Screening gene function in vivo is a powerful approach to discover novel drug targets. We present high-throughput screening （HTS） data for 3 762 distinct global gene knockout （KO） mouse lines with viable adult hom...Screening gene function in vivo is a powerful approach to discover novel drug targets. We present high-throughput screening （HTS） data for 3 762 distinct global gene knockout （KO） mouse lines with viable adult homozygous mice generated using either gene-trap or homologous recombination technologies. Bone mass was determined from DEXA scans of male and female mice at 14 weeks of age and by microCT analyses of bones from male mice at 16 weeks of age. Wild-type （WT） cagemates/littermates were examined for each gene KO. Lethality was observed in an additional 850 KO lines. Since primary HTS are susceptible to false positive findings, additional cohorts of mice from KO lines with intriguing HTS bone data were examined. Aging, ovariectomy, histomorphometry and bone strength studies were performed and possible non-skeletal phenotypes were explored. Together, these screens identified multiple genes affecting bone mass： 23 previously reported genes （Calcr, Cebpb, Crtap, Dcstamp, Dkkl, Duoxa2, Enppl, Fgf23, Kissl/Kisslr, Kl （Klotho）, Lrp5, Mstn, Neol, Npr2, Ostml, Postn, Sfrp4, S1c30a5, Sic39a13, Sost, Sumf1, Src, Wnt10b）, five novel genes extensively characterized （Cldn18, Fam20c, Lrrkl, Sgpll, Wnt16）, five novel genes with preliminary characterization （Agpat2, RassfS, Slc10a7, Stc26a7, Slc30a10） and three novel undisclosed genes coding for potential osteoporosis drug targets.展开更多
Social network structures can crucially impact complex social processes such as collective behaviour or the transmission of information and diseases. However, currently it is poorly understood how social networks chan...Social network structures can crucially impact complex social processes such as collective behaviour or the transmission of information and diseases. However, currently it is poorly understood how social networks change over time. Previous studies on primates suggest that ＇knockouts＇ （due to death or dispersal） of high-ranking individuals might be important drivers for structural changes in animal social networks. Here we test this hypothesis using long-term data on a natural population of ba- boons, examining the effects of 29 natural knockouts of alpha or beta males on adult female social networks. We investigated whether and how knockouts affected （i） changes in grooming and association rates among adult females, and （2） changes in mean degree and global clustering coefficient in these networks. The only significant effect that we found was a decrease in mean degree in grooming networks in the first month after knockouts, but this decrease was rather small, and grooming networks re- bounded to baseline levels by the second month after knockouts. Taken together our results indicate that the removal of high-ranking males has only limited or no lasting effects on social networks of adult female baboons. This finding calls into question the hypothesis that the removal of high-ranking individuals has a destabilizing effect on social network structures in social animals [Current Zoology 61 （1）： 107-113, 2015].展开更多
Effect of the specific gene knockout on the main metabolism in Escherichia coli was reviewed, and the regulation mechanisms were clarified based on different levels of information such as gene expressions, enzyme acti...Effect of the specific gene knockout on the main metabolism in Escherichia coli was reviewed, and the regulation mechanisms were clarified based on different levels of information such as gene expressions, enzyme activities, intracellular metabolite concentrations, and metabolic fluxes together with fermentation data. The effects of the knockout of such genes as pflA, pta, ppc, pykF, adhE, and ldhA on the metabolic changes were analyzed for the case under anaerobic condition. The effects of the knockout of such genes as pgi, zwf, gnd, ppc pck, pyk, and lpdA on the metabolic changes were also analyzed for the case under aerobic condition. The metabolic regulation analysis was made focusing on the roles of transcription factors.展开更多
Highlights●CRISPR/Cas9 RNP complex-based strategy demonstrates robustness and accuracy in generating gene-edited sheep.●Sheep horn development remains unaffected by partial RXFP2 knockout.●Partial RXFP2 knockout re...Highlights●CRISPR/Cas9 RNP complex-based strategy demonstrates robustness and accuracy in generating gene-edited sheep.●Sheep horn development remains unaffected by partial RXFP2 knockout.●Partial RXFP2 knockout results in unilateral cryptorchidism in sheep.展开更多
Background:C1QL3 is widely expressed in the brain and is specifically produced by a subset of excitatory neurons.However,its function is still not clear.We established C1ql3-deficient rats to investigate the role of C...Background:C1QL3 is widely expressed in the brain and is specifically produced by a subset of excitatory neurons.However,its function is still not clear.We established C1ql3-deficient rats to investigate the role of C1QL3 in the brain.Methods:C1ql3 knockout(KO)rats were generated using CRISPR/Cas9.C1ql3 KO was determined by polymerase chain reaction(PCR),DNA sequencing,and western blot-ting.Microglia morphology and cytokine expression with or without lipopolysaccha-ride(LPS)stimulus were analyzed using immunohistochemistry and real-time PCR.The brain structure changes in KO rats were examined using magnetic resonance imaging.Neuronal architecture alteration was analyzed by performing Golgi staining.Behavior was evaluated using the open field test,Morris water maze test,and Y maze test.Results:C1ql3 KO significantly increased the number of ramified microglia and decreased the number of hypertrophic microglia,whereas C1ql3 KO did not in-fluence the expression of pro-inflammatory factors and anti-inflammatory factors except IL-10.C1ql3 KO brains had more amoeboid microglia types and higher Arg-1 expression compared with the WT rats after LPS stimulation.The brain weights and HPC sizes of C1ql3 KO rats did not differ from WT rats.C1ql3 KO damaged neuronal integrity including neuron dendritic arbors and spine density.C1ql3 KO rats demonstrated an increase in spontaneous activity and an impairment in short working memory.Conclusions:C1ql3 KO not only interrupts the neuronal integrity but also affects the microglial activation,resulting in hyperactive behavior and impaired short memory in rats,which highlights the role of C1QL3 in the regulation of structure and function of both neuronal and microglial cells.展开更多
Understanding bacterial strategies for coping with heavy metal stress is essential for elucidating their resilience in contaminated environments.However,whether cell wall exfoliation contributes to bacterial tolerance...Understanding bacterial strategies for coping with heavy metal stress is essential for elucidating their resilience in contaminated environments.However,whether cell wall exfoliation contributes to bacterial tolerance under heavy metal stress,such as cadmium(Cd)exposure,remains unclear and requires further investigation.In this study,we reveal a novel self-protective mechanism in Stenotrophomonas sp.H225 isolated from a Cd-contaminated farmland soil,which underwent controlled cell wall exfoliation and regeneration in response to Cd stress up to 200 mg L^(-1).Transmission electron microscopy and energy-dispersive X-ray spectroscopy analyses revealed that the exfoliated cell wall fragments served as extracellular Cd sinks,thereby reducing intracellular Cd accumulation.Fourier-transform infrared spectroscopy and enzyme-linked immunosorbent assay indicated progressive peptidoglycan(PG)degradation,with exfoliated PG concentration in solution increasing from 148 ng mL^(-1) at 0 mg L^(-1) Cd to 240 ng mL^(-1) at 200 mg L^(-1) Cd.This degradation was counteracted by the compensatory upregulation of PG biosynthesis genes,with the enrichment ratio reaching up to 0.83,facilitating cell wall reconstruction.Transcriptomic analysis and gene knockout experiments identified mtgA(encoding a monofunctional transglycosylase)as a key determinant in cell wall repair and Cd resistance.To our knowledge,this is the first mechanistic evidence that bacteria can mitigate heavy metal toxicity through dynamic cell wall remodeling involving exfoliation and regeneration.This finding enhances our understanding of microbial survival strategies under environmental stress and highlights potential targets for engineering metal-tolerant strains for bioremediation applications.展开更多
