Inhibition of NMDA-induced protein kinase C translocation by a Zn chelator: Implication of intracellular Zn

The role of intracellular Zn²⁺ in the translocation of protein kinase C from cytosol to membrane fractions was examined by the [³H]phorbol 12,13-dibutyrate (PDBu) binding method in guinea pig cerebral synaptoneurosomes. N-methyl-d-aspartate (NMDA, 100 μM) and calcium ionophore A23187 (0.3–30 μM) decreased the binding activity in the cytosol with a concomitant increase in the membrane fractions. Pretreatment of synaptoneurosomes with a heavy metal chelator, N,N,N′,N′-tetrakis-(2-pyridylmethyl)ethylenediamine (TPEN), inhibited the NMDA- and A23187-induced changes of the distribution of [³H]PDBu binding sites in cytosol and membrane fractions. The inhibitory effect of TPEN was negated by a preincubation of TPEN with equimolar Zn²⁺ but not by that with Ca²⁺. The addition of 500 μM Zn²⁺ to the lysate of synaptoneurosomes induced an increase of [³H]PDBu binding activity in the membrane fraction with a concomitant decrease in the cytosol fraction, as did 100 μM Ca²⁺. Low concentrations of Zn²⁺ (10 μM), which alone had no effect on the distribution of the binding, significantly enhanced the effect of 10 μM Ca²⁺ in the lysate. Under those conditions TPEN inhibited the Zn²⁺-potentiated Ca²⁺-dependent changes in the binding. These results suggest that intracellular Zn²⁺ is essential for the agonist-induced translocation of protein kinase C in guinea pig synaptoneurosomes.     

瘦素、C-反应蛋白与老年急性冠脉综合征患者的相关研究

目的探讨急性冠脉综合征(ACS)患者血浆瘦素(LP)及C-反应蛋白(CRP)水平的变化及其临床意义。方法测定急性心肌梗死组(AMI组)、不稳定型心绞痛组(UA组)、对照组(健康体检者)血浆LP及CRP浓度,分析LP、CRP与病情严重程度的关系以及LP与CRP的相关性。结果AMI组、UA组患者血浆LP、CRP浓度较对照组显著升高(P<0.01),2组患者血浆LP与CRP浓度之间均呈显著正相关;AMI组血浆LP、CRP浓度比UA组明显升高(P<0.05)。结论血浆LP及CRP浓度与ACS病情严重性呈正相关,且LP与CRP浓度呈正相关。说明LP与CRP共同参与组织炎症反应,是ACS重要的危险因素及预测因子。     

Preparation of protected peptidyl thioester intermediates for native chemical ligation by Nα‐9‐fluorenylmethoxycarbonyl (Fmoc) chemistry: considerations of side‐chain and backbone anchoring strategies,and compatible protection for N‐terminal cysteine*,†

Abstract: Native chemical ligation has proven to be a powerful method for the synthesis of small proteins and the semisynthesis of larger ones. The essential synthetic intermediates, which are C‐terminal peptide thioesters, cannot survive the repetitive piperidine deprotection steps of N^α‐9‐fluorenylmethoxycarbonyl (Fmoc) chemistry. Therefore, peptide scientists who prefer to not use N^α‐t‐butyloxycarbonyl (Boc) chemistry need to adopt more esoteric strategies and tactics in order to integrate ligation approaches with Fmoc chemistry. In the present work, side‐chain and backbone anchoring strategies have been used to prepare the required suitably (partially) protected and/or activated peptide intermediates spanning the length of bovine pancreatic trypsin inhibitor (BPTI). Three separate strategies for managing the critical N‐terminal cysteine residue have been developed: (i) incorporation of N^α‐9‐fluorenylmethoxycarbonyl‐S‐(N‐methyl‐N‐phenylcarbamoyl)sulfenylcysteine [Fmoc‐Cys(Snm)‐OH], allowing creation of an otherwise fully protected resin‐bound intermediate with N‐terminal free Cys; (ii) incorporation of N^α‐9‐fluorenylmethoxycarbonyl‐S‐triphenylmethylcysteine [Fmoc‐Cys(Trt)‐OH], generating a stable Fmoc‐Cys(H)‐peptide upon acidolytic cleavage; and (iii) incorporation of N^α‐t‐butyloxycarbonyl‐S‐fluorenylmethylcysteine [Boc‐Cys(Fm)‐OH], generating a stable H‐Cys(Fm)‐peptide upon cleavage. In separate stages of these strategies, thioesters are established at the C‐termini by selective deprotection and coupling steps carried out while peptides remain bound to the supports. Pilot native chemical ligations were pursued directly on‐resin, as well as in solution after cleavage/purification.     

