骨调素和单核细胞趋化蛋白在大鼠梗阻性肾病中的表达及可能作用

目的:探讨骨调素(OPN)和单核细胞趋化蛋白(MCP-1)在大鼠梗阻性模型中的表达及其在肾脏纤维化发病机制中的作用.方法:采用-单侧输尿管结扎制造梗阻性肾病模型,分别于造模后7 d、14 d取肾组织,应用HE染色观察肾脏病理改变,免疫组化方法检测肾组织畔OPN和MCP-1蛋白的表达,应用逆转录-聚合酶链式反应(RT-PCR)法观察肾组织中OPN mRNA和MCP-1 mRNA的变化.结果:OPN、MCP-1表达主要位于肾小管上皮细胞,随着梗阻时间的延长,肾组织中OPN、MCP-1蛋白和mRNA表达明显增加.结论:OPN、MCP-1蛋白和mRNA在梗阻性肾病大鼠肾组织表达明显增加介导炎症过程,参与肾间质纤维化.     

过量氟对大鼠肾组织骨桥蛋白表达的影响

目的检测骨桥蛋白(OPN)mRNA和蛋白在氟处理大鼠肾组织中的表达差异和蛋白定位,并探讨OPN与氟中毒肾损害的关系。方法经饮水投氟喂养Wistar大鼠48只,用免疫组织化学法进行骨桥蛋白的组织学检测。提取各组大鼠肾组织mRNA,采用RT-PCR法检测骨桥蛋白mRNA的表达变化。结果免疫组织化学结果表明,骨桥蛋白主要定位于肾组织的肾小管上皮细胞质中,常食和偏食对照组大鼠肾组织OPN蛋白只是散在表达,而常食和偏食加氟组的OPN蛋白表达较相应对照组明显增强(与常食对照组比较,q=18．4,P< 0．05;与偏食对照组比较,q=12．0,P<0．01)。2个投氟组的OPN mRNA表达水平均较相对应的对照组增加,但差异无统计学意义。结论过量氟可促进大鼠。肾组织OPN表达,且随着投氟量的增加,OPN伴随着肾损伤加重而表达增强,提示OPN与氟中毒肾损伤程度密切相关,很可能作为一种氟中毒肾损害的预测标志。     

骨桥蛋白在人脑动静脉畸形的表达及意义

目的研究骨桥蛋白(OPN)在脑动静脉畸形(CAVM)的血管组织及其在放射、栓塞治疗后血管病理变化中的表达。方法采用免疫组化方法检测42例CAVM病理标本及对照组10例内减压手术所获脑组织血管中OPN的表达。结果26例无术前治疗史的畸形血管组织中22例有OPN的表达,主要见于CAVM的静脉部分,在粥样硬化样病变的动脉处也有表达。对照组脑组织的血管中未见OPN的表达;有伽玛刀治疗史的5例中,2例在早、中期放射反应的动脉中可见OPN的明显表达;在经历栓塞治疗的11例中,7例在新生内膜组织或异物巨细胞中有强阳性表达。结论人CAVM血管组织中多有OPN的表达,这可能是其适应于适应血流动力学状态而具有的血管重塑的表现,并可能在放射及栓塞治疗后的血管重塑中发挥重要作用。     

骨桥蛋白在人脑胶质瘤中的表达及其临床意义

目的探讨骨桥蛋白在人脑星形胶质细胞瘤中的表达及其临床意义。方法应用免疫组织化学方法检测58例胶质瘤和10例正常脑组织中骨桥蛋白的表达，分析其表达与肿瘤病理分级和预后的关系。结果在正常脑组织中未见骨桥蛋白的表达。Ⅰ-Ⅳ级肿瘤中骨桥蛋白的阳性细胞率分别为11.3％±6．2％、21．8％±11．2％和36．7％±18．3％和49．4％±16．5％，差异有显著性（P〈0．05）。骨桥蛋白高表达和低表达患者2年生存率分别为40．6％（13／32）和76．9％（20／26），差异显著（P〈0．05）。结论骨桥蛋白在胶质瘤的浸润性生长中可能起重要作用，其表达与肿瘤分化程度和患者预后密切相关。     

