Seaweed polysaccharide mitigates intestinal barrier dysfunction induced by enterotoxigenic Escherichia coli through NF-κB pathway suppression in porcine intestinal epithelial cells

This study aimed to investigate the protective effects and underlying mechanism of seaweed polysaccharide (SWP) on intestinal epithelial barrier dysfunction induced by E. coli in an IPEC-J2 model. A preliminary study was done to screen optimum SWP concentrations by cell viability, cytotoxicity, apoptosis and proliferation evaluation. The regular study was conducted to evaluate the protective effects of SWP against E. coli challenge via the analysis of transepithelial electrical resistance (TEER), tight junction proteins, NF-κB signalling pathway, proinflammatory cytokines and the E. coli adhesion and invasion. Our results show that 4 h E. coli challenge down-regulated tight junction proteins expression, decreased TEER, activated NF-κB signalling pathway and increased proinflammatory response, which indicates that the E. coli infection model was well-established. Pre-treatment with 240 μg/ml SWP for 24 h alleviated the 4 h E. coli -induced intestinal epithelial barrier dysfunction, as evidenced by the up-regulated expression of Occludin, Claudin-1 and ZO-1 at both mRNA and protein level and the increased TEER of IPEC-J2 cells. Pre-incubation with 240 μg/ml SWP for 24 h inhibited the activation of the NF-κB signalling pathway by 4 h E. coli challenge, including the decreased mRNA expression of TLR-4, MyD88, IκBα, p-65, as well as the reduced ratio of protein expression of p-p65/p65. Also, pre-treatment with 240 μg/ml SWP for 24 h decreased proinflammatory response (IL-6 and TNF-α) induced by 4 h E. coli challenge and decreased the E. coli adhesion and invasion. In conclusion, SWP mitigated intestinal barrier dysfunction caused by E. coli through NF-κB pathway in IPEC-J2 cells and 240 μg/ml SWP exhibited better effect. Our results also provide a fundamental basis for SWP in reducing post-weaning diarrhoea of weaned piglets, especially under E. coli -infected or in-feed antibiotic-free conditions.     

Residual effects of soil Zn fertilization on soil characteristics,yield and quality of Platycodon grandiflorum

ABSTRACT

Zinc (Zn) deficiency is common worldwide and is a major limiting factor in the cultivation of Platycodon grandiflorum because most P. grandiflorum is cultivated on soils with slight or severe levels of Zn deficiency in China. To investigate the effects of soil Zn fertilization on P. grandiflorum and its residual effects on soil characteristics, a randomized complete block design was conducted at the experimental station of Shandong Agricultural University China in 2016–2018. In 2016, ZnSO₄ · 7H₂O was used as Zn fertilizer. The fertilizer levels of Zn fertilizer were designed as 0, 4, 6, 8 and 10 kg ha^?1 and were denoted as CK, Zn₄, Zn₆, Zn₈ and Zn₁₀. No Zn fertilizer was used in 2017 and 2018. The results showed that soil Zn fertilization increased the quantity of soil bacteria, and actinobacteria, Zn₈ and Zn₁₀ treatments increased the soil fungal quantity in 2017, whereas the effect was observed only in the Z₁₀ treatment in 2018. Soil Zn fertilization enhanced the activity of polyphenol oxidase, decreased that of phosphatase, and did not influence the invertase activity in both 2017 and 2018. Both the yield and the quality of P. grandiflorum in the succeeding three years increased due to the soil Zn fertilization in 2016. Overall, soil Zn application is a suitable approach to the cultivation of P. grandiflorum, and the residual effects of a one-time soil Zn fertilization could last at least two years.     

