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
BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammato...BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammatory effects by secreting cytokines and exosomes and are widely used to treat autoimmune and inflam-mation-related diseases.Hepatocyte growth factor(HGF)is a pleiotropic cytokine that plays a key role in many inflammatory and autoimmune diseases.AIM To modify DPSCs with HGF(DPSC-HGF)and evaluate the therapeutic effect of DPSC-HGF on AS using an apolipoprotein E-knockout(ApoE-/-)mouse model and an in vitro cellular model.METHODS ApoE-/-mice were fed with a high-fat diet(HFD)for 12 wk and injected with DPSC-HGF or Ad-Null modified DPSCs(DPSC-Null)through tail vein at weeks 4,7,and 11,respectively,and the therapeutic efficacy and mechanisms were analyzed by histopathology,flow cytometry,lipid and glucose measurements,real-time reverse transcription polymerase chain reaction(RT-PCR),and enzyme-linked immunosorbent assay at the different time points of the experiment.An in vitro inflammatory cell model was established by using RAW264.7 cells and human aortic endothelial cells(HAOECs),and indirect co-cultured with supernatant of DPSC-Null(DPSC-Null-CM)or DPSC-HGF-CM,and the effect and mechanisms were analyzed by flow cytometry,RT-PCR and western blot.Nuclear factor-κB(NF-κB)activators and inhibitors were also used to validate the related signaling pathways.RESULTS DPSC-Null and DPSC-HGF treatments decreased the area of atherosclerotic plaques and reduced the expression of inflammatory factors,and the percentage of macrophages in the aorta,and DPSC-HGF treatment had more pronounced effects.DPSCs treatment had no effect on serum lipoprotein levels.The FACS results showed that DPSCs treatment reduced the percentages of monocytes,neutrophils,and M1 macrophages in the peripheral blood and spleen.DPSC-Null-CM and DPSC-HGF-CM reduced adhesion molecule expression in tumor necrosis factor-αstimulated HAOECs and regulated M1 polarization and inflammatory factor expression in lipopolysaccharide-induced RAW264.7 cells by inhibiting the NF-κB signaling pathway.CONCLUSION This study suggested that DPSC-HGF could more effectively ameliorate AS in ApoE-/-mice on a HFD,and could be of greater value in stem cell-based treatments for AS.展开更多
AIM:To investigate the role of transmembrane protein 206(TMEM206)in corneal edema in mice by knockout the TMEM206 gene using CRISPR/Cas9 editing technology.METHODS:TMEM206-knockout mice were generated using the CRISPR...AIM:To investigate the role of transmembrane protein 206(TMEM206)in corneal edema in mice by knockout the TMEM206 gene using CRISPR/Cas9 editing technology.METHODS:TMEM206-knockout mice were generated using the CRISPR-Cas9 system.Variations in ophthalmic pathology were observed using slit lamp microscope and optical coherence tomography(OCT),intraocular pressure(IOP)was measured using a TonoLab Rebound Tonometer,and the ultrastructure of the corneal was observed using a transmission electron microscope.RESULTS:Corneal opacity was observed in 4/18 homozygous TMEM206^(-/-)mice whereas a similar change was not observed in heterozygous TMEM206^(+/-)mice and wild-type littermates.OCT examination showed that the mean central cornea thickness was 125±5.4μm in 4 homozygous TMEM206^(-/-)mice developed corneal edema and 115±1.2μm in wild-type mice(t=3.468,P<0.05)at 43wk.The mean IOP was 12.08±0.07 mm Hg in four right eyes with corneal edema and 12.03±0.03 mm Hg in three normal left eyes(P>0.05).Transmission electron microscopy revealed a disruption in the organization of the collagen fibrils in the central part of the cornea in homozygous TMEM206^(-/-)mice.CONCLUSION:TMEM206 is associated with corneal edema which caused organizational disruption of collagen fibrils in corneas of mice.展开更多
Arsenic methyltransferase（As3mt） catalyzes the conversion of inorganic arsenic（i As） to its methylated metabolites, including toxic methylarsonite（MAs~Ⅲ） and dimethylarsinite（DMAs~Ⅲ）. Knockout（KO） of As3 m...Arsenic methyltransferase（As3mt） catalyzes the conversion of inorganic arsenic（i As） to its methylated metabolites, including toxic methylarsonite（MAs~Ⅲ） and dimethylarsinite（DMAs~Ⅲ）. Knockout（KO） of As3 mt was shown to reduce the capacity to methylate i As in mice. However, no data are available on the oxidation states of As species in tissues of these mice. Here, we compare the oxidation states of As species in tissues of male C57BL/6 As3mt-KO and wild-type（WT） mice exposed to arsenite（iA s~Ⅲ） in drinking water. WT mice were exposed to50 mg/L As and As3mt-KO mice that cannot tolerate 50 mg/L As were exposed to 0, 15, 20, 25 or30 mg/L As. iA s~Ⅲaccounted for 53% to 74% of total As in liver, pancreas, adipose, lung, heart, and kidney of As3mt-KO mice; tri- and pentavalent methylated arsenicals did not exceed 10% of total As. Tissues of WT mice retained iA s and methylated arsenicals： iA s~Ⅲ, MAs~Ⅲand DMAs~Ⅲ represented 55%‐68% of the total As in the liver, pancreas, and brain. High levels of methylated species, particularly MAs~Ⅲ, were found in the intestine of WT, but not As3mt-KO mice,suggesting that intestinal bacteria are not a major source of methylated As. Blood of WT mice contained significantly higher levels of As than blood of As3mt-KO mice. This study is the first to determine oxidation states of As species in tissues of As3mt-KO mice. Results will help to design studies using WT and As3mt-KO mice to examine the role of iA s methylation in adverse effects of iA s exposure.展开更多
Objective To explore the regulatory effect of fragile X mental retardation protein （FMRP） on the translation of microtubule associated protein 1B （MAP1B）. Methods The expressions of MAP1B protein and MAP1B mRNA in...Objective To explore the regulatory effect of fragile X mental retardation protein （FMRP） on the translation of microtubule associated protein 1B （MAP1B）. Methods The expressions of MAP1B protein and MAP1B mRNA in the brains of 1-week and 6-week old fragile X mental retardation-1 （FmrI） knockout （KO） mice were investigated by immunohistochemistry, Western blot, and in situ hybridization, with the age-matched wild type mice （WT） as controls. Results The mean optical density （MOD） of MAP1B was significantly decreased in each brain region in KO6W compared with WT6W, whereas in KO1W, this decrease was only found in the hippocampus and cerebellum. MAP1B in 6-week mice was much less than that in 1-week mice of the same genotype. The results of Western blot and in situ hybridization showed that MAP1B protein and MAP1B mRNA were significantly decreased in the hippocampus of both KO1W and KO6W. Conclusion The decreased MAP1B protein and MAP1B mRNA in the Fmrl knockout mice indicate that FMRP may positively regulate the expression of MAP1B.展开更多
