新牛蛙活性肽Catesbeianin-1a的抗菌抗肿瘤作用

通过筛选牛蛙皮肤cDNA文库得到具有较强抗菌抗肿瘤活性肽Catesbeianin-1a,合成其成熟肽氨基酸序列,MIC法测定抑菌活性结果表明,Catesbeianin-1a对临床常见的致病菌具有较强的抑菌活性,其中对猪链球菌2型的MIC为2.5mg/L,且对兔红细胞几乎无溶血性。MTT法测定多肽抗肿瘤细胞活性,结果 Catesbeianin-1a对胃癌细胞和肝癌细胞的抑杀活性很强,对肝癌细胞SGC7901的IC50为0.845mg/L。制作透射电镜超薄切片,利用透射电镜观察活性肽对细菌和肿瘤细胞超微结构的影响,旨在初步探讨牛蛙皮肤活性肽Catesbeianin-1a抗菌、抗肿瘤的作用机制,并以Catesbeianin-1a为基础进行分子改造来提高其抗肿瘤的靶向性并降低其毒性。     

两种牛蛙活性肽结构及体外抗肿瘤细胞增殖作用的研究

试验旨在研究新发现的两种牛蛙活性肽对人肺癌细胞NCI-H446、人乳腺癌细胞MCF-7及小鼠白血病细胞K562 3种肿瘤细胞体外增殖的影响,为新的多肽抗肿瘤药筛选提供依据。本试验利用圆二色谱(CD)法检测两种新牛蛙活性肽的二级结构,通过MTS细胞毒试验测定不同浓度的两种新牛蛙活性肽对人肺癌细胞NCI-H446、人乳腺癌细胞MCF-7及小鼠白血病细胞K562 3种瘤细胞体外增殖的影响。圆二色谱法结果显示,Temporin-Lb的二级空间结构为聚脯氨酸Ⅱ型螺旋(PPⅡ)结构,Catesbeianin-1a的二级空间结构为无规则卷曲。MTS细胞毒试验结果显示,给予Catesbeianin-1a的3种瘤细胞培养24 h后,瘤细胞形态无明显变化,正常增殖,而给予Temporin-Lb的3种瘤细胞培养24 h后,瘤细胞发生皱缩、细胞变圆、脱落甚至死亡,增殖受到抑制,其中对Temporin-Lb在4~40 μmol/L浓度范围内对小鼠白血病细胞K562的抑制效果最为明显。结果表明,新牛蛙活性肽Temporin-Lb具有一定的抑瘤作用,Catesbeianin-1a对瘤细胞的体外增殖没有明显影响。     

杆菌肽(锌)及其应用进展

随着科学技术的发展,作为一种多肽类抗菌药物,杆菌肽(锌)在人们的生产和生活中的应用逐渐合理和规范.笔者综述了近年来杆菌肽(锌)研究、应用及检测的进展情况.     

褪黑素的抗肿瘤作用及其作用机制的研究进展

褪黑素有多种生理作用，其中抗肿瘤作用引起人们广泛注意。它对多种肿瘤，特别是恶性肿瘤具有明显抑制作用，被认为是一种重要的生理性肿瘤抑制剂。其抗肿瘤机制较复杂．主要有直接抑瘤机制(调节雌激素作用，调节生长因子的作用，影响细胞生长周期，影响钙调节蛋白的功能和细胞代谢，增加细胞间缝管连接)和间接抑瘤机制(抗氧化作用，增强免疫保护作用)等。对这些机制的详细了解将为开发褪黑素作为新的抗肿瘤药物提供理论依据。     

生物活性肽的作用及其应用状况

生物活性肽是一类能够调节生物机体生物活动或具有特殊生理作用的肽类，它们分布广泛，多来源于动植物，在生物体内具有重要的作用。笔者综述了营养肽、免疫肽、促生长肽、抗高血压肽、抗菌肽的来源、种类和功能，以期为生物活性肽的研究和应用提供参考。     

牦牛胎盘肽抗氧化作用的研究

为了探讨牦牛胎盘肽的抗氧化作用,试验将20只小鼠随机分为药物组和对照组,连续15 d分别注射牦牛胎盘肽和生理盐水,0.2 m L/d,然后处死小鼠,取其心肌、肝脏、脾脏、肺脏和肾脏,制成组织匀浆,测定各组织中超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)活性和丙二醛(MDA)含量。结果表明:药物组心肌和肝脏SOD活性显著高于对照组(P0.05);药物组心肌、肝脏、脾脏、肺脏和肾脏GSH-Px活性显著高于对照组(P0.05);药物组心肌、肝脏和脾脏MDA含量显著低于对照组(P0.05)。说明牦牛胎盘肽具有一定抗氧化能力。     

