Alzheimer病的T淋巴细胞功能活性检测

目的　探讨外周Ｔ淋巴细胞功能活性与Ａｌｚｈｅｉｍ ｅｒ病（ＡＤ）病情的关系。方法　流式细胞仪测定Ｔ细胞亚群;ＭＴＴ法检测ＣｏｎＡ刺激ＡＤ外周淋巴细胞的增殖功能;ＥＬＩＳＡ法检测细胞因子及其受体表达。结果　（１）ＣＤ４＋ 、ＣＤ８＋ Ｔ细胞比例和ＣＤ４＋ ／ＣＤ８＋ 比值各组间无统计学差异。（２）淋巴细胞增值在轻、重ＡＤ组略下降,但刺激指数（ＳＩ）都大于２,与对照无明显差异。（３）ＩＬ－２的分泌在重度ＡＤ组和ｓＩＬ－２Ｒ表达在轻、重度ＡＤ组明显升高（Ｐ＜ ０．０５）,ＩＬ－２Ｒ的表达各组间差异无显著性。结论　Ｔ淋巴细胞亚群变化与ＡＤ病情的联系不明显,细胞因子分泌功能与ＡＤ进展有关。     

脑缺血病人血小板肌球蛋白轻链激酶和Ca2+、Mg2+-ATP酶活性及其与[Ca2+]i关系的研究

目的探讨急性缺血性脑血管病血小板肌球蛋白轻链激酶（ＭＬＣＫ）和Ｃａ２＋、Ｍｇ２＋－ＡＴＰ酶活性的变化及与血小板胞浆游离钙浓度［Ｃａ２＋］ｉ的关系。方法用３２Ｐ同位素掺入法和比色法分别测定５８例脑缺血病人，及３５名健康对照者血小板ＭＬＣＫ和Ｃａ２＋、Ｍｇ２＋－ＡＴＰ酶活性。用荧光钙指示剂Ｆｕｒａ－２负载血小板扫描的方法，测定脑缺血病人血小板［Ｃａ２＋］ｉ浓度。结果ＴＩＡ组和脑梗死组ＭＬＣＫ活性与对照组相比均有明显增加（Ｐ＜０．０１），而Ｃａ２＋Ｍｇ２＋－ＡＴＰ酶活性均低于对照组（Ｐ＜０．０５，Ｐ＜０．０１）。ＴＩＡ组和脑梗死组血小板静息［Ｃａ２＋］ｉ；均高于对照组；血小板静息［Ｃａ２＋］ｉ与血小板ＭＬＣＫ活性呈显著正相关（Ｐ＜０．０１），而与血小板Ｃａ２＋、Ｍｇ２＋－ＡＴＰ酶活性呈负相关（Ｐ＜０．０５）。结论血小板ＭＬＣＫ和Ｃａ２＋、Ｍｇ２＋－ＡＴＰ酶活性与急性缺血性脑血管病的发生有密切关系，ＭＬＣＫ活性的变化可能是脑缺血病人血小板活化的分子基础。     

抗CD_3抗体活化杀伤细胞免疫表型变化对抗胶质瘤活性影响

研究用抗ＣＤ３抗体刺激体外培养了８例胶质瘤病人的肿瘤浸润淋巴细胞（ＴＩＬ）和外周血淋巴细胞（ＰＢＬ细胞），从中发现了一种新型高效的广谱抗肿瘤效应细胞──ＣＤ３ＡＫ细胞，通过与ＣＤ３＋ＴＩＬ、ＣＤ３－ＴＩＬ和ＬＡＫ细胞比较分析的方法分析了这些效应细胞培养前后细胞表面表型的变化以及对自体胶质瘤细胞，异体胶质瘤细胞，Ｕ２５１ＭＧ及Ｕ９３７等的杀伤活性的影响，同时也讨论了抗ＣＤ２抗体在Ｔ细胞活化中的作用及活化机制。由于αＣＤ３ＡＫ细胞来源广泛，制备简单和抗肿瘤细胞毒活性强，所以将有可能代替ＴＩＬ细胞成为过继免疫治疗的主要抗肿瘤效应细胞。     

