胃癌患者血清中骨桥蛋白和CD44v6检测的临床意义

目的检测胃癌患者血清中骨桥蛋白(osteopontin,OPN)和CD44v6(CD44 splice variant 6)的表达水平,探讨二者在胃癌诊断和病情监测方面的价值。方法应用酶联免疫吸附试验检测60例胃癌患者(胃癌组),42例胃溃疡患者(胃溃疡组),33例健康体检正常者(健康体检组)血清中OPN和CD44v6的水平。结果胃癌组血清中OPN和CD44v6的水平均高于溃疡组及健康体检组,差异均有统计学意义(P<0.01);胃癌患者血清OPN和CD44v6的水平均与胃癌的临床分期、肿瘤大小、淋巴结转移及远处转移相关(P<0.05)。胃癌患者血清中OPN与CD44v6的水平呈正相关(r=0.8645,P<0.01);二者联合检测敏感性提高至92.5%。结论血清OPN和CD44v6的水平可作为胃癌诊断和病情监测的临床指标,二者联合检测提高了胃癌诊断敏感性,可作为筛查指标用于胃癌的筛检。     

Osteopontin expression correlates with nuclear factor-κB activation and apoptosis downregulation in clear cell renal cell carcinoma

Osteopontin (OPN) is a phosphoglycoprotein implicated in tumorigenesis and tumor cell metastasis. Apoptosis inhibition is one of the mechanisms that contribute to development and progression of cancer, and might be initiated by OPN interaction with tumor cells. The aim of this study was to analyze the relation between OPN and nuclear factor-kappa B (NF-κB) expression in clear cell renal cell carcinoma (CCRCC), as well as their relation to apoptotic activity of tumor cells.Expression of OPN protein and p65 NF-κB subunit was analyzed immunohistochemically in 87 CCRCC samples, and compared mutually and with apoptotic index. Expression of OPN mRNA was analyzed using quantitative real-time PCR and compared with OPN and NF-κB protein expression in 22 CCRCC samples.Statistical analysis showed an association of p65 NF-κB with OPN mRNA (p = 0.015) and protein (p < 0.001). Also, we found an inverse relationship of OPN with NF-κB protein expression and apoptotic activity of tumor cells (p = 0.006 and p = 0.022, respectively). Our results indicate that p65 NF-κB signaling pathway may be involved in OPN-mediated CCRCC progression, partly by protecting tumor cells from apoptosis. Therefore, both molecules can constitute potential targets for therapeutic intervention in CCRCC.     

Osteopontin upregulation in rotavirus-induced murine biliary atresia requires replicating virus but is not necessary for development of biliary atresia

Biliary atresia (BA) is a progressive fibro-inflammatory pediatric liver disease in which osteopontin (OPN), a glycoprotein with inflammatory and fibrogenic activity, may play a pathogenic role. The current studies were conducted in a mouse model of rotavirus-induced BA to test the hypotheses that live but not inactivated rotavirus causes antigenemia, upregulation of hepatic OPN expression, and induction of BA and fibrosis; and that OPN is necessary for development of BA. Prolonged or transient antigenemia developed in mice inoculated with live or inactivated virus, respectively, but only live virus upregulated hepatic OPN and caused BA and fibrosis. OPN was expressed in intra- and extrahepatic bile ducts in healthy mice and in mice with BA. OPN-deficient mice, similar to WT mice, developed BA. Together, these data show that live but not inactivated rotavirus causes upregulation of hepatic OPN expression and BA but that OPN is not necessary for development of BA.     

Downregulating osteopontin reduces angiotensin II-induced inflammatory activation in vascular smooth muscle cells

Objective:  Atherosclerosis is considered as a chronic inflammatory response in arterial blood vessels. The function of osteopontin (OPN), a proinflammatory cytokine, in the Ang II-induced inflammatory activation in vascular smooth muscle cells (VSMCs) remains poorly understood. Methods:  In the present study, the role of OPN was investigated by knocking down OPN using small interfering RNA (siRNA). VSMCs from human saphenous vein were divided into three groups according to RNAi treatment: OPN siRNA group, sham (un-transfected) treated group, and control siRNA group. RNAi effect was investigated by real time PCR, western blotting analysis and ELISA. Then all groups were stimulated with Ang II. The inflammatory activation was assessed by determining the activation of NFκB and activator protein-1 (AP-1), and the release of interleukin-6 (IL-6) and IL-1β. Results:  OPN was knocked down effectively in OPN RNAi group. Inflammatory activations, such as NFκB and AP-1 activation and IL-6 accumulation, were induced by Ang II in sham treated group and control siRNA group. However, it was abolished in OPN siRNA group by the downregulation of OPN compared to sham treated group and control siRNA group. Conclusions:  This result suggested that OPN plays an important role in Ang II-induced inflammatory activation in VSMCs. The finding further supports OPN as a potential target for atherosclerotic therapy. Received 21 February 2008; returned for revision 6 May 2008; received from final revision 16 May 2008; accepted by S. Stimpson 21 August 2008 The first three authors contributed equally to this work.     

