家兔隔核中去甲肾上腺素对皮肤与内脏痛阈的影响

本工作以电刺激内脏大神经或耳尖部皮肤测定清醒家兔内脏痛阈或皮肤痛阈,以探讨隔核去甲肾上腺素在内脏镇痛和皮肤镇痛中的作用以及与中脑导水管周围灰质(PAG)中内阿片肽系统的关系。实验观察到,双侧隔核内微量注射α受体激动剂可乐宁(10μg/2μl)或α受体阻断剂酚妥拉明(10μg/2μl)对内脏痛阈无明显影响。注入β受体激动剂异丙肾上腺素(1μg/2μl)使内脏痛阐明显升高;而注入β受体阻断剂心得安(1Cμg/2μl)则内脏痛阈明显降低。隔核内注入酚妥拉明(10μg/2μl)或心得安(10μg/2μl)均可使皮肤痛阈明显提高。提示,隔核内NA通过β受体调制内脏痛;通过α受体和β受体调制皮肤痛。隔核内注入异丙肾上腺素(1μg/2μl)明显地镇内脏痛,此作用可被PAG内注射纳洛酮(1μg/2μl)或注射抗亮啡肽抗血清(1:20,000)所减弱;但可使PAG内亮啡肽样物质释放量增加。这提示,隔核内NA的镇内脏痛作用与PAG的内阿片肽系统有关;其中亮非肽在这一过程中具有重要作用。     

以Candin为佐剂的人乳头瘤病毒多肽 治疗疫苗诱导小鼠的体液免疫应答

目的研究以Candin为佐剂的人乳头瘤病毒(Human papillomavirus,HPV)多肽治疗疫苗诱导小鼠产生的体液免疫应答。方法分别采用PBS(对照)、佐剂Candin(150μL/只)、HPV16 E7多肽[3个E7多肽片段,50μg/(段·只)]和包含佐剂Candin与HPV16 E7多肽的疫苗对C57BL/6小鼠进行免疫接种实验,在第21天和第42天各加强1次免疫,剂量同第1次免疫。第3次免疫后2周,颈脱位处死小鼠,取血清。以ELISA法检测血清总IgG、IgG1和IgG2a抗体浓度。结果经多肽(P=0.001,P0.001)和疫苗(P=0.001,P=0.008)免疫的小鼠总IgG抗体、IgG1抗体水平均较对照组高,差异有统计学意义。与对照组相比,疫苗(P=0.008)可明显提高小鼠血清中IgG2a抗体水平。结论包含佐剂Candin及HPV16 E7多肽的疫苗能诱导小鼠产生明显的体液免疫应答。     

合成多肽对结核分枝杆菌的胞内抑制作用

评价合成多肽对人白血病单核巨噬细胞THP-1的毒性和对结核分枝杆菌H37Rv的胞内抑菌作用。通过多肽的不同给药浓度,确定多肽对结核分枝杆菌的最小抑菌浓度(MIC),利用流式细胞术方法和MTT法检测2号肽对细胞THP-1的毒性作用,同时采用CFU方法检测其对H37Rv菌株的胞内抑菌作用。筛选的4条多肽均有抑菌作用,其中2号肽的最小抑菌浓度(MIC)最小,为200μg/m L。2号肽与细胞THP-1作用时,浓度为1 200μg/m L时表现出细胞毒性,与INH细胞毒性无显著差别。对于胞内H37Rv,2号肽抗结核作用具有时间和剂量依赖效应,随给药时间和剂量的增加H37Rv菌落数明显下降。2号肽不仅对巨噬细胞THP-1的毒性小,而且具有较好的抗胞内结核分枝杆菌活性,是一种潜在的抗结核新型药物。     

抗ANXA1自身抗体ELISA检测方法的建立及应用

[目的]建立检测抗膜联蛋白A1(Annexin A1,ANXA1)自身抗体的ELISA方法,初步探索其临床应用。[方法]设计ANXA1特异性抗原肽并人工合成,用抗原肽制备可检测ANXA1抗体的ELISA板,通过回归方程的建立、精密度的测定和抗干扰能力的验证,建立检测ANXA1抗体的ELISA方法。用该法检测临床乳腺癌及健康对照患者血清中的抗ANXA1自身抗体。[结果]该方法在抗体浓度0.025～0.400μg/mL范围内线性良好,R²=0.994,连续5次批内和批间变异系数%均小于10%,对乳糜标本和溶血标本的检测相对误差均小于10%,用于乳腺癌患者抗ANXA1自身抗体的诊断上,实验组抗ANXA1自身抗体表达显著高于对照组,差异具有统计学意义(P<0.05)。[结论]建立了抗ANXA1自身抗体检测的ELISA方法,线性范围0.025～0.400μg/mL,变异系数小于10%,对乳糜标本和溶血标本检测的相对误差小于10%,应用于临床初步显示乳腺癌患者血清中抗ANXA1自身抗体高表达。     