首发精神分裂症患者的细胞免疫改变

目的研究未经治疗的首发精神分裂症患者细胞免疫功能状态。方法３０例符合入组标准的首发精神分裂症患者和１１名正常人，用流式细胞仪测定外周血ＣＤ＋３、ＣＤ＋４和ＣＤ＋８Ｔ淋巴细胞，ＭＴＴ法检测患者血浆ＩＬ－２活性和酶标法检测血浆ｓＩＬ－２Ｒ水平。比较细胞免疫功能。结果发现精神分裂症患者血浆ＩＬ－２水平显著高于正常志愿者，ＣＤ＋３、ＣＤ＋４Ｔ淋巴细胞数量降低，ｓＩＬ－２Ｒ水平和ＣＤ＋８Ｔ淋巴细胞没有明显变化。结论精神分裂症患者细胞免疫功能异常活化，活化的细胞免疫功能可能与精神分裂症的发生有关，作用机制有待进一步研究。     

老年人急性脑梗塞患者红细胞变形能力与红细胞 ATP 酶活性关系的研究

检测了６０例老年人急性脑梗塞患者的红细胞变形能力（ＥＤ），红细胞ＡＴＰ酶活性和红细胞内离子浓度的变化。结果显示：老年人急性脑梗塞患者红细胞滤过指数（ＥＦＩ）较对照组显著增高（Ｐ＜０．００１）；红细胞Ｎａ＋－Ｋ＋－ＡＴＰ酶活性和Ｃａ２＋－Ｍｇ２＋－ＡＴＰ酶活性明显降低（Ｐ＜０．００１）；红细胞内Ｎａ＋、Ｃａ２＋浓度明显增高（Ｐ＜０．００１），而Ｍｇ２＋浓度明显降低（Ｐ＜０．０１）。脑梗塞患者红细胞ＥＦＩ与Ｎａ＋－Ｋ＋－ＡＴＰ酶、Ｃａ２＋－Ｍｇ２＋－ＡＴＰ酶活性、Ｍｇ２＋浓度呈负相关（ｒ＝－０．５４２、－０．４１７、－０．４３６）（Ｐ＜０．００１）；与红细胞内Ｎａ＋、Ｃａ２＋浓度呈正相关（ｒ＝０．４７３、０．４６６，Ｐ＜０．００１）；提示脑梗塞病人ＥＤ降低与红细胞ＡＴＰ酶活性降低有关。     

脑缺血病人血小板肌球蛋白轻链激酶和Ca^2＋,Mg^2＋—ATP？…

目的 探讨急性缺血性脑血管病血小板球蛋白轻链激酶（ＭＬＣＫ）和Ｃａ＾２＋Ｍｇ＾２＋－ＡＴＰ酶活性的变化及与血小板胞浆游离钙浓度（Ｃａ＾２＋）ｉ浓度,结果 ＴＩＡ组和脑梗死组ＭＬＣＫ活性与对照组相比均有明显增加（Ｐ〈０．０１）,而Ｃａ＾２＋,Ｍｇ＾２＋－ＡＴＰ酶活性均低于对照组（Ｐ〈０．０５,Ｐ〈０．０１）,ＴＩＡ组和脑梗死组血小板静息（Ｃａ＾２＋）ｉ均高于对照组;血小板静息（Ｃａ＾２＋）ｉ与血小板     

重症肌无力患者周围血中CD4+T淋巴细胞亚群的研究

目的研究ＣＤ４＋Ｔ淋巴细胞的功能亚群在重症肌无力（ｍｙａｓｔｈｅｎｉａｇｒａｖｉｓ，ＭＧ）发病中的作用。方法采用免荧光双标记技术和流式细胞仪对３９例ＭＧ患者和１８例健康对照者周围血中ＣＤ４＋Ｔ淋巴细胞的两个亚群（ＣＤ４＋ＣＤ４５ＲＡ＋、ＣＤ４＋ＣＤ４５ＲＡ－）的百分率进行测定。结果ＣＤ４＋ＣＤ４５ＲＡ＋亚群的百分率在ＭＧ组和ＡＣｈＲＡｂ阳性组明显低于对照组和ＡＣｈＲＡｂ阴性组；而ＣＤ４＋ＣＤ４５ＲＡ－细胞的百分率在ＭＧ组和ＡＣｈＲＡｂ阳性组显著高于对照组和ＡＣｈＲＡｂ阴性组。结论表明ＣＤ４＋Ｔ淋巴细胞的两个功能不同亚群在ＭＧ患者体内发生了异常改变，与ＡＣｈＲＡｂ的产生有关。     

