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Chromatin condensation but not DNA integrity of pig sperm is greater in the sperm-rich fraction

Journal of animal Science and Biotechnology 2024年第1期15卷 171-181页

作者： Estel Viñolas-Vergés Jordi Ribas-Maynou Isabel Barranco Camila Peres Rubio Sergi Bonet Jordi Roca Marc Yeste Unit of Cell Biology Department of BiologyFaculty of SciencesUniversity of GironaGironaSpain Biotechnology of animal and Human Reproduction(TechnoSperm) Institute of Food and Agricultural TechnologyUniversity of GironaGironaSpain department of medicine and animal Surgery Faculty of Veterinary ScienceUniversity of MurciaMurciaSpain Catalan Institution for Research and Advanced Studies(ICREA) S08010 BarcelonaSpain

Background Protamination and condensation of sperm chromatin as well as DNA integrity play an essential role during fertilization and embryo development.In some mammals,like pigs,ejaculates are emitted in three separate fractions:pre-sperm,sperm-rich(SRF)and post sperm-rich(PSRF).These fractions are known to vary in volume,sperm concentration and quality,as well as in the origin and composition of seminal plasma(SP),with differences being also observed within the SRF one.Yet,whether disparities in the DNA integrity and chromatin condensation and pro-tamination of their sperm exist has not been interrogated.Results This study determined chromatin protamination(Chromomycin A3 test,CMA_(3)),condensation(Dibromobi-mane test,DBB),and DNA integrity(Comet assay)in the pig sperm contained in the first 10 m L of the SRF(SRF-P1),the remaining portion of the sperm-rich fraction(SRF-P2),and the post sperm-rich fraction(PSRF).While chromatin protamination was found to be similar between the different ejaculate fractions(P>0.05),chromatin condensation was seen to be greater in SRF-P1 and SRF-P2 than in the PSRF(P=0.018 and P=0.004,respectively).Regarding DNA integrity,no differences between fractions were observed(P>0.05).As the SRF-P1 has the highest sperm concentra-tion and ejaculate fractions are known to differ in antioxidant composition,the oxidative stress index(OSi)in SP,calcu-lated as total oxidant activity divided by total antioxidant capacity,was tested and confirmed to be higher in the SRF-P1 than in SRF-P2 and PSRF(0.42±0.06 vs.0.23±0.09 and 0.08±0.00,respectively;P<0.01);this index,in addition,was observed to be correlated to the sperm concentration of each fraction(Rs=0.973;P<0.001).Conclusion While sperm DNA integrity was not found to differ between ejaculate fractions,SRF-P1 and SRF-P2 were observed to exhibit greater chromatin condensation than the PSRF.This could be related to the OSi of each fraction.

关键词： Chromatin Condensation DNA integrity Ejaculate fractions Pig Protamination Sperm

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Identifying and validating MMP family members(MMP2,MMP9,MMP12,and MMP16)as therapeutic targets and biomarkers in kidney renal clear cell carcinoma(KIRC)

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Oncology Research 2024年第4期32卷 737-752页

作者： KUNLUN LI DANDAN LI BARBOD HAFEZ MOUNIR M.SALEM BEKHIT YOUSEF A.BIN JARDAN FARS KAED ALANAZI EHAB I.TAHA SAYED H.AUDA FAIQAH RAMZAN MUHAMMAD JAMIL The Second Affiliated Hospital of Harbin Medical University Harbin Medical UniversityHarbinChina department of Pharmaceutical Engineering Jiangsu Ocean UniversityLianyungangChina department of Biological Engineering University of SalfordSalfordUK department of Pharmaceutics College of PharmacyKing Saud UniversityRiyadhSaudi Arabia department of animal and Poultry production Faculty of Veterinary and Animal SciencesGomal UniversityDera Ismail KhanPakistan department of Arid Zone Research PARC instituteDera Ismail KhanPakistan