单种群生长的广义Logistic模型

描述单种群生长的一般Logistic模型其中r>0为种群的内禀生长率,K>0为环境容纳量。它通常被称为Verhulst-Pearl模型,其基本特征是描述了种群的S型生长(如图1),当种群的初始值x_o相似文献   

利用天蚕蛾肽基因增强青鱼将对病原细菌的抵抗力

MarineBiotechnology 2 0 0 2年 5 / 6月 4卷 3期 31 0～ 32 2页报道 :天蚕蛾肽 (cecropin)是最早在天蚕蛾(Hyalophoracecropia)中发现的一组抗微生物肽。它具有广谱的抗细菌活性。最近 ,美国康涅狄格大学分子和细胞生物学系生物技术中心的科学家AliyeSarmasik等研究了 (1 )天蚕蛾肽是否可以对已知的鱼类病原细菌显示杀菌活性 ,和 (2 )将天蚕蛾肽基因转移于青鱼将 (Oryriaslatipas)是否可以增强转基因鱼对病原细菌感染的抵抗力等诸问题。Sarmasik等将前原天蚕蛾肽B、原天蚕蛾肽B和天蚕蛾肽B ,以及猪天蚕蛾肽P1,在巨细胞病毒 (CMV)启动…     

SO2代谢衍生物对大鼠海马CA1区神经元瞬间外向钾电流和延迟整流钾电流的影响

本文利用全细胞膜片钳技术研究了SO2 代谢衍生物———NaHSO3 和Na2 SO3 (二者分子比为 1∶3)对大鼠海马CA1区神经元瞬间外向钾电流 (IA)和延迟整流钾电流 (IK)的影响。结果表明 ,SO2 代谢衍生物可显著增大IA 和IK,且呈剂量依赖性关系 ,使IA 和IK 增大 5 0 %的剂量分别为 2 6 19μmol/L和 14 5 0 μmol/L。此外还与电压呈依赖性关系 ,但不具有频率依赖性。结果还表明 ,10 μmol/LSO2 代谢衍生物不影响IA 的激活过程 ,而对IK 的激活过程有非常显著的影响 ,给药前后IK 的半数激活电压分别为 17 6 4± 7 31mV和 13 43± 2 0 0mV (n=10 ,P <0 0 1) ,但不改变其斜率因子。另外 ,10 μmol/LSO2 代谢衍生物还非常显著地影响IA 的失活过程 ,给药前后其半数失活电压分别为 - 6 5 93± 1 97mV和 - 5 9 2 2± 3 83mV (n =10 ,P <0 0 1) ,但不改变其斜率因子。由此推断 ,SO2 代谢衍生物增大大鼠海马CA1区神经元的IA 和IK,促进IK 的激活过程 ,并抑制IA 的失活过程 ,可导致胞内K 通过K 通道的外流增加 ,胞内K 浓度降低 ,造成中枢神经元功能紊乱 ,诱导神经细胞凋亡。这意味着SO2 代谢衍生物对中枢神经系统具有损伤作用 ,从而提示大气SO2 污染可能与一些中枢神经系统疾病的发生以及衰老有关 [动物学报 49(1) :73     

下丘脑神经元钙激活钾通道的整流现象

目的研究SD乳鼠下丘脑神经元中钙激活钾通道的整流现象.方法采用膜片钳内面向外式记录方式.结果记录到一种大电导钙激活钾通道(KCa),在对称140mmol/L[K+]时内向电导为(171±12)pS,不随[Ca2+]变化而改变,而外向电导可受[Ca2+]调控,当[Ca2+]为500μmol/L时,外向电导为(76±14)pS.[Ca2+]越大,整流现象越明显,Mg2+对这种KCa的整流作用不明显.结论下丘脑神经元中KCa具有Ca2+依赖性整流现象,它可能与神经元的兴奋性和稳定性有关.     

