早孕和足月滋养细胞中侵袭相关性基因的表达变化

目的：探讨早孕和足月滋养细胞侵袭能力差异的机制。方法：分离并在体外培养早孕和足月的滋养细胞为，采用RT-PCR法，检测其侵袭相关性基因的表达。结果：早孕滋养细胞表达MMP-2、MMP-9、TIMP-1和UPA；而晚孕滋养细胞表达MMP-2、TIMP-1、TIMP-2和PAI。晚孕滋养细胞TTMP—1的表达显著高于早孕滋养细胞，而MMP-2的表达则无明显变化。结论：MMP-9等蛋白酶及其抑制因子表达的变化，可能是影响滋养细胞侵袭性的重要原因。     

阿司匹林对TNF-α诱导人早孕滋养细胞增殖和侵袭的影响及机制

目的:采用体外细胞模型探讨阿司匹林促进人早孕滋养细胞增殖和侵袭在预防子痫前期(preeclampsia,PE)发生中的作用及机制。方法:收集PE病例20例和正常孕妇作为对照20例;HE染色观察胎盘组织病理改变;ELISA法检测基质金属蛋白酶2(MMP-2)及MMP-9的含量。体外培养人早孕滋养细胞HTR-8/SVneo,分别加入不同浓度的阿司匹林和肿瘤坏死因子α(TNF-α);CCK-8法检测细胞活力;Transwell实验检测细胞侵袭能力;Western blot检测细胞周期蛋白D1(cyclin D1)和增殖细胞核抗原(PCNA)的蛋白表达水平。结果:PE组胎盘组织存在子宫螺旋动脉血管重铸障碍,且孕妇血清中MMP-2及MMP-9浓度明显低于对照组(P0.05)。体外早孕滋养细胞系实验表明,与对照组比较,0.1 mmol/L阿司匹林明显增加细胞活力,且可缓解TNF-α引起的细胞活力降低(P0.05);TNF-α处理滋养细胞后,细胞cyclin D1及PCNA蛋白表达水平显著下降,侵袭能力减弱,培养上清液中MMP-2和MMP-9表达水平及活性均明显下降(P0.05);同时给予0.1 mmol/L阿司匹林和TNF-α处理后,滋养细胞cyclin D1及PCNA蛋白表达水平显著增高,侵袭能力增强,培养上清液中MMP-2和MMP-9表达水平及活性均明显上升(P0.05)。结论:小剂量阿司匹林可以促进绒毛滋养细胞增殖和侵袭,可能有利于改善PE胎盘浅着床和子宫螺旋动脉重铸异常情况。这为阿司匹林预防PE的发生提供实验依据。     

研究滋养细胞侵袭的常用模型

正常妊娠绒毛滋养细胞(extravillous trophoblast,EⅥ)侵入子宫蜕膜、肌层内1/3及螺旋动脉,破坏螺旋动脉血管平滑肌、弹力纤维及神经组织,致螺旋动脉发生"血管重塑".目前,滋养细胞侵袭过程的调控机制是一个研究热点,现对滋养细胞侵袭研究的细胞模型及细胞外基质模型做一综述.     

IL-10对早孕滋养细胞MMP-9表达的影响

人类滋养细胞的侵入仅限于子宫内膜及肌层的浅层 1/ 3,并于妊娠中期停止[1] 。这一过程受到多种因素的控制 ,ＩＬ 10在整个妊娠过程中都可由胎盘滋养细胞分泌 ,且α6整合素表达阳性的滋养细胞表面也存有ＩＬ 10的受体 ,故ＩＬ 10也是参与该调节的因子之一[2 ] 。侵入并非一个由细胞增殖造成的被动事件 ,而是一个主动的生化过程。滋养细胞通过分泌蛋白酶类获得侵袭能力 ,其中基质金属蛋白酶 9(ＭＭＰ 9)是参与滋养细胞侵入子宫内膜的主要蛋白酶[3 ] 。因此 ,为探讨ＩＬ 10对滋养细胞侵袭力的影响 ,我们采用ＲＴ ＰＣＲ方法 ,观察ＩＬ 10对早…     

