vMIP对细胞膜上趋化因子受体亲和力的结合特性研究

目的通过病毒巨噬细胞炎性蛋白(vMIP)对外周血中单个核细胞(PBMC)膜上趋化因子受体结合的比较性研究,阐明vMIP与受体的结合能力.方法通过放射性配体受体结合实验,利用饱和实验、动力学实验和特异性分析,鉴定vMIP的受体结合能力.结果 vMIP与人、食蟹猴PBMC膜上受体结合的Kd分别为11.1、14.3 nmol/L,对趋化因子受体CCR5和CXCR4竞争结合的IC50分别为3.4、4.5 nmoL/L.结论提示了vMIP与膜上受体有较高的亲和力,并对CCR5和CXCR4具有高度特异性,可能对防治炎症及HIV-1感染等具有重要意义.     

病毒趋化因子vMIP封闭人趋化因子受体作用及其应用研究

来源于人疱疹病毒8的巨噬细胞炎性蛋白(viralmacrophage inflammatory proteins,vMIP)能广谱结合CCR1、CCR3、CCR5和CXCR4等多种趋化因子受体,但不激活受体,可作为这些受体的封闭剂。由于CCR5、CXCR4是HIV在人体内感染靶细胞的主要共受体,vMIP作为CCR5、CXCR4的抑制因子可拮抗H     

病毒巨噬细胞炎性蛋白与趋化因子受体结合的效应分析

目的: 探讨人疱疹病毒8K6基因编码的产物病毒巨噬细胞炎性蛋白(viral macrophage inflamm atory protein,vMIP)是否具有结合趋化因子受体以及趋化作用。方法: 受体配体交联试验检测vMIP与受体结合能力。趋化实验及细胞内钙流检测判断vMIP的生物学活性。结果: vMIP可与外周血单个核细胞(PBMCs)膜上的趋化因子受体结合,抑制hMIP-1α对PBMC的趋化能力,EC₅₀为3.39 ng/ml。其本身只有较弱的趋化能力。钙流实验证实vM IP轻度升高胞内钙离子浓度,但可明显抑制hMIP-1α所引起的胞内钙离子高峰。结论: 重组vMIP与hMIP-1α受体(CCR5)结合后,可有效的阻断人源性趋化因子的结合与信号传导,但其本身对细胞未有明显的激活作用,因此可作为趋化因子受体的天然阻断剂,可用于免疫移植中的排斥反应或HIV-1病毒感染等。     

重组趋化因子vMIPⅡ对小鼠异体移植皮肤存活时间的影响

目的　观察纯化的重组vMIPⅡ蛋白对小鼠异体皮肤移植免疫排斥反应是否有抑制作用及能否延长移植皮肤的存活时间。方法　pET3 2a为vMIPⅡ克隆基因的表达载体 ,在大肠杆菌中诱导表达出硫氧还蛋白 vMIPⅡ的融合蛋白 (2 6× 10 3 )。经金属离子亲和吸附、肠激酶消化以游离vMIPⅡ、阳离子交换层析等步骤纯化目的蛋白。用体外配体结合实验证实重组vMIPⅡ与趋化因子受体CCR5的结合能力。纯化的vMIPⅡ经尾静脉给药异体皮肤移植小鼠 (昆明鼠 /Balb/c)连续 14d。结果　经纯化可获得高纯度目的蛋白。vMIPⅡ与趋化因子受体CCR5结合的解离常数 (Kd)为 11 5 6± 1 98nmol/L。给药vMIPⅡ的异体皮肤移植小鼠组移植皮存活时间较未注射对照组延长至术后 7天以上。结论　纯化的重组vMIPⅡ对小鼠异体皮肤移植免疫排斥反应有明显的抑制作用 ,能显著延长移植皮的存活时间     