Objective The protein interacting with C kinase 1(PICK1)plays a critical role in vesicle trafficking,and its deficiency in sperm cells results in abnormal vesicle trafficking from Golgi to acrosome,which eventually di...Objective The protein interacting with C kinase 1(PICK1)plays a critical role in vesicle trafficking,and its deficiency in sperm cells results in abnormal vesicle trafficking from Golgi to acrosome,which eventually disrupts acrosome formation and leads to male infertility.Methods An azoospermia sample was filtered,and the laboratory detection and clinical phenotype indicated typical azoospermia in the patient.We sequenced all of the exons in the PICK1 gene and found that there was a novel homozygous variant in the PICK1 gene,c.364delA(p.Lys122SerfsX8),and this protein structure truncating variant seriously affected the biological function.Then we constructed a PICK1 knockout mouse model using clustered regularly interspaced short palindromic repeat cutting technology(CRISPRc).Results The sperm from PICK1 knockout mice showed acrosome and nucleus abnormalities,as well as dysfunctional mitochondrial sheath formation.Both the total sperm and motility sperm counts were decreased in the PICK1 knockout mice compared to wild-type mice.Moreover,the mitochondrial dysfunction was verified in the mice.These defects in the male PICK1 knockout mice may have eventually led to complete infertility.Conclusion The c.364delA novel variant in the PICK1 gene associated with clinical infertility,and pathogenic variants in the PICK1 may cause azoospermia or asthenospermia by impairing mitochondrial function in both mice and humans.展开更多
Early studies suggested that estrogen receptor alpha （ERa） is involved in estrogen-mediated imprinting effects in prostate development. We recently reported a more complete ERa knockout （KO） mouse model via mating...Early studies suggested that estrogen receptor alpha （ERa） is involved in estrogen-mediated imprinting effects in prostate development. We recently reported a more complete ERa knockout （KO） mouse model via mating β-actin Cre transgenic mice with floxed ERa mice. These ACTB-ERaKO male mice showed defects in prostatic branching morphogenesis, which demonstrates that ERa is necessary to maintain proliferative events in the prostate. However, within which prostate cell type ERa exerts those important functions remains to be elucidated. To address this, we have bred floxed ERa mice with either fibroblast-specific protein （FSP）-Cre or probasin-Cre transgenic mice to generate a mouse model that has deleted ERa gene in either stromal fibroblast （FSP-ERaKO） or epithelial （pes-ERaKO） prostate cells. We found that circulating testosterone and fertility were not altered in FSP.ERaKO and pes-ERaKO male mice. Prostates of FSP-ERaKO mice have less branching morphogenesis compared to that of wild-type littermates. Further analyses indicated that loss of stromal ERa leads to increased stromal apoptosis, reduced expression of insulin-likegrowth factor-1 （IGF-1） and FGFIO, and increased expression of BMP4. Collectively, we have established the first in vivo prostate stromal and epithelial selective ERaKO mouse models and the results from these mice indicated that stromal fibroblast ERa plays important roles in prostatic branching morphogenesis via a paracrine fashion. Selective deletion of the ERa gene in mouse prostate epithelial cells by probasin-Cre does not affect the regular prostate development and homeostasis.展开更多
BACKGROUND Claudin-7, one of the important components of cellular tight junctions, is currently considered to be expressed abnormally in colorectal inflammation and colorectal cancer. However, there is currently no ef...BACKGROUND Claudin-7, one of the important components of cellular tight junctions, is currently considered to be expressed abnormally in colorectal inflammation and colorectal cancer. However, there is currently no effective animal model to study its specific mechanism. Therefore, we constructed three lines of Claudin-7 knockout mice using the Cre/LoxP system.AIM To determine the function of the tumor suppressor gene Claudin-7 by generating three lines of Claudin-7 gene knockout mice.METHODS We crossed Claudin-7-floxed mice with CMV-Cre, vil1-Cre, and villin-CreERT2 transgenic mice, and the offspring were self-crossed to obtain conventional Claudin-7 knockout mice, conditional(intestinal specific) Claudin-7 knockout mice, and inducible conditional Claudin-7 knockout mice. Intraperitoneal injection of tamoxifen into the inducible conditional Claudin-7 knockout mice can induce the knockout of Claudin-7. PCR and agarose gel electrophoresis were used to identify mouse genotypes, and Western blot was used to confirm the knockout of Claudin-7. The mental state, body length, and survival time of these mice were observed. The dying mice were sacrificed, and hematoxylin-eosin(HE) staining and immunohistochemical staining were performed to observe changes in intestinal structure and proliferation markers.RESULTS We generated Claudin-7-floxed mice and three lines of Claudin-7 gene knockout mice using the Cre/LoxP system successfully. Conventional and intestinal specific Claudin-7 knockout mice were stunted and died during the perinatal period, and intestinal HE staining in these mice revealed mucosal gland structure disappearance and connective tissue hyperplasia with extensive inflammatory cell infiltration. The inducible conditional Claudin-7 knockout mice had a normal phenotype at birth, but after the induction with tamoxifen, they exhibited a dying state. Intestinal HE staining showed significant inflammatory cell infiltration, and atypical hyperplasia and adenoma were also observed. Intestinal immunohistochemistry analysis showed abnormal expression and distribution of Ki67, and the normal intestinal proliferation balance was disrupted. The intestinal crypt size in inducible conditional Claudin-7 knockout mice was increased compared with control mice(small intestine: 54.1 ± 2.96 vs 38.4 ± 1.63;large intestine: 44.7 ± 1.93 vs 27.4 ± 0.60; P < 0.001).CONCLUSION The knockout of Claudin-7 in vivo causes extensive inflammation, atypical hyperplasia, and adenoma in intestinal tissue as well as animal death in mice.Claudin-7 may act as a tumor suppressor gene in the development of colorectal cancer.展开更多