Objective To investigate the mechanism of in alleviating colonic mucosal inflammation in ten-eleven translocation(TET)protein 2 gene knockout(TET2^(-/-))mice with ulcerative colitis(UC)by regulating DNA methyltransfer...Objective To investigate the mechanism of in alleviating colonic mucosal inflammation in ten-eleven translocation(TET)protein 2 gene knockout(TET2^(-/-))mice with ulcerative colitis(UC)by regulating DNA methyltransferase(DNMT)and DNA hydroxymethylase.Methods Male specific pathogen-free(SPF)grade C57BL/6J wild-type(WT)mice(n=8)and TET2^(-/-)mice(n=20)were used to establish UC models by freely drinking 3%dextran sulfate sodium solution for 7 d.After UC model validation through histopathological examination in two mice from each type,the remaining mice were divided into four groups(n=6 in each group):WT model(WT+UC),TET2^(-/-)model(TET2^(-/-)+UC),TET2^(-/-)mild moxibustion(TET2^(-/-)+MM),and TET2^(-/-)electroacupuncture(TET2^(-/-)+EA)groups.TET2^(-/-)+MM group received mild moxibustion on Tianshu(ST25)and Qihai(CV6)for 10 min daily for 7 d.The TET2^(-/-)+EA group also applied electroacupuncture(1 mA,2/100 Hz)at the same acupoints for 10 min daily for 7 d.The disease activity index(DAI)scores of each group of mice were accessed daily.The colon lengths of mice in groups were measured following intervention.The pathological changes in the colon tissues were observed with hematoxylin and eosin(HE)staining.The concentrations of interleukin(IL)-6,C-C motif chemokine 17(CCL17),and C-X-C motif chemokine ligand 10(CXCL10)in serum were detected by enzyme-linked immunosorbent assay(ELISA).The expression of DNMT proteins(DNMT1,DNMT3A,and DNMT3B)in the colon tissues was detected by immunohistochemistry.The expression of 5-methylcytosine(5-mC),5-hydroxymethylcytosine(5-hmC),histone deacetylase 2(HDAC2),and DNA hydroxymethylase family proteins(TET 1 and TET3)was detected using immunofluorescence,which also determined the co-localization of TET1 and IL-6 protein.Results Compared with WT+UC group,TET2^(-/-)+UC group exhibited significantly higher DAI scores and shorter colon lengths(P<0.01).Both mild moxibustion and electroacupuncture significantly decreased DAI scores and ameliorated colon shortening in TET2^(-/-)mice(P<0.001).Histopathological scores of TET2^(-/-)+UC mice were significantly higher than those of WT+UC group(P<0.001)and were significantly reduced after both mild moxibustion and electroacupuncture interventions(P<0.001).Serum levels of IL-6,CCL17,and CXCL10 were significantly elevated in TET2^(-/-)+UC group compared with WT+UC group(P<0.001).Mild moxibustion significantly reduced IL-6,CCL17,and CXCL10 levels(P<0.001,P<0.001,and P<0.01,respectively),while electroacupuncture also significantly reduced IL-6,CCL17,and CXCL10 levels(P<0.05,P<0.01,and P<0.01,respectively).TET2^(-/-)+UC mice showed increased expression levels of DNMT1,DNMT3A,DNMT3B,and 5-mC(P<0.05,P<0.01 and P<0.001,respectively),with decreased expression levels of TET1,TET3,5-hmC,and HDAC2(P<0.001).Mild moxibustion significantly reduced DNMT1,DNMT3B,and 5-mC levels(P<0.05,P<0.01,and P<0.001,respectively),while increasing expression levels of TET1,TET3,5-hmC,and HDAC2(P<0.001,P<0.001,P<0.05,and P<0.001,respectively).Electroacupuncture significantly decreased 5-mC and DNMT3B levels(P<0.001 and P<0.01,respectively)and increased 5-hmC and HDAC2 levels(P<0.05 and P<0.001,respectively),but did not significantly affect TET1 and TET3 expression(P>0.05).Compared with TET2^(-/-)+MM group,TET2^(-/-)+EA group showed significantly higher 5-mC expression(P<0.001).TET2^(-/-)+UC group exhibited markedly increased IL-6 expression and higher co-localization of TET1 and IL-6 in mucosal epithelium,whereas minimal IL-6 expression was observed in the other groups.Conclusion Mild moxibustion and electroacupuncture significantly ameliorate colonic inflammation exacerbated by TET2 deficiency in UC mice via epigenetic modulation.Distinct mechanisms exist between the two interventions:mild moxibustion regulates both DNMT and hydroxymethylase,whereas electroacupuncture primarily affects DNMT.展开更多
Magnaporthe oryzae,the causal agent of rice blast,induces significant upregulation of OsPR10b,a pathogenesis-related(PR)pollen allergen(BetV-1)family gene.To investigate its role in immunity,we generated OsPR10b knock...Magnaporthe oryzae,the causal agent of rice blast,induces significant upregulation of OsPR10b,a pathogenesis-related(PR)pollen allergen(BetV-1)family gene.To investigate its role in immunity,we generated OsPR10b knockout mutants in the Zhonghua 11(ZH11)background.OsPR10b was predominantly expressed in rice calli and strongly induced by M.oryzae infection.Knockout mutants(ospr10b-1 and ospr10b-2)exhibited heightened susceptibility to both M.oryzae and Xanthomonas oryzae pv.oryzae(Xoo),demonstrating that OsPR10b positively regulates resistance to blast and bacterial blight.Our findings elucidate OsPR10b’s role in rice immunity and provide genetic resources for disease-resistant breeding.展开更多
The crosstalk between megakaryocytic lineage cells and the skeletal system has just begun to be explored but remains largely elusive.Using conditional gene knockout mouse models,we demonstrated that loss of Beclin 1(B...The crosstalk between megakaryocytic lineage cells and the skeletal system has just begun to be explored but remains largely elusive.Using conditional gene knockout mouse models,we demonstrated that loss of Beclin 1(Becn1),a major regulator of mammalian autophagy,exclusively in the megakaryocytic lineage disrupted autophagy in platelets but did not compromise megakaryopoiesis or the formation and function of platelets.Unexpectedly,conditional Becn1 deletion in male mice led to a remarkable increase in bone mass with improved bone quality,in association with a decrease in sex hormone binding globulin(SHBG)and an increase in free testosterone(FT).In vivo Becn1 overexpression in megakaryocytic lineage-specific cells reduced bone mass and quality,along with an increase in SHBG and a decrease in FT.Transplantation of wild-type bone marrow cells into megakaryocytic lineage Becn1-deficient male mice restored bone mass and normalized SHBG and FT.Furthermore,bilateral orchiectomy of Becn1^(f/f);Pf4-iCre mice,which are crippled with the production of testosterone,resulted in a reduction in bone mass and quality,whereas in vivo overexpression of SHBG,specifically in the liver of Becn1^(f/f);Pf4-iCre mice,decreased FT and reduced bone mass and quality.In addition,metformin treatment,which induces SHBG expression,reduced FT and normalized bone mass in Becn1^(f/f);Pf4-iCre mice.We thus concluded that Becn1 of the megakaryocytic lineage is dispensable locally for platelet hemostasis but limits bone mass by increasing SHBG,which in turn reduces the FT of male mice.Our findings highlight a mechanism by which Becn1 from megakaryocytic lineage cells distally balances bone growth.展开更多
Membrane-initiated estrogen receptorα(mERα)signaling has been shown to affect bone mass in murine models.However,it remains unknown which cell types mediate the mERα-dependent effects on bone.In this study,we gener...Membrane-initiated estrogen receptorα(mERα)signaling has been shown to affect bone mass in murine models.However,it remains unknown which cell types mediate the mERα-dependent effects on bone.In this study,we generated a novel mouse model with a conditional C451A mutation in Esr1,which enables selective knockout of the palmitoylation site essential for the membrane localization of ERα(C451A^(f/f)).First,we used Runx2-Cre mice to generate Runx2-C451A^(f/f)mice with conditional inactivation of mERαsignaling in Runx2-expressing osteoblast lineage cells.No significant changes were observed in body weight,weights of estrogen-responsive organs,or serum concentrations of estradiol between female Runx2-C451A^(f/f)and homozygous C451A^(f/f)littermate controls.High-resolution microcomputed tomography analysis showed a consistent decrease in cortical bone mass in the tibia,femur,and vertebra L5 of Runx2-C451A^(f/f)mice and three-point bending analysis of humerus revealed an impaired mechanical bone strength in Runx2-C451A^(f/f)female mice compared to controls.Additionally,primary osteoblast cultures from mice lacking mERαsignaling showed impaired differentiation compared to controls.展开更多