趋化因子受体CXCR4的结构与功能

目的 认识趋化因子受体CXCR4之结构与功能的关系。方法 分别建立野生型趋化因子受体CXCR4及CXCR2、5个CXCR4/CXCR2嵌合受体和2个CXCR4突变受体的CHO稳定表达细胞株,以配体-受体结合试验,细胞微生理监测术、体外细胞-细胞融合实验为手段观察各变异受体与重组人SDF-1β的结合能力、在受刺激后的信号转导能力,以及作为HIV-1辅助受体的能力。结果 有4个变异受体（2444a,4442,4222,CXCR4-Tr）保持了程度不同的具有辅助受体功能。结论 CXCR4以多个结构区域参与与SDF-1β的相互作用。其N端胞外区足以及具有与SDF-1β的高亲和性结合能力。第三环链对CXCR4的结合能力也具有其特定的贡献。CXCR4的信号传导不仅需要保守结构DRY盒,而且还需要贯穿整个分子的7个跨膜区域和受配体刺激后形成并维持一定的构象。CXCR4的辅助受体功能结构域与配体结合结构域间存在交叉重叠。     

梅毒血清固定患者趋化因子受体的研究

目的研究梅毒血清固定患者趋化因子受体CCR3、CCR5、CXCR4、CXCR3水平,探讨梅毒患者免疫功能的变化。方法检测梅毒血清固定患者（血清固定组）35例外周血趋化因子受体CCR3、CCR5、CXCR4、CXCR3水平,并以规范治疗后梅毒血清反应素试验血清转阴者梅毒患者（转阴组）40例及健康体检者（健康组）35例分别作为对照。比较3组外周血趋化因子受体水平。结果梅毒血清固定患者CCR3表达高于健康组及转阴组,CCR5、CXCR3表达低于健康组及转阴组,差异均有统计学意义（P＜0.01）。转阴组与健康组比较,趋化因子受体表达差异无统计学意义（P＞0.05）。结论趋化因子受体在梅毒血清固定患者的发病、发展中可能起着重要作用。     

CXCR4在前列腺癌组织中的表达与意义

目的:探讨趋化因子受体在前列腺癌组织中的表达及其意义.方法:采用酶标记免疫组织化学方法检测45例前列腺癌及10例前列腺增生组织中趋化因子受体CCR1、CCR3、CXCR4和CCR5的表达情况,应用ELISA双抗体夹心方法检测相应患者血清中趋化因子SDF-1的含量,并应用免疫发光方法检测相应患者血清中前列腺特异性抗原PSA的含量.结果:在前列腺癌组织上检测到趋化因子受体CXCR4的表达(表达率55.5%),PSA＞20 ng/ml与PSA＜20 ng/ml的前列腺癌组织上CXCR4的表达率分别为61.2%和42.8%,而前列腺增生组织中未发现这4种趋化因子受体的表达;与前列腺增生患者相比,在前列腺癌患者血清中SDF-1含量明显升高[(567.9±90.73)vs(169.1±46.01)pg/ml,P＜0.01].结论:在前列腺癌组织上发现有趋化因子受体CXCR4的表达,其表达可能在前列腺癌的发生、发展和转移中起重要作用.     

CCR5,CXCR3,CXCR6和CCR7在丙肝患者肝外周血NK和NKT淋巴细胞的表达及其意义

[目的]比较趋化因子受体CCR5,CXCR3,CXCR6和CCR7在丙肝患者肝外周血NK和NKT淋巴细胞上的表达及其意义,了解其与肝组织学炎症反应的关系.[方法]用荧光标记抗趋化因子受体的单克隆抗体对肝及外周血NK和NKT细胞表面的趋化因子受体染色后,用9色11参数流式细胞仪KSRⅡ检测分析.[结果]肝组织中NKT细胞比例18.2±5.8高于外周血的3.7±2.9,P＜0.01;肝组织NK细胞比例7.8±3.3低于外周血中15.1±10.1,P＜0.01.肝组织CCR5 ,CXCR3 或/和CXCR 的NK和NKT细胞频数高于外周血,P＜0.001,CCR7 NK和NKT细胞频数低于外周血,P＜0.001;肝组织学炎症明显组表达趋化因子受体CCR5,CXCR3或CXCR6的NK和NKT细胞频数高于炎症轻微组.[结论]是趋化因子受体CCR5,CXCR3和CXCR6而不是CCR7介导NK和NKT细胞向肝迁徙定植,它们并可能参与肝炎症的病理免疫学反应过程.     