Kidney Renal Clear Cell Carcinoma(KIRC)is a malignant tumor that carries a substantial risk of morbidity and mortality.The MMP family assumes a crucial role in tumor invasion and metastasis.This study aimed to uncover the mechanistic relevance of the MMP gene family as a therapeutic target and diagnostic biomarker in Kidney Renal Clear Cell Carcinoma(KIRC)through a comprehensive approach encompassing both computational and molecular analyses.STRING,Cytoscape,UALCAN,GEPIA,OncoDB,HPA,cBioPortal,GSEA,TIMER,ENCORI,DrugBank,targeted bisulfite sequencing(bisulfite-seq),conventional PCR,Sanger sequencing,and RT-qPCR based analyses were used in the present study to analyze MMP gene family members to accurately determine a few hub genes that can be utilized as both therapeutic targets and diagnostic biomarkers for KIRC.By performing STRING and Cytohubba analyses of the 24 MMP gene family members,MMP2(matrix metallopeptidase 2),MMP9(matrix metallopeptidase 9),MMP12(matrix metallopeptidase 12),and MMP16(matrix metallopeptidase 16)genes were denoted as hub genes having highest degree scores.After analyzing MMP2,MMP9,MMP12,and MMP16 via various TCGA databases and RT-qPCR technique across clinical samples and KIRC cell lines,interestingly,all these hub genes were found significantly overexpressed at mRNA and protein levels in KIRC samples relative to controls.The notable effect of the up-regulated MMP2,MMP9,MMP12,and MMP16 was also documented on the overall survival(OS)of the KIRC patients.Moreover,targeted bisulfite-sequencing(bisulfite-seq)analysis revealed that promoter hypomethylation pattern was associated with up-regulation of hub genes(MMP2,MMP9,MMP12,and MMP16).In addition to this,hub genes were involved in various diverse oncogenic pathways.The MMP gene family members(MMP2,MMP9,MMP12,and MMP16)may serve as therapeutic targets and prognostic biomarkers in KIRC.

关键词： KIRC MMP gene family Chemotherapy Overall survival

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A nanobody-based blocking enzyme-linked immunosorbent assay for detecting antibodies against pseudorabies virus glycoprotein E

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Journal of Integrative Agriculture 2024年第4期23卷 1354-1368页

作者： Huanhuan Lü Pinpin Ji Siyu Liu Ziwei Zhang Lei Wang Yani Sun Baoyuan Liu Lizhen Wang Qin Zhao department of Preventive veterinary medicine College of Veterinary MedicineNorthwest A&F University/Shaanxi Scientific Observing and Experimental Station of Veterinary Pharmacology and Diagnostic TechnologyMinistry of Agriculture and Rural AffairsYangling 712100China

Pseudorabies(PR)is an acute infectious disease of pigs caused by the PR virus(PRV)and results in great economic losses to the pig industry worldwide.PRV glycoprotein E(gE)-based enzyme-linked immunosorbent assay(ELISA)has been used to distinguish gE-deleted vaccine-immunized pigs from wild-type virus-infected pigs to eradicate PR in some countries.Nanobody has the advantages of small size and easy genetic engineering and has been a promising diagnostic reagent.However,there were few reports about developing nanobody-based ELISA for detecting anti-PRV-gE antibodies.In the present study,the recombinant PRV-gE was expressed with a bacterial system and used to immunize the Bactrian camel.Then,two nanobodies against PRV-gE were screened from the immunized camel by phage display technique.Subsequently,two nanobody-HRP fusion proteins were expressed with HEK293T cells.The PRV-gE-Nb36-HRP fusion protein was selected as the probe for developing the blocking ELISA(bELISA)to detect anti-PRV-gE antibodies.Through optimizing the conditions of bELISA,the amount of coated antigen was 200 ng per well,and dilutions of the fusion protein and tested pig sera were separately 1:320 and 1:5.The cut-off value of bELISA was 24.20%,and the sensitivity and specificity were 96.43 and 92.63%,respectively.By detecting 233 clinical pig sera with the developed bELISA and a commercial kit,the results showed that the coincidence rate of two assays was 93.99%.Additionallly,epitope mapping showed that PRV-gE-Nb36 recognized a conserved conformational epitope in different reference PRV strains.Simple,great stability and low-cost nanobody-based bELISA for detecting anti-PRV-gE antibodies were developed.The bELISA could be used for monitoring and eradicating PR.

