低強度脈沖超聲對兔髕骨—髕腱連接點損傷后修復早期的蛋白質(zhì)組影響
發(fā)布時間：2018-08-14 13:06
【摘要】：目的利用蛋白質(zhì)組學技術,獲取低強度脈沖超聲(Low intensity pulsed ultrasound stimulations,L工PUS)治療組與對照組兔髕骨-髕腱新生復合體的蛋白質(zhì)雙向凝膠電泳圖譜,比較兩組間蛋白質(zhì)表達譜的差異,鑒定差異蛋白質(zhì)并進行初步功能分析,探討LIPUS對骨腱連接點損傷后修復早期的分子機制。 方法成熟的新西蘭兔16只,體重2.5±0.5kg,建立兔髕骨-髕腱損傷動物模型。隨機分成兩組：(1)LIPUS治療組(T組,n=8)；(2)對照組(C組,n=8)。治療組動物于術后第3天開始LIPUS治療,每天1次,每次20min；對照組在同樣時間點予以LIPUS模擬治療(無超聲信號)。分別于治療1天、3天后收集兔髕骨-髕腱新生復合體,每個時間點動物數(shù)各4只。使用蛋白質(zhì)裂解液提取髕骨-髕腱新生復合體的總蛋白,在檢測蛋白提取液的濃度后,用固相PH梯度雙向凝膠電泳法分離蛋白質(zhì),考馬斯亮藍染色后掃描,獲取高清晰的蛋白質(zhì)圖譜；再用PDQuest-8.0.1圖象分析軟件對比分析,得到并切取差異表達的蛋白質(zhì)斑點；采用基質(zhì)輔助激光解吸電離飛行時間質(zhì)譜(MALDI-TOF-MS)進行質(zhì)譜分析后獲取肽質(zhì)量指紋圖譜；利用MascotWi zard分析軟件,在SWISS-PROT數(shù)據(jù)庫進行檢索鑒定差異蛋白質(zhì)；并利用Western-blot技術對骨-腱損傷后修復早期與LIPUS治療相關的差異蛋白質(zhì)進行驗證；查閱文獻對檢索鑒定的差異蛋白質(zhì)進行初步功能分析。 結(jié)果LIPUS治療組和對照組的雙向凝膠電泳圖譜分辨率高,重復性好；圖譜分析后發(fā)現(xiàn)兩組間存在較明顯的差異。1天和3天組共鑒定出6個差異蛋白質(zhì),分別是熱休克蛋白70、載脂蛋白A-Ⅰ、肌酸肌酶、肌動蛋白、血清前白蛋白、β血紅蛋白,這些差異蛋白質(zhì)在LIPUS治療組的表達均較對照組增高。差異蛋白質(zhì)與組織損傷后早期修復相關,包括應激保護、細胞骨架形成、炎性反應等功能。 結(jié)論LIPUS可影響兔髕骨-髕腱損傷后修復早期的蛋白質(zhì)表達,從而可能影響其修復的進程,本研究為進一步探討LIPUS修復骨腱連接點損傷的分子機制提供了一定的線索。
[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 鹵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，
本文編號：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 鹵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，
本文編號：2182946
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