Immunogenicity and safety of a novel yeast Hansenula polymorpha-derived recombinant Hepatitis B candidate vaccine in healthy adolescents and adults aged 10–45 years

The aim was to determine whether the immunogenicity of an investigational hepatitis B vaccine (spHB) is at least as high as that of a licensed control vaccine, Engerix B^®, and to evaluate its safety before inclusion in new pediatric combination vaccines. Two randomized, controlled, blind-observer, Phase 3 trials were performed: one in Argentina (344 participants aged 10–15 years, 10 μg HBsAg/dose) and one in Uruguay (344 participants aged 16–45 years, 20 μg HBsAg/dose). Both vaccines were given in a 0, 1, 6 month schedule to all participants with a baseline anti-Hep B antibody titer <0.6 mIU/mL. Antibody titers were measured pre-dose 1, 1 month after dose 2, pre-dose 3, and 1 month after dose 3. Statistical non-inferiority analyses were performed on seroprotection rates (SP) post-dose 3 (% with anti-Hep B titers ≥10 mIU/mL; delta non-inferiority limit of −10%). In both studies, SP for the spHB vaccine was 100% and the spHB vaccine was non-inferior in terms of SP to the licensed control vaccine. GMTs post-dose 3 were approximately 1.8- and 4.1-fold higher for spHB in the 10–15 year and 16–45 year age groups, respectively. Reactogenicity was low for each vaccine, after each dose. This highly immunogenic hepatitis B candidate vaccine was selected for further investigation as a component of new pediatric combination vaccines.     

复方积雪草有效组分对人肾小管上皮细胞补体C3表达的影响

目的：探讨复方积雪草有效组分——积雪草苷／大黄素干预肿瘤坏死因子-α（TNF-α）诱导的人肾近曲小管上皮细胞（HPTEC）补体C3 mRNA及蛋白的表达水平。方法：采用HPTEC，模型组TNF-α 10ng／ml诱导，治疗组在TNF-α诱导的同时，以不同浓度的积雪草苷、大黄素以及积雪草苷合大黄素进行干预，于24h后分别提取细胞RNA及上清，应用逆转录-聚合酶链反应（RT—PCR）和酶链免疫方法（ELISA）分别检测HPTEC C3 mRNA和蛋白的表达。结果：正常HPTEC具有C3 mRNA和蛋白表达，经TNF-α诱导后G表达明显上调，用不同浓度的积雪草苷、大黄素以及积雪草苷合大黄素干预后，C3 mRNA及蛋白水平表达出现不同程度的下调，呈一定的剂量依赖关系和协同作用。结论：复方积雪草有效组分能够抑制炎性细胞因子TNF-α上调所致的肾局部G过度产生。     

慢性乙型肝炎患者病毒e系统状态与肝纤维化关系的研究

目的:观察慢性乙型肝炎患者乙型肝炎病毒e系统状态和复制指标在肝纤维化发生过程中的变化及其与血清纤维化标志的关系,探讨它们在肝纤维化发生过程中的作用及其可能的临床意义.方法:188例慢性乙型肝炎患者根据肝纤维化程度分为S0～S4期等5组,分别用定量PCR及放免法检测患者血清中HBV-DNA及肝纤维化标志透明质酸、Ⅳ型胶原、Ⅲ型前胶原和层粘连蛋白的含量;HBeAg和抗-HBe采用酶联免疫吸附法(ELISA)检测,并观察其在不同肝病理纤维化分期时的变化.结果:随着肝纤维化程度加重,血清HBV-DNA含量逐渐升高,从S1期开始显著增加(P＜0.01);而HBeAg阳性率逐渐降低,S3、S4期较S0显著减少(P＜0.05和P＜0.01);抗-HBe阳性率呈相反的变化趋势,在S3和S4期的阳性率明显高于S0期(P＜0.05和P＜0.01).血清HBV-DNA( )HBeAg( )组血清纤维化标志最低,HBV-DNA(-)抗-HBe( )组最高,两者差异有显著性(P＜0.01).结论:HBV复制和e系统状态的改变与肝纤维化程度密切相关,肝内病毒复制标志与血清纤维化标志联合检测,对于判断肝纤维化程度和指导抗病毒治疗有重要的价值.     

Immunoadsorption in Severe C4d-Positive Acute Kidney Allograft Rejection: A Randomized Controlled Trial

Antibody-mediated rejection (AMR) frequently causes refractory graft dysfunction. This randomized controlled trial was designed to evaluate whether immunoadsorption (IA) is effective in the treatment of severe C4d-positive AMR. Ten out of 756 kidney allograft recipients were included. Patients were randomly assigned to IA with protein A (N = 5) or no such treatment (N = 5) with the option of IA rescue after 3 weeks. Enrolled recipients were subjected to tacrolimus conversion and, if indicated, 'anti-cellular' treatment. All IA-treated patients responded to treatment. One death unrelated to IA occurred after successful reversal of rejection. Four control subjects remained dialysis-dependent. With the exception of one patient who developed graft necrosis, non-responders were subjected to rescue IA, however, without success. Because of a high graft loss rate in the control group the study was terminated after a first interim analysis. Even though limited by small patient numbers, this trial suggests efficiency of IA in reversing severe AMR.     