不同期别梅毒患者外周血CD4＋T细胞表面趋化因子受体的研究

目的 研究不同期别梅毒患者趋化因子受体CCR3、CCR5、CXCR4、CXCR3水平,探讨梅毒患者免疫功能的变化.方法 检测不同期别梅毒患者外周血趋化因子受体CCR3、CCR5、CXCR4、CXCR3水平,以健康体检者作为正常对照组.结果 二、三期梅毒患者与正常对照组及一期梅毒患者比较,外周血CCR3水平增高,CCR5水平降低（P＜0.05或0.01）;三期梅毒患者与二期患者比较,外周血CCR3水平增高（P＜0.05）;二、三期梅毒患者治疗后与治疗前比较,外周血CCR3水平降低（P＜0.05）;二期梅毒患者治疗后与治疗前比较,外周血CCR5水平升高（P＜0.05）.结论 趋化因子受体在梅毒的发病和发展中可能起着重要作用.     

尖锐湿疣患者疣体组织趋化因子受体的表达及意义

目的:探讨趋化因子MIP-1α、MIP-1β和RANTES及趋化因子受体CCR1、CCR3、CCR5和CXCR4在尖锐湿疣发病机制中的作用.方法: 采用酶标记免疫组织化学方法检测78例尖锐湿疣患者疣体组织细胞上CCR1、CCR3、CCR5和CXCR4的表达,用ELISA双抗体夹心法测定78例尖锐湿疣患者血清中MIP-1α、MIP-1β和RANTES的含量.结果:在48例尖锐湿疣复发者中,疣体组织细胞表达CCR5的有19例(19/48,39.5%)、表达CCR3的11例(11/48,22.9%),5例(5/48,10.4%)同时表达这两种受体;所有待测患者疣体组织细胞中未发现有CCR1和CXCR4的阳性表达;尖锐湿疣患者MIP-1α和MIP-1β水平高于正常人组(P<0.01),两组的RANTES水平则无显著性差异.结论:尖锐湿疣患者疣体细胞趋化因子受体CCR3和CCR5的表达及MIP-1α和MIP-1β的水平升高可能与患者免疫功能低下及病毒逃避机体免疫反应有关,对于揭示尖锐湿疣的复发及病毒的免疫逃逸机制可能具有一定的意义.     

Oncol2012: 腹腔镜下CO2气腹对结直肠癌细胞表达趋化因子受体CXCR4及CCR7的影响

目的 探讨不同压强、不同作用时间下持续性CO2气腹对结直肠癌细胞表达趋化因子受体的影响。 方法 建立体外气腹模型,选用人结直肠癌细胞株SW480,分别在6、9、12、15mmHg四种不同压强CO2气体下作用1h、2h及4h后,放在与无气腹组(37℃、5%CO2常规培养)相同的环境中分别培养0、24、48、72h后,使用免疫细胞化学法及RT-PCR检测趋化因子受体CXCR4、CCR7的表达。 结果 免疫细胞化学法检测结果显示SW480在相同作用时间下,经6、9、12、15mmHg压强的持续CO2气腹处理后,CXCR4表达下降;经12、15mmHg压强的持续CO2气腹处理后,CCR7表达下降,与无气腹组表达水平的差异均有统计学意义(P＜0.05),上述趋化因子受体表达在气腹处理后常规培养24、48h均增至无气腹组水平(P＞0.05)。CXCR4及CCR7的表达在相同作用时间下,随着压强增高,其表达量逐渐降低;在相同压强下,随着作用时间延长,其表达量无明显差异。 RT-PCR结果显示SW480在相同作用时间下经6、9、12、15mmHg压强的持续CO2气腹处理后,CXCR4 mRNA、CCR7 mRNA表达下降,与无气腹组表达水平的差异均有统计学意义(P＜0.05),上述趋化因子受体的mRNA表达在气腹处理后常规培养48h均增至无气腹组水平(P＞0.05)。CXCR4及CCR7的mRNA表达在相同作用时间下,随着压强增高,其表达量无明显差异;在相同压强下,随着作用时间延长,其表达量也无明显差异。 结论 不同压强CO2气腹能对结直肠癌细胞表面的趋化因子受体表达产生一过性影响,随CO2气腹压强的增高,可抑制趋化因子受体的表达。不同作用时间CO2气腹对结直肠癌细胞表面趋化因子受体的表达无明显影响。     