流式细胞仪分离人早孕绒毛外滋养细胞及鉴定

目的用流式细胞仪分离早期绒毛外滋养细胞，以便对其功能进行进一步研究。方法用胰蛋白酶消化法得到早期绒毛细胞，用流式细胞仪分离纯化绒毛外滋养细胞。所得的细胞用免疫细胞化学，光镜，透射电镜检以及Westen-blot方法鉴定。结果使用流式细胞仪成功的分离了绒毛外滋养细胞，纯度超过98％。结论此方法可快速准确大量获得绒毛外滋养细胞。     

早孕绒毛滋养细胞FasL表达异常与自然流产的关系

通过比较正常早孕与自然流产者滋养细胞表面FasL表达,进一步从分子免疫学角度探讨自然流产的发病机理。方法：用特异性的FasL抗体进行免疫组化染色,并通过高清晰彩色病理免疫组化测量系统对其定量分析,SSPS对两组进行比较。结果：自然流产组滋养细胞表面FasL表达面积及强度均明显低于正常早孕组,两者间差异有显著性意义。结论：滋养细胞表面FasL表达减少,引起母胎间的免疫耐受的破坏,是导致自然流产的一重量免疫病因,诱导FasL的产生或调节母胎间的免疫耐受将为临床治疗自然流产提供新的方向。     

人早孕滋养细胞与蜕膜基质细胞共培养对外周NK细胞的影响

为了分析人早孕母-胎界面功能细胞对外周NK细胞表型及功能的调节作用,我们模拟母-胎界面微环境,建立滋养细胞和蜕膜基质细胞共培养系统,并将此共培养体系与外周NK细胞共培养,采用流式细胞术分析NK细胞的表型和细胞因子的表达,用细胞毒检测试剂盒检测NK细胞的细胞毒活性。研究发现,母-胎界面滋养细胞与蜕膜基质细胞共培养体系能够显著上调外周NK细胞抑制型受体KIR2DL1的表达,下调外周NK细胞杀伤性受体NKp44的表达;显著上调Th2型细胞因子IL-10的表达水平,下调外周NK细胞穿孔素perforin表达及其细胞毒活性。结果表明,母-胎界面功能细胞能够诱导外周NK细胞的耐受表型及Th2型优势,下调外周及蜕膜NK细胞的细胞毒活性,从而有利于形成母-胎界面免疫耐受微环境。     

过表达CXCR4/CXCR7影响人脐带间充质干细胞迁移和增殖

目的探讨SDF-1/CXCR4及SDF-1/CXCR7对人脐带间充质干细胞(h UC-MSCs)迁移和增殖活性的影响。方法用Ad-EASY腺病毒质粒系统分别构建表达CXCR4和CXCR7的重组腺病毒,感染h UC-MSCs后用FCM分别检测细胞膜的CXCR4和CXCR7受体的表达,Transwell法检测细胞迁移,MTT检测h UC-MSCs增殖活性,以及在H2O2诱导细胞毒性作用对细胞存活的影响。结果成功构建表达人源的CXCR4和CXCR7的重组腺病毒,感染后h UC-MSCs细胞膜上的CXCR4/CXCR7受体阳性表达率分别达到93.7%和78.5%(P0.01),高表达CXCR4和CXCR7均显著增强SDF-1诱导的h UC-MSCs细胞迁移,其中CXCR7效应稍弱;高表达CXCR7而不是CXCR4显著提高SDF-1诱导的h UC-MSCs增殖活性和氧化应激状态下的细胞存活率(P0.05)。结论 CXCR4/CXCR7均参与介导了SDF-1诱导的h UC-MSCs细胞迁移作用,同时CXCR7还介导了SDF-1诱导的h UC-MSCs增殖和H2O2处理损伤的保护。     