关键词： nanobody nanobody-HRP blocking ELISA PRV-gE antibody

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Gram-negative bacterial diversity and evidence of international clones of multidrug-resistant strains in zoo animals

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Integrative Zoology 2024年第3期19卷 417-423页

作者： João Pedro Rueda FURLAN Micaela Santana RAMOS Fábio Parra SELLERA Irys Hany Lima GONZALEZ Patrícia Locosque RAMOS Eliana Guedes STEHLING department of Clinical Analyses Toxicology and Food ScienceSchool of Pharmaceutical Sciences of Ribeirão PretoUniversity of São PauloRibeirão PretoBrazil department of Internal medicine School of Veterinary Medicine and Animal ScienceUniversity of São PauloSão PauloBrazil School of veterinary medicine Metropolitan University of SantosSantosBrazil Coordination of Wild Fauna Secretary of EnvironmentInfrastructure and LogisticsSão PauloBrazil

Antimicrobial resistance(AMR)is a growing phenomenon that poses a major public health challenge worldwide(Dhingra et al.2020;Antimicrobial Resistance Collaborators 2022).The selective pressure exerted in clinical settings has contributed to the longlasting successful dissemination of multidrug-resistant(MDR)Gram-negative bacteria in these environments(Chia et al.2020).

关键词： multidrug clinical

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Antigen epitopes of animal coronaviruses:a mini-review

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animal Diseases 2024年第1期4卷 19-26页

作者： Mingjun Su Guanghui Zheng Xiangwen Xu Houhui Song Key Laboratory of Applied Technology On Green-Eco-Healthy animal Husbandry of Zhejiang Province Zhejiang Provincial Engineering Research Center for Animal Health Diagnostics&Advanced TechnologyZhejiang International Science and Technology Cooperation Base for Veterinary Medicine and Health ManagementChina-Australia Joint Laboratory for Animal Health Big Data AnalyticsCollege of Animal Science and Technology&College of Veterinary Medicine of ZhejiangA&F University666 Wusu StreetLin'an DistrictHangzhou 311300Zhejiang ProvinceChina

Coronaviruses are widespread in nature and can infect mammals and poultry,making them a public health concern.Globally,prevention and control of emerging and re-emerging animal coronaviruses is a great challenge.The mecha-nisms of virus-mediated immune responses have important implications for research on virus prevention and control.The antigenic epitope is a chemical group capable of stimulating the production of antibodies or sensitized lympho-cytes,playing an important role in antiviral immune responses.Thus,it can shed light on the development of diagnos-tic methods and novel vaccines.Here,we have reviewed advances in animal coronavirus antigenic epitope research,aiming to provide a reference for the prevention and control of animal and human coronaviruses.

关键词： animal coronavirus Antigen epitope B-cell epitope T-cell epitope Immune responses Vaccines

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Effects of early postnatal gastric and colonic microbiota transplantation on piglet gut health

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Journal of animal Science and Biotechnology 2024年第2期15卷 645-658页

作者： Christina Larsen Simone Margaard Offersen Anders Brunse Mattia Pirolo Soumya Kanti Kar Luca Guadabassi Thomas Thymann department of veterinary and animal Science University of CopenhagenDyrlagevej 681870 Frederiksberg CDenmark animal Nutrition Wageningen Livestock ResearchWageningen University&Research1 De Elst6708 WageningenThe Netherlands

