苏太仔猪FUT1基因M307位点多态性与F18大肠杆菌抗病相关性的体外鉴定

采用PCR-RFLP方法检测了江苏苏太断奶仔猪FUT1基因M307位点等位基因多态性分布,在所检的49头仔猪中,GG基因型个体16头,AG基因型19头,AA基因型14头。在此基础上,制备上述不同基因型个体仔猪小肠上皮细胞,分别与表达F18ab菌毛的野生型大肠杆菌、表达F18ac菌毛含fed操纵子全基因的重组大肠杆菌和V型系统表面分泌表达F18abFedF亚单位的重组大肠杆菌进行体外黏附试验和黏附抑制试验。研究结果表明:FUT1基因M307位点中GG型和AG型仔猪小肠上皮细胞均能黏附上述3种大肠杆菌,而AA型个体小肠上皮细胞则不能黏附。将上述3种大肠杆菌分别与抗F18ab菌毛高免血清、F18ac菌毛高免血清及抗F18abFedF亚单位单因子血清作用后,则失去黏附仔猪肠上皮细胞能力。上述结果对苏太猪从体外试验上证明了FUT1基因M307位点多态性与断奶仔猪腹泻和水肿病存在着直接的相关性。     

Effects of PPARγ agonist on calpain expression in model of acute experimental autoimmune encephalomyelitis

AIM: To explore the effects of peroxisome proliferator-activated receptor γ （PPARγ） agonist on calcium-activated neutral proteinase, calpain, expression in the brain of rats with acute experimental autoimmune encephalomyelitis (EAE). METHODS: EAE model was established in rats by inoculating the homogenate contained spinal cord of guinea pig and complete Freunds adjuvant. Respectively, the PPARγ agonist rosiglitazone and non-steroid anti-inflammatory drug (NSAID) ibuprofen were used to treat the EAE rats. Outcome measures (Kohs scale) were applied at baseline and after treatment to assess the improvement of clinical symptoms. Calpain expression levels were detected by RT-PCR and Western blotting. RESULTS: All the groups showed significant improvements of scales scores after treatment with rosiglitazone and ibuprofen. No significant difference of the expression of calpain mRNA was found among EAE group, rosiglitazone and ibuprofen groups (P>0.05), but the expression of calpain reduced markedly in rosiglitazone and ibuprofen groups compared with that in EAE group (P<0.05). CONCLUSION: Rosiglitazone and ibuprofen inhibit the expression of calpain and improve the clinical symptoms of EAE rats. PPARγ agonist plays a neuroprotective role in EAE rats.     

荷斯坦乳牛乳腺和乳腺上皮细胞中Toll样受体及辅助因子编码基因的检测

参照牛TLR4、TLR2、CD14、MD-2基因序列设计了相应基因的引物。采用RT-PCR技术检测了体外培养的荷斯坦乳牛乳腺和乳腺上皮细胞中Toll样受体TLR4、TLR2及辅助因子CD14、MD-2基因。结果显示,乳腺上皮细胞中存在TLR4、TLR2、CD14和MD-2四个基因的表达,而乳腺中除MD-2未检测到外,其余3个基因均扩增成功。说明该受体及辅助因子可能参与了乳腺的先天性免疫防御。该研究为探讨乳腺的先天性免疫及乳腺上皮细胞在乳腺先天性免疫中的作用奠定了基础。     

烟酸对高脂血症兔瘦素、过氧化体增殖物激活型受体γ和CD36表达的影响

目的探讨烟酸对高脂血症兔血清瘦素及皮下脂肪组织瘦素、过氧化体增殖物激活型受体γ及CD36 mRNA表达的影响。方法12只健康雄性新西兰兔给予高胆固醇饮食饲养8周后,随机分为高脂组和烟酸组:高脂组继续饲以高胆固醇饲料6周;烟酸组在饲以高胆固醇饲料的基础上给予烟酸0.2g/(kg.d),共6周。另选择普通饮食14周兔(n=6)作为对照组。实验结束后,取腹股沟处皮下脂肪组织称重并冻存,用酶联免疫吸附法测定血清及脂肪细胞培养基瘦素水平;半定量逆转录聚合酶链反应测定脂肪组织瘦素、过氧化体增殖物激活型受体γ及CD36mRNA的表达。此外,在体外观察不同浓度的烟酸对高脂兔脂肪细胞瘦素、过氧化体增殖物激活型受体γ及CD36 mRNA的表达。结果高脂组兔血清及脂肪组织瘦素水平明显高于正常对照组,烟酸治疗6周后可降低血清及脂肪组织瘦素水平。逆转录聚合酶链反应表明烟酸组较高脂组瘦素mRNA表达降低,瘦素mRNA与过氧化体增殖物激活型受体γmRNA和CD36 mRNA的表达呈负相关。体外实验亦表明,烟酸呈剂量依赖性地降低脂肪细胞瘦素mRNA表达,并剂量依赖性地上调过氧化体增殖物激活型受体γ及CD36 mRNA的表达。结论烟酸治疗能降低高脂血症兔血清及脂肪分泌瘦素水平,上调过氧化体增殖物激活型受体γ及CD36 mRNA的表达。     