The distribution of the Na-K-2Cl co-transporter （NKCCl） in the cochlear K^＋ cycling pathway in cochlea and cochlear histological changes in the NKCCl knockout mice were investigated. By using immunohistochemistry a...The distribution of the Na-K-2Cl co-transporter （NKCCl） in the cochlear K^＋ cycling pathway in cochlea and cochlear histological changes in the NKCCl knockout mice were investigated. By using immunohistochemistry and toluidine blue staining, the localization of NKCCl in cochlea of the C57BL/6J mice and the cochlear histological changes in the NKCCl knockout mice were observed. It was found that the NKCCl was expressed mainly in the stria marginal cells and the fibrocytes in the inferior portion of the spiral ligament in the adult C57BL/6J mice. Subpopulation of the fibrocytes in the suprastrial region and the limbus was also moderately immunoreactive. While in the cochlea of the NKCCl knockout mice, Reissner＇s membrane was collapsed and scala media disappeared, accompanied with the loss of inner hair cells, outer hair cells and the support cells. The tunnel of Corti was often absent. All the findings suggested the localization of NKCCl in the cochlea was closely correlated with cochlear K^＋ cycling. Loss of NKCCl led to the destruction of the cochlear structures, and subsequently influenced the physiological function of cochlea.展开更多
An overt phenotype of aquaporin-1 knockout（AQP1 ko） mice is growth retardation, suggesting possible defects in bone development and metabolism. In the present study, we analyzed the bone mineral density（ BMD）, bon...An overt phenotype of aquaporin-1 knockout（AQP1 ko） mice is growth retardation, suggesting possible defects in bone development and metabolism. In the present study, we analyzed the bone mineral density（ BMD）, bone calcium and phosphorus contents, and bone metabolism in an AQP1 ko mouse model. The BMD of femurs in AQP1 ko mice was significantly lower than that of litter-matched wildtype mice as measured by dual energy X-ray absorptiometry. Consistently, the contents of bone total calcium and phosphorus were also significantly lower in AQP1 ko mice. The reduced BMD caused by AQP1 deficiency mainly affect male mice. Bone metabolic activity, as indicated by 99m^Tc-MDP absorption measurements, was remarkably reduced in AQP1 ko mice. These results provide the first evidence that AQP1 play an important role in bone structure and metabolism.展开更多
AIM: To investigate the role of host and bacterial arginases in the colonization of mice by Helicobacter pylori (H.pylori).METHODS: H.pylori produces a very powerful urease that hydrolyzes urea to carbon dioxide and a...AIM: To investigate the role of host and bacterial arginases in the colonization of mice by Helicobacter pylori (H.pylori).METHODS: H.pylori produces a very powerful urease that hydrolyzes urea to carbon dioxide and ammonium,which neutralizes acid.Urease is absolutely essential to H.pylori pathogenesis;therefore,the urea substrate must be in ample supply for urease to work efficiently.The urea substrate is most likely provided by arginase activity,which hydrolyzes L-arginine to L-ornithine and urea.Previous work has demonstrated that H.pylori arginase is surprisingly not required for colonization of wild-type mice.Hence,another in vivo source of the critical urea substrate must exist.We hypothesized that the urea source was provided by host arginase Ⅱ,since this enzyme is expressed in the stomach,and H.pylori has previously been shown to induce the expression of murine gastric arginase Ⅱ.To test this hypothesis,wild-type and arginase (rocF) mutant H.pylori strain SS1 were inoculated into arginase Ⅱ knockout mice.RESULTS: Surprisingly,both the wild-type and rocF mutant bacteria still colonized arginase Ⅱ knockout mice.Moreover,feeding arginase Ⅱ knockout mice the host arginase inhibitor S-(2-boronoethyl)L-cysteine (BEC),while inhibiting > 50% of the host arginase Ⅰ?activity in several tissues,did not block the ability of the rocF mutant H.pylori to colonize.In contrast,BEC poorly inhibited H.pylori arginase activity.CONCLUSION: The in vivo source for the essential urea utilized by H.pylori urease is neither bacterial arginase nor host arginase Ⅱ;instead,either residual host arginase Ⅰ?or agmatinase is probably responsible.展开更多
Due to their inability to generate a complete immune response, mice knockout for type I interferon （IFN） receptors （Ifnar-/-） are more susceptible to viral infections, and are thus commonly used for pathogenesis s...Due to their inability to generate a complete immune response, mice knockout for type I interferon （IFN） receptors （Ifnar-/-） are more susceptible to viral infections, and are thus commonly used for pathogenesis studies. This mouse model has been used to study many diseases caused by highly pathogenic viruses from many families, including the Flaviviridae, Filoviridae, Arenaviridae, Bunyaviridae, Henipaviridae, and Togaviridae. In this review, we summarize the findings from these animal studies, and discuss the pros and cons of using this model versus other known methods for studying pathogenesis in animals.展开更多
Microtubule-severing proteins(MTSPs),are a family of proteins which use adenosine triphosphate to sever microtubules.MTSPs have been shown to play an important role in multiple microtubule-involved cellular processes....Microtubule-severing proteins(MTSPs),are a family of proteins which use adenosine triphosphate to sever microtubules.MTSPs have been shown to play an important role in multiple microtubule-involved cellular processes.One member of this family,fidgetin(FIGN),is also involved in male fertility;however,no studies have explored its roles in female fertility.In this study,we found mouse fidgetin is rich within oocyte zona pellucida(ZP)and is the only MTSP member to do so.Fidgetin also appears to interact with all three ZP proteins.These findings prompted us to propose that fidgetin might prevent polyspermy.Results from in vitro maturation oocytes analysis showed that fidgetin knockdown did cause polyspermy.We then deleted all three fidgetin isoforms with CRISPR/Cas9 technologies;however,female mice remained healthy and with normal fertility.Of all mouse MTSPs,only the mRNA level of fidgetin-like 1(FIGNL1)significantly increased.Therefore,we assert that fidgetin-like 1 compensates fidgetin's roles in fidgetin knockout female mice.展开更多
[Objectives]This study was conducted to investigate the similarity and differences between TLR4 knockout mice and C57 BL/6 mice used in liver fibrosis research in terms of growth rate and reproduction ability.[Methods...[Objectives]This study was conducted to investigate the similarity and differences between TLR4 knockout mice and C57 BL/6 mice used in liver fibrosis research in terms of growth rate and reproduction ability.[Methods]Twenty TLR4 knockout mice and C57 BL/6 mice,half male and half female,were selected to compare the growth rates of body weight and body length of mice from the 4th to 12th weeks;and 20 pairs of male and female mice of the same strain were compared for the number of baby mice of the second litter.[Results]The growth rates of body weight and body length of the TLR4 knockout mice were significantly lower than those of C57 BL/6 mice(P<0.05)(except for the 4th and 5th weeks when there was no significant difference in body length);and in terms of reproductive ability,the TLR4 knockout mice were significantly lower than the C57 BL/6 mice(the ratio of the total number of baby mice in the second litter of the two strains,72∶147).[Conclusions]Knockout of the TLR4 gene has a significant impact on the growth and reproduction of mice.展开更多
