CXCR1、CXCR2 及CXCL8 在乙肝相关肝癌中的表达及临床意义

目的:探讨乙肝相关肝癌患者外周血单个核细胞及肝活检组织CXCR1、CXCR2 及CXCL8 表达及其临床意义。方法:用实时荧光定量PCR 法检测36 例乙肝相关肝癌患者外周血PBMCs 中CXCR1、CXCR2 及CXCL8 mRNA 水平;SP 法检测肝活检组织CXCR1、CXCR2 及CXCL8 蛋白表达水平;化学发光免疫分析法检测血清CRP 水平,并对各指标进行相关性分析。结果:乙肝相关肝癌患者PBMCs 中CXCR1、CXCR2 及CXCL8 mRNA 水平分别为(0.952 7±0.197 2)、(0.896 9±0.173 0)、(1.771 9±0.248 9),较正常对照组明显升高,差异有统计学意义(P<0.01)。SP 结果提示,CXCR1、CXCR2 及CXCL8 的蛋白水平较对照组明显升高(P<0.05)。乙肝相关肝癌患者血清CRP 水平与PBMC 内CXCR1(r =0.54,P<0.01)、CXCR2(r =0.49,P<0.01)及CXCL8(r = 0.63,P<0.01)呈正相关。结论:乙肝相关肝癌患者外周血PBMCs 及肝活检组织CXCR1、CXCR2 及CXCL8 表达明显升高,且与血清CRP 呈正相关,推测CXCR1、CXCR2 及CXCL8 介导的信号转导过程可能在乙肝相关肝癌发病过程中发挥重要作用,为肝癌免疫干扰治疗提供新的靶点。     

趋化因子CXCL12及其配体CXCR4、CXCR7对HER2阳性乳腺癌细胞生物学功能的影响北大核心CSCD

目的:探究趋化因子CXCL12及其配体CXCR4、CXCR7对HER2阳性乳腺癌细胞增殖、细胞周期、迁移、侵袭和上皮-间质转化(EMT)的影响。方法:体外培养HER2阳性乳腺癌细胞BT474,应用siRNA转染技术单独或联合沉默CXCR4、CXCR7,RT-PCR与Western blot检测转染效率;应用CXCL12刺激上述转染细胞,并将其分为5组,A组:正常培养的BT474细胞;B组:CXCL12处理的BT474细胞;C组:CXCL12处理的转染si-CXCR4细胞;D组:CXCL12处理的转染si-CXCR7细胞;E组:CXCL12处理的联合转染si-CXCR4与si-CXCR7细胞;应用CCK-8、流式细胞术、划痕实验、侵袭实验及Western blot分别检测沉默CXCR4和(或)CXCR7对CXCL12诱导的乳腺癌细胞增殖、细胞周期进展、迁移、侵袭和EMT行为的影响。结果:转染siRNA能显著降低BT474细胞中CXCR4和(或)CXCR7的mRNA和蛋白表达水平(P<0.01);单独或联合沉默CXCR4与CXCR7均能明显抑制CXCL12对乳腺癌细胞增殖、细胞周期进展、迁移、侵袭行为的促进作用(P<0.05或P<0.01),并以联合沉默CXCR4与CXCR7时效果最为显著(P<0.01),与单独沉默CXCR7相比,沉默CXCR4更能抑制CXCL12对乳腺癌EMT的促进(P<0.05)。结论:沉默CXCR4或CXCR7表达均能抑制CXCL12对HER2阳性乳腺癌细胞增殖、细胞周期进展、迁移、侵袭的促进功能,但CXCL12主要通过与CXCR4相互作用来促进乳腺癌的EMT行为,而同时抑制CXCR4或CXCR7能更有效地抑制CXCL12的上述功能。     

