Lipofectin介导转移的外源性ADA基因在反复上感儿T淋巴细胞的表达

本文对４１例健康儿童和１７例反复上呼吸道感染患儿外周血淋巴细胞腺苷脱氨酶（ＡＤＡ）活性进行了检测。在此基础上筛选出２例反复上感伴ＡＤＡ活性低下患儿。在体外对这２例患儿的外周血Ｔ淋巴细胞进行培养后，以Ｌｉｐｏｆｅｃｔｉｎ（脂质体）介导的方法对其进行了外源性ＡＤＡ基因的基因转移。结果显示：２例患儿体外培养淋巴细胞ＡＤＡ活性较转基因前升高。同步进行的标志基因ｐＢＬａｃＺ的基因转移的检测结果也直观地证实了Ｌｉｐｏｆｅｃｔｉｎ介导的基因转移是成功的。该研究为ＡＤＡ－ＳＣＩＤ淋巴细胞基因治疗的研究提供了初步的体外实验资料。     

复苏早期应用氨茶碱对腺苷及血管活性因子的影响

目的： 观察心搏骤停大鼠复苏早期应用氨茶碱对复苏成功率、血浆去甲肾上腺素(NE)、腺苷、一氧化氮(NO)水平及心肌组织内皮素-1(ET-1)、腺苷水平的影响。方法: 选60只SD大鼠，随机分为3组：手术对照组、肾上腺素治疗组和肾上腺素+氨茶碱治疗组各20只。分别测定治疗组自主循环恢复30 min后及手术对照组的血浆NE、腺苷、NO及心肌组织ET-1、腺苷的水平。结果: 肾上腺素+氨茶碱治疗组自主循环恢复时间明显少于肾上腺素治疗组(P＜0.05)。肾上腺素+氨茶碱治疗组自主循环恢复率为75%，30 min存活率为70%，肾上腺素治疗组分别为60%和55% (P>0.05)。2个治疗组自主循环恢复大鼠的血浆腺苷、NE水平及心肌组织ET-1、腺苷水平均明显高于手术对照组(P<0.05)，肾上腺素治疗组血浆NO水平也显著高于手术对照组(P＜0.01)，肾上腺素+氨茶碱治疗组血浆NO及心肌组织ET-1水平低于肾上腺素治疗组(P＜0.05)。结论: 在复苏早期应用腺苷受体拮抗剂氨茶碱不仅可提高复苏成功率，并且降低血浆NO和心肌组织ET-1水平，有利于减轻复苏后综合征。     

联合检测IFN-γ、VEGF-C、CRP及ADA对结核性与恶性胸腔积液鉴别诊断的价值

目的探讨干扰素-γ(IFN-γ)、血管内皮生长因子(VEGF-C)、C-反应蛋白(CRP)及腺苷脱氨酶(ADA)在结核性与恶性胸腔积液鉴别诊断中的应用价值。方法检测122例临床确诊的胸腔积液患者(恶性胸腔积液56例,结核性胸膜炎48例,其他类型18例)胸水和血清中的IFN-γ、VEGF-C、CRP及ADA含量。结果结核组的IFN-γ、CRP浓度及ADA活性明显高于恶性肿瘤组,差异有统计学意义(P0.01),根据受试者工作特征(ROC)曲线结果判断,以100ng/L为临界值,IFN-γ对结核性胸腔积液诊断的灵敏度、特异性分别为83.1%、92.3%;以45U/L为临界值,ADA对结核性胸腔积液诊断的灵敏度、特异性分别为85.6%、96.3%;以110mg/L为临界值,CRP对结核性胸腔积液诊断的灵敏度、特异性分别为79.1%、84.2%;三项指标联合检测,其灵敏度、特异性分别达到87.8%和86.0%。恶性胸腔积液中VEGF-C高于结核性及其他类型胸腔积液(P0.01);VEGF-C/ADA≥8对恶性胸腔积液诊断的灵敏度、特异性分别为86.3%、82.6%;VEGF-C/ADA≤3对结核性胸腔积液诊断的灵敏度、特性度分别为85.1%、87.1%。结论联合检测IFN-γ、VEGF-C、CRP及ADA可以提高结核性胸膜炎诊断的灵敏度及特异性,VEGF-C与ADA浓度比值对胸腔积液的鉴别诊断具有较好的临床价值。     