Objective-To investigate the effects of heat shock transcription factor 1) gene on the constitutivety expressed αB-CrystaUin (aBC) in mice myocardium. Methods-The expression levels of constitutive aBC in HSF1 knockou...Objective-To investigate the effects of heat shock transcription factor 1) gene on the constitutivety expressed αB-CrystaUin (aBC) in mice myocardium. Methods-The expression levels of constitutive aBC in HSF1 knockout (hsf1 - /- ) and HSFl wild type (As/1 + /+) mice myocardium were evaluated by western blot and immunohistochemistry. Results : The αBC levels in hsfl -/- and hsfl +/+ were 68. 42±4. 16, 100. 00±7. 58, respectively (P<0. 05, cytoso-lic fraction) , and 20. 53±1. 01, 37. 55±1. 91, respectively (P<0. 05, pellet fraction). The aBC signals decreased significantly in hsfl -/- myocardium when compared with those in hsfl +/+ myocardium stained with fluorescence immunohistochemistry. Conclusion-HSF1 is an important, but not the only factor, which mediates the constitutively expressed aBC.展开更多
Background Immune inflammatory response is throughout the entire process of atherosclerosis （AS）. It was unclear whether the mechanism of mesenchymal stem cells （MSCs） transplantation for treatment of AS is involv...Background Immune inflammatory response is throughout the entire process of atherosclerosis （AS）. It was unclear whether the mechanism of mesenchymal stem cells （MSCs） transplantation for treatment of AS is involved with inflammation regulation in the plaque area. The aim of this study was to explore the effects of MSCs in the formation of atherosclerosis plaque in hypercholesterolemic apoliprotein （apo）E-/- mice. Methods ApoE-/- mice MSCs were isolated and identified. At 8 weeks of age, 30 male ApoE-/- mice were randomly divided into negative control group （Neg, n = 10）, positive control group （Pos, n = 10） and mesenchymal stem cells group （MSCs, n = 10）. MSCs were injected through caudal vein into the body of Pos and MSCs group. The plaque area of all subjects were compared, the percentage of CD4^＋CD25^＋Tregs in different tissues were analyzed by fluorescence activated cell sorter （FACS）, proliferation response of splenocytes to MSCs was detected and cytokines in the supernatant were determined by enzyme linked immunosorbent assay （ELISA）. Results Compared with controls, MSCs resulted in a significant decrease of latherosclerotic plaques size （P 〈 0.05）, and a significant increase of CD4^＋CD25^＋ regulatory T cells in spleen （P 〈 0.05）. Specific proliferation response of CD4^＋CD25^＋ regulatory T cells in splenocytes to MCSs was significantly suppressed, the superanant level of TGF-[3 and IL-10 in MSCs group were increased while IFN-γ/ decreased significantly. Conclusion MSCs play an important role in regulating the inflammatory response and significantly inhibit the formation of the atherosclerosis plaque in ApoE-/-mice.展开更多
Background Subtypes of T cells, called regulatory T cells （Treg cell）, play a critical role in limiting autoimmune processes and inflammatory responses, The aim of this study was to explore functional roles of FOXP3...Background Subtypes of T cells, called regulatory T cells （Treg cell）, play a critical role in limiting autoimmune processes and inflammatory responses, The aim of this study was to explore functional roles of FOXP3 in the manifestation of atherosclerosis in Apolipoprotein E deficient （ApoE）-/- mice. Methods Lentivirus-mediated （siRNA） was used to knock down FOXP3 and FOXP3high＋CD4＋ CD25＋ T cells adoptive transfer assays in high fat diet ApoE-/- mice were done. The resulting atherosclerotie lesions were assessed by determining FOXP3 transcript levels and investigating the expression of FOXP3 protein in different tissues. Results Animals treated with siRNA of FOXP3 showed a significant increase in atherosclerotic lesion formation and a reduction in the number of FOXP3＋CD4＋CD25＋ T cells compared with other groups. Transfer of FOXP3highCD4＋CD25＋ T cells significantly decreased atherosclerotic plaque formation and increased the number of FOXP3＋ CD4＋ CD25＋ T cells. FOXP3 protein levels and FOXP3 transcript levels were lowest in the siRNA group, and were highest in tissues from the Treg transfer group. Conclusion FOXP3 plays an important role in regulating the inflammatory response within the atherosclerotic lesion. It can inhibit significantly the progression of the atherosclerosis plaque in ApoE-/- mice.展开更多
OBJECTIVE To observe the effects of serum metabolites by using ~1H-NMR-based metabonomic approach to explore the possible mechanisms of total flavonoids in Ocimum BasilicumLinn(OBL) on atherosclerosis in apolipomtein ...OBJECTIVE To observe the effects of serum metabolites by using ~1H-NMR-based metabonomic approach to explore the possible mechanisms of total flavonoids in Ocimum BasilicumLinn(OBL) on atherosclerosis in apolipomtein E knockout(ApoE-/-) mice.METHODS Six-week-old male apoE knockout mice were divided into four groups(n=10) and fed with high fet diet:model,Simv.astatin,OBL-H,OBL-M and OBL-L groups.The homogeneous male mice of C57 BL/6 J were used as the normol group and fed with normal chow diet.After 14 weeks,~1H-NMR technology was used to ex.plore the variability of serum metabolites by the method of PLS-DA and OPLS-DA.RESULTS Com.pared with normal group,Model group showed a significant increase in the serum levels of VLDL,LDL,β-hydroxyisobutyrate,lactate,myo-inositol and showed a significant decrease in the serum levels of al.anine,glutamine,proline,carnitine,methylamine,citrate,creatine,choline,taurine,pyruvate,β-glu.cose,α-glucose,glycine,lysine.Combined with model group OBL-H,OBL-M,OBL-L groups showed the effects of regulating the levels of different metabolites of the glucose,lipid and amino acid metabo.lism.CONCLUSION The anti-atheros-clerotic activity of total flavonoids in Ocimum BasilicumLinn may be related not only to regulation of lipid metabolism,but also glycometabolism and amino acid metabolism.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
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
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.
