2021YFA0805700 and 2021YFA1102000)the National Natural Science Foundation of China(U2102204)the Natural Science Foundation of Yunnan Province(202001BC070001 and 202102AA100053).
文摘Non-human primates(NHPs)are increasingly used in preclinical trials to test the safety and efficacy of biotech-nology therapies.Nonetheless,given the ethical issues and costs associated with this model,it would be highly advantageous to use NHP cellular models in clinical studies.However,developing and maintaining the naive state of primate pluripotent stem cells(PSCs)remains difficult as does in vivo detection of PSCs,thus limiting biotech-nology application in the cynomolgus monkey.Here,we report a chemically defined,xeno-free culture system for culturing and deriving monkey PSCs in vitro.The cells display global gene expression and genome-wide hypometh-ylation patterns distinct from monkey-primed cells.We also found expression of signaling pathways components that may increase the potential for chimera formation.Crucially for biomedical applications,we were also able to integrate bioluminescent reporter genes into monkey PsCs and track them in chimeric embryos in vivo and in vitro.The engineered cells retained embryonic and extra-embryonic developmental potential.Meanwhile,we generated a chimeric monkey carrying bioluminescent cells,which were able to track chimeric cells for more than 2 years in living animals.Our study could have broad utility in primate stem cell engineering and in utilizing chimeric monkey models forclinical studies.
基金This work was supported by grants from the National Key Research and Development Program of China(2017YFA0103601)the National Natural Science Foundation of China(31730056 and 31988101)to YGC.
文摘Pig and monkey are widely used models for exploration of human diseases and evaluation of drug efficiency and toxicity,but high cost limits their uses.Organoids have been shown to be promising models for drug test as they reasonably preserve tissue structure and functions.However,colonic organoids of pig and monkey are not yet established.Here,we report a culture medium to support the growth of porcine and monkey colonic organoids.Wnt signaling and PGE2 are important for long-term expansion of the organoids,and their withdrawal results in lineage differentiation to mature cells.Furthermore,we observe that porcine colonic organoids are closer to human colonic organoids in terms of drug toxicity response.Successful establishment of porcine and monkey colonic organoids would facilitate the mechanistic investigation of the homeostatic regulation of the intestine of these animals and is useful for drug development and toxicity studies.
基金the National Natural Science Foundation of China(grant numbers 81930121 and 82125008)STI2030-Major projects(grant number 2021ZD0200900)+1 种基金the National Key Research and Development Program of China(grant numbers 2018YFA0801403 and 2018YFA0107902)the Natural Science Foundation of Yunnan Province(grant numbers 202001BC070001 and 202102AA100053)。
文摘Splice-switching antisense oligonucleotides(ASOs)and engineered U7 small nuclear ribonucleoprotein(U7 Sm OPT)are the most commonly used methods for exon skipping.However,challenges remain,such as limited organ delivery and repeated dosing for ASOs and unknown risks of by-products produced by U7 Sm OPT.Here,we showed that antisense circular RNAs(AS-circRNAs)can effectively mediate exon skipping in both minigene and endogenous transcripts.We also showed a relatively higher exon skipping efficiency at the tested Dmd minigene than U7 Sm OPT.AS-circRNA specifically targets the precursor mRNA splicing without off-target effects.Moreover,AS-circRNAs with adeno-associated virus(AAV)delivery corrected the open reading frame and restored the dystrophin expression in a mouse model of Duchenne muscular dystrophy.In conclusion,we develop an alternative method for regulating RNA splicing,which might be served as a novel tool for genetic disease treatment.
基金supported by the Natural Science Foundation of China (Grant Nos 31600920,91232718,and 91132301)the Strategic Priority Research Program of the Chinese Academy of Sciences (Grant No.XDB02030300)+1 种基金the Natural Science Foundation of Guangdong Province (No.2019A1515012134)the.Shenzhen Basic Research Project (Grant No.20220526002851001,JCYJ2017081802123707 and 20200807225058001)。
文摘Background:Aggression is a commonly hostile behavior linked to the hippocampal activity.Childhood trauma(CT)exposure has been associated with altered sensitization of the hypothalamic-pituitary-adrenal(HPA)axis and hippocampal volumewhich could increase violent aggressive behaviors.Additionally,Catechol-O-methyltransferase(COMT),the major dopamine metabolism enzyme,is impli-cated in stress responsivity,including aggression.Hence,CT exposure may affect aggression through the effect on the hippocampal function,which might also be modulated by the COMT variations.Objectives:This study examined whether both CT and haplotypes of COMT moderate hippocampal function and thus affect human aggressive behavior.Methods:We obtained bilateral hippocampal functional connectivity maps using resting state functional magnetic resonance imag-ing(MRI)data.COMT haplotype estimation was performed using Haploview 4.2 and PHASE 2.1.Then we constructed a moderated mediation model to study the effect of the CTQ×COMT on aggressive behavior.Results:Three major haplotypes were generated from thirteen single nucleotide polymorphisms(SNPs)within the COMT gene and formed three haplotypes corresponding to high,medium,and low enzymatic activity of COMT.The results showed interactive re-lationships between the Childhood Trauma Questionnaire(CTQ)and COMT with respect to the functional connectivity(FC)of the bilateral hippocampus(HIP)-orbital frontal cortex(OFC).Specifically,CT experience predicted lower negative HIP-OFC coupling in the APS and HPS haplotypes corresponding to the medium and high enzymatic activity of COMT,but greater FC in the LPS haplotypes corresponding to the low enzymatic activity.We also observed a conditional mediation effect of the right HIP-OFC coupling in the link between COMT and aggressive behavior that was moderated by CT experience.Conclusions:These results suggest that CT and COMT have a combined effect on aggressive behavior through hippocampal function.This mediation analysis sheds light on the influence of childhood experience on aggressive behavior in different genetic backgrounds.
