P2X7受体表达在肺炎支原体肺炎早期诊断中的价值

目的：探讨肺炎支原体肺炎患儿中P2X7受体表达水平及与TNF-α、IL-1β水平的相关性。方法：选取300例肺炎支原体肺炎患儿作为研究组并选取健康儿童50例作为对照组,检测两组间P2X7受体、TNF-α和IL-1β表达水平的差异及相关性。结果：研究组P2X7受体表达水平为（151.4±14.3）ng/mL,显著高于对照组的（64.7±10.8）ng/mL（P<0.01）;研究组TNF-α表达水平为（1.72±0.14）ng/mL,显著高于对照组的（0.74±0.16）ng/mL（P<0.01）;研究组IL-1β表达水平为（0.89±0.14）ng/mL,显著高于对照组的（0.23±0.15）ng/mL（P<0.01）。肺炎支原体肺炎患儿中P2X7受体表达与TNF-α、IL-1β表达水平呈正相关（R2分别为0.533、0.666,P<0.05）。结论：肺炎支原体肺炎患儿中存在P2X7受体及TNF-α、IL-1β过度表达,且P2X7受体与TNF-α、IL-1β表达呈正相关。     

孟鲁司特对哮喘患者可溶性白介素-2受体及肿瘤坏死因子α表达水平的影响

目的探讨孟鲁司特通过影响SIL-2R和TNF-α的表达从而引发哮喘患者体内巨噬细胞和T细胞的抗炎症反应的机制。为临床选择用药提供指导。方法对48例轻中度稳定期哮喘患者进行8周孟鲁司特治疗试验,同时选取48列正常成人作为对照。试验前后均测定其1秒量(FEV1)值。同时测定血清中SIL-2R、TNF-α及嗜酸粒细胞阳离子蛋白水平。结果药物治疗后,嗜酸粒细胞阳离子蛋白量明显降低(32.1±11.8VS20.2±11.2,P<0.01),FEV1显著升高(78.9±10.9VS103±13.7,P<0.05)。蛋白水平和检测显示:患者治疗前SIL-2R表达量均显著高于治疗后(1034.77±175.18,773±230.19,P<0.05)及对照组(571.11±123.12;P<0.01)TNF-α表达水平治疗前亦显著高于治疗后与对照组(8.60±1.79,5.89±2.16,5.10±1.07;P<0.05)。Spearman相关分析法分析后显示孟鲁司特疗效与SIL-2RuL及TNF-αTE4有相关性。结论我们的研究显示,孟鲁司特通过降低SIL-2R与TNF-α的表达水平从而引发抗炎症反应。同时,SIL-2R与TNF-α的表达水平亦与孟鲁司特临床疗效密切相关,可作为选择此类药物的参考标志物。     

P2X7R/NLRP3信号通路在酒精性肝损伤中的作用

目的探讨嘌呤P2X7R及其介导的NLRP3炎性体信号通路在酒精诱导的肝损伤中的作用。方法采用NIAAA法建立小鼠急性酒精性肝损伤模型,将30只♂C57BL/6小鼠随机分为3组(n=10):对照组、模型组、P2X7特异性阻断剂A438079干预组,最后1周,分组进行以下处理,对照组和模型组:给予等剂量的生理盐水腹腔注射(每只约0.2 mL),每日1次;A438079组:根据小鼠体质量,腹腔注射200μmol·kg~(-1)的A438079(按7 g·L~(-1)配制A438079,每只约0.2mL),每日1次。最后1天清晨给予单次31.5%酒精溶液灌胃,剂量为10 mL·kg~(-1)。9 h后小鼠眼眶取血,测定谷丙转氨酶(ALT)、谷草转氨酶(AST)、胆固醇(TCHO)、甘油三酯(TG)。HE染色观察肝脏病理变化;免疫组化方法检测肝组织中P2X7R的表达;Western blot法检测肝组织中的P2X7、NLRP3、ASC、IL-1β、IL-18水平。结果与对照组相比,模型组ALT、AST、TG、TCHO含量明显增强,且肝组织损伤明显;与模型组相比,A438079组ALT、AST、TG、TCHO含量明显降低。与对照组相比,模型组P2X7、NLRP3、ASC、IL-1β、IL-18的表达明显升高;与模型组相比,A438079组P2X7、NLRP3、ASC、IL-1β、IL-18的表达水平明显降低。结论酒精诱导的肝损伤可能与P2X7R-NLRP3信号通路有关。     