Expression of chemokine receptors CCR3,CCR5 and CXCR3 on CD4+ T cells in CBA/JxDBA/2 mouse model,selectively induced by IL-4 and IL-10,regulates the embryo resorption rate

Background Chemokines and their receptors have been a research focus in transplantation immunology.Chemokines and their receptors play a role in lymphocyte recruitment and differentiation process.This study aimed to observe whether IL-4 and IL-10 may regulate the expression of chemokine receptors CCR3,CCR5 and CXCR3 on CD4+ T cells in CBA/JxDBA/2 mouse model and to explore the role of CCR3,CCR5,CXCR3 in immune tolerance in pregnancy.Methods The mouse model of spontaneous abortion (CBA/JxDBA/2) and the normal pregnant mouse model (CBA/JxBALB/c) were used.CBA/JxDBA/2 mice were injected with IL-4 (CBA/JxDBA/2-1L-4),IL-4 and IL-10 (CBA/JxDBA/2-1L-4+IL-10),or normal saline (CBA/JxDBA/2-NS) as a control.The expression of CCR3,CCR5 and CXCR3 on CD4+ T cells from mouse peripheral blood was measured by the double-labelled FCM method,and the embryo resorption rate was also examined.Results The embryo resorption rate in the CBA/JxDBA/2 group without any treatment was significantly higher than that in the CBA/JxBALB/c group (17.9% vs 3.7%,P＜0.01).The embryo resorption rate in the CBA/JxDBA/2 group immunized with IL-4 or IL-4 together with IL-10 was significantly decreased,compared with that in the control and NS groups respectively.CCR3 expression on CD4+ T cells in the CBA/JxDBA/2 group without any treatment was significantly lower than that in the CBA/JxBALB/c group (0.3738±0.3575 vs 1.2190±0.2772,P＜0.01 );both CCR5 (3.0900±1.5603 vs 1.2390±0.6361,P ＜0.01)and CXCR3 (2.4715±0.9074 vs 0.9200±0.5585,P ＜0.01 ) expressions on CD4+ T cells of the CBA/JxDBA/2 group without any treatment were significantly higher than those of the CBA/JxBALB/c group.Significant up-regulation of CCR3 and down-regulation of CXCR3 were found in the CBA/JxDBA/2 group treated with IL-4 (CCR3:2.0360±0.6944,CXCR3:1.3510±0.5263,P ＜0.01) or IL-4 and IL-10 (CCR3:1.8160±1.0947,CXCR3:1.0940±0.7168,P＜0.01).Because of the CCR5,IL-4 and IL-10 (1.9400±0.8504 vs 3.0900±1.5603,P ＜0.05),but IL-4 alone (2.5310±1.3595 vs 3.0900±1.5603,P ＞0.05)treatment significantly decreased the expression of CCR5 in CBA/JxDBA/2.Conclusions The abnormal expression of CCR3,CCR5 and CXCR3 on CD4+ T cells may play an important role in the pathogenesis of spontaneous abortion.The pregnancy immune tolerance may be induced through selective induction of CCR3,CCR5 and CXCR3 expressions by IL-4 together with IL-10.     