Intervertebral disc degeneration(IDD)is a progressive and dynamic process in which the senescence-associated secretory phenotype(SASP)of nucleus pulposus cells(NPC)plays a significant role.While impaired chaperone-med...Intervertebral disc degeneration(IDD)is a progressive and dynamic process in which the senescence-associated secretory phenotype(SASP)of nucleus pulposus cells(NPC)plays a significant role.While impaired chaperone-mediated autophagy(CMA)has been associated with inflammation and cellular senescence,its specific involvement in the self-perpetuating feedback loop of NPC senescence remains poorly understood.Through LAMP2A knockout in NPC,we identified a significant upregulation of DYRK1A,a core mediator of premature senescence in Down syndrome.Subsequent validation established DYRK1A as the critical driver of premature senescence in CMA-deficient NPC.Combinatorial transcription factor analysis revealed that under IL1B stimulation or CMA inhibition,elevated DYRK1A promoted FOXC1 phosphorylation and nuclear translocation,initiating transcriptional activation of cell cycle arrest.Intriguingly,CMA impairment concurrently enhanced glutamine metabolic flux in senescent NPC,thereby augmenting their survival fitness.Transcriptomic profiling demonstrated that CMA reactivation in senescent NPC facilitated fate transition from senescence to apoptosis,mediated by decreased glutamine flux via GLUL degradation.Therefore,CMA exerts protective effects against IDD by maintaining equilibrium between premature senescence and senolysis.This study elucidates CMA’s regulatory role in SASP-mediated senescence amplification circuits,providing novel therapeutic insights for IDD and other age-related pathologies.展开更多
Background Genome editing has been considered as powerful tool in agricultural fields.However,genome editing progress in cattle has not been fast as in other mammal species,for some disadvantages including long gestat...Background Genome editing has been considered as powerful tool in agricultural fields.However,genome editing progress in cattle has not been fast as in other mammal species,for some disadvantages including long gestational periods,single pregnancy,and high raising cost.Furthermore,technically demanding methods such as microinjection and somatic cell nuclear transfer(SCNT)are needed for gene editing in cattle.In this point of view,electroporation in embryos has been risen as an alternative.Results First,editing efficiency of our electroporation methods were tested for embryos.Presence of mutation on embryo was confirmed by T7E1 assay.With first combination,mutation rates for MSTN and PRNP were 57.6%±13.7%and 54.6%±13.5%,respectively.In case of MSTN/BLG,mutation rates were 83.9%±23.6%for MSTN,84.5%±18.0%for BLG.Afterwards,the double-KO embryos were transferred to surrogates and mutation rate was identified in resultant calves by targeted deep sequencing.Thirteen recipients were transferred for MSTN/PRNP,4 calves were delivered,and one calf underwent an induction for double KO.Ten surrogates were given double-KO embryos for MSTN/BLG,and four of the six calves that were born had mutations in both genes.Conclusions These data demonstrated that production of genome edited cattle via electroporation of RNP could be effectively applied.Finally,MSTN and PRNP from beef cattle and MSTN and BLG from dairy cattle have been born and they will be valuable resources for future precision breeding.展开更多
Aging is an inevitable physiological process,often accompanied by age-related bone loss and subsequent bone-related diseases that pose serious health risks.Research on skeletal diseases caused by aging in humans is ch...Aging is an inevitable physiological process,often accompanied by age-related bone loss and subsequent bone-related diseases that pose serious health risks.Research on skeletal diseases caused by aging in humans is challenging due to lengthy study durations,difficulties in sampling,regional variability,and substantial investment.Consequently,mice are preferred for such studies due to their similar motor system structure and function to humans,ease of handling and care,low cost,and short generation time.In this review,we present a comprehensive overview of the characteristics,limitations,applicability,bone phenotypes,and treatment methods in naturally aging mice and prematurely aging mouse models(including SAMP6,POLG mutant,LMNA,SIRT6,ZMPSTE24,TFAM,ERCC1,WERNER,and KL/KL-deficient mice).We also summarize the molecular mechanisms of these aging mouse models,including cellular DNA damage response,senescence-related secretory phenotype,telomere shortening,oxidative stress,bone marrow mesenchymal stem cell(BMSC)abnormalities,and mitochondrial dysfunction.Overall,this review aims to enhance our understanding of the pathogenesis of aging-related bone diseases.展开更多
Neonatal hypoxic-ischemic encephalopathy is often associated with permanent cerebral palsy,neurosensory impairments,and cognitive deficits,and there is no effective treatment for complications related to hypoxic-ische...Neonatal hypoxic-ischemic encephalopathy is often associated with permanent cerebral palsy,neurosensory impairments,and cognitive deficits,and there is no effective treatment for complications related to hypoxic-ischemic encephalopathy.The therapeutic potential of human placental chorionic plate-derived mesenchymal stem cells for various diseases has been explored.However,the potential use of human placental chorionic plate-derived mesenchymal stem cells for the treatment of neonatal hypoxic-ischemic encephalopathy has not yet been investigated.In this study,we injected human placental chorionic plate-derived mesenchymal stem cells into the lateral ventricle of a neonatal hypoxic-ischemic encephalopathy rat model and observed significant improvements in both cognitive and motor function.Protein chip analysis showed that interleukin-3 expression was significantly elevated in neonatal hypoxic-ischemic encephalopathy model rats.Following transplantation of human placental chorionic plate-derived mesenchymal stem cells,interleukin-3 expression was downregulated.To further investigate the role of interleukin-3 in neonatal hypoxic-ischemic encephalopathy,we established an in vitro SH-SY5Y cell model of hypoxic-ischemic injury through oxygen-glucose deprivation and silenced interleukin-3 expression using small interfering RNA.We found that the activity and proliferation of SH-SY5Y cells subjected to oxygen-glucose deprivation were further suppressed by interleukin-3 knockdown.Furthermore,interleukin-3 knockout exacerbated neuronal damage and cognitive and motor function impairment in rat models of hypoxic-ischemic encephalopathy.The findings suggest that transplantation of hpcMSCs ameliorated behavioral impairments in a rat model of hypoxic-ischemic encephalopathy,and this effect was mediated by interleukin-3-dependent neurological function.展开更多
Tumor necrosis factorα(TNFα)exhibits diverse biological functions;however,its regulatory roles in myogenesis are not fully understood.In the present study,we explored the function of TNFαin myoblast proliferation,d...Tumor necrosis factorα(TNFα)exhibits diverse biological functions;however,its regulatory roles in myogenesis are not fully understood.In the present study,we explored the function of TNFαin myoblast proliferation,differentiation,migration,and myotube fusion in primary myoblasts and C2C12 cells.To this end,we constructed TNFαmuscle-conditional knockout(TNFα-CKO)mice and compared them with flox mice to assess the effects of TNFαknockout on skeletal muscles.Results indicated that TNFα-CKO mice displayed phenotypes such as accelerated muscle development,enhanced regenerative capacity,and improved exercise endurance compared to flox mice,with no significant differences observed in major visceral organs or skeletal structure.Using label-free proteomic analysis,we found that TNFα-CKO altered the distribution of several muscle development-related proteins,such as Hira,Casz1,Casp7,Arhgap10,Gas1,Diaph1,Map3k20,Cfl2,and Igf2,in the nucleus and cytoplasm.Gene set enrichment analysis(GSEA)further revealed that TNFαdeficiency resulted in positive enrichment in oxidative phosphorylation and MyoD targets and negative enrichment in JAK-STAT signaling.These findings suggest that TNFα-CKO positively regulates muscle growth and development,possibly via these newly identified targets and pathways.展开更多
Neuropeptide Y receptor Y8(NPY8R)is a fish-specific receptor with two subtypes,NPY8AR and NPY8BR.Changes in expression levels during physiological processes or in vivo regulation after ventricular injection suggest th...Neuropeptide Y receptor Y8(NPY8R)is a fish-specific receptor with two subtypes,NPY8AR and NPY8BR.Changes in expression levels during physiological processes or in vivo regulation after ventricular injection suggest that NPY8BR plays an important role in feeding regulation;this has been found in only a few fish,at present.In order to better understand the physiological function of npy8br,especially in digestion,we used clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)technology to generate npy8br-/-japanese medaka(Oryzias latipes).We found that the deletion of npy8br in medaka larvae affected their feeding and digestion ability,ultimately affecting their growth.Specifically,npy8br deficiency in medaka larvae resulted in decreased feed intake and decreased expression levels of orexigenic genes(npy and agrp).npy8br-/-medaka larvae fed for 10 d(10th day of feeding)still had incompletely digested brine shrimp(Artemia nauplii)in the digestive tract 8 h after feeding,the messenger RNA(mRNA)expression levels of digestion-related genes(amy,lpl,ctra,and ctrb)were significantly decreased,and the activity of amylase,trypsin,and lipase also significantly decreased.The deletion of npy8br in medaka larvae inhibited the growth and significantly decreased the expression of growth-related genes(gh and igf1).Hematoxylin and eosin(H&E)sections of intestinal tissue showed that npy8br-/-medaka larvae had damaged intestine,thinned intestinal wall,and shortened intestinal villi.So far,this is the first npy8br gene knockout model established in fish and the first demonstration that npy8br plays an important role in digestion.展开更多