慢性丙肝患者干扰素治疗前后血清细胞因子的变化

目的 :探讨细胞因子在慢性丙型肝炎发病机制中的作用。方法:2 0例慢性丙型肝炎患者应用干扰素联合利巴韦林抗病毒治疗 2 4周。于治疗前后用 EL ISA法检测血清 IL- 2、IFN- γ、IL- 4、IL- 10水平 ,同时检测生化和病毒指标并观察不良反应。结果:抗病毒治疗后 IL- 2、IFN- γ、IL- 4、IL- 10水平发生了明显变化。与治疗前比较 ,IL- 2、IFN- γ水平逐渐升高 ,IL- 4、IL- 10水平逐渐降低 ,其中 IL- 2、IL- 10水平变化具有统计学意义 (P<0 .0 1)。对干扰素完全应答者 15例 ,IL- 2、IL- 10水平变化具有统计学意义 (P<0 .0 5 ) ;无应答者 5例 ,IL- 2、IFN- γ、IL- 4、IL- 10水平变化无统计学意义 (P >0 .0 5 )。抗病毒治疗过程中无严重不良反应。 结论:IL - 2、IL - 4、IL - 10、IFN-γ共同参与了丙型肝炎的发病。干扰素联合利巴韦林通过调整机体的免疫状态而发挥抗病毒效应 ,IL - 2、IL - 10可以作为预测抗病毒疗效的指标之一。     

Triterpenoid saponins, new metalloprotease snake venom inhibitors isolated from Pentaclethra macroloba.

We report here the antiproteolytic and antihemorrhagic properties of triterpenoid saponin inhibitors, named macrolobin-A and B, from Pentaclethra macroloba, against Bothrops snake venoms. The inhibitors were able to neutralize the hemorrhagic, fibrin(ogen)olytic, and proteolytic activities of class P-I and P-III metalloproteases isolated from B. neuwiedi and B. jararacussu venoms. Clotting and fibrinogenolytic activities induced by snake venoms and isolated thrombin-like enzymes were partially inhibited. Furthermore, the potential use of these inhibitors to complement antivenom therapy as an alternative treatment and/or used as molecular models for development of new therapeutical agents in the treatment of snake bite envenomations needs to be evaluated in future studies.     

早期大剂量维生素C对重症急性胰腺炎大鼠核因子-κB的影响

目的:观察早期大剂量应用抗坏血酸(维生素C)对重症急性胰腺炎(SAP)大鼠的核因子-!B(NF-!B)的影响,研究其作用机制。方法:将72只SAP模型SD大鼠随机分成3组,每组各24只。A组:由大鼠股静脉滴注生理盐水5 ml/kg。B组:大鼠股静脉滴注Vit C 15 mg/kg加生理盐水至5 ml/kg。C组:由大鼠股静脉滴注Vit C 150 mg/kg加生理盐水至5 ml/kg。另取8只SD大鼠作为正常对照组。各组分别于8 h和24 h处死8只大鼠,采血测淀粉酶、脂肪酶、维生素C(PV-C)、超氧化物歧化酶(SOD)、TNF-αI、L-6。大鼠处死时分别取胰头组织3份,一份组织HE染色,行光镜检查,按Kusske的方法,对水肿、炎症、出血和坏死分别评分;一份制成超薄切片,行电镜检查;另一份SP法进行免疫组化染色,检测NF-!B的表达。每组另外8只大鼠观察3 d内存活情况,计算3 d成活率。结果:各组大鼠3 d内的生存率为正常对照组100%(8/8),A组0%(0/8),B组12.5%(1/8),C组50%(4/8),C组的3 d生存率显著高于其他两组(P<0.05)。各组的4项病理学评分均高于正常对照组(P<0.01),C组的4项病理学评分均低于A、B组(P<0.05)。透射电镜检查示C组中分泌颗粒较少,其包膜完整、内质网轻度肿胀、线粒体清晰,未见大片坏死。SAP大鼠体内淀粉酶、脂肪酶、细胞因子TNF-α和IL-6的水平明显增高,血清SOD和P-VC降低,胰腺组织中NF-!B活化阳性胰腺细胞数明显增多。C组的血清淀粉酶和脂肪酶低于A、B组(P<0.05),SOD和P-VC水平高于A、B组(P<0.05),血清TNF-αI、L-6水平低于A、B组(P<0.05),胰腺组织NF-!B活化水平低于A、B组(P<0.05)。结论:早期大剂量应用Vit C有助于及时提高SAP大鼠的P-VC、E-SOD水平,降低体内淀粉酶、脂肪酶、TNF-αI、L-6水平,其作用机制可能与大剂量Vit C抑制SAP大鼠体内NF-!B活化、在整体水平上抑制细胞因子基因表达、有助于机体免受自由基和过量细胞因子的损伤及减轻胰腺组织的病理性改变等因素有关。