癫痫患者的血清细胞因子水平变化及其临床意义

目的探讨白细胞介素-2（IL-2）、白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）对癫痫患者的神经免疫调节作用。方法采用双抗体夹心酶联免疫吸附试验（ELISA）检测45例癫痫患者血清中IL-2、IL-6和TNF-α的水平,并与年龄和性别相匹配的40例正常对照组进行比较。结果癫痫组血清IL-2、IL-6和TNF-α的水平均明显高于对照组（P〈0.01）。相关分析发现癫痫患者血清IL-2、IL-6与TNF-α水平之间呈显著正相关（r=0.49,P〈0.01;r=0.56,P〈0.01）。结论癫痫患者的免疫系统处于活化状态,细胞因子水平的失衡参与了癫痫的免疫病理过程。     

P2X7受体参与疼痛调节机制的研究进展

ATP门控离子通道受体P2X7受体近年来被广泛关注。P2X7受体受ATP及其衍生物激活后,诱导一系列反应如PANX1的激活和IL-1β的释放。阐明P2X7受体在疼痛中的作用及机制,为新型高效镇痛药物的研发提供思路。该文从炎性痛、神经病理痛、癌痛和吗啡镇痛耐受等方面阐述P2X7受体在疼痛中的研究进展,并探讨P2X7受体参与疼痛调节过程的可能机制。     

血清IL-2、IL-8及TNF-α水平在2型糖尿病肾病患者中的变化及意义

目的探讨2型糖尿病肾病患者血清IL-2、IL-8及TNF-α水平的变化及其在2型糖尿病肾病病理生理中的作用。方法2型糖尿病110例,其中糖尿病肾病72例,分为早期肾病（DN1）组35例,临床肾病（DN2）组37例,2型糖尿病无白蛋白尿组（DM）38例,健康对照组3O例;分别测定血清IL-2、IL-8及TNF-α水平。结果2型糖尿病肾病患者组血清中IL-2水平地低于正常人组及2型糖尿病非肾病组,而IL-8及TNF-α水平高于正常人组及2型糖尿病非肾病组,且随糖尿病肾病的加重差异更为显著;相关分析发现,IL-2与尿白蛋白排泄率（UAER）呈负相关,IL-8及TNF-α水平与UAER呈正相关。结论在2型糖尿病肾病的病理生理过程中,IL-2、IL-8及及TNF-α水平其发生的重要炎症介质,在2型糖尿病肾病的发生及发展过程中可能起重要作用。     

P2X7受体参与神经炎症机制及潜在药物靶点研究进展

嘌呤能P2 X7受体是非选择性ATP门控阳离子通道受体,在中枢神经系统中主要在小胶质细胞上大量表达,P2X7受体表达失调和过度激活参与许多中枢神经系统疾病的病理过程.近年来研究发现,脑卒中后脑损伤程度与P2X7受体数量及活化程度密切相关,P2X7受体可能成为脑卒中后多种急慢性神经炎症的重要干预靶点.本文对P2 X7受体...     

离子通道型嘌呤受体P2X4在中枢神经系统中的作用及其作为神经系统疾病干预靶标的前景

ATP除了作为细胞内能量来源物质,也可充当细胞外信号分子激活离子通道型的嘌呤受体P2X受体和G蛋白偶联受体型的P2Y受体。P2X4亚型是一类在中枢神经系统(CNS)广泛表达的P2X受体,通过感受胞外ATP的刺激而参与CNS生理过程,包括慢性痛、乙醇滥用、癫痫、阿尔茨海默病、帕金森病和卒中缺血等。因此,靶向P2X4受体的小分子拮抗剂或增强剂因为其药用前景而受到广泛关注。本文总结了P2X4受体在CNS中相关研究的进展情况,概述了近年来新发现的P2X4受体高选择性拮抗剂。     