Butyrylcholinesterase(BChE;EC 3.1.1.8),an enzyme structurally related to acetylcholinesterase,is widely distributed in the human body.It plays a role in the detoxification of chemicals such as succinylcholine,a muscle...Butyrylcholinesterase(BChE;EC 3.1.1.8),an enzyme structurally related to acetylcholinesterase,is widely distributed in the human body.It plays a role in the detoxification of chemicals such as succinylcholine,a muscle relaxant used in anesthetic practice.BChE is well-known due to variant forms of the enzyme with little or no hydrolytic activity which exist in some endogamous communities and result in prolonged apnea following the administration of succinylcholine.Its other functions include the ability to hydrolyze acetylcholine,the cholinergic neurotransmitter in the brain,when its primary hydrolytic enzyme,acetylcholinesterase,is absent.To assess its potential roles,BChE was studied in relation to insulin resistance,type 2 diabetes mellitus,cognition,hepatic disorders,cardiovascular and cerebrovascular diseases,and inflammatory conditions.Individuals who lack the enzyme activity of BChE are otherwise healthy,until they are given drugs hydrolyzed by this enzyme.Therefore,BChE is a candidate for the study of loss-of-function mutations in humans.Studying individuals with variant forms of BChE can provide insights into whether they are protected against metabolic diseases.The potential utility of the enzyme as a biomarker for Alzheimer’s disease and the response to its drug treatment can also be assessed.展开更多
BACKGROUND Currently,intrahepatic cholangiocarcinoma(ICC)poses a continuing,significant health challenge,but the relationship has yet to be established between ICC and the proteasome 26S subunit non-ATPase 6(PSMD6).AI...BACKGROUND Currently,intrahepatic cholangiocarcinoma(ICC)poses a continuing,significant health challenge,but the relationship has yet to be established between ICC and the proteasome 26S subunit non-ATPase 6(PSMD6).AIM To investigate the protein expression and clinicopathological significance of PSMD6 in ICC.METHODS The potential impact of the PSMD6 gene on the growth of ICC cell lines was analyzed using clustered regularly interspaced short palindromic repeat knockout screening technology.Forty-two paired specimens of ICC and adjacent noncancerous tissues were collected.PSMD6 protein expression was determined by immunohistochemistry.Receiver operating characteristic curve analysis was performed to validate PSMD6 expression level,and its association with ICC patients’various clinicopathological characteristics was investigated.RESULTS The PSMD6 gene was found to be essential for the growth of ICC cell lines.PSMD6 protein was significantly overexpressed in ICC tissues(P<0.001),but showed no significant association with patient age,gender,pathological grade,or tumor-node-metastasis stage(P>0.05).CONCLUSION PSMD6 can promote the growth of ICC cells,thus playing a pro-oncogenic role.展开更多
Background:A multitude of mouse models are utilized to emulate and study intestinal inflammation.T-cell receptor alpha chain(TCRα)-deficient mice are used as a model of spontaneous colitis that has similarities with ...Background:A multitude of mouse models are utilized to emulate and study intestinal inflammation.T-cell receptor alpha chain(TCRα)-deficient mice are used as a model of spontaneous colitis that has similarities with human ulcerative colitis.However,colitis is triggered late in the life of the mouse(age:4-5 months),and inflammation does not develop at the same time in different mice.A previously conducted study reported that the administration of the drug piroxicam triggered predictable and early colitis in TCRα-deficient mice at the age of 6-8 weeks.However,a detailed characterization of ensuing inflammation was not provided.Methods:We conducted an in-depth examination of piroxicam-triggered colitis in TCRα-deficient mice,with emphasis on spatial histopathologic changes and analysis of expression of inflammatory markers.Furthermore,we tested amelioration of colitis with dexamethasone.Results:We confirmed that piroxicam induced a time-prescribed colitis and did so in the proximal colon as well as the cecum of TCRα-deficient mice.Piroxicam adminis-tration was observed to induce epithelial hyperplasia,goblet cell loss,and leukocyte infiltration with occasional ulceration.A Swiss roll technique was used to examine the colon and cecum in its entirety.Importantly,we observed that inflammation was mul-tifocal segmental,with areas of tissue damage in between healthy tissue.In addition,we observed variability in the severity of inflammation among replicate animals and treatments,and that the administration of dexamethasone only partially ameliorated inflammation in the proximal colon.Conclusions:Piroxicam consistently induced multifocal segmental colitis in the proxi-mal colon and cecum,although the degree of inflammation was reduced in the latter.Importantly,spatial variability in inflammation in the large intestine and the inter-replicate variation in the severity of inflammation must be taken into consideration when utilizing this murine model of synchronized colitis.展开更多
Williams syndrome(WS)is a rare multisystemic disorder caused by recurrent microdeletions on 7q11.23,characterized by intellectual disability,distinctive craniofacial and dental features,and cardiovascular problems.Pre...Williams syndrome(WS)is a rare multisystemic disorder caused by recurrent microdeletions on 7q11.23,characterized by intellectual disability,distinctive craniofacial and dental features,and cardiovascular problems.Previous studies have explored the roles of individual genes within these microdeletions in contributing to WS phenotypes.Here,we report five patients with WS with 1.4 Mb-1.5 Mb microdeletions that include RFC2,as well as one patient with a 167-kb microdeletion involving RFC2 and six patients with intragenic variants within RFC2.To investigate the potential involvement of RFC2 in WS pathogenicity,we generate a rfc2 knockout(KO)zebrafish using CRISPR-Cas9 technology.Additionally,we generate a KO zebrafish of its paralog gene,rfc5,to better understand the functions of these RFC genes in development and disease.Both rfc2 and rfc5 KO zebrafish exhibit similar phenotypes reminiscent of WS,including small head and brain,jaw and dental defects,and vascular problems.RNA-seq analysis reveals that genes associated with neural cell survival and differentiation are specifically affected in rfc2 KO zebrafish.In addition,heterozygous rfc2 KO adult zebrafish demonstrate an anxiety-like behavior with increased social cohesion.These results suggest that RFC2 may contribute to the pathogenicity of WS,as evidenced by the zebrafish model.展开更多
文摘Screening gene function in vivo is a powerful approach to discover novel drug targets. We present high-throughput screening （HTS） data for 3 762 distinct global gene knockout （KO） mouse lines with viable adult homozygous mice generated using either gene-trap or homologous recombination technologies. Bone mass was determined from DEXA scans of male and female mice at 14 weeks of age and by microCT analyses of bones from male mice at 16 weeks of age. Wild-type （WT） cagemates/littermates were examined for each gene KO. Lethality was observed in an additional 850 KO lines. Since primary HTS are susceptible to false positive findings, additional cohorts of mice from KO lines with intriguing HTS bone data were examined. Aging, ovariectomy, histomorphometry and bone strength studies were performed and possible non-skeletal phenotypes were explored. Together, these screens identified multiple genes affecting bone mass： 23 previously reported genes （Calcr, Cebpb, Crtap, Dcstamp, Dkkl, Duoxa2, Enppl, Fgf23, Kissl/Kisslr, Kl （Klotho）, Lrp5, Mstn, Neol, Npr2, Ostml, Postn, Sfrp4, S1c30a5, Sic39a13, Sost, Sumf1, Src, Wnt10b）, five novel genes extensively characterized （Cldn18, Fam20c, Lrrkl, Sgpll, Wnt16）, five novel genes with preliminary characterization （Agpat2, RassfS, Slc10a7, Stc26a7, Slc30a10） and three novel undisclosed genes coding for potential osteoporosis drug targets.