Transient expression of osteopontin mRNA and protein in amoeboid microglia in developing rat brain

To investigate a potential role of osteopontin (OPN) in developing rat brain, the expression of OPN mRNA and protein in the developing rat brain relative to the distribution of brain macrophages was investigated using in situ hybridization and immunohistochemistry, and the phagocytic capability of OPN-expressing cells was accessed using rhodamine isothiocyanate (RhIc) as a tracer. OPN-expressing cells appeared from embryonic day 16. During the first week of postnatal life, OPN-labeled cells increased markedly, and peaked around P7, then declined and had completely disappeared by the end of the second postnatal week. The spatiotemporal distribution pattern of OPN mRNA closely matched that of OPN protein. Their morphology and localization were compared with those of cells expressing the established microglial marker OX-42 in adjacent sections, and double-labeling studies demonstrated that OPN was localized to the amoeboid microglia which stain with the lectin GSI-B₄, another marker for microglia. Furthermore, OPN-labeled cells were confirmed to be active phagocytes emitting RhIc fluorescence indicating that the tracer into the brain tissues was engulfed by phagocytosis. Therefore, these results provide the first evidence that OPN is transiently expressed in active brain macrophages in the embryonic and early postnatal brain, and suggest that OPN may contribute to the migration and phagocytic function of brain macrophages in the developing brain.This work was supported by a Korea Research Foundation grant (KRF-2002-015-EP0106)     

低氧和高二氧化碳作用下肺动脉高压形成过程中骨桥蛋白表达与分布的实验研究

目的： 观察慢性低O2高CO2大鼠肺动脉骨桥蛋白（OPN）及其基因的表达与分布情况，探讨OPN在肺动脉高压发病机制中的作用。方法： 48只雄性SD大鼠随机分为4组：即正常对照组（NC组），低O2高CO2 1周、2周和4周组（1HH、2HH和4HH）。采用RT-PCR法测定不同低O2高CO2 组大鼠离体肺动脉和肺组织骨桥蛋白（OPN）mRNA，免疫组织化学染色法测定肺细小动脉骨桥蛋白分布表达情况，ELISA 法定量测定肺组织匀浆OPN水平，Western blotting 法测定离体肺动脉OPN表达水平。结果： ①低O2高CO2各组大鼠mPAP和RV/LV+S均明显高于NC组（P<0.01），4组大鼠间mCAP比较无显著差异（P>0.05）；②低O2高CO2 各组大鼠肺组织和离体肺主动脉OPN mRNA表达水平显著高于对照组（均P<0.01）；③免疫组化法显示NC组大鼠肺组织OPN表达主要见于支气管、肺泡上皮细胞内，1HH组大鼠肺细小动脉内皮细胞、平滑肌细胞以及周围巨噬细胞均见有OPN的表达，其中以中膜平滑肌细胞最为明显，而2HH和4HH组大鼠OPN表达更加明显, 低O2高CO2 各组肺细小动脉OPN相对含量与对照组比较有显著差异（均P<0.01），且随缺氧时间的延长，骨桥蛋白表达也呈增高趋势；④ 低O2高CO2 各组大鼠肺组织匀浆OPN含量分别为正常对照组的1.69倍、2.28倍和2.87倍（均P<0.01）；⑤ Western blotting 分析1HH、2HH和4HH各组大鼠离体肺动脉OPN表达明显增强，与NC组比较有显著差异（均P<0.01）。结论： 慢性低O2高CO2能够刺激大鼠肺组织和肺动脉OPN及其基因表达增强，OPN可能在慢性低O2高CO2性肺动脉高压发病的病理和病理生理过程中起着重要的作用。     