E.coli ZB43生成Microcin B17的发酵研究

Microcin B17(简称MccB17)是核糖体合成的肽类DNA螺旋酶抑制剂,其独特的分子结构是抗菌剂未来发展的重要的新颖结构,MccB17产生菌的杂环形成机理对于制药工业的药物设计具有重要意义。MccB17产生菌E.coliZB43适宜在M63合成培养基中积累胞内MccB17,低溶氧水平利于MccB17的积累。当葡萄糖浓度超过1g/L时,MccB17的合成受到阻遏。采用均匀设计表U11(1110)进行发酵培养基配比调整,确定了最佳的含葡萄糖合成培养基配比:葡萄糖1g/L,KH2PO43g/L,K2HPO47g/L,(NH4)2SO41g/L,MgSO4.7H2O 1.8mmol/L,盐酸硫胺1.8μg/ml。丁二酸钠能解除葡萄糖对E.coliZB43生成MccB17的代谢阻遏。以10g/L丁二酸钠替代葡萄糖为唯一碳源,在上述最佳发酵培养基配比条件下,37℃24h,MccB17的产量可达559.6μg/ml。     

黄芩苷介导PI3K/AKT通路减轻脂多糖诱导的心肌细胞凋亡及炎症反应

为探讨黄芩苷对脂多糖(LPS)诱导的大鼠心肌细胞凋亡、炎症及磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(AKT)信号通路的调控作用,该研究体外培养大鼠心肌H9C2细胞,将其分为对照组(不做干预)、LPS组(10μg/m L LPS)、实验组(10μg/m L LPS+10、20、40、80μmol/L黄芩苷)、黄芩苷+Y组(10μg/m L LPS+10μmol/L黄芩苷+5μmol/L PI3K/AKT通路抑制剂LY294002)、抑制剂组(10μg/m L LPS+5μmol/L LY294002)和黄芩苷+A组[10μg/m L LPS+10μmol/L黄芩苷+100 ng/mL PI3K/AKT通路激活剂胰岛素样生长因子-I(IGF-I)]。用细胞计数试剂盒-8测定细胞活力;酶联免疫吸附试验检测炎症因子白细胞介素-1β(IL-1β)、IL-6和IL-10的含量; Hoechst33258染色法测定细胞凋亡率; 5-乙炔基-2’脱氧尿嘧啶核苷测定细胞增殖率;蛋白免疫印迹法测定PI3K/AKT相关蛋白、细胞周期蛋白D1(Cyclin D1)和半胱氨酸天冬氨酸蛋白酶-3(Caspas...     

镇痛多肽——内吗啡肽-1的人工合成及活性研究

 用液相合成方法合成了具有镇痛作用的μ阿片受体的内源性配体——内吗啡肽 - 1(endomorphin- 1 ) ,该四肽为 Tyr- Pro- Trp- Phe NH2 .液相合成法是在氨基酸的 N端用 Boc(叔丁氧羰酰基 )作保护基 ,C端用 HOSu(N-羟基琥珀酰亚胺 )活化 ,与未加保护基的氨基酸在碱性条件下接肽 .先分别合成 C端二肽和 N端二肽 ,再缩合为四肽 ,产物的保护基用盐酸脱帽去除 .中间产物用薄层层析和熔点鉴定其纯度 ,最终得到了高纯度的四肽 .小白鼠脑室注射 (i.c.v)测定表明 ,8.2 5nmol剂量给药 ,其镇痛活性为 87% ,明显高于吗啡 (morphine) .     

Joseph Rudinger memorial lecture: Unexpected functions of angiotensin converting enzyme,beyond its enzymatic activity

Angiotensin converting enzyme (ACE) is a well‐known enzyme, largely studied for its action on hypertension, as it produces angiotensin II from angiotensin I. This paper describes two original behaviours of ACE. We showed that ACE could hydrolyse gastrin, a neuropeptide from the gastrointestinal tract, releasing the C‐terminal amidated dipeptide H‐Asp‐Phe‐NH₂. This dipeptide is believed to be involved in the gastrin‐induced acid secretion in the stomach. This hypothetic mechanism of action of gastrin resulted in a strategy to rationally design gastrin receptor antagonists. Beyond, we showed that the brain renin angiotensin system (RAS) could be activated by a new characterized peptide named acein, resulting in stimulation of dopamine release within the striatum. This new and original ‘receptor‐like’ activity for brain membrane‐bound ACE is quite significant taking into account the role of dopamine in the brain, particularly in neurodegenerative diseases. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.     