CXCL8及其受体CXCR1、CXCR2在慢性乙肝患者外周血PMNs中的表达

目的:探讨慢性乙肝患者外周血中性粒细胞(PMNs)上CXCL8及其受体CXCR1、CXCR2的表达。方法:以中性粒细胞分离液分离、纯化PMNs,检测患者血清HBe Ag及PMNs内HBV DNA,入选患者依据检测结果进行分组,SABC免疫细胞化学染色法检测各组患者PMNs内CXCL8及其受体CXCR1、CXCR2的表达。结果:SABC免疫细胞化学染色结果显示,CXCL8主要位于PMNs胞浆中,CXCR1、CXCR2多见于胞浆和胞膜上。其中HBe Ag(+)者CXCL8、CXCR1免疫着色较深,而CXCR2免疫着色较浅;PMNs内HBV DNA(+)者CXCL8、CXCR1免疫着色亦较深,而CXCR2免疫着色亦较浅。患者CXCL8和CXCR1的水平均显著升高,与正常对照相比,差异均有显著性(P0.05),而CXCR2的表达无统计学意义(P0.05)。结论:HBV侵染中性粒细胞后可促进CXCL8分泌,使胞膜CXCR1表达进一步增强。CXCL8、CXCR1、CXCR2免疫组化染色程度与患者HBe Ag表达、HBV DNA载量密切相关。高表达CXCR1的中性粒细胞与CXCL8相互作用,趋化吸引更多PMNs至病灶,参与局部炎性损伤和组织修复。     

前列腺酸性磷酸酶在慢性痛大鼠脊髓背角和背根神经节的表达变化

目的:观察前列腺酸性磷酸酶(prostatic acid phosphatase,PAP)在多种慢性痛大鼠脊髓背角(spinaldorsal horn,SDH)和背根神经节(dorsal root ganglion,DRG)内的表达变化。方法:应用免疫组织化学染色法以及免疫荧光多重染色技术在多种慢性痛模型大鼠观察PAP的表达变化。结果:在正常大鼠,PAP阳性反应产物主要位于DRG的中、小型的非肽能神经元,PAP阳性神经元约占DRG神经元总数的64±4.3%;在脊髓背角,PAP阳性纤维和终末主要位于Ⅱ层。在神经病理性痛模型大鼠,术侧脊髓背角Ⅱ层的PAP阳性初级传入终末较对侧减少甚至消失,DRG内PAP阳性神经元较对侧明显减少。在慢性炎性痛模型大鼠,双侧脊髓背角和DRG内PAP的表达未见明显改变。结论:PAP特异地定位于DRG神经元以及脊髓背角Ⅱ层,可能与神经病理性痛信号的传递和加工密切相关。     

小鼠中央杏仁核中GluR1的表达变化参与骨癌痛形成

目的:观察GluR1在骨癌痛(bone cancer pain,BCP)模型小鼠中央杏仁核中的表达变化。方法:健康C57小鼠分为假手术对照组(n=100)和骨癌痛组(n=100),每组均在术前和术后7、14、21、28 d先进行行为学检测,检测完毕后取材,进行免疫组化染色和Western Blot检测,观察GluR1在中央杏仁核中的表达变化。结果:从癌细胞接种后第7 d开始,BCP组出现自发缩足次数增多、PWT值降低,14 d后与Sham组比较开始有明显差异,表明模型建立成功;免疫组织化学染色显示GluR1在正常C57小鼠中央杏仁核中的表达水平较低,但在建模术后小鼠中央杏仁核中GluR1的表达开始逐渐升高,图像分析表明GluR1的光密度与对照组比较,第14 d时差异有统计学意义(P<0.05),BCP组21 d时GluR1的表达达高峰(P<0.01),Western Blot检测结果亦与之相符。结论:骨癌痛小鼠GluR1可能在杏仁核参与神经病理性痛的过程中具有重要作用。     