A1 and A2A receptor activation by endogenous adenosine is required for VIP enhancement of K+ -evoked [3H]-GABA release from rat hippocampal nerve terminals

Vasoactive intestinal peptide (VIP) modulates GABA release from hippocampal nerve terminals and enhances hippocampal synaptic transmission through a pathway dependent on GABAergic transmission. Since VIP modulation of hippocampal synaptic transmission is dependent on the tonic actions of adenosine we investigated if endogenous adenosine could influence VIP enhancement of GABA release from isolated hippocampal nerve endings, and which adenosine receptors could be mediating this influence. When extracellular endogenous adenosine was removed using adenosine deaminase (ADA, 1 U/ml), the enhancement (57.2 ± 3.7%) caused by VIP on GABA release was prevented. Blockade of adenosine A₁ receptors with 1,3-dipropyl-8-cyclopentylxanthine (DPCPX, 10 nM) or of A_2A receptors with ZM241385 (50 nM) abolished the effect of VIP. In the presence of ADA, selective A_2A receptor-activation with CGS21680 (10 nM) readmitted most of the enhancement caused by VIP on GABA release (50.7 ± 5.3%). Also in the presence of ADA, A₁ receptor activation with N⁶-cyclopentyladenosine (CPA, 50 nM) partially readmitted that effect of VIP (32.6 ± 3.8%). In conclusion, the enhancement of GABA release caused by VIP in hippocampal nerve terminals is dependent on the tonic actions of adenosine on both A₁ and A_2A receptors, and this action of adenosine is essential to VIP modulation of GABA release.     

Quality validation of platelets obtained from the Haemonetics and Trima Accel automated blood-collection systems

BackgroundPlatelet transfusion is required to treat haemo-oncology or trauma patients. Platelet apheresis (PA) performed with apheresis equipment has increased rapidly in recent years. Leucocyte-reduced platelet apheresis (LRPA) can reduce the risk of platelet refractoriness and febrile nonhemolytic transfusion reactions (FNHTRs) for transfusion. Accordingly, this study aimed to investigate and compare the platelet metabolic and functional responses between PA performed with Haemonetics and LRPA performed with Trima Accel cell separator.MethodsThe qualities of platelets collected through PA and LRPA were evaluated in terms of visual appearance, morphology, platelet-aggregation changes, metabolic activities, and bacterium-screening test during 5-day storage. Statistical analyses included two-sample t-test and generalised estimating equation(GEE) method.ResultsDuring 5-day storage in LRPA, residual leucocytes were all <1.0×10⁶, and the parameters of platelet function were as follows: platelet aggregated to agonists such as adenosine 5′-diphosphate (ADP) and collagen, and the extent of shape change and pO₂ showed no statistically significant difference between PA and LRPA. The hypotonic shock reaction (HSR) on days 0, 1, and 3 were significantly higher in LRPA than in PA (71.78±6.92 vs. 64.10±7.42; P=0.002; 71.53±8.98 vs. 62.96±9.84; P=0.007; 68.05±7.28 vs. 57.76±6.80; P<0.0001, respectively). Values of mean platelet volume (MPV) were statistically larger in PA than in LRPA on days 0, 1, and 3. On day 5, the swirling score was higher in LRPA than in PA. The mean lactate levels had no statistically significant difference between PA and LRPA. Moreover, no growth was observed through bacterium-screening test conducted on 40 samples.ConclusionComparison of LRPA and PA products collected from the Trima Accel and Haemonetics automated blood-collection systems, respectively, revealed that both products possessed good platelet qualities even though additional processes are needed to reduce leucocytes. Furthermore, investigating the outcomes of other apheresis instruments with focus on the safety of donors, products, and recipients is necessary.     