文摘Social network structures can crucially impact complex social processes such as collective behaviour or the transmission of information and diseases. However, currently it is poorly understood how social networks change over time. Previous studies on primates suggest that ＇knockouts＇ （due to death or dispersal） of high-ranking individuals might be important drivers for structural changes in animal social networks. Here we test this hypothesis using long-term data on a natural population of ba- boons, examining the effects of 29 natural knockouts of alpha or beta males on adult female social networks. We investigated whether and how knockouts affected （i） changes in grooming and association rates among adult females, and （2） changes in mean degree and global clustering coefficient in these networks. The only significant effect that we found was a decrease in mean degree in grooming networks in the first month after knockouts, but this decrease was rather small, and grooming networks re- bounded to baseline levels by the second month after knockouts. Taken together our results indicate that the removal of high-ranking males has only limited or no lasting effects on social networks of adult female baboons. This finding calls into question the hypothesis that the removal of high-ranking individuals has a destabilizing effect on social network structures in social animals [Current Zoology 61 （1）： 107-113, 2015].
文摘Effect of the specific gene knockout on the main metabolism in Escherichia coli was reviewed, and the regulation mechanisms were clarified based on different levels of information such as gene expressions, enzyme activities, intracellular metabolite concentrations, and metabolic fluxes together with fermentation data. The effects of the knockout of such genes as pflA, pta, ppc, pykF, adhE, and ldhA on the metabolic changes were analyzed for the case under anaerobic condition. The effects of the knockout of such genes as pgi, zwf, gnd, ppc pck, pyk, and lpdA on the metabolic changes were also analyzed for the case under aerobic condition. The metabolic regulation analysis was made focusing on the roles of transcription factors.
基金supported by the National Key Research and Development Program of China(2022YFD1300200)the National Natural Science Foundation of China(32161143010,32202646,and 32272848)+2 种基金the China Agriculture Research System(CARS-39)the Key Special Project of Ningxia Science and Technology Department,China(2021BEF02024)the local grants,China(NXTS2021-001,2022GD-TSLD-46,NK2022010207,and NXTS2022-001)。
文摘Highlights●CRISPR/Cas9 RNP complex-based strategy demonstrates robustness and accuracy in generating gene-edited sheep.●Sheep horn development remains unaffected by partial RXFP2 knockout.●Partial RXFP2 knockout results in unilateral cryptorchidism in sheep.
基金The present work was supported by the National Natural Science Foundation(31970508)the National Key Research and Development Program of China(2022YFF0710702).
文摘Background:C1QL3 is widely expressed in the brain and is specifically produced by a subset of excitatory neurons.However,its function is still not clear.We established C1ql3-deficient rats to investigate the role of C1QL3 in the brain.Methods:C1ql3 knockout(KO)rats were generated using CRISPR/Cas9.C1ql3 KO was determined by polymerase chain reaction(PCR),DNA sequencing,and western blot-ting.Microglia morphology and cytokine expression with or without lipopolysaccha-ride(LPS)stimulus were analyzed using immunohistochemistry and real-time PCR.The brain structure changes in KO rats were examined using magnetic resonance imaging.Neuronal architecture alteration was analyzed by performing Golgi staining.Behavior was evaluated using the open field test,Morris water maze test,and Y maze test.Results:C1ql3 KO significantly increased the number of ramified microglia and decreased the number of hypertrophic microglia,whereas C1ql3 KO did not in-fluence the expression of pro-inflammatory factors and anti-inflammatory factors except IL-10.C1ql3 KO brains had more amoeboid microglia types and higher Arg-1 expression compared with the WT rats after LPS stimulation.The brain weights and HPC sizes of C1ql3 KO rats did not differ from WT rats.C1ql3 KO damaged neuronal integrity including neuron dendritic arbors and spine density.C1ql3 KO rats demonstrated an increase in spontaneous activity and an impairment in short working memory.Conclusions:C1ql3 KO not only interrupts the neuronal integrity but also affects the microglial activation,resulting in hyperactive behavior and impaired short memory in rats,which highlights the role of C1QL3 in the regulation of structure and function of both neuronal and microglial cells.
基金partially supported by the National Natural Science Foundation of China (Nos. 42377004 and 41991334)the Fundamental Research Funds for the Central Universities (No. 226-2025-0004)+1 种基金the China Agriculture Research System (No. CARS-01)the opportunity granted by the China Scholarship Council (No. 202406320448)
文摘Understanding bacterial strategies for coping with heavy metal stress is essential for elucidating their resilience in contaminated environments.However,whether cell wall exfoliation contributes to bacterial tolerance under heavy metal stress,such as cadmium(Cd)exposure,remains unclear and requires further investigation.In this study,we reveal a novel self-protective mechanism in Stenotrophomonas sp.H225 isolated from a Cd-contaminated farmland soil,which underwent controlled cell wall exfoliation and regeneration in response to Cd stress up to 200 mg L^(-1).Transmission electron microscopy and energy-dispersive X-ray spectroscopy analyses revealed that the exfoliated cell wall fragments served as extracellular Cd sinks,thereby reducing intracellular Cd accumulation.Fourier-transform infrared spectroscopy and enzyme-linked immunosorbent assay indicated progressive peptidoglycan(PG)degradation,with exfoliated PG concentration in solution increasing from 148 ng mL^(-1) at 0 mg L^(-1) Cd to 240 ng mL^(-1) at 200 mg L^(-1) Cd.This degradation was counteracted by the compensatory upregulation of PG biosynthesis genes,with the enrichment ratio reaching up to 0.83,facilitating cell wall reconstruction.Transcriptomic analysis and gene knockout experiments identified mtgA(encoding a monofunctional transglycosylase)as a key determinant in cell wall repair and Cd resistance.To our knowledge,this is the first mechanistic evidence that bacteria can mitigate heavy metal toxicity through dynamic cell wall remodeling involving exfoliation and regeneration.This finding enhances our understanding of microbial survival strategies under environmental stress and highlights potential targets for engineering metal-tolerant strains for bioremediation applications.
基金National Natural Science Foundation of China(82274641,81873372,and 82105012).