系统性红斑狼疮患者外周血IL-17、TNF-α和INF-γ表达水平及临床意义

目的 探讨系统性红斑狼疮(SLE)患者外周血白介素-17(IL-17)、干扰素-γ(INF-γ)和肿瘤坏死因子-α(TNF-α)表达水平及其临床意义.方法 选取2012年7月-2015年12月治疗的80例SLE患者及同期40例正常健康体检人群为研究对象,分别记为SLE组和健康组,再根据SLE活动指数评分将SLE分为活动期组和非活动期组,每组40例.测定并比较各组IL-17、INF-γ和TNF-α的表达水平,分析其临床意义.结果 活动期组、非活动期组、SLE组的IL-17、INF-γ和TNF-α水平均高于健康组(P＜0.05).活动期组IL-17、INF-γ和TNF-α水平高于非活动期组(P＜0.05).结论 IL-17、INF-γ和TNF-α在SLE患者外周血中存在高表达,且随着病情活动度增强而升高.     

P2X2/3受体在大鼠眶下神经慢性压迫性损伤中表达水平的动态变化

目的探讨P2X2/3受体在三叉神经痛(TN)模型大鼠三叉神经节(TG)神经元内表达的动态变化及其作用。方法采用行为学观察、HE染色评价慢性压迫性损伤大鼠眶下神经模型造模结果,免疫组化方法测定每组TG神经元不同时期P2X2/3的表达情况。结果正常对照组P2X2/3在中、小直径神经元中有阳性表达;P2X2/3的表达量在手术侧组各时间段均明显高于正常对照组(P<0.05);假手术组术后早期,P2X2/3表达量明显高于对照组(P<0.05),晚期两者无明显差异;手术对侧组与假手术组相比,P2X2/3表达量在术后早期无明显差异,术后晚期明显增多(P<0.05)。结论TG神经元中P2X2/3参与了TN的诱发、痛觉传递过程,其中P2X2主要参与急性痛或炎性痛,而P2X3对急性痛及痛疼的维持都起重要作用;手术侧的痛觉神经传导对对侧TG产生了影响。     

Mammalian P2X7 receptor pharmacology: comparison of recombinant mouse,rat and human P2X7 receptors

Background and purpose:

Acute activation of P2X7 receptors rapidly opens a non-selective cation channel. Sustained P2X7 receptor activation leads to the formation of cytolytic pores, mediated by downstream recruitment of hemichannels to the cell surface. Species- and single-nucleotide polymorphism-mediated differences in P2X7 receptor activation have been reported that complicate understanding of the physiological role of P2X7 receptors. Studies were conducted to determine pharmacological differences between human, rat and mouse P2X7 receptors.

Experimental approach:

Receptor-mediated changes in calcium influx and Yo-Pro uptake were compared between recombinant mouse, rat and human P2X7 receptors. For mouse P2X7 receptors, wild-type (BALB/c) and a reported loss of function (C57BL/6) P2X7 receptor were also compared.

Key results:

BzATP [2,3-O-(4-benzoylbenzoyl)-ATP] was more potent than ATP in stimulating calcium influx and Yo-Pro uptake at rat, human, BALB/c and C57BL/6 mouse P2X7 receptors. Two selective P2X7 receptor antagonists, A-740003 and A-438079, potently blocked P2X7 receptor activation across mammalian species. Several reported P2X1 receptor antagonists [e.g. MRS 2159 (4-[(4-formyl-5-hydroxy-6-methyl-3-[(phosphonooxy)methyl}-2-pyridinyl)azo]-benzoic acid), PPNDS and NF279] blocked P2X7 receptors. NF279 fully blocked human P2X7 receptors, but only partially blocked BALB/c P2X7 receptors and was inactive at C57BL/6 P2X7 receptors.

Conclusions and implications:

These data provide new insights into P2X7 receptor antagonist pharmacology across mammalian species. P2X7 receptor pharmacology in a widely used knockout background mouse strain (C57BL/6) was similar to wild-type mouse P2X7 receptors. Several structurally novel, selective and competitive P2X7 receptor antagonists show less species differences compared with earlier non-selective antagonists.     