1,8-桉叶素对背根神经节内P2X3受体介导神经病理痛的作用

目的:探讨1,8-桉叶素对大鼠背根神经节(DRG)神经元P2X3受体介导神经病理痛的作用。方法:建立大鼠坐骨神经慢性压迫性损伤模型(CCI)。SD大鼠随机分为假手术(Sham)组,坐骨神经慢性压迫性损伤(模型组,CCI)组、低剂量1,8-桉叶素治疗组、高剂量1,8-桉叶素治疗组、二甲亚砜对照组。检测大鼠术后7、14 d机械缩足反射(MWT)及热缩足反射潜伏期(TWL),观察大鼠行为学变化。免疫组织化学和原位杂交观察神经病理痛大鼠第4~5腰(L_(4-5))DRG神经元P2X3受体表达变化。结果:术后第7和14天,模型组大鼠MWT和TWL明显低于假手术组,低、高剂量治疗组较模型组明显升高,二甲亚砜组与模型组比较无差别;L_(4-5)DRG内P2X3受体表达模型组明显高于假手术组,低、高剂量治疗组较模型组均明显降低,二甲亚砜组与模型组比较无明显区别。结论:1,8-桉叶素抑制CCI大鼠L_(4-5)DRG神经元P2X3受体过表达,从而缓解神经病理性疼痛症状。     

CXCL12、CXCR4、MMP-2在宫颈鳞癌组织的表达及意义

目的探讨趋化因子CXCL12与其受体CXCR4及金属蛋白酶MMP-2在宫颈鳞癌组织中的表达及临床意义。方法采用免疫组织化学SP法检测50例宫颈鳞癌组织及50例癌旁正常组织中CXCL12、CXCR4、MMP-2的表达。结果 CXCL12、CXCR4、MMP-2在宫颈鳞癌组织中的阳性表达率(68.0%、72.0%、78.0%)明显高于正常宫颈组织(12.0%、24.0%、30.0%),在中/低度分化的宫颈鳞癌组织阳性表达率分别为84.4%、75.0%、87.5%。CXCL12、CXCR4、MMP-2在宫颈鳞癌淋巴结转移组中阳性表达率(86.2%、82.8%、79.3%)也明显高于无淋巴结转移组(52.4%、57.1%、52.4%)。在宫颈鳞癌组织中CXCL12与CXCR4、MMP-2表达都呈正相关(r=0.355,P=0.004;r=0.310,P=0.036);MMP-2与CXCR4表达也呈正相关(r=0.297,P=0.042)。结论 CXCL12和受体CXCR4、MMP-2与宫颈鳞癌的病变进展和转移密切相关,可能对宫颈鳞癌的预后和复发具有指导意义。     

小鼠背根神经节慢性压迫引起的痛敏化机制

目的 探讨背根神经节慢性压迫（CCD）引起的低剂量辣椒素诱发痛敏化机制。 方法 将光滑L型钢柱插入小鼠的L4椎间孔，对背根神经节（DRG）造成持续压迫，建立小鼠CCD模型。于CCD术前1 d及术后第1、3、5、7 d向小鼠胫骨前区皮下注射不同浓度的辣椒素（0.01、0.1、1 g/L）10 μl后采用行为学检测动物的痛行为学表现并录像，以寻找CCD术后辣椒素诱发痛行为差异最显著的浓度。利用Pirt-GCaMP3转基因小鼠，胫骨前区皮下注射最佳浓度辣椒素，利用激光扫描共焦显微镜在体记录DRG神经元对外周体表感受野辣椒素刺激的反应。利用免疫荧光染色技术观察正常及CCD模型压迫5 d后DRG内辣椒素受体（TRPV1）的表达变化。 结果 行为学检测结果显示，与正常小鼠相比，低浓度（0.1 g/L）辣椒素在CCD模型小鼠上引起的诱发痛显著提升（n=8; 术后1 d，P<0.01；术后5 d，P<0.05；术后7 d，P<0.05；术后3 d，P> 0.05）。在体激光扫描共焦成像结果表明，在记录的398（n=4）个正常小鼠L4 DRG神经元中，对皮下注射0.1 g/L辣椒素有反应的神经元为75个，在382（n=6）个CCD术后5 d的小鼠L4 DRG神经元中，对皮下注射0.1 g/L辣椒素有反应的神经元为169个，差异有显著性（P<0.01）。免疫荧光染色结果显示，在653（n=10）个正常神经元中，TRPV1阳性的细胞数为148，在611（n=6）个CCD神经元中，TRPV1阳性的细胞数为237，差异有显著性（P<0.01）。 结论 在小鼠CCD模型中，受到压迫DRG神经元TRPV1表达上调，导致相应的体表感受区域对辣椒素诱发痛敏化。     