文摘Objective To investigate the mechanism of in alleviating colonic mucosal inflammation in ten-eleven translocation(TET)protein 2 gene knockout(TET2^(-/-))mice with ulcerative colitis(UC)by regulating DNA methyltransferase(DNMT)and DNA hydroxymethylase.Methods Male specific pathogen-free(SPF)grade C57BL/6J wild-type(WT)mice(n=8)and TET2^(-/-)mice(n=20)were used to establish UC models by freely drinking 3%dextran sulfate sodium solution for 7 d.After UC model validation through histopathological examination in two mice from each type,the remaining mice were divided into four groups(n=6 in each group):WT model(WT+UC),TET2^(-/-)model(TET2^(-/-)+UC),TET2^(-/-)mild moxibustion(TET2^(-/-)+MM),and TET2^(-/-)electroacupuncture(TET2^(-/-)+EA)groups.TET2^(-/-)+MM group received mild moxibustion on Tianshu(ST25)and Qihai(CV6)for 10 min daily for 7 d.The TET2^(-/-)+EA group also applied electroacupuncture(1 mA,2/100 Hz)at the same acupoints for 10 min daily for 7 d.The disease activity index(DAI)scores of each group of mice were accessed daily.The colon lengths of mice in groups were measured following intervention.The pathological changes in the colon tissues were observed with hematoxylin and eosin(HE)staining.The concentrations of interleukin(IL)-6,C-C motif chemokine 17(CCL17),and C-X-C motif chemokine ligand 10(CXCL10)in serum were detected by enzyme-linked immunosorbent assay(ELISA).The expression of DNMT proteins(DNMT1,DNMT3A,and DNMT3B)in the colon tissues was detected by immunohistochemistry.The expression of 5-methylcytosine(5-mC),5-hydroxymethylcytosine(5-hmC),histone deacetylase 2(HDAC2),and DNA hydroxymethylase family proteins(TET 1 and TET3)was detected using immunofluorescence,which also determined the co-localization of TET1 and IL-6 protein.Results Compared with WT+UC group,TET2^(-/-)+UC group exhibited significantly higher DAI scores and shorter colon lengths(P<0.01).Both mild moxibustion and electroacupuncture significantly decreased DAI scores and ameliorated colon shortening in TET2^(-/-)mice(P<0.001).Histopathological scores of TET2^(-/-)+UC mice were significantly higher than those of WT+UC group(P<0.001)and were significantly reduced after both mild moxibustion and electroacupuncture interventions(P<0.001).Serum levels of IL-6,CCL17,and CXCL10 were significantly elevated in TET2^(-/-)+UC group compared with WT+UC group(P<0.001).Mild moxibustion significantly reduced IL-6,CCL17,and CXCL10 levels(P<0.001,P<0.001,and P<0.01,respectively),while electroacupuncture also significantly reduced IL-6,CCL17,and CXCL10 levels(P<0.05,P<0.01,and P<0.01,respectively).TET2^(-/-)+UC mice showed increased expression levels of DNMT1,DNMT3A,DNMT3B,and 5-mC(P<0.05,P<0.01 and P<0.001,respectively),with decreased expression levels of TET1,TET3,5-hmC,and HDAC2(P<0.001).Mild moxibustion significantly reduced DNMT1,DNMT3B,and 5-mC levels(P<0.05,P<0.01,and P<0.001,respectively),while increasing expression levels of TET1,TET3,5-hmC,and HDAC2(P<0.001,P<0.001,P<0.05,and P<0.001,respectively).Electroacupuncture significantly decreased 5-mC and DNMT3B levels(P<0.001 and P<0.01,respectively)and increased 5-hmC and HDAC2 levels(P<0.05 and P<0.001,respectively),but did not significantly affect TET1 and TET3 expression(P>0.05).Compared with TET2^(-/-)+MM group,TET2^(-/-)+EA group showed significantly higher 5-mC expression(P<0.001).TET2^(-/-)+UC group exhibited markedly increased IL-6 expression and higher co-localization of TET1 and IL-6 in mucosal epithelium,whereas minimal IL-6 expression was observed in the other groups.Conclusion Mild moxibustion and electroacupuncture significantly ameliorate colonic inflammation exacerbated by TET2 deficiency in UC mice via epigenetic modulation.Distinct mechanisms exist between the two interventions:mild moxibustion regulates both DNMT and hydroxymethylase,whereas electroacupuncture primarily affects DNMT.
基金supported by the Special Fund for Agro-scientific Research in the Public Interest of Fujian Province,China(Grant No.2023R1021006)the National Natural Science Foundation of China(Grant No.32402387)+1 种基金the extended research project of the National Natural Science Foundation of China(Grant No.GJYS202511)the 5511 Collaborative Engineering Project,China(Grant No.XTCXGC2021001).
文摘Magnaporthe oryzae,the causal agent of rice blast,induces significant upregulation of OsPR10b,a pathogenesis-related(PR)pollen allergen(BetV-1)family gene.To investigate its role in immunity,we generated OsPR10b knockout mutants in the Zhonghua 11(ZH11)background.OsPR10b was predominantly expressed in rice calli and strongly induced by M.oryzae infection.Knockout mutants(ospr10b-1 and ospr10b-2)exhibited heightened susceptibility to both M.oryzae and Xanthomonas oryzae pv.oryzae(Xoo),demonstrating that OsPR10b positively regulates resistance to blast and bacterial blight.Our findings elucidate OsPR10b’s role in rice immunity and provide genetic resources for disease-resistant breeding.
基金supported in part by grants from the National Natural Science Foundation of China(No.81673093,No.82170227,No.91649113,No.82470165,No.82000121,No.31771640)the Jiangsu Science and Technology Department(No.SBK20200191)+1 种基金the State Key Laboratory of Radiation Medicine and Protection of Soochow University(No.GZC00201)a project funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions.
文摘The crosstalk between megakaryocytic lineage cells and the skeletal system has just begun to be explored but remains largely elusive.Using conditional gene knockout mouse models,we demonstrated that loss of Beclin 1(Becn1),a major regulator of mammalian autophagy,exclusively in the megakaryocytic lineage disrupted autophagy in platelets but did not compromise megakaryopoiesis or the formation and function of platelets.Unexpectedly,conditional Becn1 deletion in male mice led to a remarkable increase in bone mass with improved bone quality,in association with a decrease in sex hormone binding globulin(SHBG)and an increase in free testosterone(FT).In vivo Becn1 overexpression in megakaryocytic lineage-specific cells reduced bone mass and quality,along with an increase in SHBG and a decrease in FT.Transplantation of wild-type bone marrow cells into megakaryocytic lineage Becn1-deficient male mice restored bone mass and normalized SHBG and FT.Furthermore,bilateral orchiectomy of Becn1^(f/f);Pf4-iCre mice,which are crippled with the production of testosterone,resulted in a reduction in bone mass and quality,whereas in vivo overexpression of SHBG,specifically in the liver of Becn1^(f/f);Pf4-iCre mice,decreased FT and reduced bone mass and quality.In addition,metformin treatment,which induces SHBG expression,reduced FT and normalized bone mass in Becn1^(f/f);Pf4-iCre mice.We thus concluded that Becn1 of the megakaryocytic lineage is dispensable locally for platelet hemostasis but limits bone mass by increasing SHBG,which in turn reduces the FT of male mice.Our findings highlight a mechanism by which Becn1 from megakaryocytic lineage cells distally balances bone growth.
基金supported by the Swedish Research Council(2017-01286,2020-01840)the Swedish state under the agreement between the Swedish government and the county councils(ALF-agreement)(ALFGBG721581)+2 种基金the Gustaf V 80-years fund(FAI-2018-0466)the IngaBritt and Arne Lundberg Foundation(LU2017-0076)the Novo Nordisk Foundation(26844).