文摘BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammatory effects by secreting cytokines and exosomes and are widely used to treat autoimmune and inflam-mation-related diseases.Hepatocyte growth factor(HGF)is a pleiotropic cytokine that plays a key role in many inflammatory and autoimmune diseases.AIM To modify DPSCs with HGF(DPSC-HGF)and evaluate the therapeutic effect of DPSC-HGF on AS using an apolipoprotein E-knockout(ApoE-/-)mouse model and an in vitro cellular model.METHODS ApoE-/-mice were fed with a high-fat diet(HFD)for 12 wk and injected with DPSC-HGF or Ad-Null modified DPSCs(DPSC-Null)through tail vein at weeks 4,7,and 11,respectively,and the therapeutic efficacy and mechanisms were analyzed by histopathology,flow cytometry,lipid and glucose measurements,real-time reverse transcription polymerase chain reaction(RT-PCR),and enzyme-linked immunosorbent assay at the different time points of the experiment.An in vitro inflammatory cell model was established by using RAW264.7 cells and human aortic endothelial cells(HAOECs),and indirect co-cultured with supernatant of DPSC-Null(DPSC-Null-CM)or DPSC-HGF-CM,and the effect and mechanisms were analyzed by flow cytometry,RT-PCR and western blot.Nuclear factor-κB(NF-κB)activators and inhibitors were also used to validate the related signaling pathways.RESULTS DPSC-Null and DPSC-HGF treatments decreased the area of atherosclerotic plaques and reduced the expression of inflammatory factors,and the percentage of macrophages in the aorta,and DPSC-HGF treatment had more pronounced effects.DPSCs treatment had no effect on serum lipoprotein levels.The FACS results showed that DPSCs treatment reduced the percentages of monocytes,neutrophils,and M1 macrophages in the peripheral blood and spleen.DPSC-Null-CM and DPSC-HGF-CM reduced adhesion molecule expression in tumor necrosis factor-αstimulated HAOECs and regulated M1 polarization and inflammatory factor expression in lipopolysaccharide-induced RAW264.7 cells by inhibiting the NF-κB signaling pathway.CONCLUSION This study suggested that DPSC-HGF could more effectively ameliorate AS in ApoE-/-mice on a HFD,and could be of greater value in stem cell-based treatments for AS.
基金Supported by National Natural Science Foundation of China(No.31571294).
文摘AIM:To investigate the role of transmembrane protein 206(TMEM206)in corneal edema in mice by knockout the TMEM206 gene using CRISPR/Cas9 editing technology.METHODS:TMEM206-knockout mice were generated using the CRISPR-Cas9 system.Variations in ophthalmic pathology were observed using slit lamp microscope and optical coherence tomography(OCT),intraocular pressure(IOP)was measured using a TonoLab Rebound Tonometer,and the ultrastructure of the corneal was observed using a transmission electron microscope.RESULTS:Corneal opacity was observed in 4/18 homozygous TMEM206^(-/-)mice whereas a similar change was not observed in heterozygous TMEM206^(+/-)mice and wild-type littermates.OCT examination showed that the mean central cornea thickness was 125±5.4μm in 4 homozygous TMEM206^(-/-)mice developed corneal edema and 115±1.2μm in wild-type mice(t=3.468,P<0.05)at 43wk.The mean IOP was 12.08±0.07 mm Hg in four right eyes with corneal edema and 12.03±0.03 mm Hg in three normal left eyes(P>0.05).Transmission electron microscopy revealed a disruption in the organization of the collagen fibrils in the central part of the cornea in homozygous TMEM206^(-/-)mice.CONCLUSION:TMEM206 is associated with corneal edema which caused organizational disruption of collagen fibrils in corneas of mice.
基金supported by NIH grant No. 2 R01 ES010845 to M.S.the UNC Nutrition Obesity Research Center grant no. DK056350,+1 种基金NIH grant No. P30ES010126 to the UNC Center for Environmental Health and Susceptibilitysupported by a pre-doctoral traineeship (National Research Service Award T32 ES007126) from the National Institute of Environmental Health Sciences, NIH
文摘Arsenic methyltransferase（As3mt） catalyzes the conversion of inorganic arsenic（i As） to its methylated metabolites, including toxic methylarsonite（MAs~Ⅲ） and dimethylarsinite（DMAs~Ⅲ）. Knockout（KO） of As3 mt was shown to reduce the capacity to methylate i As in mice. However, no data are available on the oxidation states of As species in tissues of these mice. Here, we compare the oxidation states of As species in tissues of male C57BL/6 As3mt-KO and wild-type（WT） mice exposed to arsenite（iA s~Ⅲ） in drinking water. WT mice were exposed to50 mg/L As and As3mt-KO mice that cannot tolerate 50 mg/L As were exposed to 0, 15, 20, 25 or30 mg/L As. iA s~Ⅲaccounted for 53% to 74% of total As in liver, pancreas, adipose, lung, heart, and kidney of As3mt-KO mice; tri- and pentavalent methylated arsenicals did not exceed 10% of total As. Tissues of WT mice retained iA s and methylated arsenicals： iA s~Ⅲ, MAs~Ⅲand DMAs~Ⅲ represented 55%‐68% of the total As in the liver, pancreas, and brain. High levels of methylated species, particularly MAs~Ⅲ, were found in the intestine of WT, but not As3mt-KO mice,suggesting that intestinal bacteria are not a major source of methylated As. Blood of WT mice contained significantly higher levels of As than blood of As3mt-KO mice. This study is the first to determine oxidation states of As species in tissues of As3mt-KO mice. Results will help to design studies using WT and As3mt-KO mice to examine the role of iA s methylation in adverse effects of iA s exposure.
