低强度脉冲超声对兔髌骨—髌腱连接点损伤后修复早期的蛋白质组影响
发布时间：2018-08-14 13:06
【摘要】：目的利用蛋白质组学技术,获取低强度脉冲超声(Low intensity pulsed ultrasound stimulations,L工PUS)治疗组与对照组兔髌骨-髌腱新生复合体的蛋白质双向凝胶电泳图谱,比较两组间蛋白质表达谱的差异,鉴定差异蛋白质并进行初步功能分析,探讨LIPUS对骨腱连接点损伤后修复早期的分子机制。 方法成熟的新西兰兔16只,体重2.5±0.5kg,建立兔髌骨-髌腱损伤动物模型。随机分成两组：(1)LIPUS治疗组(T组,n=8)；(2)对照组(C组,n=8)。治疗组动物于术后第3天开始LIPUS治疗,每天1次,每次20min；对照组在同样时间点予以LIPUS模拟治疗(无超声信号)。分别于治疗1天、3天后收集兔髌骨-髌腱新生复合体,每个时间点动物数各4只。使用蛋白质裂解液提取髌骨-髌腱新生复合体的总蛋白,在检测蛋白提取液的浓度后,用固相PH梯度双向凝胶电泳法分离蛋白质,考马斯亮蓝染色后扫描,获取高清晰的蛋白质图谱；再用PDQuest-8.0.1图象分析软件对比分析,得到并切取差异表达的蛋白质斑点；采用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)进行质谱分析后获取肽质量指纹图谱；利用MascotWi zard分析软件,在SWISS-PROT数据库进行检索鉴定差异蛋白质；并利用Western-blot技术对骨-腱损伤后修复早期与LIPUS治疗相关的差异蛋白质进行验证；查阅文献对检索鉴定的差异蛋白质进行初步功能分析。 结果LIPUS治疗组和对照组的双向凝胶电泳图谱分辨率高,重复性好；图谱分析后发现两组间存在较明显的差异。1天和3天组共鉴定出6个差异蛋白质,分别是热休克蛋白70、载脂蛋白A-Ⅰ、肌酸肌酶、肌动蛋白、血清前白蛋白、β血红蛋白,这些差异蛋白质在LIPUS治疗组的表达均较对照组增高。差异蛋白质与组织损伤后早期修复相关,包括应激保护、细胞骨架形成、炎性反应等功能。 结论LIPUS可影响兔髌骨-髌腱损伤后修复早期的蛋白质表达,从而可能影响其修复的进程,本研究为进一步探讨LIPUS修复骨腱连接点损伤的分子机制提供了一定的线索。
[Abstract]:Objective to obtain the protein two-dimensional gel electrophoresis patterns of patellar and patellar tendon neonate complex in rabbits of low intensity pulsed ultrasound (Low intensity pulsed ultrasound) stimulationsl (PUS) treatment group and control group by proteomics technique, and to compare the differences of protein expression profiles between the two groups. Differential proteins were identified and preliminary functional analysis was carried out to explore the early molecular mechanism of LIPUS repair after bone tendon junction injury. Methods Sixteen mature New Zealand rabbits, weighing 2.5 卤0.5 kg, were used to establish patellar tendon injury model. They were randomly divided into two groups: (1) LIPUS treatment group (T group) and control group (C group). The animals in the treatment group were treated with LIPUS once a day for 20 minutes on the 3rd day after operation, and the control group were treated with LIPUS at the same time (no ultrasound signal). The patellar-patellar tendon neovascularization complex was collected at 1 day and 3 days after treatment with 4 animals at each time point. Protein lysate was used to extract the total protein of patellar and patellar tendon neonate complex. After detecting the concentration of protein extract, protein was separated by solid phase PH gradient two dimensional gel electrophoresis. Coomassie brilliant blue staining was used to scan the protein. High definition protein map was obtained, and then the protein spots of differential expression were obtained and analyzed by PDQuest-8.0.1 image analysis software. Matrix assisted laser desorption / ionization time of flight mass spectrometry (MALDI-TOF-MS) was used to obtain the peptide mass fingerprint, and MascotWi zard software was used to search and identify the differential proteins in the SWISS-PROT database. Western-blot technique was used to verify the differential proteins related to LIPUS treatment in the early stage of repair after bone-tendon injury, and the preliminary functional analysis of the differentially identified proteins was made by consulting the literature. Results the two dimensional gel electrophoresis patterns of LIPUS treatment group and control group had high resolution and good reproducibility. Heat shock protein 70, apolipoprotein A- 鈪，
本文编号：2182946
[Abstract]:Objective to obtain the protein two-dimensional gel electrophoresis patterns of patellar and patellar tendon neonate complex in rabbits of low intensity pulsed ultrasound (Low intensity pulsed ultrasound) stimulationsl (PUS) treatment group and control group by proteomics technique, and to compare the differences of protein expression profiles between the two groups. Differential proteins were identified and preliminary functional analysis was carried out to explore the early molecular mechanism of LIPUS repair after bone tendon junction injury. Methods Sixteen mature New Zealand rabbits, weighing 2.5 卤0.5 kg, were used to establish patellar tendon injury model. They were randomly divided into two groups: (1) LIPUS treatment group (T group) and control group (C group). The animals in the treatment group were treated with LIPUS once a day for 20 minutes on the 3rd day after operation, and the control group were treated with LIPUS at the same time (no ultrasound signal). The patellar-patellar tendon neovascularization complex was collected at 1 day and 3 days after treatment with 4 animals at each time point. Protein lysate was used to extract the total protein of patellar and patellar tendon neonate complex. After detecting the concentration of protein extract, protein was separated by solid phase PH gradient two dimensional gel electrophoresis. Coomassie brilliant blue staining was used to scan the protein. High definition protein map was obtained, and then the protein spots of differential expression were obtained and analyzed by PDQuest-8.0.1 image analysis software. Matrix assisted laser desorption / ionization time of flight mass spectrometry (MALDI-TOF-MS) was used to obtain the peptide mass fingerprint, and MascotWi zard software was used to search and identify the differential proteins in the SWISS-PROT database. Western-blot technique was used to verify the differential proteins related to LIPUS treatment in the early stage of repair after bone-tendon injury, and the preliminary functional analysis of the differentially identified proteins was made by consulting the literature. Results the two dimensional gel electrophoresis patterns of LIPUS treatment group and control group had high resolution and good reproducibility. Heat shock protein 70, apolipoprotein A- 鈪，
本文编号：2182946
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