人绒毛滋养细胞体外对T淋巴细胞的调节作用

妊娠是子宫和成熟的囊胚之间复杂的相互作用的结果[1].随着对滋养细胞研究的深入,发现母胎免疫是发生在母胎之间的免疫对话,且贯穿整个妊娠过程[2].作为半同种移植物的胎儿,在母体内安然成长至足月,得益于母胎之间的免疫耐受.母胎界面的胎儿面主要由滋养细胞组成,其起源于上皮细胞,是唯一与母体免疫系统直接接触的胎儿细胞,主要分泌β类趋化因子参与蜕膜免疫细胞的募集.近年来研究发现,RANTES(regulated upon activation normal T-cell expressed and secreted)、 CXCL12和CCL2是母胎界面起关键作用的趋化因子[3],但关于RANTES在妊娠早期对外周血T淋巴细胞Th1/Th2的影响却未见报道.本实验通过体外分离共培养滋养细胞和外周血T淋巴细胞,观察滋养细胞及其分泌的趋化因子RANTES对T淋巴细胞Th1/Th2型细胞因子的影响,为探讨早孕期母胎免疫机制提供实验依据.     

CXCL12/CXCR4/CXCR7轴在胎盘滋养细胞中作用机制的探讨

CXCL12介导的信号通路参与调节免疫细胞的活化和募集、造血干细胞的动员和归巢、新生血管形成、胚胎发育、肿瘤侵袭转移等过程.最近,CXCL12/CXCR4/CXCR7生物轴在胎盘植入和发育中的重要作用越来越受到关注.研究发现,CXCL12介导的信号通路参与调节胎盘滋养细胞的分化、增殖和侵袭,同时又可以趋化滋养细胞定向移动到蜕膜间质及血管周围,促进胎盘的植入、子宫-胎盘血管重塑和免疫耐受,进而维持妊娠的正常进行.深入理解CXCL12/CXCR4/CXCR7轴在胎盘滋养细胞中的作用机制,将有助于进一步认识胎盘植入的内在机制,为滋养细胞功能紊乱相关疾病的治疗提供新的思路.     

早孕期人滋养细胞CXCR4/CXCL12的表达及低氧对其的影响

目的：研究人早孕期滋养细胞CXCR4/CXCL12的表达情况及低氧对其表达的调节作用.方法：免疫组织化学法检测早孕期绒毛组织中及原代培养的滋养细胞CXCR4/CXCL12的表达情况;实时定量PCR检测低氧状态下滋养细胞CXCR4 mRNA水平.结果：胎盘绒毛柱、滋养细胞及血管内均有CXCR4/CXCL12表达;低氧培养12、24、48和72小时后滋养细胞CXCR4 mRNA表达增加,显著高于正常氧条件下的表达情况.结论：早孕期胎盘表达CXCR4/CXCL12对于维系正常妊娠的顺利进行可能发挥重要作用;低氧是CXCR4表达的重要调节因子,可能参与了妊娠的病理生理过程.     

结直肠肿瘤中CXCR7、NDRG1的表达

目的 探讨CXCR7、NDRG1蛋白在结直肠肿瘤中的表达及其对结直肠肿瘤发生、发展的影响.方法 采用免疫组化染色技术观察人结直肠正常黏膜组织、结直肠腺瘤、无淋巴结转移(无LNM)结直肠腺癌组织及伴有淋巴结转移(有LNM)结直肠腺癌组织中CXCR7、NDRG1蛋白的表达水平.结果 (1)结直肠正常黏膜组织、腺瘤组织、无LNM结直肠腺癌组织及有LNM结直肠腺癌组织中CXCR7蛋白表达逐渐增高,其中后者CXCR7的表达与其他组别相比,差异有统计学意义(P＜0.05);CXCR7的表达与淋巴结转移呈正相关关系(rs=0.341),与肿瘤TNM分期有关(P＜0.05).(2)NDRG1蛋白表达逐渐增高,差异均有统计学意义(P＜0.05);NDRG1的表达与淋巴结转移呈正相关关系(rs＝0.402),与肿瘤TNM分期有关(P＜0.05).(3)CXCR7与NDRG1在结直肠肿瘤中的表达呈正相关关系(P=0.003,rs=0.334),2者均具有诊断准确性.结论 CXCR7、NDRG1在结直肠腺癌发生发展整个过程的不同阶段起着重要的作用,两者在促进肿瘤转移方面具有正相关性.     