Alzheimer病sIL-2R释放与T细胞活化测定

目的　探讨外周淋巴细胞可溶性白介素－２ 受体（ｓＩＬ－２Ｒ）释放与Ａｌｚｈｅｉｍ ｅｒ 病（ＡＤ）病情的关系。方法　流式细胞仪测定、ＭＴＴ 法和ＥＬＩＳＡ 法检测了刀豆蛋白Ａ（ＣｏｎＡ）和ＣＤ３ 单抗（ＯＫＴ３）刺激 ＡＤ 外周淋巴细胞白介素－２ 受体（ＩＬ－２Ｒ）表达、增殖和ｓＩＬ－２Ｒ 的释放。结果　（１）在ＣｏｎＡ 和ＯＫＴ３刺激下淋巴细胞增殖在轻、重ＡＤ 组均略下降,但刺激指数（ＳＩ）都大于２,与对照组无明显差异。（２）ｓＩＬ－２Ｒ 释放在轻、重度ＡＤ组均较对照组明显升高（Ｐ＜ ０．０５）,ＩＬ－２Ｒ 的表达各组间无显著性差异。结论　ｓＩＬ－２Ｒ作为ＡＤ 的诊断、病情观察指标值得进一步探讨。     

阿尔茨海默病患者外周淋巴细胞对Aβ1-40刺激反应性研究

目的 探讨阿尔茨海默病（ＡＤ）外周淋巴细胞免疫功能的变化。方法 淀粉样蛋白肽（Ａβ１－４０）等刺激ＡＤ患者外周淋巴细胞,测定其增殖功能和凋亡细胞比例;异硫氰酸荧光素（ＦＩＴＣ）标记单抗直接标记,流式细胞仪测定Ｔ细胞亚群。结果 轻、中～重ＡＤ组淋巴细胞对Ａβ１－４０刺激的增殖能力都显著低于对照;白介素－２（ＩＬ－２）分泌、白介素－２受体（ＩＬ－２Ｒ,ＣＤ２５）表达和Ｔ细胞亚群及细胞凋亡在各组间无差异。结论 ＡＤ病人淋巴细胞对Ａβ１－４０刺激增殖反应低下,提示ＡＤ患者对Ａβ１－４０清除能力减弱可能与ＡＤ发病有关。     

精神分裂症免疫指标与精神症状的关系

为探讨精神分裂症精神病理与免疫指标的相关性、评估抗精神病药对免疫指标的影响及其与疗效的关系，用固定剂量氟哌啶醇治疗５０例慢性精神分裂症患者１２周，在治疗前后测查Ｔ细胞亚群和白细胞介素２（ＩＬ－２）分泌细胞，并采用简明精神病评定量表（ＢＰＲＳ）、阳性症状评定量表（ＳＡＰＳ）和阴性症状评定量表（ＳＡＮＳ）进行评定。结果显示，治疗前ＣＤ３阳性细胞（ＣＤ＋３）、ＣＤ４阳性细胞（ＣＤ＋４）、ＣＤ４／ＣＤ８阳性细胞比值（ＣＤ４／ＣＤ８）和ＩＬ－２分泌细胞均明显低于正常人，治疗后ＣＤ＋４呈显著性增高；治疗前ＣＤ４／ＣＤ８与ＳＡＰＳ总分呈显著负相关，ＢＰＲＳ、ＳＡＰＳ和ＳＡＮＳ减分率与治疗前ＣＤ＋３细胞数均呈显著正相关，ＳＡＮＳ减分率与治疗前ＣＤ＋４细胞数亦呈正相关。研究表明，抗精神病药在改善患者精神症状的同时，也使其免疫功能得到改善；临床症状改善程度与治疗前的免疫功能状态相关。     

An alternatively spliced form of glycogen synthase kinase-3β is targeted to growing neurites and growth cones