抗菌肽Temprine-La(FS)抑制2型猪链球菌生物被膜形成的研究

试验旨在研究抗菌肽Temprine-La(S)(T-La(S))、Temprine-La(FS)(T-La(FS))、RGD-T-La(S)和RGD-T-La(FS)对2型猪链球菌(SS2)生物被膜形成的抑制作用。通过结晶紫染色法(CV)检测SS2生物被膜形成能力;微量稀释法测定抗菌肽对SS2生物被膜的最小生物被膜抑菌浓度(MBIC)和最小生物被膜杀菌浓度(MBEC);结晶紫染色法和扫描电镜(SEM)检测抗菌肽对SS2生物被膜形成的影响;XTT法检测抗菌肽对SS2生物被膜代谢活性的影响;苯酚硫酸法检测抗菌肽对SS2生物被膜胞外多糖含量的影响;建立猪链球菌-斑马鱼感染模型,HE染色法观察T-La(FS)对斑马鱼脑组织病理变化的影响;实时荧光定量PCR法分析抗菌肽对SS2生物被膜相关基因及对斑马鱼炎性细胞因子基因转录水平的影响。结果显示,SS2具有良好的生物被膜形成能力;T-La(S)、RGD-T-La(S)、T-La(FS)和RGD-T-La(FS)对SS2生物被膜的MBIC分别为31.3、15.6、7.8和15.6μg/mL;MBEC分别为62.6、31.2、15.6和31.2μg/mL;结晶紫染色结果表明,抗菌肽对SS2生物被膜的形成有抑制作用;扫描电镜结果显示,抗菌肽使SS2生物被膜中的细菌数量和生物被膜的形态发生明显变化,细胞外基质大量减少;XTT法结果显示,抗菌肽可显著降低SS2生物被膜的代谢活性;苯酚硫酸法结果显示,抗菌肽能有效抑制SS2生物被膜合成胞外多糖;实时荧光定量PCR结果表明,抗菌肽作用后降低了SS2生物被膜基因的转录水平;T-La(FS)作用后TLR2、MyD88及促炎性细胞因子基因的转录水平显著或极显著降低(P<0.05;P<0.01),抗炎性细胞因子基因的转录水平显著或极显著升高(P<0.05;P<0.01)。抗菌肽主要通过影响生物被膜相关基因转录水平和阻断胞外多糖的合成与分泌来抑制SS2生物被膜的形成,其中T-La(FS)可能通过抑制TLR2信号通路中TLR2和MyD88分子表达,抑制炎性细胞因子的释放,减轻脑膜炎的炎性反应。     

抗菌肽Temprine-La(FS)抑制2型猪链球菌生物被膜形成的研究

试验旨在研究抗菌肽Temprine-La（S）（T-La（S））、Temprine-La（FS）（T-La（FS））、RGD-T-La（S）和RGD-T-La（FS）对2型猪链球菌（SS2）生物被膜形成的抑制作用。通过结晶紫染色法（CV）检测SS2生物被膜形成能力;微量稀释法测定抗菌肽对SS2生物被膜的最小生物被膜抑菌浓度（MBIC）和最小生物被膜杀菌浓度（MBEC）;结晶紫染色法和扫描电镜（SEM）检测抗菌肽对SS2生物被膜形成的影响;XTT法检测抗菌肽对SS2生物被膜代谢活性的影响;苯酚硫酸法检测抗菌肽对SS2生物被膜胞外多糖含量的影响;建立猪链球菌-斑马鱼感染模型,HE染色法观察T-La（FS）对斑马鱼脑组织病理变化的影响;实时荧光定量PCR法分析抗菌肽对SS2生物被膜相关基因及对斑马鱼炎性细胞因子基因转录水平的影响。结果显示,SS2具有良好的生物被膜形成能力;T-La（S）、RGD-T-La（S）、T-La（FS）和RGD-T-La（FS）对SS2生物被膜的MBIC分别为31.3、15.6、7.8和15.6 μg/mL;MBEC分别为62.6、31.2、15.6和31.2 μg/mL;结晶紫染色结果表明,抗菌肽对SS2生物被膜的形成有抑制作用;扫描电镜结果显示,抗菌肽使SS2生物被膜中的细菌数量和生物被膜的形态发生明显变化,细胞外基质大量减少;XTT法结果显示,抗菌肽可显著降低SS2生物被膜的代谢活性;苯酚硫酸法结果显示,抗菌肽能有效抑制SS2生物被膜合成胞外多糖;实时荧光定量PCR结果表明,抗菌肽作用后降低了SS2生物被膜基因的转录水平;T-La（FS）作用后TLR2、MyD88及促炎性细胞因子基因的转录水平显著或极显著降低（P<0.05;P<0.01）,抗炎性细胞因子基因的转录水平显著或极显著升高（P<0.05;P<0.01）。抗菌肽主要通过影响生物被膜相关基因转录水平和阻断胞外多糖的合成与分泌来抑制SS2生物被膜的形成,其中T-La（FS）可能通过抑制TLR2信号通路中TLR2和MyD88分子表达,抑制炎性细胞因子的释放,减轻脑膜炎的炎性反应。     

