牛羊胎盘免疫调节因子的提取及对PANC-Ⅰ增殖和迁移影响的比较

胎盘免疫调节因子(placenta immunoregulating factor,PF)是从胎盘提取的小分子多肽,对病毒性疾病、免疫缺陷疾病及恶性肿瘤等具有潜在的临床应用价值.为检测牛(Bos taurus)和绵羊(Ovis aries)胎盘免疫调节因对人(Homo sapiens)胰腺癌细胞(pancreatic cancer cells-Ⅰ,PANC-Ⅰ)增殖与迁移的影响,本研究对从屠宰场获得的牛和绵羊胎盘进行分离,利用双酶水解法制备PF,将质量浓度为100～2 000 ng/mL的牛、羊PF作用于体外培养的PANC-Ⅰ细胞,以3-(4,5-二甲基噻唑-2-基)-5-(3-羧甲酯基)-2-(4-磺苯基)-2H-四唑(3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium,MTS)法观察细胞增殖抑制率,应用流式细胞技术检测其对PANC-Ⅰ细胞周期的影响.并利用Transwell迁移实验检测添加PF对PANC-Ⅰ细胞迁移能力的影响,同时通过实时荧光定量PCR法检测肿瘤抑制基因(tumor suppressor gene,PS3)和细胞周期依赖性蛋白激酶抑制因子1A基因(cyclin-dependent kinaseinhibitor 1A,P21)的表达情况.结果显示,17.11 g绵羊胎盘制得0.588 mg PF,6.61 g牛胎盘制得0.312 mgPF,胎盘质量与所得PF的质量比约为20 000∶1.在体外培养的PANC-Ⅰ细胞培养液中添加PF,从形态上观察,牛羊PF对PANC-Ⅰ细胞没有明显的影响.绵羊PF处理后PANC-Ⅰ细胞增殖抑制率为9.75％～22.89％,牛PF处理后PANC-Ⅰ细胞增殖抑制率为-3.81％～23.56％,增殖抑制率最显著的绵羊PF浓度为500 ng/mL,牛PF浓度为1 000 ng/mL.绵羊PF在浓度为500 ng/mL时,PANC-Ⅰ细胞的PI和SPF最低,而牛PF各浓度下的胰腺癌细胞增殖指数(proliferation index,PI)和增殖活性(S-phase fraction,SPF)没有明显的变化.同时,牛羊PF对PANC-Ⅰ细胞的迁移能力无显著影响.牛羊PF使PANC-Ⅰ细胞P53表达上调而P21表达下调.上述结果表明,在一定浓度下,PF对体外培养的PANC-Ⅰ细胞增殖有负调控作用,且牛羊胎盘免疫调节因子对胰腺癌细胞的影响不同,可能因为其存在的活性成分有区别.本研究为胰腺癌的治疗提供了新的思路.     

TDZ对杏香兔耳风组培苗复壮的影响

为了了解TDZ对组培苗增殖的影响情况,探讨了TDZ在衰退组培苗复壮过程中的作用,以增殖衰退的杏香兔耳风组培苗为试验材料,就TDZ的不同添加方式(抽滤灭菌和高压灭菌)和不同浓度(0.0、0.2、0.4、0.6、0.8、1.0 mg/L)处理对组培苗增殖系数、有效芽及苗高生长的影响情况进行了试验。结果表明:当TDZ的浓度0.2 mg/L时,TDZ的添加方式对组培苗的增殖系数和有效芽数均有较大影响,抽滤灭菌后的增殖系数和有效芽数要明显高于高压灭菌的;不同浓度的TDZ对有效芽数和髙生长的影响均有极显著的差异性(P0.01),当TDZ浓度为0.8 mg/L时,苗质量总体达到最优,增殖系数为5.11,达到了预期的复壮效果;最优生根培养基配方为1/2MS+NAA 0.02 mg/L+IBA 0.1 mg/L。在杏香兔耳风组培苗复壮过程中,TDZ可作为一种有效的植物生长调节剂使用。     

橡胶树花药愈伤组织继代培养与胚状体发生能力研究

利用橡胶树花药愈伤组织进行继代培养与胚状体诱导,结果表明,在诱导愈伤组织、继代培养和分化胚状体中MS培养基稍优于改良MS培养基;以继代1代的愈伤组织胚状体的诱导率最高,分别达到70.8%和67.5%;在继代0代、2代和3代之间差异达到极显著,在继代4代时差异达到显著,其他继代次数之间差异不显著。在继代8代后,两种培养基的诱导率逐渐升高,分别达到44.6%和35.3%。     