人早孕期绒毛组织和滋养细胞趋化因子受体转录水平

目的：研究早孕期绒毛组织及滋养细胞18种趋化因子受体的转录水平，以揭示趋化因子受体在母一胎界面生理性调节作用。方法：提取人早孕期绒毛组织及滋养细胞总RNA，半定量RT-PCR检测绒毛组织和滋养细胞18种趋化因子受体mRNA的表达水平。结果：CXCR4及CXCR6在绒毛组织中普遍高表达；CCR6、CCR7、XCR1及CX3CR1呈普遍中等表达；CCR1～CCR5、CCR8～CCR10、CXCR1～CXCR3在部分绒毛组织中表达，部分绒毛组织不表达或表达量很低；早孕人绒毛组织不表达CXCR5。早孕期滋养细胞表达CCR1、CCR3～CCR5、CCR8～CCR9、CXCR1～CXCR4、CXCR6、XCR1、CX3CR1；不表达CCR2、CCR6、CCR7、CCR10及CXCR5。结论：早孕期绒毛组织及滋养细胞表达多种趋化因子受体，它们在正常妊娠中具有重要的生理学意义。     

甲状腺癌组织中趋化因子4受体和趋化因子7受体的表达情况及临床病理意义

目的 研究甲状腺癌组织中趋化因子4受体(CXCR4)和趋化因子7受体(CXCR7)的表达情况及临床病理恿义.方法 选取2012年5月至2014年6月期间在本院进行外科手术的83例甲状腺癌患者及2014年45例甲状腺腺瘤患者,采用免疫组织化学染色方法检测甲状腺腺瘤及甲状腺癌中CXCR4和CXCR7的表达.结果 CXCR4和CXCR7在甲状腺癌中的表达阳性率均明显高于甲状腺腺瘤(P＜0.05).CXCR4和CXCR7在临床分期为Ⅲ～Ⅳ期的甲状腺癌患者中的表达阳性率均明显高于临床分期为Ⅰ～Ⅱ期者(P ＜0.05);CXCR7在有淋巴结转移的甲状腺癌患者中的表达阳性率明显高于无淋巴结转移者(P＜0.05),而CXCR4的表达阳性率与甲状腺癌患者有无淋巴结转移无关(P＞0.05);CXCR4和CXCR7在甲状腺癌组织中的表达阳性率与甲状腺癌患者性别无关(P＞0.05).在甲状腺癌患者中,CXCR4阳性表达和CXCR7阳性表达呈正相关(r =0.49,P＜0.01);在甲状腺腺瘤患者中,CXCR4阳性表达和CXCR7阳性表达不相关(r=0.14,P=0.21).结论 CXCR4和CXCR7参与了甲状腺癌的进展,并为临床上甲状腺癌诊断及靶向治疗提供理论基础.     

The expression of CXCR4, CXCL12 and CXCR7 in malignant pleural mesothelioma

The chemokine CXCL12 and its receptors, CXCR4 and CXCR7, are involved in tumour progression, metastasis, and survival. We investigated the expression of CXCR4, CXCL12, and CXCR7 in malignant pleural mesothelioma to determine if they are possible biomarkers and potential therapeutic targets. Forty-one mesothelioma tumour tissues, ten normal human pleural tissues, and two mesothelioma cell lines were stained with anti-CXCR4, anti-CXCL12, anti-CXCR7, and anti-p-Akt antibodies. RT-PCR was performed to determine the expression of CXCR4, CXCL12, and CXCR7 in six human mesothelioma cell lines (H28, 211H, H2052, ms-1, H290, and H513) and one human normal mesothelial cell line, LP9. These seven cell lines were also stained with anti-CXCR7. We found that CXCR4 and CXCL12 were expressed in 97.6% and 78.0% mesothelioma tissue samples, concurrently with strong expression of p-Akt (R(2) = 0.739 and 0.620, respectively). In addition, CXCR7 expression was weaker than CXCR4 expression in mesothelioma tissues. Furthermore, RT-PCR showed that CXCR4 and CXCL12 were overexpressed in 5/6 mesothelioma cell lines (211H, H2052, ms-1, H290, and H513), whereas CXCR7 was overexpressed in only 2/6 (H513 and H2052). Moreover, we found that the CXCR4 antagonist AMD3100 inhibited the growth of all five mesothelioma cell lines that overexpress CXCR4 and CXCL12. Our results suggest that the Akt-mTOR pathway is involved during the interruption of the CXCL12/CXCR4 axis in these five mesothelioma cell lines. In conclusion, CXCR4 and CXCL12 are highly expressed in most mesothelioma cell lines and tumour tissues, suggesting that CXCR4 and CXCL12 may be used as biomarkers for patients with mesothelioma. The CXCL12-CXCR4 interaction may be a potential therapeutic target for mesothelioma.     