基金This work was supported by grants from the National Natural Science Foundation of China(31988101 and 31730056)the National Key Research and Development Program of China(2017YFA0103601)to YGC.
文摘Animal models are widely used for biomedical studies and drug evaluation.The small intestine plays key roles in nutrient absorption,hormone secretion,microbiota defense and drug absorption and metabolism.Although the intestinal structure of mammals is conserved,the differences on epithelial cell composition,functional assignments and drug absorption among mammals are largely unknown.Here,cross-species analysis of single-cell transcriptomic atlas of the ileum epithelium from mouse,rat,pig,macaque and human reveals the conserved and differential cell types and functions among species,identifies a new CA7+cell type in pig,macaque and human ileum,uncovers the distinct expression pattern in enterocytes,enteroendocrine cells and Paneth cells,and defines the conserved and species-specific intestinal stem cell signature genes.The examination of drug absorption across species suggests that drug metabolism in mouse ileum is closer to human while drug transport in macaque ileum is more similar to human.Together,our data provide the comprehensive information about cell composition and functional assignments in five species,and offer the valuable guidance for animal model selection and drug testing.
基金supported by the National Natural Science Foundation of China(81930121)the National Key Research and Development Program of China(2018YFA0107902)the Major Basic Research Project of Science and Technology of Yunnan(202001BC070001).
文摘Animal models of human diseases are vital in better understanding the mechanism of pathogenesis and essential for evaluating and validating potential therapeutic interventions.As close relatives of humans,nonhuman primates(NHPs)play an increasingly indispensable role in advancing translational medicine research.In this review,we summarized the progress of NHP models generated by embryo engineering,analyzed their unique advantages in mimicking clinical patients,and discussed the remaining gap between basic research of NHP models to translational medicine.
基金supported by the Natural Science Foundation of Yunnan Province(202001BC070001,202102AA100053)the National Natural Science Foundation of China General Program(No.32070580)。
文摘Cynomolgus macaques(Macaca fascicularis)belong to the genus Macaca.They are the most widespread nonhuman primate(NHP)and share a common ancestor with humans from about 25 million years ago(Kumar and Hedges,1998).Because of their phylogenetic proximity to humans,macaques are an attractive NHP research model for a wide range of biomedical research(Yan et al.,2011).
基金supported by the National Natural Science Foundation of China(81930121)the National Key Research and Development Program of China(2018YFA0107902)the Major Basic Research Project of Science and Technology of Yunnan(202001BC070001).
文摘Animal models of human diseases are vital in better understanding the mechanism of pathogenesis and essential for evaluating and validating potential therapeutic interventions.As close relatives of humans,nonhuman primates(NHPs)play an increasingly indispensable role in advancing translational medicine research.In this review,we summarized the progress of NHP models generated by embryo engineering,analyzed their unique advantages in mimicking clinical patients,and discussed the remaining gap between basic research of NHP models to translational medicine.
基金the financial support of the National Science Foundation(ECCS-1916839 and CBET-1931777)the support of the National Institute of Health under grant number R21 HD090680-01support by the U.S.Army Research Office through the Institute for Soldier Nanotechnologies at MIT,under Contract Number W911NF-13-D-0001.
文摘Direct Ink Writing(DIW)has demonstrated great potential as a versatile method to 3D print multifunctional structures.In this work,we report the implementation of hydrogel meta-structures using DIW at room temperature,which seamlessly integrate large specific surface areas,interconnected porous characteristics,mechanical toughness,biocompatibility,and water absorption and retention capabilities.Robust but hydrophobic polymers and weakly crosslinked nature-origin hydrogels form a balance in the self-supporting ink,allowing us to directly print complex meta-structures without sacrificial materials and heating extrusion.Mechanically,the mixed bending or stretching of symmetrical re-entrant cellular lattices and the unique curvature patterns are combined to provide little lateral expansion and large compressive energy absorbance when external forces are applied on the printed meta-structures.In addition,we have successfully demonstrated ear,aortic valve conduits and hierarchical architectures.We anticipate that the reported 3D meta-structured hydrogel would offer a new strategy to develop functional biomaterials for tissue engineering applications in the future.