外周神经损伤大鼠背根神经节中Ephrin B1及RYK 受体表达的变化

目的研究外周神经损伤后背根神经节细胞中Ephrin B1及其相关受体的表达变化。方法建立一侧坐骨神经夹伤的大鼠动物模型,通过免疫荧光组织化学方法检测受损侧背根神经节细胞中Ephrin B1及其相关受体Eph B1、Eph B2、Eph B3和Eph A4、RYK等的表达,并分析阳性细胞数和不同大小阳性细胞的构成比例。结果外周坐骨神经受损侧背根神经节细胞中Ephrin B1的表达明显减弱,而Eph B1、Eph B2、Eph B3和Eph A4受体的表达无明显变化,但RYK受体的表达则明显加强。结论Ephrin B1和RYK受体在一侧外周坐骨神经夹伤后的大鼠背根神经节细胞中表达的变化,说明它很有可能参与了损伤后的功能活动。     

Down-regulation of mu opioid receptor expression within distinct subpopulations of dorsal root ganglion neurons in a murine model of bone cancer pain

Although micro opioid receptor (MOR) agonists are used for treatment of most types of pain, a recent study has suggested that the sensitivity of bone cancer pain to systemic morphine was lower than that of inflammatory pain. However, the reasons for this have remained unclear. In this study, MOR expression and the analgesic effects of morphine in a bone cancer model were compared with those in an inflammatory pain model. A bone cancer pain model and an inflammatory pain model were made by implantation of sarcoma cells into the intramedullary space of the femur and hind-paw injection of complete Freund's adjuvant (CFA), respectively. In a behavioral study, sarcoma-implanted mice showed flinching behavior of magnitude comparable to that induced by CFA injection. The flinching behavior of sarcoma-implanted mice was less sensitive to intrathecal morphine than that of CFA-injected mice. Western blot analysis showed that MOR expression in the dorsal root ganglion (DRG) ipsilateral to sarcoma implantation was significantly reduced, while that in the DRG ipsilateral to CFA injection was increased. In sarcoma-implanted mice, the percentage of MOR-positive DRG neuronal profiles was lower than that in control mice (30.3% vs. 45.2%). In particular, MOR expression was reduced among calcitonin gene-related peptide- and transient receptor potential vanilloid subfamily 1-positive DRG neuronal profiles, which are considered to be involved in the generation of bone cancer pain (from 61.5% to 41.5% and from 72.1% to 48.4%, respectively). These results suggest that down-regulation of MOR in the distinct populations of DRG neurons contributes to the fact that higher doses of morphine are needed to produce analgesia in bone cancer as compared with those used in non-malignant inflammatory situations.     

胃癌组织中CXCL12和CXCR4的表达及意义

目的观察趋化因子CXCL12及其特异性受体CXCR4在人胃癌组织中的表达,探讨其与临床病理参数、预后的关系。方法选择120例胃癌标本,应用免疫组化SP法检测CXCL12和CXCR4在人胃癌组织中的表达,分析CXCL12和CXCR4的表达与患者临床病理参数、术后生存率之间的关系。结果胃癌组织及正常胃黏膜组织中均可检测到CXCL12、CXCR4的表达,但胃癌组织中的表达水平均明显高于正常胃黏膜组织,表达差异有显著性(P<0.05)。CXCL12阳性与CXCR4阳性呈正相关(r=0.276,P<0.05)。胃癌CXCL12和CXCR4的表达水平与肿瘤细胞淋巴结转移及分化程度密切相关(P<0.05),与患者的年龄、性别、肿瘤的大小、浸润深度及远处转移等无关(P>0.05)。CXCL12和CXCR4阳性表达的患者其五年生存率明显低于其阴性表达的患者。结论胃癌中CXCL12和CXCR4的高表达与胃癌的生物学行为及预后密切相关,检测其表达对预测胃癌的转移及判断预后有一定价值。     

Expressions of chemokine CXCL12 and its receptor CXCR4 in gastric cancer and their significance