Background Diarrhea is a major cause of reduced growth and mortality in piglets during the suckling and weaning periods and poses a major threat to the global pig industry.Diarrhea and gut dysbiosis may in part be prevented via improved early postnatal microbial colonization of the gut.To secure better postnatal gut colonization,we hypothesized that transplantation of colonic or gastric content from healthy donors to newborn recipients would prevent diarrhea in the recipients in the post-weaning period.Our objective was to examine the impact of transplanting colonic or gastric content on health and growth parameters and paraclinical parameters in recipient single-housed piglets exposed to a weaning transition and challenged with enterotoxigenic Escherichia coli(ETEC).Methods Seventy-two 1-day-old piglets were randomized to four groups:colonic microbiota transplantation(CMT,n=18),colonic content filtrate transplantation(CcFT,n=18),gastric microbiota transplantation(GMT,n=18),or saline(CON,n=18).Inoculations were given on d 2 and 3 of life,and all piglets were milk-fed until weaning(d 20)and shortly after challenged with ETEC(d 24).We assessed growth,diarrhea prevalence,ETEC concentration,organ weight,blood parameters,small intestinal morphology and histology,gut mucosal function,and microbiota composition and diversity.Results Episodes of diarrhea were seen in all groups during both the milk-and the solid-feeding phase,possibly due to stress associated with single housing.However,CcFT showed lower diarrhea prevalence on d 27,28,and 29 compared to CON(all P<0.05).CcFT also showed a lower ETEC prevalence on d 27(P<0.05).CMT showed a higher alpha diversity and a difference in beta diversity compared to CON(P<0.05).Growth and other paraclinical endpoints were similar across groups.Conclusion In conclusion,only CcFT reduced ETEC-related post-weaning diarrhea.However,the protective effect was marginal,suggesting that higher doses,more effective modalities of administration,longer treatment periods,and better donor quali

关键词： Colonic content filtrate transplantation Colonic microbiota transplantation Gastric microbiota transplantation Gut microbiota Mucosa Neonatal Post-weaning diarrhea

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KLF4 facilitates chromatin accessibility remodeling in porcine early embryos

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Science China(Life Sciences) 2024年第1期67卷 96-112页

作者： Wei Zhu Guowei Bu Ruifeng Hu Jixiang Zhang Lianyong Qiao Kai Zhou Tingting Wang Qiao Li Jingjing Zhang Linhui Wu Yali Xie Taotao Hu Shichun Yang Jiaqi Guan Xiaoyu Chu Juanjuan Shi Xia Zhang Falong Lu Xin Liu Yi-Liang Miao Institute of Stem Cell and Regenerative Biology College of Animal Science and Veterinary MedicineHuazhong Agricultural UniversityWuhan 430070China Key Laboratory of Agricultural animal Genetics Breeding and Reproduction(Huazhong Agricultural University)Ministry of EducationWuhan 430070China State Key Laboratory of Molecular Developmental Biology Institute of Genetics and Developmental BiologyInnovative Academy of Seed DesignChinese Academy of SciencesBeijing 100101China Frontiers Science Center for animal Breeding and Sustainable production Huazhong Agricultural UniversityWuhan 430070China Hubei Hongshan Laboratory Wuhan 430070China

Chromatin accessibility remodeling driven by pioneer factors is critical for the development of early embryos.Current studies have illustrated several pioneer factors as being important for agricultural animals,but what are the pioneer factors and how the pioneer factors remodel the chromatin accessibility in porcine early embryos is not clear.By employing low-input DNase-seq(liDNase-seq),we profiled the landscapes of chromatin accessibility in porcine early embryos and uncovered a unique chromatin accessibility reprogramming pattern during porcine preimplantation development.Our data revealed that KLF4 played critical roles in remodeling chromatin accessibility in porcine early embryos.Knocking down of KLF4 led to the reduction of chromatin accessibility in early embryos,whereas KLF4 overexpression promoted the chromatin openness in porcine blastocysts.Furthermore,KLF4 deficiency resulted in mitochondrial dysfunction and developmental failure of porcine embryos.In addition,we found that overexpression of KLF4 in blastocysts promoted lipid droplet accumulation,whereas knockdown of KLF4 disrupted this process.Taken together,our study revealed the chromatin accessibility dynamics and identified KLF4 as a key regulator in chromatin accessibility and cellular metabolism during porcine preimplantation embryo development.

