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Lactobacillus rhamnosus GR-1 attenuates foodborne Bacillus cereus-induced NLRP3 inflammasome activity in bovine mammary epithelial cells by protecting intercellular tight junctions

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Journal of Animal Science and Biotechnology 2023年第1期14卷 307-321页

作者： Qiang Shan Ning Liu Xue wang Yaohong Zhu Jinhua Yin jiufeng wang Department of Veterinary Clinical Sciences College of Veterinary MedicineChina Agricultural UniversityBeijing 100193China. College of Animal Science and Technology Tarim UniversityAlar 843300China.

Background:Bacillus cereus is an important pathogen that causes human food poisoning,specifically diarrhea and *** can also induce mastitis in dairy cows and has a strong survival ability in milk,as it cannot be inactivated by high-temperature short-time ***,*** can enter the market through pasteurized milk and other dairy products,imposing enormous hidden dangers on food safety and human ***:In this study,*** 2101(BC)was isolated from milk samples of cows with *** grew rapidly with strong hemolysis,making it difficult to prevent mastitis and ensure food ***-T cells were treated with BC and/or Lactobacillus rhamnosus GR-1(LGR-1).Pretreatment with LGR-1 protected the integrity of tight junctions and the expression of zonula occludens-1(ZO-1)and occludin destroyed by ***,LGR-1 pretreatment reduced the expression of NOD-like receptor family member pyrin domain-containing protein 3(NLRP3),caspase recruitment and activation domain(ASC),Caspase-1 p20,gasdermin D(GSDMD)p30,inflammatory factors(interleukin(IL)-1βand IL-18),and cell death induced by ***,LGR-1 pretreatment reduced NLRP3 inflammasome activity and increased expressions of ZO-1 and occludin induced by lipopolysaccharides(LPS)+ATP ***-T cells were transfected with NLRP3 si RNA or MCC950 and/or treated with BC and/or ***3-si RNA transfection and MCC950 attenuated BC-induced NLRP3 inflammasome *** of inflammatory cytokines and cell death suggested that the inflammatory pathway might play an important role in the induction of the NLRP3 inflammasome by BC and the protection of ***:These results suggest that LGR-1 might be a probiotic alternative to antibiotics and could be administered to prevent mastitis in dairy cows,thus ensuring food security.

关键词： Bacillus cereus Intercellular tight junctions Lactobacillus rhamnosus GR-1 NLRP3 inflammasome

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Effects of feeding untreated, pasteurized and acidified waste milk and bunk tank milk on the performance, serum metabolic profiles, immunity, and intestinal development in Holstein calves

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Journal of Animal Science and Biotechnology 2017年第4期8卷 934-944页

作者： Yang Zou Yajing wang Youfei Deng Zhijun Cao Shengli Li jiufeng wang State Key Laboratory of Animal Nutrition Beijing Engineering Technology Research Center of Raw Milk Quality and Safety Control College of Animal Science and Technology China Agricultural University Beijing 100193 China Beijing Dairy Cattle Center Beijing 100192 China College of Veterinary Medicine China Agricultural University Beijing 100193 China

Background: The present experiment was performed to assess the effects of different sources of milk on the growth performance, serum metabolism, immunity, and intestinal development of calves. Eighty-four Holstein male neonatal calves were assigned to one of the following four treatment groups: those that received bunk tank milk(BTM), untreated waste milk(UWM), pasteurized waste milk(PWM), and acidified waste milk(AWM) for 21 ***: Calves in the BTM and AWM groups consumed more starter(P < 0.05) than those in the UWM *** daily gain in the UWM group was the highest(P < 0.05). Calves exhibited the highest(P < 0.05) serum total protein, albumin, total cholesterol, high density lipoprotein, triglycerides, growth hormone, immunoglobulin(Ig) A and Ig M concentrations in the UWM group, highest malondialdehyde and tumor necrosis factor-α in the PWM group(P < 0.05), and highest glutathione peroxidase and Ig G in the BTM group(P < 0.05). The jejunum and ileum of the calves in all treatments presented a slight inflammatory response. The jejunal inflammation scores were higher(P < 0.05) in the UWM and AWM groups than the BTM group; the ileal inflammation scores increased more(P < 0.05) in the AWM group than the BTM group. Jejunal immunohistochemical scores(IHS) were higher(P < 0.05) in the PWM and AWM groups than the BTM group. Compared to the other three groups, calves feeding on BTM had lower(P < 0.05) ileal IHS. Jejunal interleukin(IL)-1β, IL-8, and IL-10 mRNA expression in the UWM group was the highest(P < 0.05). Calves fed AWM increased(P < 0.05) mRNA expression of IL-8 and toll like receptor 4(TLR-4) in the jejunum and IL-8, IL-1β, IL-6, IL-8, and IL-10 in the mesenteric lymph ***: Overall, bunk tank milk is the best choice for calf raising compared to waste milk. The efficiency of feeding pasteurized and acidified waste milk are comparable, and the acidification of waste milk is an acceptable labor-saving and diarrhea-preventing feed for youn