目的 观察趋化因子CXCL12及其特异性受体CXCR4在人胃癌组织中的表达,探讨其与临床病理参数、预后的关系.方法 选择120例胃癌标本,应用免疫组化SP法检测CXCL12和CXCR4在人胃癌组织中的表达,分析CXCL12和CXCR4的表达与患者临床病理参数、术后生存率之间的关系.结果 胃癌组织及正常胃黏膜组织中均可检测到CXCL12、CXCR4的表达,但胃癌组织中的表达水平均明显高于正常胃黏膜组织,表达差异有显著性(P＜0.05).CXCL12阳性与CXCR4阳性呈正相关(r=0.276,P＜0.05).胃癌CXCL12和CXCR4的表达水平与肿瘤细胞淋巴结转移及分化程度密切相关(P＜0.05),与患者的年龄、性别、肿瘤的大小、浸润深度及远处转移等无关(P＞0.05).CXCL12和CXCR4阳性表达的患者其五年生存率明显低于其阴性表达的患者.结论 胃癌中CXCL12和CXCR4的高表达与胃癌的生物学行为及预后密切相关,检测其表达对预测胃癌的转移及判断预后有一定价值.     

CB_1受体对坐骨神经损伤导致的神经痛及背根节HCN2通道表达的调节作用

目的:观察大麻素CB_1受体在坐骨神经缩窄性损伤(CCI)所致的神经痛中的作用及其对背根节HCN2通道表达的调节。方法:7~8周龄SD大鼠分为4组:(1)Sham组(假手术组);(2)CCI组;(3)CP55940+CCI组;(4)AM251+CP55940+CCI组。采用von Frey测痛仪测定各组大鼠神经损伤侧机械缩足阈值(MWT);免疫印迹技术检测损伤侧L_4~L_6背根节HCN2通道表达。结果:CCI术后1 d即形成稳定的机械痛敏,MWT降低;鞘内给予大麻素受体激动剂CP55940(0.05 mg/kg)可显著升高CCI大鼠MWT(P0.05);预先鞘内注射CB_1R拮抗剂AM251(0.05 mg/kg)可明显阻断CP55940的镇痛效果(P0.05)。免疫印迹实验结果显示,CCI大鼠损伤侧L_4~L_6背根节CB_1受体、HCN2通道表达明显增加(P0.05);鞘内给予CP55940可显著降低HCN2表达(P0.05),AM251可明显抑制CP55940降低HCN2表达的效应(P0.05)。结论:背根节CB_1受体激活对CCI所致的神经痛具有良好的镇痛作用,其镇痛效应可能与抑制CCI大鼠背根节HCN2通道表达有关。     

The expression of the chemokine receptor CXCR3 and its ligand, CXCL10, in human breast adenocarcinoma cell lines

The acquisition of a metastatic phenotype in breast epithelial cells is a progressive process, influenced by a large variety of cellular and soluble factors. Of these, members of the chemokine superfamily, such as CCL2, CCL5, CXCL8 and CXCL12 have been recently suggested to promote breast cancer progression. A pre-requisite for elucidation of the role of other chemokines in breast cancer progression is the characterization of chemokine and chemokine receptor expression by breast tumor cells. The present study focuses on CXCL10, a CXC chemokine that was recently suggested to have anti-malignant properties, and its corresponding receptor CXCR3. CXCR3 expression was detected in three human breast adenocarcinoma cell lines, MDA-MB-231, MCF-7 and T47D. CXCR3 expression was potently up-regulated by growing the cells under stress conditions, imposed by serum starvation. Unlike many other chemokine receptors, CXCR3 expression was not down-regulated by exposure to high concentrations (500ng/ml) of its ligand, CXCL10, but rather was promoted. CXCL10-induced up-regulation of CXCR3 expression in the three cell lines was inhibited by cycloheximide, indicating that de novo protein synthesis is required for this process. In addition to CXCR3, the secretion of CXCL10 was noted in the MDA-MB-231, MCF-7 and T47D cells. CXCL10 secretion was found to be down-regulated by IL-6, a potentially pro-malignant cytokine in breast cancer. The concomitant expression of CXCR3 and CXCL10 in breast tumor cells suggests that a CXCR3-CXCL10 axis may function in these cells, and paves the way for an in depth analysis of CXCL10-CXCR3 interactions in breast tumor cells.     