Brain-derived neurotrophic factor mediates neuroprotection against Aβ-induced toxicity through a mechanism independent on adenosine 2A receptor activation

Abstract

Brain-derived neurotrophic factor (BDNF) promotes neuronal survival through TrkB-FL activation. The activation of adenosine A_2A receptors (A_2AR) is essential for most of BDNF-mediated synaptic actions, such as synaptic plasticity, transmission and neurotransmitter release. We now aimed at evaluating the A_2AR influence upon BDNF-mediated neuroprotection against Aβ_25–35 toxicity in cultured neurons. Results showed that BDNF increases cell survival and reduces the caspase-3 and calpain activation induced by amyloid-β (Aβ) peptide, in a mechanism probably dependent on PLCγ pathway. This BDNF-mediated neuroprotection is not affected by A_2AR activation or inhibition. Moreover neither activation nor inhibition of A_2AR, per se, significantly influenced Aβ-induced neuronal death on calpain-mediated cleavage of TrkB induced by Aβ. In conclusion, these results suggest that, in opposition to the fast synaptic actions of BDNF, the neuroprotective actions of this neurotrophin against a strong Aβ insult do not require the activation of A_2AR.     

Changes in the infectivity, pyruvate kinase activity, acid phosphatase activity and P-glycoprotein expression in glibenclamide-resistant Leishmania mexicana

We previously demonstrated susceptibility of Leishmania sp. to glibenclamide, a K⁺-ATP transport blocker which interacts with members of the superfamily of adenosine 5′ triphosphate-binding cassette transporters. In order to characterize the molecular differences between a sensitive Leishmania strain, NR(Gs), and an experimentally selected glibenclamide-resistant strain, NR(Gr), specific biochemical and functional parameters have been evaluated both in the wild type and in the resistant strain. Most noteworthy, NR(Gr) exhibit an increased expression of P-glycoprotein and a decreased activity of functional key enzymes such as acid phosphatase, a prominent virulent factor of the parasite, and pyruvate kinase, a key control enzyme for both carbohydrate and protein metabolism. The specific biochemical, metabolic and functional changes observed in the resistant strain correlated with a reduced infectivity of stationary phase NR(Gr) in J774 macrophages and suggested a mechanism to overcome the effect of glibenclamide. Received: 21 January 2000 / Accepted: 1 March 2000     

冠心病患者血小板聚集功能及噻氯匹啶作用的观察

目的:探讨冠心病患者血小板聚集功能以及噻氯匹啶对此过程的抑制作用。方法:应用比浊法测量冠心病患者服药前后,由二磷酸腺苷(ADP)、肾上腺素(EPI)、胶原(Coll)和磷酯花生四烯酸(ACA)诱导的血小板聚集功能。结果:患病组ADP、EPI诱导的最大血小板聚集率(0.78±0.23,0.86±0.25)明显高于对照组(0.65±0.19,P＜0.05;0.73±0.21,P＜0.05),且被噻氯匹啶明显抑制(0.68±0.18,P＜0.05;0.75±0.20,P＜0.05)。而两组间Coll和ACA诱导的最大血小板聚集率无明显差异,且患病组治疗前后无明显差异。结论:冠心病患者血小板聚集功能明显增强,噻氯匹啶可明显抑制这一过程。     

ATP对人不死化成纤维细胞增殖及其细胞膜蛋白表达的影响

目的：探讨ATP对不死化人成纤维细胞增殖及其细胞膜蛋白表达的影响。方法：将正常人TIG-7和OUMS-36细胞株,不死化人KMST-6和SUSM-1细胞株在不同浓度的ATP、ADP、AMP条件下分别进行24和96 h的常规细胞培养,观察存活细胞数目,DNA的合成和[³²P]-ATP标记膜蛋白的表达。结果：培养96 h后,0.4 mmol/L ATP对不死化细胞KMST-6的抑制率为77%且DNA合成明显地被抑制,而正常OUMS-36细胞抑制率为41%且DNA合成无明显改变。在1 mmol/L ATP时,多数KMST-6细胞发生死亡,而正常OUMS-36细胞的增殖无明显影响。当ADP、AMP和腺苷或磷酸处理的细胞,仅有ADP处理的不死化细胞存活数目减少（P<0.01）。与正常细胞比较,不死化细胞30 kD、31 kD、33 kD和40 kD的[³²P]-ATP标记膜蛋白呈高表达。结论：ATP对不死化人成纤维细胞的增殖有明显的抑制作用,并且使磷酸化细胞膜蛋白表达增高。