罗勒多糖抗卵巢癌侵袭转移的体外实验研究

目的： 探讨罗勒多糖抗卵巢癌侵袭转移的作用机制及其肿瘤乏氧微环境对其效应的影响,为临床应用研究提供依据。方法：罗勒多糖分别在常氧（21%O2、5%CO2）和乏氧（1%O2、5%CO2和94%N2）环境中作用于人卵巢癌SKOV3细胞,形态学观察不同氧环境下各组细胞形态差异;Transwell小室实验检测各组细胞的侵袭运动能力;明胶酶谱法分析各组细胞分泌的基质金属蛋白酶-2（MMP-2）活性差异;RT-PCR技术检测骨桥蛋白（OPN）mRNA表达水平;免疫细胞化学法观察各组细胞胞内OPN表达情况。结果：与对照组相比,罗勒多糖在常氧和乏氧环境下都能够降低SKOV3细胞的侵袭运动能力,抑制细胞中MMP-2的分泌,抑制人卵巢癌SKOV3细胞中OPN的表达（转录水平、蛋白水平）,且乏氧环境下罗勒多糖的上述作用更为显著。结论：低氧显著增强人卵巢癌SKOV3细胞的迁移能力,罗勒多糖可能通过下调骨桥蛋白表达及基质金属蛋白酶-2的分泌,抑制其侵袭运动能力。     

一种骨桥蛋白小分子肽的原核表达与生物学活性鉴定

目的： 用大肠杆菌（E.coli）表达骨桥蛋白13肽(OPN 13),经亲和层析纯化后检测其生物学活性。方法：运用基因重组技术,将骨桥蛋白分子中含黏附序列的13肽cDNA片段与携带His编码序列的原核表达载体连接构建融合蛋白表达质粒pET-32c-OPN13。将重组质粒转化E.coli DH 5α宿主菌后,对诱导融合蛋白表达的条件进行优化。表达产物His-OPN13经Ni-NTA His Bind Resin金属离子螯合层析纯化后,分别检测其对骨桥蛋白诱导的血管平滑肌细胞黏附和迁移的影响。结果：所表达的His-OPN13融合蛋白在宿主菌中以胞浆可溶性的形式存在。经亲和层析可得到高纯度的His-OPN13融合蛋白。产物活性分析表明,His-OPN13融合蛋白能特异性地抑制骨桥蛋白诱导的血管平滑肌细胞的黏附和迁移。结论：OPN13肽可在大肠杆菌中高效表达,并可剂量依赖性地抑制血管平滑肌细胞黏附和迁移活性。     

骨桥蛋白在肝细胞癌中的表达及其临床病理意义

C中OPN表达评分明显高于高分化HCC(P<0.05).存在转移的HCC中OPN表达评分明显高于无转移者(2.65±1.83比1.30±1.71,P<0.01).结论 HCC中OPN的表达与其临床病理有关,提示OPN可能是HCC转移复发的一个潜在指标.     

骨桥蛋白mRNA在大肠癌患者外周血中的表达及其临床意义

目的检测大肠癌外周血中的骨桥蛋白（OPN）mRNA的表达,探讨其与临床病理分期及淋巴转移的关系。方法应用逆转录聚合酶链反应技术检测52例大肠癌患者（大肠癌组）外周血单个核细胞中OPN mRNA的表达,并以33例大肠腺瘤患者（大肠腺瘤组）及20例健康人（健康对照组）外周血作为对照。结果 OPN mRNA在大肠癌组外周血单个核细胞中表达的阳性率为65.4%,大肠腺瘤组OPN mRNA阳性率为33.3%,健康对照组外周血中均无靶OPN mRNA表达。大肠癌组与大肠腺瘤组OPN mRNA阳性表达率均高于健康对照组（P均〈0.01）,大肠癌组OPN mRNA阳性表达率高于大肠腺瘤组（P〈0.05）。随着大肠癌Dukes分期的升高,其外周血单个核细胞中OPN mRNA表达阳性率逐渐增高（P〈0.05）。有淋巴结转移的表达阳性率高于无淋巴结转移及对照组（P均〈0.01）;无淋巴结转移患者与大肠腺瘤组之间差异无统计学意义。结论外周血单个核细胞中OPN mRNA阳性表达对大肠癌的淋巴转移及预后判断具有一定价值。     

一氧化氮对大鼠主动脉血管平滑肌细胞骨桥蛋白表达的影响

目的 观察一氧化氮(NO)对培养的大鼠主动脉血管平滑肌细胞骨桥蛋白表达的影响.方法 用含10%小牛血清的DMEM培养液体外培养大鼠主动脉血管平滑肌细胞,随机分为对照组和不同浓度NO供体S-亚硝基-N-乙酰青霉胺(0.5、1、2、5mmol/L)干预组,应用反转录一聚合酶链反应及Western blot技术结合光密度扫描分析,观察NO对血管平滑肌细胞的骨桥蛋白表达的影响.结果 不同浓度NO供体S-亚硝基-N-乙酰青霉胺均明显抑制血管平滑肌细胞的骨桥蛋白mRNA和蛋白的表达,且具有剂量依赖性促进作用.结论 NO能抑制血管平滑肌细胞骨桥蛋白的表达.     

Osteopontin is extensively expressed by macrophages following CNS demyelination but has a redundant role in remyelination

Osteopontin (OPN) is a key immunoregulator in the autoimmune-mediated demyelinating disease multiple sclerosis. OPN may also play a role in the remyelination since it is 1) a ligand for αV integrins, several of which regulate the properties of the oligodendrocyte precursor cells (OPCs) primarily responsible for remyelination, and 2) enhances myelin membrane formation in OPC lines. Here we show that OPN is expressed at high levels during remyelination of toxin-induced demyelination. The increased expression is due to mRNA expression in macrophages and follows differences in macrophage responses to demyelination in young and old adult animals. To identify the role of OPN in remyelination focal demyelination was induced in wild-type and OPN^−/− mice. There was no difference in the rate of remyelination between the two groups indicating that OPN is not a critical component of remyelination.