Relationship between expression of chemokine receptors CCR3, CCR5 and CXCR3 on CD4+ T cells and spontaneous abortion in mice

Background Previous studies have shown that local immune cells in the feto-maternal interface are recruited from peripheral blood, and that chemokines and their receptors play an initial and key role in this recruitment process. In this study, we aimed to determine whether spontaneous abortion is associated with the expression of chemokine receptors CCR3, CCR5, and CXCR3 on CD4^＋ T cells.
Methods Peripheral blood, spleen, and thymus were collected from the spontaneous abortion mouse model CBA/JxDBA/2 （SA group, n=14）, the normal pregnant mouse model CBA/JxBALB/c （NP group, n=13）, and normal non-pregnant CBA/J mice （NNP group, n=11）. The number of chemokine receptors CCR3, CCR5, and CXCR3 expressed on CD4^＋ T cells was measured by double-label flow cytometry （FCM） method.
Results In peripheral blood, the SA group had significantly lower CCR3 expression （P 〈0.01） and higher CCR5 and CXCR3 expression （P 〈0.01） on CD4^＋ T cells than did the NP group. But comparing these chemokines between the SA and NNP groups, there was no significant difference （P 〉0.05）. In spleen, the SA group expressed significantly lower CCR3 expression （P 〈0.01） and higher CCR5 and CXCR3 expression （P 〈0.05） on CD4^＋ T cells than did the NP group. When compared with the NNP group, the SA group had significantly higher CCR3 expression （P 〈0.01）, but was not statistically different with regards to the other two chemokines （P 〉0.05）. In thymus, the SA group had significantly lower CCR3 expression （P 〈0.05） and higher CXCR3 expression （P 〈0.05） on CD4^＋ T cells than the NP group, with no significant difference in CCR5 expression （P 〉0.05）. Compared with the NNP group, the SA group had higher CCR3 expression （P 〈0.01）, but there was no statistical difference in CXCR3 and CCR5 expression （P 〉0.05） between the two groups.
Conclusion The abnormal expression of CCR3, CCR5 and CXCR3 on CD4^＋ T cells may play an important role in the pathogenesis of spontaneous abortion.     

高效抗逆转录病毒治疗对T细胞亚群上第二受体影响的研究

目的；研究高效抗逆转录病毒治疗(HAART)后T细胞亚群上第二受体的变化及其意义。方法：采用流式细胞仪检测CD4^ 、CD8^-、CD4^ CD45RA^ 以及CD4^ CD45RO^ 细胞表面CCR5及CXCR4的表达水平。结果：HIV感染后，第二受体在CD4和CD8以及记忆型和处女型CD4T细胞亚群上的表达水平，与正常对照没有显著性差异。HAART治疗仪能在一定程度上改变细胞表面的第二受体水平。结论：第二受体参与了HIV的致病过程，B期之前开始抗病毒治疗将有较好的疗效。     

Expression of chemokine receptors CCR3, CCR5 and CXCR3 on CD4^＋ T cells in CBA/J×DBA/2 mouse model,selectively induced by IL-4 and IL-10, regulates the embryo resorption rate

Background Chemokines and their receptors have been a research focus in transplantation immunology. Chemokines and their receptors play a role in lymphocyte recruitment and differentiation process. This study aimed to observe whether IL-4 and IL-10 may regulate the expression of chemokine receptors CCR3, CCR5 and CXCR3 on CD4^＋ T cells in CBA/J×DBA/2 mouse model and to explore the role of CCR3, CCR5, CXCR3 in immune tolerance in pregnancy. Methods The mouse model of spontaneous abortion （CBA/J×DBA/2） and the normal pregnant mouse model （CBA/J×BALB/c） were used. CBA/J×DBA/2 mice were injected with IL-4 （CBA/J×DBA/2-IL-4）, IL-4 and IL-10 （CBA/J×DBA/2-IL-4＋IL-10）, or normal saline （CBA/J×DBA/2-NS） as a control. The expression of CCR3, CCR5 and CXCR3 on CD4^＋ T cells from mouse peripheral blood was measured by the double-labelled FCM method, and the embryo resorption rate was also examined. Results The embryo resorption rate in the CBA/J×DBA/2 group without any treatment was significantly higher than that in the CBA/J×BALB/c group （17.9% vs 3.7%, P 〈0.01）. The embryo resorption rate in the CBA/J×DBA/2 group immunized with IL-4 or IL-4 together with IL-10 was significantly decreased, compared with that in the control and NS groups respectively. CCR3 expression on CD4^＋ T cells in the CBA/J×DBA/2 group without any treatment was significantly lower than that in the CBA/J×BALB/c group （0.3738±0.3575 vs 1.2190±0.2772, P 〈0.01）; both CCR5 （3.0900±1.5603 vs 1.2390±0.6361, P〈0.01） and CXCR3 （2.4715±0.9074 vs 0.9200±0.5585, P 〈0.01） expressions on CD4^＋ T cells of the CBA/J×DBA/2 group without any treatment were significantly higher than those of the CBA/J×BALB/c group. Significant up-regulation of CCR3 and down-regulation of CXCR3 were found in the CBA/J×DBA/2 group treated with IL-4 （CCR3： 2.0360±0.6944, CXCR3： 1.3510±0.5263, P〈0.01） or IL-4 and IL-10 （CCR3： 1.8160±1.0947, CXCR3：1.0940±0.7168, P〈0.01）. Because of the CCR5, IL-4 and IL-10 （1.9400±0.8504 vs 3.0900±1.5603, P 〈0.05）, but IL-4 alone （2.5310±1.3595 vs 3.0900±1.5603, P 〉0.05） treatment significantly decreased the expression of CCR5 in CBA/J×DBA/2. Conclusions The abnormal expression of CCR3, CCR5 and CXCR3 on CD4^＋ T cells may play an important role in the pathogenesis of spontaneous abortion. The pregnancy immune tolerance may be induced through selective induction of CCR3, CCR5 and CXCR3 expressions by IL-4 together with IL-10.     