血清脂联素、骨桥蛋白和HE4在子宫内膜癌患者中的表达及意义

目的：探讨血清脂联素（APN）、骨桥蛋白（OPN）和人附睾分泌蛋白4(HE4)在子宫内膜癌患者中的表达及意义。方法：选取2019年11月至2022年9月郴州市第一人民医院收治的子宫内膜癌患者62例（子宫内膜癌组）,另选取同期50例子宫内膜不典型增生患者（内膜增生组）和50例健康志愿者（正常内膜组）。检测并比较三组血清APN、OPN、HE4水平,采用受试者工作特征（ROC）曲线分析血清APN、OPN、HE4对子宫内膜癌的诊断效能,并采用Spearman相关性分析探讨血清APN、OPN、HE4与临床病理特征的相关性。结果：子宫内膜癌组患者血清APN水平低于内膜增生组和正常内膜组,OPN、HE4水平高于内膜增生组和正常内膜组。国际妇产科联盟（FIGO）分期为Ⅲ期患者血清APN水平低于Ⅱ期和Ⅰ期患者,OPN、HE4水平高于Ⅱ期和Ⅰ期患者;肌层浸润深度≥1/2肌层患者血清APN水平低于<1/2肌层患者,OPN、HE4水平高于<1/2肌层患者;淋巴结转移患者血清APN水平低于无淋巴结转移患者,OPN、HE4水平高于无淋巴结转移患者,差异均具有统计学意义（P <0.05）。ROC曲...     

Microsomal casein kinase II in endoplasmic reticulum- and Golgi apparatus-rich fractions of ROS 17/2.8 osteoblast-like cells: An enzyme that modifies osteopontin

Osteopontin is an acidic phosphoprotein containing casein kinase II (CKII) phosphorylatable sites and an acidic amino acid cluster. The metabolically ³²P-labelings of both serines and threonines in vitro in osteopontin immunoprecipitated from rat osteoblast-like ROS 17/2.8 cells may suggest that casein kinase II catalyzes this modification. The enzyme occurs in microsomal fractions of rat osteoblast-like ROS 17/2.8 cells. Subcellular fractions containing endoplasmic reticulum and Golgi apparatus were isolated by differential centrifugation and were identified according to their ultrastructures and the presence of marker enzymes such as glucose-6-phosphatase and thiamine pyrophosphatase, respectively. Both fractions phosphorylated the partially dephosphorylated osteopontin and the specific substrate peptide RRREEETEEE. Endoplasmic reticulum-catalyzed peptide phosphorylation was 2.7 times lower than that of Golgi although both endoplasmic reticulum- and Golgi-catalyzed peptide reactions were 50% inhibited by 20 and 100 ng/ml heparin, respectively. Western blot analysis revealed that both fractions contained osteopontin and microsomal CKII. Furthermore, microsomal CKII was immunogold-labeled in endoplasmic reticulum and Golgi apparatus. Heparin inhibition and utilization of [-³²P]GTP as a phosphate donor by both fractions confirmed their capacity to phosphorylate osteopontin. The results suggest that microsomal CKII modifies the acidie matrix proteins during transportation. These matrix phosphoproteins may participate in the mineralization process of hard tissues.     

骨桥蛋白反义基因重组质粒的构建

目的 构建反义骨桥蛋白(OPN)基因重组质粒，以用于肝癌基因治疗的研究。方法 通过设计含有特定酶切位点的引物，以含有全长骨桥蛋白基因的质粒(pBlueScript—OPN)为模板，将PCR产物反向克隆至pcDNA3．1( )真核表达载体上，并经酶切鉴定及测序分析。结果重组质粒经酶切后出现913bp长的片段。测序分析结果与献报道结果完全一致。结论 反义骨桥蛋白基因表达质粒克隆成功，为恶性肿瘤的基因治疗提供了一种有效手段。