The serine/threonine kinase glycogen synthase kinase-3β (GSK-3β) is expressed in two, alternatively spliced, isoforms: a short form (GSK-3β1) and a long form containing a 13 amino acid insert in the catalytic domain (GSK-3β2). We examined the expression of these isoforms in the rat using specific antibodies and found that GSK-3β2, in contrast to GSK-3β1, is only expressed in the nervous system. The highest levels of GSK-3β2 are found in the developing nervous system but expression persists into adulthood. In the adult central nervous system the highest expression of GSK-3β2 occurs in regions with a high proportion of white matter, suggesting that GSK-3β2 is expressed in axons. Consistent with this finding, sub-cellular fractionation of neonatal rat brain showed that GSK-3β2 is present in fractions enriched in neurites and growth cones. Furthermore, we found that when we separated neuronal cell bodies from neurites by culturing embryonic cortical neurons in neurite outgrowth inserts, GSK-3β2 was present in both compartments. Finally, a rabbit polyclonal antibody raised to the 13 amino acid insert of GSK-3β2 (anti-8A) that specifically recognises GSK-3β2, labels the cell body, including the nucleus, neurites and growth cones of embryonic neurons in culture. To compare functionally the two isoforms, we performed in vitro kinase assays. These showed that GSK-3β1 is more efficient at phosphorylating the microtubule-associated protein MAP1B than GSK-3β2, consistent with previous findings with the microtubule-associated protein tau. However, when co-expressed with MAP1B in COS-7 cells, both GSK-3β isoforms equally efficiently phosphorylated MAP1B and had a similar influence on the regulation of microtubule dynamics by MAP1B in these cells. We conclude that the alternatively spliced isoform of GSK-3β, GSK-3β2, is neuron-specific and has overlapping activities with GSK-3β1.     

Differential effects of hexachlorocyclohexane isomers on the GABA receptor subunits expressed in human embryonic kidney cell line

We have recently demonstrated by patch clamp experiments that the four isomers of hexachlorocyclohexane (HCH), α-, β-, γ- and δ-HCH insecticides, modulate the kinetics of the GABA_A receptor-chloride channel complex of rat dorsal root ganglion neurons. The present paper reports the differential effects of HCH isomers on the GABA-induced chloride currents in three combinations of α, β and γ subunits of GABA_A receptor expressed in human embryonic kidney cells. When co-applied with GABA, γ-HCH strongly suppressed the peak amplitude of GABA-induced current, and δ-HCH strongly enhanced it in the α1β2γ2s, α1β2, α6β2γ2s combinations in a dose-dependent manner. There was little or no difference in the dose dependence of the effects between γ- and δ-HCH in any of the three subunit combinations. However, α- and β-HCH showed differential effects on GABA-induced chloride currents in the three subunit combinations tested. α-HCH showed enhancing effects on the peak current in α1β2γ2s, small enhancing effects on α1β2, and biphasic effects on α6β2γ2s subunit combinations. β-HCH had little or no effect on the peak current in α1β2γ2s and α1β2 combinations, but suppressed currents in the α6β2γ2s subunit combination in a dose-dependent manner. The differential actions of HCH isomers may produce variable effects on different regions of the nervous systems and in different species of animals.     

Selective effects of dieldrin on the GABAA receptor-channel subunits expressed in human embryonic kidney cells

We have recently demonstrated that the cyclodiene insecticide dieldrin modulates the kinetics of the GABA_A receptor-chloride channel complex of rat dorsal root ganglion neurons in a complex manner, causing both stimulatory and inhibitory effects. We now report that the differential effects of dieldrin on the GABA-induced chloride current of human embryonic kidney cells expressing three different combinations of α, β and γ subunits. The EC₅₀ values for GABA induction of current were estimated to be 9.8 μM for the ±1²2γ2s combination, 2.0 μM for the ±1β2 combination and 3.0 μM for the ±6²2γ2s combination. When co-applied with GABA, dieldrin exerted a dual effect, enhancement and suppression, on the GABA-induced chloride currents in the ±1²2γ2s and ±6²2γ2s combinations. However, only suppression was observed in the ±1β2 combination, indicating that the γ subunit is necessary for dieldrin's enhancing effect. Dieldrin was more efficacious in enhancing the current in the ±6²2γ2s combination than in the ±1²2γ2s combination, indicating some specific role of α subunits in the dieldrin enhancement of current. Dieldrin suppressed the GABA-induced current in a non-competitive manner, with an EC₅₀ value of 2.1 μM for ±1²2γ2s, 2.8 μM for ±1β2 and 1.0 μM for ±6²2γ2s combination. These results indicated that dieldrin suppression did not require specific subunit combinations among the three tested.     