关键词： Acidified waste milk Calf Intestinal development Pasteurized waste milk Serum metabolism Waste milk

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Association of H-FABP gene polymorphisms with intramuscular fat content in Threeyellow chickens and Hetian-black chickens

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Journal of Animal Science and Biotechnology 2016年第3期7卷 366-373页

作者： Yong wang Xiaohong Hui Huie wang Tursunjan Kurban Chao Hang Ying Chen Jinming Xing jiufeng wang College of Veterinary Medicine China Agricultural University Key Laboratory of Tarim Animal Husbandry Science & Technology of Xinjiang Production and Construction Groups/College of Animal Science Tarim University

Background： To explore the relationship between the heart-type fatty acid binding protein（H-FABP） gene and intramuscular fat（IMF）, a polymorphism of the second exon of the H-FABP gene was investigated in 60 Three-yellow chickens（TYCs） and 60 Hetian-black chickens（HTBCs）.Results： The IMF contents of the cardiac, chest and leg muscles in HTBC were increased compared with TYC. Both TYC and HTBC populations exhibited Hardy-Weinberg Equilibrium（HWE） according to the χ2test. Three variations of the two birds were detected, namely, G939 A, G982 A and C1014 T. HTBCs with the TT genotypes exhibit increased IMF content in the chest muscles compared with the TC genotype. Thus, the G982 A site could be considered a genetic marker for selecting increased IMF content in the chest muscles of HTBC. The correlation coefficients revealed that H-FABP m RNA expression was negatively correlated with the IMF content in the cardiac, chest and leg muscles of HTBC and in the cardiac and chest muscles of TYC. The relative m RNA expression of H-FABP was reduced in the cardiac and leg muscles of HTBC compared with TYC, but this difference was not observed at the protein level, as assessed by Western blot ***： These findings offer essential data that can be useful in the breeding program of HTBC and future research exploring the role of H-FABP in IMF deposition and regulation in chickens.

关键词： Hetian-black Chicken H-FABP m RNA Expression Polymorphism Three-yel ow Chicken

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Prepartum body conditions affect insulin signaling pathways in postpartum adipose tissues in transition dairy cows

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Journal of Animal Science and Biotechnology 2019年第3期10卷 648-657页

作者： Fanjian Zhang Dan Li Qiong Wu Jian Sun Wenyi Guan Yinxu Hou Yaohong Zhu jiufeng wang Beijing Vocational College of Agriculture Beijing 102442People’s Republic of China College of Veterinary Medicine China Agricultural UniversityBeijing 100193People’s Republic of China Animal Science and Technology College Beijing University of AgricultureBeijing 102206People’s Republic of China

Background: Overconditioned dairy cows are susceptible to excessive lipolysis and increased insulin resistance during the transition *** associations among body fat reserve,insulin resistance,and lipolysis in adipose tissues(AT) remain to be ***,this study aimed to investigate whether excessive fat reserves influence the insulin signaling pathway in AT ***: Twenty multiparous dairy cows were selected and assigned to one of two groups,according to prepartum body condition score(BCS): Control group(BCS = 3.0–3.5;n = 10) and Overconditioned group(BCS ≥ 4.0;n = 10).Blood samples were collected on days-14,-7,-4,-2,-1,0,1,2,4,7,and 14 relative to *** AT were collected on day 2 following parturition for quantitative real-time polymerase chain reaction and western blot *** differences were observed between the two groups in serum glucose,non-esterified fatty acids,β-hydroxybutyric acid,tumor necrosis factor(TNF)α,insulin,or leptin concentrations during the experimental *** with the control cows,the overconditioned cows had lower serum triglyceride levels and higher adiponectin *** the AT postpartum,insulin receptor mRNA and protein levels were lower in the overconditioned cows than in the control cows,and no differences were found in glucose transporter 4 *** with the control cows,the overconditioned cows had lower mRNA levels of TNFα and higher mRNA levels of peroxisome proliferator-activated receptor gamma(PPARγ) in AT *** phosphorylated protein kinase B(AKT) content and phosphorylation rate of AKT were increased in the overconditioned cows compared with the control cows,which suggested that the downstream insulin signaling in AT was ***: In the present study,transition dairy cows with higher BCS did not show more fat *** changes of insulin signaling pathway in AT postpartum of overconditioned cows may be partly related to the ex

关键词： Adipose tissue Body condition score Insulin signaling pathway Transition dairy cow

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Antimicrobial Effect of Zophobas morio defensin against Bovine Mastitis Pathogens

Antimicrobial Effect of Zophobas morio defensin against Bovi...