美国动物营养学的最新研究进展-肽产品Peptiva(R)

生物活性肽产品PeptivaR(派肽威)(由美国华达公司提供)的最新研究及应用情况,它代表了动物营养学中一些最新的研究成果.研究表明,它能改善球虫抑制动物生长的贻害作用和起到解除大肠杆菌毒素引起的厌食作用.在农场使用方面,它也被证明能促进营养吸收,增加体质量,增加蛋重超过10%的功能.但最主要的作用是它能大大增加农场的投资回报率.     

槲寄生在荷瘤小鼠体内的抗肿瘤作用研究

为了探讨槲寄生在荷瘤小鼠体内的抗肿瘤作用,将槲寄生提取物1.65～15 mg/(kg·d)经皮下分别注射于荷肉瘤S180的实体瘤和腹水瘤小鼠体内,连续注射9d后,分别对其肿瘤重量、抑瘤率、脾有核细胞数、脾指数、生存期以及肿瘤组织等病理变化进行观测.试验结果表明,槲寄生对体内肿瘤生长的抑制率为14.7%～28.3%,经其治疗后,小鼠的存活时间及脾指数均明显高于对照组(P<0.01),且其瘤组织中为较致密的结缔组织,对照为疏松的结缔组织.槲寄生在荷瘤小鼠体内有明显的抗肿瘤作用,对脾脏的保护功能可能与其抗瘤机制有关.     

Isolation and immunomodulatory activity of bursal peptide,a novel bursal peptide from the chicken bursa of Fabricius

The bursa of Fabricius (BF), which is unique to birds, serves as the central humoral immune organ and plays a significant role in B lymphocyte differentiation. In this study, a new bursal peptide (BP-IV) was isolated from BF, which promoted colony-forming unit pre-B formation and regulated B cell differentiation. BP-IV also exerted immunomodulatory effects on antigen-specific immune responses via both humoral and cellular immunity in chicken and mice that had been immunized with inactivated avian influenza virus (AIV; H9N2 subtype), including enhancing AIV-specific antibody and cytokine production. The results of this study provided novel insights into the use of a potential candidate reagent for B cell development and future immuno-pharmacological use.     

中国鲎(Tachypleus tridentatus)基因工程抗菌肽的制备及其抗菌活性

从中国鲎血细胞RT-PCR扩增抗菌肽tachyplesins基因,经改造构建原核和真核表达载体pET-KRN、pPIC9-KRY,并分别在大肠杆菌DE3和毕赤酵母GSll5中诱导表达.用Tris-Tricine SDS-PAGE检测并进一步电洗脱纯化表达产物,经体外生物活性测定表明,表达产物KRN对大肠杆菌、金黄色葡萄球菌、枯草杆菌、伤寒沙门菌的抑菌环为25、22、25、16mm;MIC值为3.57、7.14、3.57、14.28mg/L.表达产物KRY对大肠杆菌、金黄色葡萄球菌、枯草杆菌、伤寒沙门菌和李氏杆菌的抑菌环为26、22、25、18、16mm;MIC值为1.57、3.14、1.57、6.28、6.28mg/L,对水霉的抑菌效果尤其显著,抑菌环达35mm.     

Production of loach (Misgurnus anguillicaudatus) germ-line chimera using transplantation of primordial germ cells isolated from cryopreserved blastomeres