Alzheimer's disease genetic mutation evokes ultrastructural alterations: Correlation to an intracellular Aβ deposition and the level of GSK-3β-P(Y216) phosphorylated form

Herein we demonstrate that PC12 cells, which overexpress human wild-type amyloid-β precursor protein (AβPPwt) or AβPP bearing double Swedish mutation (AβPPsw), reveal phenotype characteristic for Alzheimer's disease (AD). The examination of cell ultrastructure showed the presence of peptide aggregates within the cells, activation of endosomal–lysosomal system and extensive exocytosis. Furthermore, the autophagy induction was also characteristic hallmark of amyloid-β-induced cytotoxicity. Morphological changes were positively correlated with the extent of phosphorylated glycogen synthase kinase-3β (phospho-Tyr²¹⁶-GSK-3β, GSK-3β-P(Y216)). The activity of GSK-3β is believed to cause tau protein hyper-phosphorylation, increased amyloid-β production and local plaque-associated microglial-mediated inflammatory responses. All of them are symptomatic for AD. In our studies, the highly significant Y216 phosphorylation and over-expression of total GSK-3β were observed in AβPPsw-transfected PC12 cells. In addition, the immuocytochemical analysis showed co-localization of GSK-3β-P(Y216) and amyloid-β deposits. Thus, our data support a functional role of GSK-3β in AβPP processing, further implicating this kinase in the amyloid-β-dependent pathogenesis.     

Organizational strategy use in obsessive-compulsive disorder

It has been reported that the balance between T-helper type 1 (Th1) cytokines and T-helper type 2 (Th2) cytokines plays a role in psychiatric disorders such as bipolar disorder. The T-helper type 3 (Th3) cytokine, which transforming growth factor beta-1 (TGF-β1), has been shown to modulate the production of Th1 and Th2 cytokines. However, the role of TGF-β1 in bipolar disorder has not yet been explored. A total of 70 manic patients with bipolar disorder and 96 normal controls was recruited. The plasma levels of IFN-γ, IL-4, and TGF-β1 were studied at the time of admission and 8 weeks after mood stabilizer treatment. The detection rate and plasma concentrations of IFN-γ and IL-4 and the IFN-γ/TGF-β1 and IL-4/TGF-β1 ratios were significantly higher in patients than in controls, while the TGF-β1 level was significantly lower. The TGF-β1 level increased significantly after treatment and the IFN-γ/TGF-β1 and IL-4/TGF-β1 ratios returned to control values. TGF-β1 may play a role in the pathophysiology of bipolar disorder through the action of TGF-β1 in modulating the IL-4/TGF-β1 ratio.     

Costimulatory Effects of Interferon-γ and Interleukin-1β or Tumor Necrosis Factor α on the Synthesis of Aβ1-40 and Aβ1-42 by Human Astrocytes

Chronic inflammation and astrocytosis are characteristic histopathological features of Alzheimer's Disease (AD). Astrocytes are one of the predominant cell types in the brain. In AD they are activated and produce inflammatory components such as complement components, acute phase proteins, and cytokines. In this study we analyzed the effect of cytokines on the production of amyloid β (Aβ) in the astrocytoma cell line U373 and in primary human astrocytes isolated postmortem from healthy aged persons as well as from patients with AD. Astrocytes did not produce Aβ in the absence of stimuli or following stimulation with IL-1β, TNFα, IL-6, and TGF-β1. Neither did combinations of TNFα and IL-1β, IL-6 or TGF-β1, or the coadministration of IFNγ and IL-6 or TGF-β1 induce Aβ production. In contrast, pronounced production of Aβ1-40 and Aβ1-42 was observed when primary astrocytes or astrocytoma cells were stimulated with combinations of IFNγ and TNFα or IFNγ and IL-1β. Induction of Aβ production was accompanied by decreased glycosylation of APP as well as by increased secretion of APPsβ. Our results suggest that astrocytes may be an important source of Aβ in the presence of certain combinations of inflammatory cytokines. IFNγ in combination with TNFα or IL-1β seems to trigger Aβ production by supporting β-secretase cleavage of the immature APP molecule.     