文摘Objective To explore the regulatory effect of fragile X mental retardation protein （FMRP） on the translation of microtubule associated protein 1B （MAP1B）. Methods The expressions of MAP1B protein and MAP1B mRNA in the brains of 1-week and 6-week old fragile X mental retardation-1 （FmrI） knockout （KO） mice were investigated by immunohistochemistry, Western blot, and in situ hybridization, with the age-matched wild type mice （WT） as controls. Results The mean optical density （MOD） of MAP1B was significantly decreased in each brain region in KO6W compared with WT6W, whereas in KO1W, this decrease was only found in the hippocampus and cerebellum. MAP1B in 6-week mice was much less than that in 1-week mice of the same genotype. The results of Western blot and in situ hybridization showed that MAP1B protein and MAP1B mRNA were significantly decreased in the hippocampus of both KO1W and KO6W. Conclusion The decreased MAP1B protein and MAP1B mRNA in the Fmrl knockout mice indicate that FMRP may positively regulate the expression of MAP1B.
基金supported by grants from Zhejiang Provincial Natural Science Foundation of China(No.LQ21H200007)National Natural Science Foundation of China(No.82202605,No.81772664 and No.82172363)+1 种基金Zhejiang Provincial People’s Hospital Excellent Scientific Research Start-up Fundation of China(No.ZRY2019C008)Hangzhou Medical College Fundamental Scientific Research Project of China(No.KYQN202116).
文摘Objective The protein interacting with C kinase 1(PICK1)plays a critical role in vesicle trafficking,and its deficiency in sperm cells results in abnormal vesicle trafficking from Golgi to acrosome,which eventually disrupts acrosome formation and leads to male infertility.Methods An azoospermia sample was filtered,and the laboratory detection and clinical phenotype indicated typical azoospermia in the patient.We sequenced all of the exons in the PICK1 gene and found that there was a novel homozygous variant in the PICK1 gene,c.364delA(p.Lys122SerfsX8),and this protein structure truncating variant seriously affected the biological function.Then we constructed a PICK1 knockout mouse model using clustered regularly interspaced short palindromic repeat cutting technology(CRISPRc).Results The sperm from PICK1 knockout mice showed acrosome and nucleus abnormalities,as well as dysfunctional mitochondrial sheath formation.Both the total sperm and motility sperm counts were decreased in the PICK1 knockout mice compared to wild-type mice.Moreover,the mitochondrial dysfunction was verified in the mice.These defects in the male PICK1 knockout mice may have eventually led to complete infertility.Conclusion The c.364delA novel variant in the PICK1 gene associated with clinical infertility,and pathogenic variants in the PICK1 may cause azoospermia or asthenospermia by impairing mitochondrial function in both mice and humans.
文摘Early studies suggested that estrogen receptor alpha （ERa） is involved in estrogen-mediated imprinting effects in prostate development. We recently reported a more complete ERa knockout （KO） mouse model via mating β-actin Cre transgenic mice with floxed ERa mice. These ACTB-ERaKO male mice showed defects in prostatic branching morphogenesis, which demonstrates that ERa is necessary to maintain proliferative events in the prostate. However, within which prostate cell type ERa exerts those important functions remains to be elucidated. To address this, we have bred floxed ERa mice with either fibroblast-specific protein （FSP）-Cre or probasin-Cre transgenic mice to generate a mouse model that has deleted ERa gene in either stromal fibroblast （FSP-ERaKO） or epithelial （pes-ERaKO） prostate cells. We found that circulating testosterone and fertility were not altered in FSP.ERaKO and pes-ERaKO male mice. Prostates of FSP-ERaKO mice have less branching morphogenesis compared to that of wild-type littermates. Further analyses indicated that loss of stromal ERa leads to increased stromal apoptosis, reduced expression of insulin-likegrowth factor-1 （IGF-1） and FGFIO, and increased expression of BMP4. Collectively, we have established the first in vivo prostate stromal and epithelial selective ERaKO mouse models and the results from these mice indicated that stromal fibroblast ERa plays important roles in prostatic branching morphogenesis via a paracrine fashion. Selective deletion of the ERa gene in mouse prostate epithelial cells by probasin-Cre does not affect the regular prostate development and homeostasis.
基金the National Natural Science Foundation of China,No.81372585 and No.81772557Beijing Health System High Level Training Plan of Health Technical Personnel,No.2014-3-048
文摘BACKGROUND Claudin-7, one of the important components of cellular tight junctions, is currently considered to be expressed abnormally in colorectal inflammation and colorectal cancer. However, there is currently no effective animal model to study its specific mechanism. Therefore, we constructed three lines of Claudin-7 knockout mice using the Cre/LoxP system.AIM To determine the function of the tumor suppressor gene Claudin-7 by generating three lines of Claudin-7 gene knockout mice.METHODS We crossed Claudin-7-floxed mice with CMV-Cre, vil1-Cre, and villin-CreERT2 transgenic mice, and the offspring were self-crossed to obtain conventional Claudin-7 knockout mice, conditional(intestinal specific) Claudin-7 knockout mice, and inducible conditional Claudin-7 knockout mice. Intraperitoneal injection of tamoxifen into the inducible conditional Claudin-7 knockout mice can induce the knockout of Claudin-7. PCR and agarose gel electrophoresis were used to identify mouse genotypes, and Western blot was used to confirm the knockout of Claudin-7. The mental state, body length, and survival time of these mice were observed. The dying mice were sacrificed, and hematoxylin-eosin(HE) staining and immunohistochemical staining were performed to observe changes in intestinal structure and proliferation markers.RESULTS We generated Claudin-7-floxed mice and three lines of Claudin-7 gene knockout mice using the Cre/LoxP system successfully. Conventional and intestinal specific Claudin-7 knockout mice were stunted and died during the perinatal period, and intestinal HE staining in these mice revealed mucosal gland structure disappearance and connective tissue hyperplasia with extensive inflammatory cell infiltration. The inducible conditional Claudin-7 knockout mice had a normal phenotype at birth, but after the induction with tamoxifen, they exhibited a dying state. Intestinal HE staining showed significant inflammatory cell infiltration, and atypical hyperplasia and adenoma were also observed. Intestinal immunohistochemistry analysis showed abnormal expression and distribution of Ki67, and the normal intestinal proliferation balance was disrupted. The intestinal crypt size in inducible conditional Claudin-7 knockout mice was increased compared with control mice(small intestine: 54.1 ± 2.96 vs 38.4 ± 1.63;large intestine: 44.7 ± 1.93 vs 27.4 ± 0.60; P < 0.001).CONCLUSION The knockout of Claudin-7 in vivo causes extensive inflammation, atypical hyperplasia, and adenoma in intestinal tissue as well as animal death in mice.Claudin-7 may act as a tumor suppressor gene in the development of colorectal cancer.