Metalloendopeptidase (EC 3.4.24.11) but not angiotensin converting enzyme is involved in the inactivation of substance P by synaptic membranes of the rat substantia nigra

Substance P is a neuropeptide released in vivo from the substantia nigra, the principal substance P nerve terminal region in the rat brain. Its inactivation was investigated in a purified nigral synaptic membrane preparation. The membrane-bound enzyme shares many features with the endopeptidase 24-11 (EC 3.4.24.11): 1) hydrolysis of peptide bonds Gln6-Phe7, Phe7-Phe8 and Gly9-Leu10, 2) sensitivity to the inhibition by phosphoramidon and 3) relative affinity for substance P. Bestatine and captopril inhibit only the hydrolysis of the metabolites. These results suggest that substance P is inactivated in substantia nigra by endopeptidase 24-11 and that a bestatin-sensitive aminopeptidase and angiotensin converting enzyme may play a role in subsequent degradation of the substance P metabolites.     

A dipeptidyl carboxypeptidase in brain synaptic membranes active in the metabolism of enkephalin containing peptides

A dipeptidyl carboxypeptidase activity has been localized in synaptic plasma membranes which have been prepared from isolated rat brain cortical synaptosomes. The specificity of this proteolytic activity towards various synthetic and biological active peptides is compared to the peptidase activities of intact synaptosomes. In contrast to the synaptosomal peptidases which are capable of cleaving all peptide bonds of Met-enkephalin-Arg6-Phe7 the peptidase activity associated with the synaptic plasma membrane exclusively hydrolyses a dipeptide from the carboxyl terminus of all hepta- and hexapeptides tested. The fact that this dipeptidyl carboxypeptidase does not cleave the Gly3-Phe4 peptide bond of Met-enkephalin suggests that this enzyme is different from "enkephalinase". The synaptic membrane dipeptidyl carboxypeptidase is inhibited by metal chelating agents and thiols but is not affected by compounds known to inhibit serine proteases, thermolysin and "enkephalinase".     

Characterization of a membrane-specific tubulin isoform by peptide mapping

A membrane-specific tubulin-like protein, found in preparations of synaptic plasma membranes and brain mitochondria, was analyzed by chemical and proteolytic peptide mapping to determine which part of the molecule was different from cytoplasmic tubulin. The membrane polypeptide was identical to alpha tubulin in the first two-thirds of the molecule containing the amino terminal, as found by peptide mapping. However, some differences were observed in the peptide maps of the carboxy terminal one third of the molecule which includes a domain that is important in the regulation of tubulin self-assembly.     

Function of neutral endopeptidase on the cell membrane of human neutrophils

Intact human neutrophils hydrolyzed N-formyl-Met-Leu-[3H]Phe (fMLP) and released Leu-[3H]Phe, cleaving 45-50% of the peptide within 20 min at 37 degrees C. The dipeptide after its release was then hydrolyzed to free amino acids by a dipeptidase (EC 3.4.13.11). This activity, present in plasma membrane-enriched fractions of neutrophil lysates, was also inhibited over 90% by phosphoramidon, an inhibitor of neutral endopeptidase (NEP, EC 3.4.24.11). Dithiothreitol and EDTA inhibited the activity to a comparable degree, suggesting the requirement for a heavy metal cofactor. Bestatin and amastatin, inhibitors of aminopeptidases (but not human kidney NEP), did not inhibit the rate of fMLP degradation but prevented the production of free phenylalanine and enhanced the accumulation of Leu-Phe. Of other inhibitors, alpha 1-antitrypsin and alpha 2-macroglobulin slightly enhanced the rate of fMLP hydrolysis by neutrophils, and others tested were ineffective. Rabbit antiserum to homogeneous human kidney NEP reacted specifically with a 100-kDa protein present in sodium dodecyl sulfate-solubilized neutrophils. The Mr of this protein was slightly larger than that of the kidney enzyme in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The antiserum incubated with intact cells specifically inhibited the degradation of fMLP over 70%. First, we confirm that NEP present on the plasma membrane cleaves fMLP at the Met-Leu bond; then the dipeptide Leu-Phe is cleaved by a dipeptidase. Finally, inhibition of NEP completely blocks fMLP-mediated chemotaxis. Thus, the enzyme may play an important role in modulating chemotactic responses.     