An efficient procedure for the cryopreservation of fish blastomeres followed by restoration through germ-line chimera formation was established. Blastomeres of the loach (Misgurnus anguillicaudatus) were cryopreserved in 250-μL straws in Eagle's minimum essential medium with various concentrations of dimethyl-sulfoxide (0, 5, 10, 15, and 20%), and the best concentration was combined with glycerol (1, 2, and 4%) and external cryoprotectants (1 or 2% sucrose; 2, 5, or 10% fetal bovine serum; 1 or 2% BSA). Postthaw viability of the blastomeres was used to optimize cryopreservation conditions. Donor blastomeres were injected with zebrafish green fluorescence protein-nos1 3' untranslated region mRNA and biotin dextran before cryopreservation in the optimal freeze medium. Host embryos were injected with zebrafish DsRed-nos1 3' untranslated region mRNA and reared to the blastula stage. Donor blastomeres were thawed at 25 °C for 10 s and transplanted to the host embryos either immediately or after incubation for 16 h at 20 °C. Donor and host primordial germ cell migration was visualized with fluorescent imaging during the early stages of embryogenesis, and also by histology in 4-d-old embryos. Transplantation of blastomeres immediately after thawing gave decreased hatching rates (approximately 3%) and generated a smaller percentage of germ-line chimeras (approximately 1.1%). In contrast, incubation of a cryopreserved sample for 16 h followed by transplantation of the green fluorescence protein-positive blastomeres improved the hatching rate to 90%, and successfully produced presumable germ-line chimeras at a rate of 16.5%. The improved survival rates and germ-line chimerism may be an effective method for gene banking and subsequent reconstitution of endangered fish genotypes.     

A peptide derived from human bactericidal/permeability-increasing protein (BPI) exerts bactericidal activity against Gram-negative bacterial isolates obtained from clinical cases of bovine mastitis

Gram-negative bacteria are responsible for approximately one-third of the clinical cases of bovine mastitis and can elicit a life-threatening, systemic inflammatory response. Lipopolysaccharide (LPS) is a membrane component of Gram-negative bacteria and is largely responsible for evoking the inflammatory response. Antibiotic and anti-inflammatory therapy for treating Gram-negative infections remains suboptimal. Bactericidal/permeability-increasing protein (BPI) is a neutrophil-derived protein with antimicrobial and LPS-neutralizing properties. Select peptide derivatives of BPI are reported to retain these properties. The objective of this study was to evaluate the antimicrobial activity of a human BPI-derived synthetic peptide against clinical bovine mastitis isolates of Gram-negative bacteria. A hybrid peptide was synthesized corresponding to two regions of human BPI (amino acids 90-99 and 148-161), the former of which has bactericidal activity and the latter of which has LPS-neutralizing activity. The minimum inhibitory (MIC) and bactericidal (MBC) concentrations of this peptide against various genera of bacteria were determined using a broth microdilution assay. The MIC's were determined to be: 16-64 microg/ml against Escherichia coli; 32-128 microg/ml against Klebsiella pneumoniae and Enterobacter spp.; and 64-256 microg/ml against Pseudomonas aeruginosa. The MBC's were equivalent to or 1-fold greater than corresponding MIC's. The peptide had no growth inhibitory effect on Serratia marcescens. The antimicrobial activity of the peptide was retained in the presence of serum, but severely impaired in milk. Further functional evaluation of the peptide demonstrated its ability to completely neutralize LPS. Together, these data support additional investigations into the therapeutic application of BPI to the treatment of Gram-negative infections in cattle.     

蝉新型抗菌肽Cryptonin在毕赤酵母中的分泌表达及活性鉴定

根据毕赤酵母(Pichia pastoris)密码子的偏嗜性,通过SOEing法合成改造后的Cryptonin基因片段。基因克隆到pPICZα-C质粒中,获得分泌型重组酵母表达载体pPICZα-C-Cryptonin。pPICZα-C-Cryptonin通过限制性内切酶SacⅠ酶切线性化后,经电穿孔法转入毕赤酵母细胞X-33内。PCR检测Cryptonin基因与毕赤酵母染色体稳定结合,Zeocin抗性筛选,得到高拷贝转化子。在AOX(醇氧化酶)启动子调控下,Cryptonin蛋白获得分泌表达,其相对分子质量约为2700。表达产物耐热性强,在100℃条件下10min内抗菌活性不变,煮沸30min以上仍具有活性。琼脂糖孔穴扩散法检测显示表达产物对革兰氏阳性菌和革兰氏阴性菌都具有很好的抑菌活性。     

Pyrimidinones: inducers of NK cell activity and antitumor immunity

We have shown that pyrimidinone molecule, 2 amino-5-iodo-6-phenyl-4-pyrimidinone (AIPP) after regional administration, displayed both prophylactic and therapeutic effect on ascitic mammary adenocarcinoma, ACA-755. The antitumor effect was mediated by natural killer (NK) cells, since depletion of this lymphocyte population resulted in abrogation of AIPP-induced tumor resistance. This observation confirms the role of NK cells in antitumor immunity and suggests that AIPP may be therapeutically beneficial also in man.