文摘The distribution of the Na-K-2Cl co-transporter （NKCCl） in the cochlear K^＋ cycling pathway in cochlea and cochlear histological changes in the NKCCl knockout mice were investigated. By using immunohistochemistry and toluidine blue staining, the localization of NKCCl in cochlea of the C57BL/6J mice and the cochlear histological changes in the NKCCl knockout mice were observed. It was found that the NKCCl was expressed mainly in the stria marginal cells and the fibrocytes in the inferior portion of the spiral ligament in the adult C57BL/6J mice. Subpopulation of the fibrocytes in the suprastrial region and the limbus was also moderately immunoreactive. While in the cochlea of the NKCCl knockout mice, Reissner＇s membrane was collapsed and scala media disappeared, accompanied with the loss of inner hair cells, outer hair cells and the support cells. The tunnel of Corti was often absent. All the findings suggested the localization of NKCCl in the cochlea was closely correlated with cochlear K^＋ cycling. Loss of NKCCl led to the destruction of the cochlear structures, and subsequently influenced the physiological function of cochlea.
基金Supported by the National Natural Science Foundation of China(Nos30470405 and 30670477)National Natural ScienceFund for Distinguished Young Scholars(No30325011)+1 种基金Distinguished Young Scholars Fund of Jilin Province(No20030112)Excellent Young Tea
文摘An overt phenotype of aquaporin-1 knockout（AQP1 ko） mice is growth retardation, suggesting possible defects in bone development and metabolism. In the present study, we analyzed the bone mineral density（ BMD）, bone calcium and phosphorus contents, and bone metabolism in an AQP1 ko mouse model. The BMD of femurs in AQP1 ko mice was significantly lower than that of litter-matched wildtype mice as measured by dual energy X-ray absorptiometry. Consistently, the contents of bone total calcium and phosphorus were also significantly lower in AQP1 ko mice. The reduced BMD caused by AQP1 deficiency mainly affect male mice. Bone metabolic activity, as indicated by 99m^Tc-MDP absorption measurements, was remarkably reduced in AQP1 ko mice. These results provide the first evidence that AQP1 play an important role in bone structure and metabolism.
基金Supported by Public Health Service grant R01-CA101931 (to DJM) from the National Institutes of Health
文摘AIM: To investigate the role of host and bacterial arginases in the colonization of mice by Helicobacter pylori (H.pylori).METHODS: H.pylori produces a very powerful urease that hydrolyzes urea to carbon dioxide and ammonium,which neutralizes acid.Urease is absolutely essential to H.pylori pathogenesis;therefore,the urea substrate must be in ample supply for urease to work efficiently.The urea substrate is most likely provided by arginase activity,which hydrolyzes L-arginine to L-ornithine and urea.Previous work has demonstrated that H.pylori arginase is surprisingly not required for colonization of wild-type mice.Hence,another in vivo source of the critical urea substrate must exist.We hypothesized that the urea source was provided by host arginase Ⅱ,since this enzyme is expressed in the stomach,and H.pylori has previously been shown to induce the expression of murine gastric arginase Ⅱ.To test this hypothesis,wild-type and arginase (rocF) mutant H.pylori strain SS1 were inoculated into arginase Ⅱ knockout mice.RESULTS: Surprisingly,both the wild-type and rocF mutant bacteria still colonized arginase Ⅱ knockout mice.Moreover,feeding arginase Ⅱ knockout mice the host arginase inhibitor S-(2-boronoethyl)L-cysteine (BEC),while inhibiting > 50% of the host arginase Ⅰ?activity in several tissues,did not block the ability of the rocF mutant H.pylori to colonize.In contrast,BEC poorly inhibited H.pylori arginase activity.CONCLUSION: The in vivo source for the essential urea utilized by H.pylori urease is neither bacterial arginase nor host arginase Ⅱ;instead,either residual host arginase Ⅰ?or agmatinase is probably responsible.
基金supported by the National Natural Science Foundation of China International Cooperation and Exchange Program(8161101193)the National Science and Technology Major Project(2016ZX10004222)to G.Wong
文摘Due to their inability to generate a complete immune response, mice knockout for type I interferon （IFN） receptors （Ifnar-/-） are more susceptible to viral infections, and are thus commonly used for pathogenesis studies. This mouse model has been used to study many diseases caused by highly pathogenic viruses from many families, including the Flaviviridae, Filoviridae, Arenaviridae, Bunyaviridae, Henipaviridae, and Togaviridae. In this review, we summarize the findings from these animal studies, and discuss the pros and cons of using this model versus other known methods for studying pathogenesis in animals.
基金supported by the National Natural Science Foundation of China(Grant No.31671561)to Dong Zhang.
文摘Microtubule-severing proteins(MTSPs),are a family of proteins which use adenosine triphosphate to sever microtubules.MTSPs have been shown to play an important role in multiple microtubule-involved cellular processes.One member of this family,fidgetin(FIGN),is also involved in male fertility;however,no studies have explored its roles in female fertility.In this study,we found mouse fidgetin is rich within oocyte zona pellucida(ZP)and is the only MTSP member to do so.Fidgetin also appears to interact with all three ZP proteins.These findings prompted us to propose that fidgetin might prevent polyspermy.Results from in vitro maturation oocytes analysis showed that fidgetin knockdown did cause polyspermy.We then deleted all three fidgetin isoforms with CRISPR/Cas9 technologies;however,female mice remained healthy and with normal fertility.Of all mouse MTSPs,only the mRNA level of fidgetin-like 1(FIGNL1)significantly increased.Therefore,we assert that fidgetin-like 1 compensates fidgetin's roles in fidgetin knockout female mice.