关键词： porcine embryos liDNase-seq chromatin accessibility KLF4 mitochondrion lipid droplets

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Mulberry leaf supplementation inhibits skatole deposition by regulating gut microbiota and upregulating liver cytochrome P4501A1 expression in finishing pigs

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animal Nutrition 2024年第1期16卷 34-44页

作者： Yuqing Sun Xiaoming Men Tianbao Lin Bo Deng Shi Zhong Jinxi Huo Kaipeng Qin Zhiqiang Lv Ziwei Xu Yougui Li Institute of Sericulture and Tea Zhejiang Academy of Agricultural Sciences Institute of animal Husbandry and veterinary Science Zhejiang Academy of Agricultural Sciences

Skatole, a strong fecal odor substance, is generated through microbial degradation of tryptophan in the animal hindgut. It easily accumulates in adipose tissue and affects meat quality. In this study, the effect of mulberry leaf supplementation on skatole in finishing pigs was studied. In a 35-day trial, 20 finishing pigs (barrows and gilts) were fed with a basal diet or basal diet with 6% mulberry leaves. Growth performance of the pigs (n=10) was automatically recorded by a performance-testing feeder system and 8 pigs in each treatment were slaughtered and sampled for the remaining tests. Skatole and short-chain fatty acids were detected using HPLC and gas chromatography, respectively. Fecal microbiota were analyzed using 16S r RNA gene sequencing. The metabolomics analysis of feces and serum was performed with UHPLC-MS/MS. The major cytochrome P450 (CYP) enzymes that catalyze skatole degradation in the liver were tested by using RT-PCR and Western blot. Effects of major bioactive compounds in mulberry leaves on the CYP genes were verified in the hepatic cell line Hep G2 in an in vitro test (n=3). In finishing pigs, mulberry leaf supplementation had no significant effect on the average daily gain, average daily feed intake, and feed conversion ratio (P>0.05), but reduced skatole levels in feces, serum, and backfat (Pproduction, was significantly reduced in feces by mulberry leaf supplementation (P<0.05) and was positively correlated with skatole content in feces (P=0.004). In pigs treated with mulberry leaves, liver CYP1A1 expression was increased (P<0.05) and was negatively correlated with skatole content in backfat (P=0.045). The in vitro test demonstrated that mulberry leaf polyphenols and polysaccharides could directly stimulate CYP1A1 expression in h

关键词： Pig Skatole Mulberry leaf Microbiota Cytochrome P450

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Newcastle disease virus suppresses antigen presentation via inhibiting IL-12 expression in dendritic cells

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Journal of Zhejiang University-Science B(Biomedicine & Biotechnology) 2024年第3期25卷 254-270页

作者： Fulong NAN Wenlong NAN Xin YAN Hui WANG Shasha JIANG Shuyun ZHANG Zhongjie YU Xianjuan ZHANG Fengjun LIU Jun LI Xiaoqiong ZHOU Delei NIU Yiquan LI Wei WANG Ning SHI Ningyi JIN Changzhan XIE Xiaoni CUI He ZHANG Bin WANG Huijun LU department of Special medicine Department of Pathogenic BiologySchool of Basic MedicineQingdao UniversityQingdao 266071China China animal Health and Epidemiology Center Qingdao 266032China Academician Workstation of Jilin Province Changchun University of Chinese MedicineChangchun 130117China Institute of Virology Wenzhou UniversityWenzhou 325035China College of veterinary medicine Jilin UniversityChangchun 130012China Changchun veterinary Research Institute Chinese Academy of Agricultural SciencesChangchun 130122China Sino-Cell Biomed Co. Ltd.Qingdao 266000China