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中国畜牧兽医学会兽医产科学分会第八届会员代表大会暨第十五次学术研讨会

作者： wang Xue LI Shuxian DU Mengze wang jiufeng ZHU Yaohong College of Veterinary Medicine China Agricultural University

Objective:Mastitis has always been considered as a great threat for dairy cow industry around the ***,mastitis treatment mainly relies on ***,many countries restrict the use of antibiotics since the bacterial resistance and drug residues have become a serious issue in the past decades of ***,it is urgent to develop a broad-spectrum,low residual anti-bacterial drug against bacterial resistance and drug ***:We analyzed the active ingredient of Zophobas morio(***) hemolymph and obtained the full-length complete coding sequence(CDs) of *** defensin through 5’ and 3’ RACE *** target gene was cloned into pGEX 6 p-1 and transformed into *** Rosetta(DE3) *** protein expression was induced with isopropyl-p-D-1-thiogalactopyranoside(IPTG),and purified through GSTrap 4 ***,the recombinant protein was treated with Prescission Protease and purified by Superdex 30 to remove glutathione S-transferase(GST)-***-zone assay was carried out to evaluate the antimicrobial activity of recombinant *** assay and cytotoxicity test were used to evaluate the safety of recombinant ***:Our results showed that the gene of Z. morio defensin encodes 89 amino acids,and the structure of *** defensin protein includes a conformationally flexible amino-terminal loop(n-loop) followed by a cysteinestabilized α-helical and β-sheet(CSαβ) domain,which is a structural scaffold shared by defensins of other *** addition,recombinant Z. morio defensin protein could inhibit multiple bacteria growths in vitro,such as Escherichia coli,Staphylococcus aureus,Klebsiella pneumonia,bacillus cereus,et ***,recombinant Z. morio defensin protein also maintains non-hemolytic and non-cytotoxicity as ***:In summary,recombinant Z. morio defensin protein which showed excellent antibacterial activity and safety,is a promising drug candidate to treat mastitis for dairy cows.

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A selected Lactobacillus rhamnosus strain promotes EGFR-independent Akt activation in an enterotoxigenic Escherichia coli K88-infected IPEC-J2 cell model

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畜牧与饲料科学 2015年第Z1期 27-42页

作者： ZHANG Wei ZHU Yaohong YANG Jincai YANG Guiyan ZHOU Dong wang jiufeng Department of Veterinary Clinical Sciences College of Veterinary Medicine China Agricultural University

Enterotoxigenic Escherichia coli（ETEC） are important intestinal pathogens that cause diarrhea in humans and *** probiotic bacteria may protect against ETEC-induced enteric infections,the underlying mechanisms are *** this study,porcine intestinal epithelial J2 cells（IPEC-J2） were pre-incubated with and without Lactobacillus rhamnosus ATCC 7469 and then exposed to F4+ *** in TLR4 and NOD2 mRNA expression were observed at 3 h after F4+ ETEC challenge,but these increases were attenuated by *** *** of TLR2 and NOD1 mRNA was upregulated in cells pre-treated with ***-treatment with *** counteracted F4+ ETEC-induced increases in TNF-α*** *** were observed in cells infected with F4+ ETEC and in cells treated with *** only.A decrease in phosphorylated epidermal growth factor receptor（EGFR） was observed at 3 h after F4+ ETEC challenge in cells treated with ***-treatment with *** enhanced Akt phosphorylation and increased ZO-1 and occludin protein *** findings suggest that *** protects intestinal epithelial cells from F4+ ETEC-induced damage,partly through the anti-inflammatory response involving synergism between TLR2 and *** addition,*** promotes EGFR-independent Akt activation,which may activate intestinal epithelial cells in response to bacterial infection,in turn increasing tight junction integrity and thus enhancing the barrier function and restricting pathogen ***-incubation with *** was superior to co-incubation in reducing the adhesion of F4+ ETEC to IPEC-J2 cells and subsequently attenuating F4+ ETEC-induced mucin layer destruction and suppressing *** data indicate that a selected *** strain interacts with porcine intestinal epithelial cells to maintain the epithelial barrier and promote intestinal epithelial cell activation in response to bacterial infection,thus

关键词： ETEC porcine mucin incubation destruction diarrhea incubated maintain tight infected

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Lactobacillus rhamnosus GR-1 attenuates foodborne Bacillus cereus-induced NLRP3 inflammasome activity in bovine mammary epithelial cells by protecting intercellular tight junctions

Lactobacillus rhamnosus GR-1 attenuates foodborne Bacillus c...