鸵鸟红细胞免疫功能研究

应用红细胞Ｃ３ｂ受体花环（Ｃ３ｂＲＲ）和免疫复合物花环（ＩＣＲ）试验，证实鸵鸟红细胞表面存在补体Ｃ３ｂ受体（Ｃ３ｂｒｅｃｅｐｔｏｒ，Ｃ３ｂＲ），表明红细胞免疫系统（Ｒｃｄｃｅｌｉｍｍｕｎｅｓｙｓｔｅｍ，ＲＣＩＳ）的概念亦适用于这种动物；鸵鸟红细胞不但有传统医学观念上的功能，而且还可借助ＣＲ１（ＣｏｍｐｌｅｍｅｎｔｒｅｃｃｐｔｏｒｔｙｐｅＩ）进行免疫粘附，并以此为基础，表现出各种重要的免疫功能。     

Exendin-4的研究进展

在介绍Exendin-4的基础上,综述了目前最新的Exenatide用于治疗糖尿病的研究进展,并基于大量临床前数据,阐释了其功能、作用机制和毒理学研究。     

植物中γ-亚麻酸合成关键酶——△^6-脂肪酸脱氢酶

γ-亚麻酸作为人体必需的不饱和脂肪酸，对人体的激素调节及脂肪酸代谢发挥着重要的生理作用。△^6-脂肪酸脱氢酶是多不饱和脂肪酸合成途径中的限速酶。本文介绍了不饱和脂肪酸γ-亚麻酸代谢途径中的关键酶△^6-脂肪酸脱氢酶的结构功能与目前△^6-脂肪酸脱氢酶的基因工程研究进展，并对其应用进行了展望，以期为利用基因工程手段生产γ-亚麻酸提供参考。     

Adenovirus-mediated mPPARγ1 gene overexpression inhibits IFN-γ-induced galectin-9 gene and protein expression in ECV304

AIM:To investigate whether adenovirus-mediated mPPARγ1 gene overexpression inhibits IFN-γ-induced galectin-9 gene and protein expression in ECV304. METHODS:A replication-deficient recombinant adenovirus expression vector of mPPARγ1 was constructed by using the AdEasy system. ECV304 were incubated for 24 h with 1×104 U/L, 5×104 U/L, 1×105 U/L and 2×105 U/L IFN-γ, respectively. ECV304 stimulated with 1×105 U/L IFN-γ were divided into 4 groups in random: P group (PPARγ1 gene overexpression), T group (treated with troglitazone 40 μmol/L in DMSO), PT group (PPARγ1 gene overexpression+troglitazone treatment) and control group. Changes of PPARγ and galectin-9 in mRNA and protein levels in different groups and subgroups were investigated by RT-PCR and immunoblotting. RESULTS: Galectin-9 expression was very few in normal ECV304. IFN-γ induced the expression of galectin-9 in ECV304. Degree of galectin-9 expression increased with the dose of IFN-γ. PPARγ1 gene overexpression inhibited IFN-γ-induced galectin-9 expression in ECV304. Galectin-9 mRNA and protein expressions from PT group and P group were inhibited in similar degree (P>0.05). However, this effect was not observed in troglitazone intervention (P>0.05). PPARγ expression was also very few in normal ECV304. PPARγ1 gene overexpression/activation had no effect on endogenous mPPARγ expression. CONCLUSION: This may partly contributed to the anti-inflammatory and immuno-regulatory effect of PPARγ1 gene overexpression by inhibiting IFN-γ-induced galectin-9 gene and protein expression in ECV304.     

大黄鱼C型凝集素受体Clec4e的分子鉴定及凝集特性

C型凝集素受体（C-type lectin receptor，CTLR）是可以特异性结合糖类病原体相关分子模式（pathogen-associated molecular patterns，PAMPs）的模式识别受体（pattern recognition receptors，PPRs），在先天性免疫中发挥着重要作用。为了揭示硬骨鱼CTLR的生物学功能，本研究以从大黄鱼（Larimichthys crocea）转录组数据库中筛选出的一个CTLR基因—C型凝集素结构域家族4成员E基因（C-type lectin domain family 4 member E gene，Clec4e）为研究对象，研究其分子特征、表达分布和凝集特性。结果显示，LcClec4e cDNA全长1546 bp，开放阅读框（open reading frame，ORF）771 bp，编码254个氨基酸。LcClec4e的N端有一个跨膜区，无信号肽，C端含有一个糖识别结构域（carbohydrate recognition domain，CRD），其中含有糖结合位点EPN和WFD以及6个可形成二硫键的保守半胱氨酸。系统发育分析表明，LcClec4e与多种鲈形目鱼类Clec4e具有较近的亲缘关系。荧光定量PCR结果显示，LcClec4e在所检测的10种组织中呈组成型分布，且在肝脏中表达量最高；LcClec4e在来源于大黄鱼头肾组织的原代巨噬细胞、淋巴细胞和粒细胞中均有表达，且在巨噬细胞中表达量最高；经灭活溶藻弧菌刺激后，LcClec4e在3种免疫细胞中的表达均极显著上调。原核表达的重组LcClec4e胞外段（recombinant LcClec4e-extracellular domain，rLcClec4e-ex）具有Ca2+依赖性的凝集活性，可凝集小鼠、家兔的红细胞，以及嗜水气单胞菌、变形假单胞菌、溶藻弧菌和坎氏弧菌等4种水产常见的革兰氏阴性菌。D-葡萄糖、D-果糖、D-甘露糖、D-麦芽糖、α-乳糖和脂多糖均可抑制rLcClec4e-ex对大黄鱼重要病原菌变形假单胞菌的凝集作用，说明LcClec4e可能与变形假单胞菌表面的糖类物质结合。这些研究结果提示，LcClec4e可能作为一种PPR，通过结合病原菌表面的糖类PAMPs来识别病原，参与大黄鱼抗细菌感染的免疫防御。