Delivery of a lactose derivative of endomorphin 1 to the brain via the olfactory epithelial pathway

The rapid and direct delivery of a neuroactive endomorphin 1 derivative to the brain via nasal delivery is reported. A synthetic derivative of the native opioid peptide, endomorphin 1 bearing a lactose unit on the N-terminus of the peptide has been previously reported to exhibit antinoceceptive activity similar to morphine after both intravenous and oral administration. This compound has been administered nasally to rats and appeared in the olfactory bulb within 10 min of administration with negligible levels appearing in the circulating blood or in the rest of the brain. These results indicate that the peptide is absorbed into the brain via the olfactory epithelial pathway suggesting nasal delivery may be a viable alternative route of delivery in clinical applications.     

Search of the human proteome for endomorphin-1 and endomorphin-2 precursor proteins

Based on the promising opioid pharmacological profile of the peptide, Tyr-Pro-Trp-Gly-NH(2) (Tyr-W-MIF), Zadina et al. [Zadina, J.E., Hackler, L., Ge, L.-J., Kastin, A.J., 1997. A potent and selective endogenous agonist for the mu-opiate receptor. Nature 386, 499-5502] synthesized and screened other Gly(4)-substituted peptides, culminating in the synthesis of Tyr-Pro-Trp-Phe-NH(2) (endomorphin-1), which displayed high affinity and selectivity for the mu-opioid receptor. The amidated peptide was then isolated from bovine brain frontal cortex, as was a related peptide, Tyr-Pro-Phe-Phe-NH(2) (endomorphin-2), that displayed similar high affinity and selectivity for the mu-opioid receptor. The biosynthesis of the endomorphins in the brain remains obscure, since the putative precursor proteins for the peptides have not been identified. With the completion of the human genome sequencing project, we hypothesized that we should uncover the biological precursors of the peptides using a bioinformatic approach to search the current human proteome for proteins that contained the endomorphin peptide sequences followed by Gly-Lys/Arg, the consensus sequence for peptide alpha-amidation and precursor cleavage. Twelve proteins were identified that contained the endomorphin-1 Tyr-Pro-Trp-Phe sequence, however none contained the Tyr-Pro-Trp-Phe-Gly sequence necessary for alpha-amidation. Twenty-two distinct proteins contained the endomorphin-2 tetrapeptide sequence, and two of those contained the sequence, Tyr-Pro-Phe-Phe-Gly, however, none contained the requisite peptide-Gly-Lys/Arg sequence. Western blot analysis using an endomorphin-2 antibody detected 4 prominent proteins in mouse brain, necessitating reinterpretation of previous immunocytolocalization studies in the brain. Screening of the current human proteome yielded no evidence for endomorphin precursor proteins based on accepted biochemical criteria.     

ω-芋螺毒素MVIIC的N及C端修饰对折叠及活性影响

 合成了 ω-芋螺毒素 MVIIC的三种 N及 C端修饰肽 ,应用高压液相色谱、CD及生物体内活性实验 ,研究了其 N及 C端修饰对折叠及活性的影响 .结果表明 :MVIIC N端用 Phe及 Ser修饰后降低其线性肽形成正确折叠的比例及结构的稳定性 ,对小鼠的脑室给药活性也相应降低 ;C端酰胺转为电负性羧基端后活性降低 ,CD谱存在显著差异 .     

Characterization of a distinct membrane bound dipeptidyl carboxypeptidase inactivating enkephalin in brain

A dipeptidyl carboxypeptidase distinct from the angiotensin converting enzyme (EC 3.4.15.1) was isolated from membrane preparations of rabbit brain. The enzyme cleaved enkephalin at the Gly-Phe bond, releasing either Phe-Leu from Leu-enkephalin or Phe-Met from Met-enkephalin, and also acted on bradykinin, releasing the terminal dipeptide Phe-Arg. In contrast to the converting enzyme, however, this dipeptidyl carboxypeptidase did not act on angiotensin-1, and it did not degrade hippuryl-His-Leu. Chloride ions did not affect its activity, but the enzyme was inhibited by metal chelating agents. The enzyme was not inhibited by captopril (SQ 14225) or by SQ 20881. Kinetic studies indicated a Km for this enzyme of 0.14 mM with Leu-enkephalin and 0.12 mM with bradykinin as substrates. Present data indicate that more than one enzyme is present in brain membrane fractions acting as dipeptidyl carboxypeptidases inactivating enkephalin; these data suggest multiple roles for such enzymes in the regulation of peptide metabolism.