Effects of calpain-2 and autophagy-related protein 5 on hepatocyte apoptosis induced by endoplasmic reticulum stress

AIMTo investigate the effects of calpain-2 and autophagy-related protein 5 (Atg5) on apoptosis of BRL-3A rat normal liver cells during endoplasmic reticulum stress (ERS) induced by dithiothreitol (DTT). METH?ODS: BRL-3A cells were treated with DTT at 2.0 mmol/L for 0, 6, 12 and 24 h to induce ERS. Real-time cell analysis (RTCA) was used to measure the effect of DTT on BRL-3A cell proliferation. Apoptosis and cell cycle distribution were analyzed by flow cytometry. The mRNA expression of calpain-2 and Atg5 was detected by real-time PCR. The protein levels of calpain-2, Atg5, Atg7, Atg12 and microtubule-associated protein 1 light chain 3 (LC3) were determined by Western blot. The interaction between calpain-2 and Atg5 was investigated by co-immunoprecipitation (Co-IP). RESULTSThe proliferation of BRL-3A cells treated with DTT was significantly inhibited. The apoptosis of BRL-3A cells was significantly increased after DTT treatment for 6, 12 and 24 h as compared with 0 h group (P<0.05). The cell cycle was arrested in G₁ phase after DTT treatment (P<0.05). After DTT treatment for 6, 12 and 24 h, the mRNA expression of calpain-2 and Atg5 in the BRL-3A cells was significantly increased as compared with 0 h group (P<0.05). The protein levels of calpain-2, Atg12 and Atg7 in the cells treated with DTT for 6, 12 and 24 h were significantly higher than those in 0 h group, and the ratio of LC3-II/LC3-I was also significantly higher than that in 0 h group, while Atg5 expression was significantly lower than that in 0 h group (P<0.05). The results of Co-IP found that the anti-calpain-2 antibody precipitated Atg5 protein from the cell lysates, and the anti-Atg5 antibody also precipitated calpain-2 from the cell lysates, which confirmed the interaction between calpain-2 and Atg5. CONCLUSION Calpain-2 may participate in ERS-induced hepatocyte apoptosis by interacting with Atg5.     

Autophagy induces mitochondrial dysfunction in neurons from neonatal rats with hypoxic ischemic encephalopathy

AIM: To explore the influence of autophagy on the induction of mitochondrial dysfunction in the neurons in a neonatal rat hypoxic ischemic encephalopathy (HIE) model. METHODS: Ten-day-old rat pups (n=30) were randomly divided into sham group and model group. The rats in the latter group were subject to hypoxia-ischemia treatment via unilateral common carotid artery ligation. The rats were sacrificed for brain pathological examination, and the protein levels of cleaved caspase-3 and LC3B-II were detected by immunohistochemical analysis. For the in vitro experiments, the autophagy of primarily cultured rat neurons was observed after hypoxia, and Western blot and mitochondrial function testing were also performed. RESULTS: Compare with sham group, the hypoxia-ischemia treatment caused atrophy and apoptosis of neurons, and ventricular area enlargement of rat brains. Immunohistochemical results demonstrated significantly higher levels of apoptosis- and autophagy-associated proteins, such as cleaved caspase-3 and LC3B-II (P<0.01). In vitro experiments demonstrated that hypoxia induced autophagy and apoptosis in the neurons. Compared with sham group, there were higher levels of reactive oxygen species and mitochondrial superoxide, and lower mitochondrial membrane potential in the model group (P<0.01). CONCLUSION: In neonatal HIE rat model, the hypoxia-induced mitochondrial dysfunction is related to apoptosis and autophagy. It will provide a new idea for administration of autopahgy inducer agents in treatment of HIE.     

虎雪兰玻璃化超低温法脱毒技术的研究

以兰属花卉虎雪兰组培原球茎为试材,采用玻璃化超低温法对兰花病毒脱除进行了研究,以期为虎雪兰玻璃化超低温法脱毒体系的建立提供参考依据。结果表明:在蔗糖浓度0.5 mol·L-1预培养4 d,然后在蔗糖0.6 mol·L-1加载液冰上处理50 min,之后转入PVS2溶液冰上玻璃化处理120 min,再液氮冷冻40 min,37℃水浴解冻3 min,最后卸载液(1/2MS+1.2 mol·L-1蔗糖)卸载20 min,待恢复培养后,原球茎成活率可达到65%以上,随机检测不同处理样品的脱毒率可达97%。     

基于系统图法的低碳城市建设对策研究

指出了低碳城市建设是发展低碳经济的重要载体,是建设生态文明的必由之路,中国建设低碳城市是应对全球气候变化,适应全球经济发展趋势,转变城市发展方式,树立积极的国际形象的必然选择,而系统地探寻适宜中国低碳城市建设对策是当前的重中之重。系统图法有助于掌握低碳城市建设问题的全貌,明确问题的重点,系统地找到低碳城市建设的对策。基于系统图法原理分析了低碳城市建设对策问题,确定了4个二级目标,8个三级目标,16个最终对策。     