文摘Membrane-initiated estrogen receptorα(mERα)signaling has been shown to affect bone mass in murine models.However,it remains unknown which cell types mediate the mERα-dependent effects on bone.In this study,we generated a novel mouse model with a conditional C451A mutation in Esr1,which enables selective knockout of the palmitoylation site essential for the membrane localization of ERα(C451A^(f/f)).First,we used Runx2-Cre mice to generate Runx2-C451A^(f/f)mice with conditional inactivation of mERαsignaling in Runx2-expressing osteoblast lineage cells.No significant changes were observed in body weight,weights of estrogen-responsive organs,or serum concentrations of estradiol between female Runx2-C451A^(f/f)and homozygous C451A^(f/f)littermate controls.High-resolution microcomputed tomography analysis showed a consistent decrease in cortical bone mass in the tibia,femur,and vertebra L5 of Runx2-C451A^(f/f)mice and three-point bending analysis of humerus revealed an impaired mechanical bone strength in Runx2-C451A^(f/f)female mice compared to controls.Additionally,primary osteoblast cultures from mice lacking mERαsignaling showed impaired differentiation compared to controls.
基金supported by the National Natural Science Foundation of China (NSFC) (No.82172497)
文摘Intervertebral disc degeneration(IDD)is a progressive and dynamic process in which the senescence-associated secretory phenotype(SASP)of nucleus pulposus cells(NPC)plays a significant role.While impaired chaperone-mediated autophagy(CMA)has been associated with inflammation and cellular senescence,its specific involvement in the self-perpetuating feedback loop of NPC senescence remains poorly understood.Through LAMP2A knockout in NPC,we identified a significant upregulation of DYRK1A,a core mediator of premature senescence in Down syndrome.Subsequent validation established DYRK1A as the critical driver of premature senescence in CMA-deficient NPC.Combinatorial transcription factor analysis revealed that under IL1B stimulation or CMA inhibition,elevated DYRK1A promoted FOXC1 phosphorylation and nuclear translocation,initiating transcriptional activation of cell cycle arrest.Intriguingly,CMA impairment concurrently enhanced glutamine metabolic flux in senescent NPC,thereby augmenting their survival fitness.Transcriptomic profiling demonstrated that CMA reactivation in senescent NPC facilitated fate transition from senescence to apoptosis,mediated by decreased glutamine flux via GLUL degradation.Therefore,CMA exerts protective effects against IDD by maintaining equilibrium between premature senescence and senolysis.This study elucidates CMA’s regulatory role in SASP-mediated senescence amplification circuits,providing novel therapeutic insights for IDD and other age-related pathologies.
基金financially supported by the National Research Foundation of Korea(NRF-2021R1A5A1033157 for SRC program:382 Comparative medicine Disease Research Center,NRF-2021R1F1A105195313)the Research Institute of Veterinary Science,the BK21 Four for Future Veterinary Medicine Leading Education and Research Center,and a Seoul National University(SNU)grant(#550e2020005)。
文摘Background Genome editing has been considered as powerful tool in agricultural fields.However,genome editing progress in cattle has not been fast as in other mammal species,for some disadvantages including long gestational periods,single pregnancy,and high raising cost.Furthermore,technically demanding methods such as microinjection and somatic cell nuclear transfer(SCNT)are needed for gene editing in cattle.In this point of view,electroporation in embryos has been risen as an alternative.Results First,editing efficiency of our electroporation methods were tested for embryos.Presence of mutation on embryo was confirmed by T7E1 assay.With first combination,mutation rates for MSTN and PRNP were 57.6%±13.7%and 54.6%±13.5%,respectively.In case of MSTN/BLG,mutation rates were 83.9%±23.6%for MSTN,84.5%±18.0%for BLG.Afterwards,the double-KO embryos were transferred to surrogates and mutation rate was identified in resultant calves by targeted deep sequencing.Thirteen recipients were transferred for MSTN/PRNP,4 calves were delivered,and one calf underwent an induction for double KO.Ten surrogates were given double-KO embryos for MSTN/BLG,and four of the six calves that were born had mutations in both genes.Conclusions These data demonstrated that production of genome edited cattle via electroporation of RNP could be effectively applied.Finally,MSTN and PRNP from beef cattle and MSTN and BLG from dairy cattle have been born and they will be valuable resources for future precision breeding.
基金National Natural Science Foundation of China(82272608)2021 Capacity Building of Shanghai Universities(21010503600)Shanghai Key Lab of Human Performance(Shanghai University of Sport)(11DZ2261100)。
文摘Aging is an inevitable physiological process,often accompanied by age-related bone loss and subsequent bone-related diseases that pose serious health risks.Research on skeletal diseases caused by aging in humans is challenging due to lengthy study durations,difficulties in sampling,regional variability,and substantial investment.Consequently,mice are preferred for such studies due to their similar motor system structure and function to humans,ease of handling and care,low cost,and short generation time.In this review,we present a comprehensive overview of the characteristics,limitations,applicability,bone phenotypes,and treatment methods in naturally aging mice and prematurely aging mouse models(including SAMP6,POLG mutant,LMNA,SIRT6,ZMPSTE24,TFAM,ERCC1,WERNER,and KL/KL-deficient mice).We also summarize the molecular mechanisms of these aging mouse models,including cellular DNA damage response,senescence-related secretory phenotype,telomere shortening,oxidative stress,bone marrow mesenchymal stem cell(BMSC)abnormalities,and mitochondrial dysfunction.Overall,this review aims to enhance our understanding of the pathogenesis of aging-related bone diseases.
基金supported by the National Natural Science Foundation of China,No.82001604Guizhou Provincial Higher Education Science and Technology Innovation Team,No.[2023]072+1 种基金Guizhou Province Distinguished Young Scientific and Technological Talent Program,No.YQK[2023]040Guizhou Provincial Basic Research Program(Natural Science),No.ZK[2021]-368(all to LXiong),and Zunyi City Innovative Talent Team Training Plan,No.[2022]-2.
文摘Neonatal hypoxic-ischemic encephalopathy is often associated with permanent cerebral palsy,neurosensory impairments,and cognitive deficits,and there is no effective treatment for complications related to hypoxic-ischemic encephalopathy.The therapeutic potential of human placental chorionic plate-derived mesenchymal stem cells for various diseases has been explored.However,the potential use of human placental chorionic plate-derived mesenchymal stem cells for the treatment of neonatal hypoxic-ischemic encephalopathy has not yet been investigated.In this study,we injected human placental chorionic plate-derived mesenchymal stem cells into the lateral ventricle of a neonatal hypoxic-ischemic encephalopathy rat model and observed significant improvements in both cognitive and motor function.Protein chip analysis showed that interleukin-3 expression was significantly elevated in neonatal hypoxic-ischemic encephalopathy model rats.Following transplantation of human placental chorionic plate-derived mesenchymal stem cells,interleukin-3 expression was downregulated.To further investigate the role of interleukin-3 in neonatal hypoxic-ischemic encephalopathy,we established an in vitro SH-SY5Y cell model of hypoxic-ischemic injury through oxygen-glucose deprivation and silenced interleukin-3 expression using small interfering RNA.We found that the activity and proliferation of SH-SY5Y cells subjected to oxygen-glucose deprivation were further suppressed by interleukin-3 knockdown.Furthermore,interleukin-3 knockout exacerbated neuronal damage and cognitive and motor function impairment in rat models of hypoxic-ischemic encephalopathy.The findings suggest that transplantation of hpcMSCs ameliorated behavioral impairments in a rat model of hypoxic-ischemic encephalopathy,and this effect was mediated by interleukin-3-dependent neurological function.
基金Xizang Major Science and Technology Project(XZ202101ZD0005N)Yunnan Major Science and Technology Project(202302AE090015)+1 种基金National Key R&D Program of China(2023ZD04044-04)National Natural Science Foundation of China(32060736)。
文摘Tumor necrosis factorα(TNFα)exhibits diverse biological functions;however,its regulatory roles in myogenesis are not fully understood.In the present study,we explored the function of TNFαin myoblast proliferation,differentiation,migration,and myotube fusion in primary myoblasts and C2C12 cells.To this end,we constructed TNFαmuscle-conditional knockout(TNFα-CKO)mice and compared them with flox mice to assess the effects of TNFαknockout on skeletal muscles.Results indicated that TNFα-CKO mice displayed phenotypes such as accelerated muscle development,enhanced regenerative capacity,and improved exercise endurance compared to flox mice,with no significant differences observed in major visceral organs or skeletal structure.Using label-free proteomic analysis,we found that TNFα-CKO altered the distribution of several muscle development-related proteins,such as Hira,Casz1,Casp7,Arhgap10,Gas1,Diaph1,Map3k20,Cfl2,and Igf2,in the nucleus and cytoplasm.Gene set enrichment analysis(GSEA)further revealed that TNFαdeficiency resulted in positive enrichment in oxidative phosphorylation and MyoD targets and negative enrichment in JAK-STAT signaling.These findings suggest that TNFα-CKO positively regulates muscle growth and development,possibly via these newly identified targets and pathways.