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中国畜牧兽医学会兽医产科学分会第十六次学术研讨会

作者： SHAN Qiang LIU Ning wang Xue ZHU Yaohong YIN Jinhua wang jiufeng

Objective:This study aimed to explore LGR-1 attenuation of ***-induced intercellular tight junction damage and NLRP3 inflammasome activity in bovine MAC-T ***:In this study,*** was isolated from mastitis milk *** bloting,cytosolic indirect immunofluorescence assay,RT-qPCR,scanning electron microscopy,and RNA interference were used to study the effect of LGR-1 on ***-induced bovine mammary epithelial intercellular tight junction damage and attenuation of NLRP3 inflammasome ***:In this study,*** 2101(BC) was isolated from milk samples of cows with *** grew rapidly with strong hemolysis,making it difficult to prevent mastitis and ensure food ***-T cells were treated with BC and/or Lactobacillus rhamnosus GR-1(LGR-1).Pretreatment with LGR-1 protected the integrity of tight junctions and the expression of ZO-1 and Occludin destroyed by ***,LGR-1 pretreatment reduced the expression of NOD-like receptor family member pyrin domain-containing protein 3(NLRP3),caspase recruitment and activation domain(ASC),Caspase-1 p20,gasdermin D(GSDMD) p30,inflammatory factors(IL-1β and IL-18),and cell death induced by ***,LGR-1 pretreatment reduced NLRP3 inflammasome activity and increased expressions of ZO-1 and occludin induced by lipopolysaccharides(LPS)+ATP ***-T cells were transfected with NLRP3 siRNA or MCC950 and/or treated with BC and/or ***3-siRNA transfection and MCC950 attenuated BC-induced NLRP3 inflammasome *** of inflammatory cytokines and cell death suggested that the inflammatory pathway might play an important role in the induction of the NLRP3 inflammasome by BC and the protection of ***:These results suggest that LGR-1 might be a probiotic alternative to antibiotics and could be administered to prevent mastitis in dairy cows,thus ensuring food security.

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Lactobacillus rhamnosus GR-1 prevents Escherichia coli-induced apoptosis through PINK1/Parkin-mediated mitophagy in MAC-T cells

Lactobacillus rhamnosus GR-1 prevents Escherichia coli-induc...

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中国畜牧兽医学会兽医产科学分会第八届会员代表大会暨第十五次学术研讨会

作者： Yanan Li Yaohong Zhu Bingxin Chu Ning Liu Shiyan Chen jiufeng wang College of Veterinary Medicine China Agricultural University

Objectives:We tested two major hypotheses in this ***,Lactobacillus rhamnosus GR-1(LGR-1),a probiotic,exerts protective efficacy during *** ***,LGR-1 provides protective effects via mitophagy activation,leading to improvement in mitochondrial morphology,reduced excessive ROS production and NLRP3 inflammasome activation,resulting in decreasing ***:MAC-T cells divided into 4 intervention groups:control group,*** infection group,pretreatment with LGR-1 group,LGR-1 *** addition,autophagy activators(Rapa) and inhibitors(3-MA),reactive oxygen species(ROS) scavenger(N-acetyl-L-cysteine,NAC) and inducer(H O),NLRP3 inhibitor(MCC950),or PINK1-siRNA was given to MAC-T ***:*** infection induced NOD-like receptor family member pyrin domain-containing protein 3(NLRP3) inflammasome assembly,Caspase-1 activation and *** also causes mitochondrial damage and subsequent increase in ROS ***,inhibition of ROS release by NAC abrogated the importance of ROS in NLRP3 assembly and apoptosis in MAC-T *** with LGR-1 alleviated ***-induced NLRP3 inflammasome activation and apoptosis via ROS ***,*** infection inhibited mitophagy while LGR-1 pretreatment augmented PINK1/Parkinmediated mitophagy activation,which further blocked ROS ***:LGR-1 pretreatment induced PINK1/Parkin-mediated mitophagy that eliminated damaged mitochondria and reduced ROS production and NLRP3 inflammasome activation,which subsequently decreased ***-induced apoptosis.