负载As2O3壳聚糖纳米粒的药代动力学及其毒性观察

摘要 背景：As2O3作为抗肿瘤药物具有不同程度的不良反应,目前尚没有可以较好降低As2O3不良反应的方法。 目的：观察负载As2O3壳聚糖纳米粒在体内是否有缓释作用,是否可以延长药物作用时间,减低As2O3毒副作用。 方法：将20只SD大鼠以抽签法随机分为As2O3组和壳聚糖纳米粒-As2O3组进行药代动力学测定,于给药前和给药后不同时间点测定血液中砷浓度;将40只SD大鼠随机分为壳聚糖纳米粒组、壳聚糖纳米粒- As2O3组、As2O3组、生理盐水组进行毒理学检测,检测血浆中谷丙转氨酶、谷草转氨酶、肌酸激酶、肌酐、尿素氮水平,并结合苏木精-伊红染色形态学观察心、肝、肾组织形态学变化。 结果与结论：壳聚糖纳米粒- As2O3组较As2O3组半衰期明显延长(P < 0.05)。除肌酸激酶外,As2O3组其他各指标均高于其他各组(P < 0.05);而含相同浓度As2O3的壳聚糖纳米粒As2O3组与壳聚糖纳米粒组、生理盐水组各项指标差异无显著性意义(P > 0.05)。As2O3组大鼠肝脏和肾脏均有较明显的病理学改变,壳聚糖纳米粒- As2O3组未见明显形态学改变。4组大鼠心脏均未见明显形态学改变。说明负载As2O3壳聚糖纳米粒在体内有缓释作用,可以延长药物作用时间,减轻As2O3的毒副作用。 关键词：壳聚糖纳米粒;As2O3;负载As2O3壳聚糖纳米粒;药代动力学;药物毒理学 doi:10.3969/j.issn.1673-8225.2011.12.019     

THE EFFECT OF CHOLESTEROL OXIDATION PRODUCTS ON HUMAN PLATELET AGGREGATION

The cholesterol oxidation products (oxysterols) cholest-3,5-diene-7-one, cholestan-5, 6-epoxy-3β-ol (cholesterol 5-epoxide), cholestan-5β,6β-epoxy-3β-ol (cholesterol 5β-epoxide), cholest-5-ene-3β-ol-7-one (7-ketocholesterol), cholest-5-ene-3β,7-diol (7-hydroxycholesterol), cholestan-3β,5,6β-triol (choestane triol), and cholest-5-ene-3β,26-diol (27-hydroxycholesterol) potentiated platelet aggregation and increased thromboxane A₂ formation in platelets challenged with thrombin, ADP or collagen. These effects were observed at oxysterol concentrations in the range 5–100 μM. Cholesterol 5β-epoxide and 7-ketocholesterol increased the mobilization of ³H-arachidonic acid from prelabelled platelet phospholipids in response to thrombin and collagen.     

Temperature-dependent effect of zolpidem on the GABAA receptor-mediated response at recombinant human GABAA receptor subtypes

The effects of zolpidem on the two forms of recombinant human GABA_A receptors (α1β2γ2s and α3β2γ2s) at different temperatures were functionally investigated, using the whole-cell patch recording configuration. In both forms, zolpidem potentiated the response to GABA in a concentration-dependent manner. At 16°C, the apparent dissociation constant (K_D) values for the α1β2γ2s and α3β2γ2s forms were 3.7×10⁻⁸ and 5.6×10⁻⁷ M, respectively. When the temperature was increased to 36°C, the K_D values for the α1β2γ2s and α3β2γ2s forms were 2.1×10⁻⁷ and 1.5×10⁻⁶ M, respectively. Although the affinity ratio was reduced from 15.1 to 7.1-fold the selectivity of zolpidem for the α1β2γ2s still remained at 36°C.