As a potential vectored vaccine,Newcastle disease virus(NDV)has been subject to various studies for vaccine development,while relatively little research has outlined the immunomodulatory effect of the virus in antigen presentation.To elucidate the key inhibitory factor in regulating the interaction of infected dendritic cells(DCs)and T cells,DCs were pretreated with the NDV vaccine strain LaSota as an inhibitor and stimulated with lipopolysaccharide(LPS)for further detection by enzyme-linked immunosorbent assay(ELISA),flow cytometry,immunoblotting,and quantitative real-time polymerase chain reaction(qRT-PCR).The results revealed that NDV infection resulted in the inhibition of interleukin(IL)-12p40 in DCs through a p38 mitogen-activated protein kinase(MAPK)-dependent manner,thus inhibiting the synthesis of IL-12p70,leading to the reduction in T cell proliferation and the secretion of interferon-(IFN-),tumor necrosis factor-α(TNF-α),and IL-6 induced by DCs.Consequently,downregulated cytokines accelerated the infection and viral transmission from DCs to T cells.Furthermore,several other strains of NDV also exhibited inhibitory activity.The current study reveals that NDV can modulate the intensity of the innate-adaptive immune cell crosstalk critically toward viral invasion improvement,highlighting a novel mechanism of virus-induced immunosuppression and providing new perspectives on the improvement of NDV-vectored vaccine.

关键词： Newcastle disease virus Dendritic cells Interleukin-12(IL-12) T cells Immunosuppression

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AP-1 and SP1 trans-activate the expression of hepatic CYP1A1 and CYP2A6 in the bioactivation of AFB1 in chicken

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Science China(Life Sciences) 2024年第7期67卷 1468-1478页

作者： Jiang Deng Jia-Cheng Yang Yue Feng Ze-Jing Xu Kamil Kuča Meng Liu Lv-Hui Sun State Key Laboratory of Agricultural Microbiology Hubei Hongshan LaboratoryFrontiers Science Center for Animal Breeding and Sustainable ProductionCollege of Animal Sciences and TechnologyHuazhong Agricultural UniversityWuhan 430070China department of Chemistry Faculty of ScienceUniversity of Hradec KraloveHradec Kralove 50003Czech Republic

Dietary exposure to aflatoxin B1(AFB1)is harmful to the health and performance of domestic animals.The hepatic cytochrome P450s(CYPs),CYP1A1 and CYP2A6,are the primary enzymes responsible for the bioactivation of AFB1to the highly toxic exo-AFB1-8,9-epoxide(AFBO)in chicks.However,the transcriptional regulation mechanism of these CYP genes in the liver of chicks in AFB1metabolism remains unknown.Dual-luciferase reporter assay,bioinformatics and site-directed mutation results indicated that specificity protein 1(SP1)and activator protein-1(AP-1)motifs were located in the core region-1,063/-948,-606/-541 of the CYP1A1 promoter as well as-636/-595,-503/-462,-147/-1 of the CYP2A6 promoter.Furthermore,overexpression and decoy oligodeoxynucleotide technologies demonstrated that SP1 and AP-1 were pivotal transcriptional activators regulating the promoter activity of CYP1A1 and CYP2A6.Moreover,bioactivation of AFB1to AFBO could be increased by upregulation of CYP1A1 and CYP2A6 expression,which was trans-activated owing to the upregulation of AP-1,rather than SP1,stimulated by AFB1-induced reactive oxygen species.Additionally,nano-selenium could reduce ROS,downregulate AP-1 expression and then decrease the expression of CYP1A1 and CYP2A6,thus alleviating the toxicity of AFB1.In conclusion,AP-1 and SP1 played important roles in the transactivation of CYP1A1 and CYP2A6 expression and further bioactivated AFB1to AFBO in chicken liver,which could provide novel targets for the remediation of aflatoxicosis in chicks.

关键词： aflatoxin B1 CYP1A1 CYP2A6 SP1 AP-1 transcriptional activation

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