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A Novel β-Hairpin Peptide Z-d14CFR Enhances Multidrug-Resistant Bacterial Clearance in a Murine Model of Mastitis

A Novel β-Hairpin Peptide Z-d14CFR Enhances Multidrug-Resis...

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中国畜牧兽医学会兽医产科学分会第十六次学术研讨会

作者： wang Siyu wang Xue Li Shuxian Liu Ning Shan Qiang Zou Yunjing wang jiufeng Zhu Yaohong College of Veterinary Medicine Chinese Agricultural University

Objective:This study aimed to investigate the antimicrobial mechanisms of a novel β-hairpin peptide,Z-d14 CFR,derived from Zophobas atratus defensin,and evaluate the therapeutic effect on multidrug-resistant Escherichia coli infection in a murine model of ***:Broth microdilution method was used to measure the antimicrobial activity of Z-d14 CFR and hemolysis and cytotoxicity analysis were applied to evaluate its safety;Through observing bacterial ultrastructure by scanning electron microscopy and measuring the changes of N-phenyl-1-naphthylamine(NPN),3,3'-dipropylthiadicarbocyanine iodide(DiSC3(5)),SYTO 9/PI and β-galactosidase in bacteria,we further investigated the effect of Z-d14 CFR on bacterial cell membrane structure and permeability;The effect of Z-d14 CFR on bacterial biofilm formation was determined by crystalline violet ***:In this study,we designed a novel β-hairpin peptide,Z-d14 CFR,derived from Zophobas atratus *** vitro studies illustrated that Z-d14 CFR showed broad-spectrum antimicrobial activity with disrupting bacterial membrane,promoting cellular contents release and inhibiting biofilm formation significantly,but had low hemolysis and cytotoxicity.Z-d14 CFR also enhanced bacterial clearance,decreased neutrophils infiltration and down-regulated TNF-α and IL-1βexpressions in a murine model of ***:The study indicated that Z-d14 CFR performed antimicrobial effect via inhibiting biofilm formation,disrupting bacterial membrane integrity and promoting cellular contents release,and showed a great therapeutic effect on multidrug-resistant Escherichia coli infection in *** present findings suggested that Z-d14 CFR might be a promising candidate against multidrugresistant bacteria.

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Dose-dependent effects of Lactobacillus rhamnosus on serum interleukin-17 production and intestinal T-cell responses in pigs challenged with Escherichia coli

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畜牧与饲料科学 2015年第Z1期 43-58页

作者： ZHU Yaohong LI Xiaoqiong ZHANG Wei ZHOU Dong LIU Haoyu wang jiufeng College of Veterinary Medicine China Agricultural University Department of Animal Nutrition and Management Faculty of Veterinary Medicine and Animal Science Swedish University of Agricultural Sciences

The mechanism underlying the dose effect of probiotics on ameliorating diarrhea has not been fully ***,low（1×109 CFU/mL） or high(1×1011 CFU/mL) doses of Lactobacillus rhamnosus ATCC 7469 were administered orally to piglets for 1 week before F4（K88）-positive enterotoxigenic Escherichia coli（F4+ ETEC）*** of a low,but not a high,dose of *** decreased the percentage of CD3+ CD4+CD8- T cells in the peripheral ***,transiently increased serum concentrations of interleukin-17A（IL—17A） were observed after F4+ ETEC challenge in pigs pretreated with a high dose of *** of *** increased the percentage of the small intestinal lamina propria CD3+ CD4+ CD8- cells and Peyer’s patch CD3+ CD4- CD8- and CD3- CD4- CD8+ *** percentage of ileal intraepithelial CD3+ CD4- CD8+ cells increased only in the high-dose *** of *** downregulated expression of ileal IL-17 A after F4+ ETEC challenge but had no effect on expression of gamma interferon（IFN-7）,IL-12,IL-4,and FOXP3 mRNA in the small *** of jejunal IL-2,ileal transforming growth factor β1（TGF-β1）,and ileal IL-10 was upregulated in the low-dose piglets after F4+ ETEC *** findings suggest that amelioration of infectious diarrhea in piglets by *** is associated with the generation of lamina propria CD3+ CD4+ CD8T cells,the expansion of Peyer’s patch CD3+ CD4- CD8- and CD3- CD4- CD8+ cells,and the attenuation of F4+ETEC-induced increase in CD3+ CD4+ CD8+ T cells in the small ***,consumption of high doses of *** may increase levels of serum IL-17 A after F4+ ETEC challenge,thus eliciting a strong proinflammatory response.

关键词： ETEC interleukin ileal diarrhea intestine patch challenge lamina probiotics pretreatment

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