糖尿病酮症酸中毒患者血清中炎性细胞因子水平增高的临床意义

目的探讨糖尿病酮症酸中毒患者血清白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、C反应蛋白(CRP)水平变化与病情加重的关系。方法选取46例糖尿病酮症酸中毒患者,其中合并感染者28例,非感染者18例,同时选取同期非酮症糖尿病患者32例作为对照组。留取清晨空腹静脉血,检查血常规同时检测各组患者空腹血糖(FPG)、糖化血红蛋白(Hb A1c)生化指标的变化情况;收集患者尿液检测尿酮体变化;ELISA法检测血清IL-6,TNF-α,CRP水平。结果 DKA感染组和非感染组血清IL-6、TNF-α、CRP水平均明显高于对照组,差异具有统计学意义(均P<0.05)。与DKA非感染组相比,DKA感染组IL-6、TNF-α、CRP水平也有明显升高,差异也具有统计学意义(P<0.05)。DKA合并感染患者空腹血糖与IL-6、TNF-α、CRP均成正相关(r分别为0.534、0.652、0.493,均P<0.01),而DKA合并感染患者CRP与IL-6、TNF-α亦成正相关(r分别为0.678、0.624,均P<0.01)。结论糖尿病酮症酸中毒患者的血清IL-6、TNF-α、CRP水平变化明显升高,与疾病严重程度有一定的关系。     

IL-1、IL-6、TNF-在肩峰撞击综合征患者肩峰下滑囊的表达及意义

目的探讨IL-1、IL-6、TNF-在肩峰撞击综合征患者肩峰下滑囊的表达及临床意义。方法 26例患者被列入实验,分为实验组和对照组,实验组肩峰撞撞击综合征患者16例,对照组非肩峰撞撞击综合征肩部损伤10例,实验组采用关节镜微创术式取SAB标本,对照组采取开放手术方式取得标本,采用ELISA法测SAB中IL-1、IL-6、TNF-含量。结果与空白组相比,实验组的IL-1、IL-6、TNF-在肩峰下滑囊的表达水平明显升高（P〈0.05）有统计学意义。结论肩峰下撞击综合征导致的肩峰下滑囊炎的慢性炎症反应是导致患者肩痛及功能障碍的重要病理基础,在肩峰撞击综合征患者行肩峰下成形术时应广泛、彻底地切除肩峰下滑囊。     

Synthesis,SAR, and Pharmacological Characterization of Brain Penetrant P2X7 Receptor Antagonists

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再生障碍性贫血患者血清IL-8、TNF-α水平的检测及其意义

目的 探讨再生障碍性贫血(AA)患者血清IL-8、TNF-α的变化及其意义.方法 应用酶联免疫吸附试验检测45例AA患者及30名健康人血清中IL-8、TNF-α水平.结果 血清IL-8、TNF-α水平AA组及急性AA(AAA)亚组均明显高于对照组(P＜0.05),慢性AA(CAA)亚组与对照组差异无统计学意义(P＞0.05);AAA亚组明显高于CAA亚组(P＜0.05).AA组血清IL-8与TNF-α水平呈正相关(r=0.682,P＜0.05).结论 AA患者存在细胞免疫功能紊乱,IL-8和TNF-α过量分泌可能在AA特别是在AAA的发病中起一定的作用.     

Pharmacological characterization of a novel centrally permeable P2X7 receptor antagonist: JNJ-47965567

BACKGROUND AND PURPOSE

An increasing body of evidence suggests that the purinergic receptor P2X, ligand-gated ion channel, 7 (P2X7) in the CNS may play a key role in neuropsychiatry, neurodegeneration and chronic pain. In this study, we characterized JNJ-47965567, a centrally permeable, high-affinity, selective P2X7 antagonist.

EXPERIMENTAL APPROACH

We have used a combination of in vitro assays (calcium flux, radioligand binding, electrophysiology, IL-1β release) in both recombinant and native systems. Target engagement of JNJ-47965567 was demonstrated by ex vivo receptor binding autoradiography and in vivo blockade of Bz-ATP induced IL-1β release in the rat brain. Finally, the efficacy of JNJ-47965567 was tested in standard models of depression, mania and neuropathic pain.

KEY RESULTS

JNJ-47965567 is potent high affinity (pK_i 7.9 ± 0.07), selective human P2X7 antagonist, with no significant observed speciation. In native systems, the potency of the compound to attenuate IL-1β release was 6.7 ± 0.07 (human blood), 7.5 ± 0.07 (human monocytes) and 7.1 ± 0.1 (rat microglia). JNJ-47965567 exhibited target engagement in rat brain, with a brain EC₅₀ of 78 ± 19 ng·mL⁻¹ (P2X7 receptor autoradiography) and functional block of Bz-ATP induced IL-1β release. JNJ-47965567 (30 mg·kg⁻¹) attenuated amphetamine-induced hyperactivity and exhibited modest, yet significant efficacy in the rat model of neuropathic pain. No efficacy was observed in forced swim test.