香果树带芽茎段不定芽高频增殖的优化

以香果树(Emmenopterys henryi Oliv.)带芽茎段为外植体,以MS为基本培养基,采用正交试验和单因子试验方法研究了不同因素对带芽茎段不定芽增殖的影响。结果表明:香果树带芽茎段不定芽增殖的最佳培养基为MS+KT2.0mg/L+6-BA2.0mg/L+NAA0.1mg/L,最佳蔗糖和琼脂浓度分别为40和7.5g/L,最佳pH值6、培养温度25℃,培养条件为光培养。证明利用组织培养技术能够实现香果树带芽茎段不定芽的高频增殖,为珍稀濒危植物香果树的快速繁殖提供了一条简单.有效的途径。     

MSTN基因对牛肌动蛋白细胞骨架调节通路的影响

为探究肌肉生长抑制素（myostatin,MSTN）对牛骨骼肌生长发育的作用机制,本研究前期利用定量蛋白质组学与磷酸化蛋白质组学分析野生型蒙古牛（MG.WT）和MSTN^+/-蒙古牛（MG.MSTN^+/-）腿臀肌肌肉组织中蛋白质水平和磷酸化修饰水平的差异变化,使用已建立的牛骨骼肌卫星细胞体外诱导成肌分化模型,检测设计合成的MSTN siRNA （si-MSTN）干扰效果;采用实时荧光定量PCR和Western blotting方法检测转染si-MSTN的增殖期（GM）和分化第3天（DM3）牛骨骼肌卫星细胞中肌动蛋白细胞骨架调节通路相关基因的mRNA和蛋白水平的表达变化,研究敲低MSTN表达对肌动蛋白细胞骨架调节通路的影响。结果显示,在MSTN^+/-蒙古牛肌肉组织中共鉴定到16个肌动蛋白细胞骨架调节通路相关基因表达丰度上调;转染si-MSTN细胞中的MSTN表达水平极显著降低（P<0.01）;在转染si-MSTN的GM期牛骨骼肌卫星细胞中,肌动蛋白细胞骨架调节通路相关基因ENAH、ACTN4和Cdc42的mRNA水平均显著升高（P<0.05）,PFN1、RhoA和ACTN4的蛋白水平均显著或极显著升高（P<0.05;P<0.01）;在转染si-MSTN的DM3牛骨骼肌卫星细胞中,ENAH、CFL1、SCIN和Cdc42基因mRNA水平均显著升高（P<0.05）,RhoA基因mRNA水平极显著升高（P<0.01）,PFN1和ACTN4的蛋白水平均显著升高（P<0.05）。结果表明,干扰MSTN可以促进肌动蛋白细胞骨架调节通路相关基因的表达,探明了MSTN可能通过介导肌动蛋白细胞骨架调节通路影响牛骨骼肌卫星细胞增殖和成肌分化的分子机制,为进一步研究MSTN对牛成肌分化的调控机制提供参考。     

猴头菌子实体提取物对人肺癌细胞SPCA-1的抑制作用

猴头菌(Hericium erinaceus)子实体的醇提物用不同极性有机溶剂分部提取,得到石油醚、氯仿、乙酸乙酯和正丁醇分部,其中仅石油醚分部能抑制SPCA-1细胞增殖和促进其释放ROS(reactive oxygen species),对石油醚分部的进一步研究发现石油醚分部可诱导SPCA-1细胞早期凋亡和降低G_0/G_1期细胞数量。     

Direct somatic embryogenesis in Epipactis veratrifolia,a temperate terrestrial orchid

Most temperate terrestrial orchids are endangered species. Attempts to produce plantlets from plants of the genus Epipactis by asexual methods have totally failed. This study was conducted using somatic embryogenesis as a rapid vegetative propagation technique for conservation of E. veratrifolia. For these purposes, effects of different types of plant growth regulators (PGRs), different types of explants, light and dark conditions, and the effect of gibberellic acid (GA₃) and Paclobutrazol (PBZ) were investigated on somatic embryogenesis induction. Abscisic acid (ABA) pre-treatment effectiveness on inducting somatic embryogenesis and increasing the number of embryos per explant were investigated. Subsequently, 16 media were tested to find the best medium for plant regeneration and shoot and root proliferation. BA (3 mg L^?1) resulted in a better response than the other PGRs by supporting the development of 17 embryos per protocorm. PBZ, which resulted in 11 embryos per explant, was better than GA₃. FAST medium supplemented with organic substances was recognized as the best medium for plant regeneration and shoot and root proliferation. ABA pre-treatment had positive effect on somatic embryogenesis initiation. This study, for the first time, succeeded in finding a rapid and suitable protocol for propagation and genetic resource conservation of E. veratrifolia.     