CXC chemokine ligand 12 (CXCL12) and its receptor CXCR4

Chemokines were recognized originally for their ability to dictate the migration and activation of leukocytes. However, CXC chemokine ligand 12 (CXCL12, also known as stromal cell-derived factor-1) and its receptor CXCR4 are the first chemokine and receptor that have been shown to be critical for developmental processes, including homing and maintenance of hematopoietic stem cells (HSCs), production of immune cells, homing of primordial germ cells (PGCs), cardiogenesis, arterial vessel branching in some organs, and appropriate assemblies of particular types of neurons. This review focuses on the pathophysiological relevance of CXCL12-CXCR4 signaling in mammals.     

Serotonin receptor mRNA expression in rat dorsal root ganglion neurons

In the present study, we have used in situ hybridization to examine the distribution of serotonin (5-HT) receptors in rat dorsal root ganglion (DRG) neurons. Within DRG neurons, mRNAs for 5-HT1B, 5-HT1D, 5-HT2A, 5-HT2B, 5-HT3B and 5-HT4 receptors were readily detected in small (<25 microm), medium (25-45 microm) and large (>45 microm) diameter neurons. In contrast mRNAs for 5-HT1A, 5-HT1E, 5-HT2C, 5-HT5A, 5-HT5B, 5-HT6 and 5-HT7 receptors were undetectable in these neurons. The present study provides an insight into the molecular profile of 5-HT receptor subtypes in neurons responsible for modulating sensory information.     

Substance P expression in TRPV1 and trkA-positive dorsal root ganglion neurons innervating the mouse lung

In the present study, the co-localisation of substance P (SP) with the vanilloid receptor TRPV1 and the neurotrophin receptor tyrosine kinase trkA was analysed in airway-specific murine dorsal root ganglion (DRG) neurons. DRG neurons labelled with Fast Blue were predominantly found at the segmental levels T2-T5. Immunoreactivity for the receptor TRPV1 was localized to 12% of Fast Blue labelled DRG neurons. Double-labelling immunohistochemistry revealed that a substantial number of them also co-express SP (7.6 +/- 1.1% (mean +/- S.E.M.)), whereas neurons with immunoreactivity for TRPV1 only were found in 4.4 +/- 1.3% of the retrogradely labelled neuronal population. Further analysis of retrogradely labelled neurons showed that their majority expressed trkA (62.8 +/- 1.4%), neurofilament protein 68-kDa (64.8 +/- 1.5%) or glutamate alone (19.5 +/- 1.9%). SP was always expressed in trkA-positive neurons. Based on the extent of co-localization of SP with the receptors TRPV1 and trkA in DRG airway neurons, the present study indicates that the DRG pathway may have effects on the magnitude of neurogenic inflammation in airway diseases such as asthma.     

Role of signals from the dorsal root ganglion in neuropathic pain in a rat model

Vascular endothelial growth factor (VEGF), known as an endothelial cell-specific mitogen, has been reported to be linked also to the NO/cGMP-pathway, which has been notified in the inner ear. Up to now, VEGF has not yet been described in the inner ear. We performed immunohistochemical analysis using specific antibodies to VEGF and to both known VEGF-receptors Flt-1 and KDR/Flk-1 on paraffin-sections of temporal bones from guinea pigs (n=5). Immunoreactivity of VEGF, Flt-1 and KDR/Flk-1 was detectable in a subpopulation of vestibular ganglion cells. VEGF could be found also in the endothelium of blood vessels, in fibrocytes of the lamina propria and in the neuroepithelium. Strong immuno-labelling to Flt-1 was evident in nerve fibres, vascular endothelium and in the neuroepithelium. Fibrocytes, endothelium of blood vessels, supporting cells and calyces in the sensory epithelium revealed immunoreactivity to KDR/Flk-1. These findings give evidence that VEGF, Flt-1 and KDR/Flk-1 are constitutively expressed in the vestibule.