低聚木糖和低聚壳聚糖及其复合制剂对肉鸡生产性能、免疫机能和盲肠菌群的影响

本试验旨在研究低聚木糖(XOS)和低聚壳聚糖(COS)及其复合制剂对肉鸡生产性能、免疫机能和盲肠菌群的影响。将240只1日龄爱拔益加(AA)肉鸡随机分为5组,每组6个重复,每重复8只。对照组(CON)饲喂基础日粮,其他处理组分别在基础日粮中添加100 mg/kg XOS(XOS),30 mg/kg COS(COS),50 mg/kg XOS+10 mg/kg COS(XCⅠ),75 mg/kg XOS+15 mg/kg COS(XCⅡ)。结果表明:与对照组相比,XOS、XCⅠ、XCⅡ均能显著提高肉鸡21 d、42 d平均体重(BW)和1²1 d平均日增重(ADG)(P<0.05);XCⅠ、XCⅡ较COS 42 d体重和121 d平均日增重(ADG)(P<0.05);XCⅠ、XCⅡ较COS 42 d体重和1⁴2 d的ADG显著提高(P<0.05)。与对照组相比,XCⅠ、XCⅡ14 d、28 d肉鸡胸腺指数及14 d肉鸡十二指肠和空肠黏膜IgG含量;且XCⅠ14 d肉鸡盲肠乳酸杆菌数量较CON和XOS也有提高。结果提示,XOS和COS能改善肉鸡生产性能,改善肉鸡微生物区系,提高机体免疫功能,且XOS与COS低剂量复合添加效果优于单一寡糖,两者具有协同效应,本试验条件下以50 mg/kg XOS+10 mg/kg COS复合效果为最佳。     

“一带一路”背景下云南民族竹文化的传承与传播

云南民族竹文化在维系民族的生存和发展、保护生态环境、引领精神价值方面具有重要作用。在“一带一路”倡议下，民族竹文化的传播不但能加强文明间的对话、谋求更多共识，更能维护文化多样性、提升中国话语权。文章分析了云南民族竹文化的内容体系、新时代下传播与传承中存在的问题，从加大政策支持、加快人才培养、培育文化产业等方面提出了云南民族竹文化的建设路径。     

基于侯选基因标记的四倍体马铃薯休眠QTL关联分析

休眠期是马铃薯(SolanumtuberosumL.)重要的块茎性状之一,寻找调控马铃薯块茎休眠的关键基因,揭示其分子机制以选育具有适宜休眠期长度的马铃薯品种,对于解决当前马铃薯产业中过长或过短休眠期带来的经济损失和食品安全隐患等问题十分关键。前期研究在二倍体马铃薯连锁群体中定位了6个加性休眠QTL,本研究拟在四倍体马铃薯育种材料中验证这些休眠QTL。基于休眠QTL连锁的候选基因标记,采用混合线性模型(MLM),模型中考虑群体结构和亲缘关系(Q+K),在四倍体马铃薯自然群体St-hzau中对马铃薯块茎休眠期进行了关联分析。5号染色体上休眠QTL DorB5.3连锁的候选基因标记S199_300和GWD (根据葡聚糖水双激酶α-glucan water dikinase基因设计)与马铃薯块茎休眠期具有显著的关联(P0.05),分别解释了休眠期表型变异的7.8%和3.2%,分别能增加休眠期7.1 d和4.5 d,即在二倍体马铃薯连锁群体中定位的稳定主效休眠QTL DorB5.3在四倍体马铃薯关联群体St-hzau中也表现显著, DorB5.3的稳定性在关联分析结果中得到了验证,表明候选基因标记策略在马铃薯块茎休眠QTL关联分析中是一种有效的策略。本研究所验证的主效休眠QTL DorB5.3及相应连锁标记可以直接用于马铃薯休眠育种。据此可以推测GWD可能在控制还原糖含量和块茎休眠2个方面均发挥作用,马铃薯块茎休眠机制与还原糖含量变化机制可能存在着部分交叉。