Th1/Th2趋化因子及其受体在大疱性类天疱疮皮损中的表达

目的 探讨 Th1 趋化因子 MIG/CXCL9、I-TAC/CX-CL11与其受体 CXCR3,Th2 趋化因子 TARC/CCL17、MDC/CCL22及其受体CCR4在大疱性类天疱疮( BP)患者发病中的作用.方法 应用免疫组化及光密度(OD)技术检测趋化因子CXCL9、CXCL11、CCL17、CCL22 及受体CXCR3 、CCR4在50例BP患者皮损及30例正常皮肤中的表达.结果 50例BP患者皮损中4种趋化因子及其受体阳性表达的OD值均高于正常皮肤.其中, BP 皮损和对照组 Th1 趋化因子CXCL9、CXCL11 及其受体 CXCR3 表达的 OD 值分别为(0.55 ± 0.09) vs (0.26 ± 0.08)、(0.38 ± 0.06) vs (0.18 ± 0.04)和(0.37 ± 0.04) vs (0.22 ± 0.02),Th2 趋化因子CCL17、CCL22及其受体CCR4表达的OD值分别为(0.42 ± 0.10) vs (0.25 ± 0.12)、(0.35 ± 0.04) vs (0.21 ± 0.03)和(0.42 ± 0.05) vs (0.24 ± 0.07),且表达差异均有统计学意义(P＜0.01).结论 Th1趋化因子CXCL9、CXCL11和Th2趋化因子CCL17、CCL22及其受体CXCR 3、CCR4可能与BP的发病相关,对探讨BP发病机制有一定的价值.     

防感煎剂对甲1型流感病毒感染小鼠Th细胞特异性趋化因子受体的影响

[目的]研究防感煎剂对甲1型流感病毒感染小鼠Th细胞特异性趋化因子受体的影响。[方法]建立H1N1型流感病毒感染小鼠模型,流式细胞仪直接免疫荧光法检测Th细胞特异性趋化因子受体CXCR3、CCR4、CCR5,动态观察。[结果]在感染后第1、5、9天,防感煎剂各剂量组可提高下降的CD4＋CCR5＋%、CD4＋CXCR3＋%,第13天基本恢复正常;防感煎剂似可减少感染后9～13天染毒小鼠CD4＋CCR4＋%表达,中高剂量组明显。[结论]防感煎剂主要促进感染早期CCR5、CXCR3表达增加,Th1细胞活化并向炎症部位趋化,引起Th1型抗病毒免疫炎症反应,在感染后期减少CCR4表达,提示有一定的抑制高反应性哮喘和特异性过敏反应的作用。