基金This study was supported by National Natural Science Foundation of China(Grant No.62176044)Sichuan Science and Technology Program(Grant Nos.2021YJ0186,2022YFS0178)Natural Science Foundation of Yunnan Province(GrantNos.202001BC070001,202102AA100053).
文摘Background Although postpartum depression(PPD)and non-peripartum major depressive disorder(MDD)occurring within and outside the postpartum period share many clinical characteristics,whether PPD and MDD are the same or not remains controversial.Methods The current study was devoted to identify the shared and different neural circuits between PPD and MDD by resting-state functionalmagnetic resonance imaging data from 77 participants(22 first-episodic drug-naleMDD,26 drug-nale PPD,and 29 healthy controls(HC)).Results Both the PPD andMDD groups exhibited higher fractional amplitude of low-frequency fluctuation(fALFF)in left temporal pole relative to the HC group;the MDD group showed specifically increased degree centrality in the right cerebellum while PPD showed specifically decreased fALFF in the left supplementary motor area and posterior middle temporal gyrus(pMTG_L),and specifically decreased functional connectivities between pMTG and precuneus and between left subgeneual anterior cingulate cortex(sgACC_L)and right sgACC.Moreover,sgACC and left thalamus showed abnormal regional homogeneity of functional activities between any pair of HC,MDD,and PPD.Conclusions These results provide initial evidence that PPD and MDD have common and distinct neural circuits,which may facilitate understanding the neurophysiological basis and precision treatment for PPD.
基金supported by the National Natural Science Foundation of China(82171620 and 81830043)the National Key R&D Program of China(2021YFC2701403 and 2018YFC2002201)the National High Level Hospital Clinical Research Funding(2022-PUMCH-A-205 and 2022-PUMCH-A-114)。
基金supported by the National Natural Science Foundation of China(Grant Nos.31960093,81973210,81873872,82071781,32160153)the Natural Science Foundation of Yunnan Province(Grant Nos.202001BC070001,202102AA100053,202105AD160008,202207AA110003)the Innovation Team of Chronic Kidney Disease with Integrated Traditional Chinese and Western Medicine(No.2019KCXTD014).
文摘The twenty-first century has already recorded more than ten major epidemics or pandemics of viral disease,including the devastating COVID-19.Novel effective antivirals with broad-spectrum coverage are urgently needed.Herein,we reported a novel broad-spectrum antiviral compound PAC5.Oral administration of PAC5 eliminated HBV cccDNA and reduced the large antigen load in distinct mouse models of HBV infection.Strikingly,oral administration of PAC5 in a hamster model of SARS-CoV-2 omicron(BA.1)infection significantly decreases viral loads and attenuates lung inflammation.Mechanistically,PAC5 binds to a pocket near Asp49 in the RNA recognition motif of hnRNPA2B1.PAC5-bound hnRNPA2B1 is extensively activated and translocated to the cytoplasm where it initiates the TBK1-IRF3 pathway,leading to the production of type I IFNs with antiviral activity.Our results indicate that PAC5 is a novel small-molecule agonist of hnRNPA2B1,which may have a role in dealing with emerging infectious diseases now and in the future.
基金the National Key Research and Development Program of China(2018YFA0108503)the National Natural Science Foundation of China(81760251 and 81560234)the Yunnan Provincial Natural Science Foundation(2018FB118 and KKSY201626001).
文摘Rapid detection and response to visual threats are critical for survival in animals.The amygdala(AMY)is hypothesized to be involved in this process,but how it interacts with the visual system to do this remains unclear.By recording flash-evoked potentials simultaneously from the superior colliculus(SC),lateral posterior nucleus of the thalamus,AMY,lateral geniculate nucleus(LGN)and visual cortex,which belong to the cortical and subcortical pathways for visual fear processing,we investigated the temporal relationship between these regions in visual processing in rats.A quick flash-evoked potential(FEP)component was identified in the AMY.This emerged as early as in the LGN and was approximately 25 ms prior to the earliest component recorded in the SC,which was assumed to be an important area in visual fear.This quick P1 component in the AMY was not affected by restraint stress or corticosterone injection,but was diminished by RU38486,a glucocorticoid receptor blocker.By injecting a monosynaptic retrograde AAV tracer into the AMY,we found that it received a direct projection from the retina.These results confirm the existence of a direct connection from the retina to the AMY,that the latency in the AMY to flashes is equivalent to that in the sensory thalamus,and that the response is modulated by glucocorticoids.