基金supported by the National Natural Science Foundation of China(No.31972809)the Key Research&Development Program of Hubei Province(No.2022BBA0051),China.
文摘Neuropeptide Y receptor Y8(NPY8R)is a fish-specific receptor with two subtypes,NPY8AR and NPY8BR.Changes in expression levels during physiological processes or in vivo regulation after ventricular injection suggest that NPY8BR plays an important role in feeding regulation;this has been found in only a few fish,at present.In order to better understand the physiological function of npy8br,especially in digestion,we used clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)technology to generate npy8br-/-japanese medaka(Oryzias latipes).We found that the deletion of npy8br in medaka larvae affected their feeding and digestion ability,ultimately affecting their growth.Specifically,npy8br deficiency in medaka larvae resulted in decreased feed intake and decreased expression levels of orexigenic genes(npy and agrp).npy8br-/-medaka larvae fed for 10 d(10th day of feeding)still had incompletely digested brine shrimp(Artemia nauplii)in the digestive tract 8 h after feeding,the messenger RNA(mRNA)expression levels of digestion-related genes(amy,lpl,ctra,and ctrb)were significantly decreased,and the activity of amylase,trypsin,and lipase also significantly decreased.The deletion of npy8br in medaka larvae inhibited the growth and significantly decreased the expression of growth-related genes(gh and igf1).Hematoxylin and eosin(H&E)sections of intestinal tissue showed that npy8br-/-medaka larvae had damaged intestine,thinned intestinal wall,and shortened intestinal villi.So far,this is the first npy8br gene knockout model established in fish and the first demonstration that npy8br plays an important role in digestion.
文摘Butyrylcholinesterase(BChE;EC 3.1.1.8),an enzyme structurally related to acetylcholinesterase,is widely distributed in the human body.It plays a role in the detoxification of chemicals such as succinylcholine,a muscle relaxant used in anesthetic practice.BChE is well-known due to variant forms of the enzyme with little or no hydrolytic activity which exist in some endogamous communities and result in prolonged apnea following the administration of succinylcholine.Its other functions include the ability to hydrolyze acetylcholine,the cholinergic neurotransmitter in the brain,when its primary hydrolytic enzyme,acetylcholinesterase,is absent.To assess its potential roles,BChE was studied in relation to insulin resistance,type 2 diabetes mellitus,cognition,hepatic disorders,cardiovascular and cerebrovascular diseases,and inflammatory conditions.Individuals who lack the enzyme activity of BChE are otherwise healthy,until they are given drugs hydrolyzed by this enzyme.Therefore,BChE is a candidate for the study of loss-of-function mutations in humans.Studying individuals with variant forms of BChE can provide insights into whether they are protected against metabolic diseases.The potential utility of the enzyme as a biomarker for Alzheimer’s disease and the response to its drug treatment can also be assessed.
文摘BACKGROUND Currently,intrahepatic cholangiocarcinoma(ICC)poses a continuing,significant health challenge,but the relationship has yet to be established between ICC and the proteasome 26S subunit non-ATPase 6(PSMD6).AIM To investigate the protein expression and clinicopathological significance of PSMD6 in ICC.METHODS The potential impact of the PSMD6 gene on the growth of ICC cell lines was analyzed using clustered regularly interspaced short palindromic repeat knockout screening technology.Forty-two paired specimens of ICC and adjacent noncancerous tissues were collected.PSMD6 protein expression was determined by immunohistochemistry.Receiver operating characteristic curve analysis was performed to validate PSMD6 expression level,and its association with ICC patients’various clinicopathological characteristics was investigated.RESULTS The PSMD6 gene was found to be essential for the growth of ICC cell lines.PSMD6 protein was significantly overexpressed in ICC tissues(P<0.001),but showed no significant association with patient age,gender,pathological grade,or tumor-node-metastasis stage(P>0.05).CONCLUSION PSMD6 can promote the growth of ICC cells,thus playing a pro-oncogenic role.
基金Alberta Innovates,Grant/Award Number:CD-60Canadian Glycomics Network,Grant/Award Number:CD-50Agriculture and Agri-Food Canada,Grant/Award Number:2991。
文摘Background:A multitude of mouse models are utilized to emulate and study intestinal inflammation.T-cell receptor alpha chain(TCRα)-deficient mice are used as a model of spontaneous colitis that has similarities with human ulcerative colitis.However,colitis is triggered late in the life of the mouse(age:4-5 months),and inflammation does not develop at the same time in different mice.A previously conducted study reported that the administration of the drug piroxicam triggered predictable and early colitis in TCRα-deficient mice at the age of 6-8 weeks.However,a detailed characterization of ensuing inflammation was not provided.Methods:We conducted an in-depth examination of piroxicam-triggered colitis in TCRα-deficient mice,with emphasis on spatial histopathologic changes and analysis of expression of inflammatory markers.Furthermore,we tested amelioration of colitis with dexamethasone.Results:We confirmed that piroxicam induced a time-prescribed colitis and did so in the proximal colon as well as the cecum of TCRα-deficient mice.Piroxicam adminis-tration was observed to induce epithelial hyperplasia,goblet cell loss,and leukocyte infiltration with occasional ulceration.A Swiss roll technique was used to examine the colon and cecum in its entirety.Importantly,we observed that inflammation was mul-tifocal segmental,with areas of tissue damage in between healthy tissue.In addition,we observed variability in the severity of inflammation among replicate animals and treatments,and that the administration of dexamethasone only partially ameliorated inflammation in the proximal colon.Conclusions:Piroxicam consistently induced multifocal segmental colitis in the proxi-mal colon and cecum,although the degree of inflammation was reduced in the latter.Importantly,spatial variability in inflammation in the large intestine and the inter-replicate variation in the severity of inflammation must be taken into consideration when utilizing this murine model of synchronized colitis.
基金supported by the National Research Foundation of Korea grant funded by the Korean government(MIST)(2020R1A5A8017671,2022R1A2C100677813,and RS-2024-00349650)。
文摘Williams syndrome(WS)is a rare multisystemic disorder caused by recurrent microdeletions on 7q11.23,characterized by intellectual disability,distinctive craniofacial and dental features,and cardiovascular problems.Previous studies have explored the roles of individual genes within these microdeletions in contributing to WS phenotypes.Here,we report five patients with WS with 1.4 Mb-1.5 Mb microdeletions that include RFC2,as well as one patient with a 167-kb microdeletion involving RFC2 and six patients with intragenic variants within RFC2.To investigate the potential involvement of RFC2 in WS pathogenicity,we generate a rfc2 knockout(KO)zebrafish using CRISPR-Cas9 technology.Additionally,we generate a KO zebrafish of its paralog gene,rfc5,to better understand the functions of these RFC genes in development and disease.Both rfc2 and rfc5 KO zebrafish exhibit similar phenotypes reminiscent of WS,including small head and brain,jaw and dental defects,and vascular problems.RNA-seq analysis reveals that genes associated with neural cell survival and differentiation are specifically affected in rfc2 KO zebrafish.In addition,heterozygous rfc2 KO adult zebrafish demonstrate an anxiety-like behavior with increased social cohesion.These results suggest that RFC2 may contribute to the pathogenicity of WS,as evidenced by the zebrafish model.