Conclusion and Implications

JNJ-47965567 is centrally permeable, high affinity P2X7 antagonist that can be used to probe the role of central P2X7 in rodent models of CNS pathophysiology.     

Positive allosteric modulation by ivermectin of human but not murine P2X7 receptors

BACKGROUND AND PURPOSE

In mammalian cells, the anti-parasitic drug ivermectin is known as a positive allosteric modulator of the ATP-activated ion channel P2X4 and is used to discriminate between P2X4- and P2X7-mediated cellular responses. In this paper we provide evidence that the reported isoform selectivity of ivermectin is a species-specific phenomenon.

EXPERIMENTAL APPROACH

Complementary electrophysiological and fluorometric methods were applied to evaluate the effect of ivermectin on recombinantly expressed and on native P2X7 receptors. A biophysical characterization of ionic currents and of the pore dilation properties is provided.

KEY RESULTS

Unexpectedly, ivermectin potentiated currents in human monocyte-derived macrophages that endogenously express hP2X7 receptors. Likewise, currents and [Ca²⁺]_i influx through recombinant human (hP2X7) receptors were potently enhanced by ivermectin at submaximal or saturating ATP concentrations. Since intracellular ivermectin did not mimic or prevent its activity when applied to the bath solution, the binding site of ivermectin on hP2X7 receptors appears to be accessible from the extracellular side. In contrast to currents through P2X4 receptors, ivermectin did not cause a delay in hP2X7 current decay upon ATP removal. Interestingly, NMDG⁺ permeability and Yo-Pro-1 uptake were not affected by ivermectin. On rat or mouse P2X7 receptors, ivermectin was only poorly effective, suggesting a species-specific mode of action.

CONCLUSIONS AND IMPLICATIONS

The data indicate a previously unrecognized species-specific modulation of human P2X7 receptors by ivermectin that should be considered when using this cell-biological tool in human cells and tissues.     

Ionic effects on human recombinant P2X7 receptor function

The actions of monovalent and divalent ions on the P2X₇ receptor have been assessed by measuring their effect on responses to the P2 receptor agonist, 2’- and 3’-O-(4-benzoyl-benzoyl)-ATP (DbATP), in HEK293 cells expressing the human recombinant P2X₇ receptor. In these cells, DbATP increased the cellular accumulation of the DNA binding, fluorescent dye, YO-PRO-1. The potency of DbATP to elicit this effect was decreased by both calcium and magnesium ions. In addition, when the pH was increased above 8 or reduced below 6.5, the potency of DbATP was less than obtained at pH 7.5. Monovalent ions also affected the P2X₇ receptor such that the potency of DbATP was 19-fold higher in NaCl-free buffer containing 280 mM sucrose (pEC₅₀=6.48) than in 140 mM NaCl containing buffer (pEC₅₀=5.19). Monovalent cations differentially affected the potency of DbATP. Thus, when the chloride concentration was maintained at 140 mM, pEC₅₀ values for DbATP were 6.14, 5.87 and 5.19 when the counter cation was 140 mM choline, potassium or sodium, respectively. Monovalent anions also differentially affected the potency of DbATP and in the presence of 140 mM sodium ions, pEC₅₀ values for DbATP were 6.14, 6.07, 5.19 and 4.53, respectively, when the counter anion was 140 mM aspartate, glutamate, chloride or iodide. The inhibitory effect of monovalent anions on P2X₇ receptor function was also observed in electrophysiological studies. Thus in sodium glutamate containing buffer the potency of DbATP (pEC₅₀=5.55) was approximately 22-fold higher than in NaCl containing buffer (pEC₅₀=4.20). This study has demonstrated that P2X₇ receptor function can be markedly affected by a wide range of ions and that physiological concentrations of sodium and chloride ions, as well as divalent cations, contribute to the low potency of ATP as an agonist at this receptor. Received: 27 July 1998 / Accepted: 18 November 1998