Effects of STK15 overexpression on growth of human esophageal carcinoma cell line KYSE150

AIM:To investigate the effects of serine/threonine kinase 15 (STK15) overexpression on the growth of human esophageal squamous-cell carcinoma (ESCC) cell line KYSE150. METHODS:Recombinant pEGFP-C1-STK15 expression vector was transfected into KYSE150 cells using LipofectamineTM 2000 and the expression of STK15 was detected by fluorescence microscopy and Western blotting. The proliferation of the cells in vitro was measured by MTT assay. The cell cycle distribution and apoptosis were detected by flow cytometry. The proliferation of the cells in vivo was measured by tumorigenicity experiment in nude mice. RESULTS:After recombinant pEGFP-C1-STK15 expression vector was stably transfected into KYSE150 cells, GFP-STK15 fusion protein localized to centrosome and spindle. The STK15-overexpressing colonies were further confirmed by Western blotting. MTT assay showed that the proliferation of the cells in STK15 overexpression group was increased compared with control group (P<0.01). Flow cytometry analysis showed that the percentage of the cells in G0/G1 phase and the cell apoptosis in STK15 overexpression group were decreased compared with control group (P<0.01). The tumorigenicity experiment in nude mice showed that the proliferation of the cells in STK15 overexpression group was increased compared with control group (P<0.01). CONCLUSION: Overexpression of STK15 in human ESCC KYSE150 cells promotes the cell growth in vitro and in vivo, indicating that STK15 may serve as a novel therapeutic target for esophageal carcinoma.     

成纤维上皮细胞炎症反应模型的建立

利用培养金黄色葡萄球菌培养基过滤液建立成纤维上皮细胞炎症反应模型，用于观察和研究炎症反应过程中细胞形态的变化。将培养金黄色葡萄球菌培养基除菌过滤冻干，用细胞培养基稀释，按照一定蛋白含量加到上皮细胞的培养基中，制作细胞炎症反应模型。结果显示，成纤维上皮细胞发生炎症反应，高浓度细菌滤过液的细胞形态发生变化，细胞边缘模糊不清，细胞出现空泡，随着培养时间的延长低浓度的大量大型细胞开始增加；在添加金黄色葡萄球菌培养基滤过液培养24h后利用^3H—TdR方法测定细胞增殖率，细胞增殖差异是显著的。结果表明，建立细胞炎症反应模型是可行的，并为以后炎症反应机理研究和抗炎性反应药物研究奠定了基础。     

连花柴芩可溶性粉对鸡风热犯肺证的临床试验研究

【目的】 研究连花柴芩可溶性粉对风热犯肺所导致的鸡发热、咳嗽、食欲减退、口渴喜饮等证候的临床疗效。【方法】 采用鸡传染性支气管炎病毒人工攻毒建立风热犯肺证肉鸡病例模型,将出现典型风热犯肺证临床症状的150只鸡纳入到试验中,并随机分为5组,30只/组,分别为连花柴芩可溶性粉高、中、低剂量组（连花柴芩可溶性粉浓度分别为4、2、1 g/L）,麻杏石甘颗粒药物对照组（1 g/L）,模型对照组,另设30只为健康对照组。除健康对照组和模型对照组外,各试验组连续饮水给药5 d,记录给药前后鸡的精神状态、体温、呼吸道症状、采食及饮水的变化,并检测各组鸡的免疫器官指数、外周血免疫细胞亚群指标,综合考察连花柴芩可溶性粉的临床疗效。【结果】 连花柴芩可溶性粉高、中剂量组对风热犯肺所致的鸡发热、咳嗽的总有效率分别为90.00%和86.66%,显著高于模型对照组（P<0.05）,且对病鸡采食量下降、饮水量增多具有较好的缓解效果（P<0.05）;连花柴芩可溶性粉能提高病鸡免疫器官指数,促进病鸡外周血中CD3⁺、CD4⁺、CD8⁺ T淋巴细胞增殖,且连花柴芩可溶性粉高、中剂量显著高于模型对照组（P<0.05）。【结论】 连花柴芩可溶性粉对风热犯肺所致的鸡发热、咳嗽、食欲减退、口渴喜饮具有较好的缓解和治疗效果,结合药物量效关系和经济成本等因素综合考虑,推荐连花柴芩可溶性粉临床应用剂量为2 g/L,饮水给药,连用5 d。