基金National Natural Science Foundation of China(81960761,81960751,81902764)。
文摘[Objectives]This study was conducted to investigate the similarity and differences between TLR4 knockout mice and C57 BL/6 mice used in liver fibrosis research in terms of growth rate and reproduction ability.[Methods]Twenty TLR4 knockout mice and C57 BL/6 mice,half male and half female,were selected to compare the growth rates of body weight and body length of mice from the 4th to 12th weeks;and 20 pairs of male and female mice of the same strain were compared for the number of baby mice of the second litter.[Results]The growth rates of body weight and body length of the TLR4 knockout mice were significantly lower than those of C57 BL/6 mice(P<0.05)(except for the 4th and 5th weeks when there was no significant difference in body length);and in terms of reproductive ability,the TLR4 knockout mice were significantly lower than the C57 BL/6 mice(the ratio of the total number of baby mice in the second litter of the two strains,72∶147).[Conclusions]Knockout of the TLR4 gene has a significant impact on the growth and reproduction of mice.
文摘Objective-To investigate the effects of heat shock transcription factor 1) gene on the constitutivety expressed αB-CrystaUin (aBC) in mice myocardium. Methods-The expression levels of constitutive aBC in HSF1 knockout (hsf1 - /- ) and HSFl wild type (As/1 + /+) mice myocardium were evaluated by western blot and immunohistochemistry. Results : The αBC levels in hsfl -/- and hsfl +/+ were 68. 42±4. 16, 100. 00±7. 58, respectively (P<0. 05, cytoso-lic fraction) , and 20. 53±1. 01, 37. 55±1. 91, respectively (P<0. 05, pellet fraction). The aBC signals decreased significantly in hsfl -/- myocardium when compared with those in hsfl +/+ myocardium stained with fluorescence immunohistochemistry. Conclusion-HSF1 is an important, but not the only factor, which mediates the constitutively expressed aBC.
基金supported by the grand from the Department of Education Project of Hubei Province to WANG Zhi-xiao (B20102108)
文摘Background Immune inflammatory response is throughout the entire process of atherosclerosis （AS）. It was unclear whether the mechanism of mesenchymal stem cells （MSCs） transplantation for treatment of AS is involved with inflammation regulation in the plaque area. The aim of this study was to explore the effects of MSCs in the formation of atherosclerosis plaque in hypercholesterolemic apoliprotein （apo）E-/- mice. Methods ApoE-/- mice MSCs were isolated and identified. At 8 weeks of age, 30 male ApoE-/- mice were randomly divided into negative control group （Neg, n = 10）, positive control group （Pos, n = 10） and mesenchymal stem cells group （MSCs, n = 10）. MSCs were injected through caudal vein into the body of Pos and MSCs group. The plaque area of all subjects were compared, the percentage of CD4^＋CD25^＋Tregs in different tissues were analyzed by fluorescence activated cell sorter （FACS）, proliferation response of splenocytes to MSCs was detected and cytokines in the supernatant were determined by enzyme linked immunosorbent assay （ELISA）. Results Compared with controls, MSCs resulted in a significant decrease of latherosclerotic plaques size （P 〈 0.05）, and a significant increase of CD4^＋CD25^＋ regulatory T cells in spleen （P 〈 0.05）. Specific proliferation response of CD4^＋CD25^＋ regulatory T cells in splenocytes to MCSs was significantly suppressed, the superanant level of TGF-[3 and IL-10 in MSCs group were increased while IFN-γ/ decreased significantly. Conclusion MSCs play an important role in regulating the inflammatory response and significantly inhibit the formation of the atherosclerosis plaque in ApoE-/-mice.
基金supported by a grant from theNational Natural Science Foundation of Chinato Dr LI Dazhu (No.C03030201)
文摘Background Subtypes of T cells, called regulatory T cells （Treg cell）, play a critical role in limiting autoimmune processes and inflammatory responses, The aim of this study was to explore functional roles of FOXP3 in the manifestation of atherosclerosis in Apolipoprotein E deficient （ApoE）-/- mice. Methods Lentivirus-mediated （siRNA） was used to knock down FOXP3 and FOXP3high＋CD4＋ CD25＋ T cells adoptive transfer assays in high fat diet ApoE-/- mice were done. The resulting atherosclerotie lesions were assessed by determining FOXP3 transcript levels and investigating the expression of FOXP3 protein in different tissues. Results Animals treated with siRNA of FOXP3 showed a significant increase in atherosclerotic lesion formation and a reduction in the number of FOXP3＋CD4＋CD25＋ T cells compared with other groups. Transfer of FOXP3highCD4＋CD25＋ T cells significantly decreased atherosclerotic plaque formation and increased the number of FOXP3＋ CD4＋ CD25＋ T cells. FOXP3 protein levels and FOXP3 transcript levels were lowest in the siRNA group, and were highest in tissues from the Treg transfer group. Conclusion FOXP3 plays an important role in regulating the inflammatory response within the atherosclerotic lesion. It can inhibit significantly the progression of the atherosclerosis plaque in ApoE-/- mice.
基金supported by National Natural Science Foundation of China (81560586) and Xinjiang Natural Science Foundation (2016D01C161)
文摘OBJECTIVE To observe the effects of serum metabolites by using ~1H-NMR-based metabonomic approach to explore the possible mechanisms of total flavonoids in Ocimum BasilicumLinn(OBL) on atherosclerosis in apolipomtein E knockout(ApoE-/-) mice.METHODS Six-week-old male apoE knockout mice were divided into four groups(n=10) and fed with high fet diet:model,Simv.astatin,OBL-H,OBL-M and OBL-L groups.The homogeneous male mice of C57 BL/6 J were used as the normol group and fed with normal chow diet.After 14 weeks,~1H-NMR technology was used to ex.plore the variability of serum metabolites by the method of PLS-DA and OPLS-DA.RESULTS Com.pared with normal group,Model group showed a significant increase in the serum levels of VLDL,LDL,β-hydroxyisobutyrate,lactate,myo-inositol and showed a significant decrease in the serum levels of al.anine,glutamine,proline,carnitine,methylamine,citrate,creatine,choline,taurine,pyruvate,β-glu.cose,α-glucose,glycine,lysine.Combined with model group OBL-H,OBL-M,OBL-L groups showed the effects of regulating the levels of different metabolites of the glucose,lipid and amino acid metabo.lism.CONCLUSION The anti-atheros-clerotic activity of total flavonoids in Ocimum BasilicumLinn may be related not only to regulation of lipid metabolism,but also glycometabolism and amino acid metabolism.
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 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.