CXCR7 influences the migration of B cells during maturation

The atypical chemokine receptor CXCR7 binds the chemokines CXCL12 and CXCL11. The receptor is widely expressed and was shown to tune CXCR12‐induced responses of CXCR4. Here, the function of CXCR7 was examined at late stages of human B‐cell maturation, when B cells differentiate into Ab‐secreting plasmablasts. We identified two populations of CXCR7⁺ cells in tonsillar lymphocytes, one being presumably memory B cells or early plasmablasts (FSC^lowCD19⁺CD38^mid) and the other being plasmablasts or early plasma cells (FSC^highCD19⁺CD38⁺). CXCR7 is expressed on CD19⁺CD27⁺ memory B cells, on CD19⁺CD38⁺CD138^? and intracellular immunoglobulin high plasmablasts, but not on CD19⁺CD138⁺icIg^high plasma cells. The differential expression pattern suggests a potential contribution of the scavenger receptor in final B‐cell maturation. On in vitro differentiating B cells, we found a marked inverse correlation between CXCR7 and CXCR5 cell surface levels, whereas expression of CXCR4 remained almost constant. Migration assays performed with tonsillar mononuclear cells or in vitro differentiated cells revealed that inhibition of CXCR7 markedly increases chemotaxis toward CXCL12, especially at late stages of B‐cell maturation. Chemotaxis was attenuated in the presence of CXCR4 antagonists, confirming that migration is CXCR4 mediated. Our findings unequivocally demonstrate a novel role for CXCR7 in regulating the migration of plasmablasts during B‐cell maturation.     

CXCR4 expression affects overall survival of HCC patients whereas CXCR7 expression does not

Hepatocellular carcinoma (HCC) is a heterogeneous disease with a poor prognosis and limited markers for predicting patient survival. Because chemokines and chemokine receptors play numerous and integral roles in HCC disease progression, the CXCR4–CXCL12–CXCR7 axis was studied in HCC patients. CXCR4 and CXCR7 expression was analyzed by immunohistochemistry in 86 HCC patients (training cohort) and validated in 42 unrelated HCC patients (validation cohort). CXCR4 levels were low in 22.1% of patients, intermediate in 30.2%, and high in 47.7%, whereas CXCR7 levels were low in 9.3% of patients, intermediate in 44.2% and high in 46.5% of the patients in the training cohort. When correlated to patient outcome, only CXCR4 affected overall survival (P=0.03). CXCR4–CXCL12–CXCR7 mRNA levels were examined in 33/86 patients. Interestingly, the common CXCR4–CXCR7 ligand CXCL12 was expressed at significantly lower levels in tumor tissues compared to adjacent normal liver (P=0.032). The expression and function of CXCR4 and CXCR7 was also analyzed in several human HCC cell lines. CXCR4 was expressed in Huh7, Hep3B, SNU398, SNU449 and SNU475 cells, whereas CXCR7 was expressed in HepG2, Huh7, SNU449 and SNU475 cells. Huh7, SNU449 and SNU475 cells migrated toward CXCL12, and this migration was inhibited by AMD3100/anti-CXCR4 and by CCX771/anti-CXCR7. Moreover, SNU449 and Huh7 cells exhibited matrix invasion in the presence of CXCL12 and CXCL11, a ligand exclusive to CXCR7. In conclusion, CXCR4 affects the prognosis of HCC patients but CXCR7 does not. Therefore, the CXCR4–CXCL12–CXCR7 axis plays a role in the interaction of HCC with the surrounding normal tissue and represents a suitable therapeutic target.