本文關鍵詞： 兔 骨缺損 植骨 同種異體 富血小板血漿 修復　出處：《河北醫(yī)科大學》2017年碩士論文　論文類型：學位論文

【摘要】：目的:本實驗旨在觀察不同時間同種異體富血小板血漿聯(lián)合同種異體骨植入對骨缺損修復的影像學和組織學變化,以探討同種異體富血小板血漿對骨缺損植骨修復的療效,從而為臨床工作提供實驗理論依據(jù)及指導意見。方法:1實驗分組:選取健康新西蘭大白兔36只,于兔的雙下肢股骨內(nèi)側髁建立骨缺損的模型后按分籠飼養(yǎng)編號隨機分為A、B兩組,A組(實驗組,n=18),同種異體骨復合同種異體富血小板血漿植入;B組(對照組,n=18),單純植入同種異體骨,另取6只單純建立骨缺損模型,缺損不做處理作為模型組。2同種異體PRP的制備:心臟取血50ml,分別移入10ml離心管進行二次離心,第一次以200g離心力,離心10min,第二次同樣以200g離心力離心,時間同上,制備PRP約1ml,將PRP與同種異體骨按2:1比例混合激活后,制成復合型PRP凝膠。3血小板計數(shù):心臟取血后,使用血細胞分析儀,取少量血液進行血小板計數(shù),記錄全血中血小板數(shù),參考Landesberg法通過兩次離心法制備PRP,同樣對PRP進行血小板計數(shù),驗證二次離心法制備出的PRP是否滿足實驗要求。4觀察及評價指標:第4、8、12周時,采用過量麻醉的方法處死各組實驗動物,首先設定特定的曝光條件攝股骨髁側位X光片(距離1m,條件220KV,30mA.曝光時間0.15s),觀察骨缺損植骨愈合情況。參照Yang氏放射學評分標準進行評分,并通過X線平片測量記錄其骨缺損區(qū)上、中、下三點的阻射密度值,取平均值進行統(tǒng)計分析。而后截取標本對其進行大體標本觀察,觀察植入材料的表面變化和骨組織缺損區(qū)域植入骨的生長修復情況。制作病理組織切片及免疫組化染色,光學顯微鏡下觀察兔骨缺損再生與修復情況;并根據(jù)Lane氏組織學評分標準對每組動物的骨修復情況進行評分;應用顯微圖像分析系統(tǒng)(HMIAS-2000)和專業(yè)圖像分析軟件(Image-Pro Plus 6.0)計算其免疫組化切片的染色陽性細胞平均光密度(Mean Optical Density,MOD)值,進行統(tǒng)計學分析。結果:1一般情況及大體觀察:術后所有動物切口愈合良好,從缺損邊界和骨痂形成情況,術后4周和8周時,A組比B組缺損區(qū)組織修復骨缺損較平整,修復較好,術后12周,A組和B組骨組織缺損區(qū)域均得到完全修復,邊界已無法分辨,部分可見異位骨化;2全血及PRP血小板計數(shù)情況:PRP中血小板數(shù)量為全血中的(4.30±0.28)倍,符合Landesberg法通過兩次離心制備PRP的要求,滿足了目前實驗需求;3影像學評分及觀察結果:術后4、8、12周各時間點A組與B組兩組放射學評分比較差異具有統(tǒng)計學意義(P0.05),A組評分顯著高于B組;而A組與B組兩組術后各期X線阻射密度比較,A組阻射密度值優(yōu)于B組,差異具有統(tǒng)計學意義(P0.05);4組織學評分及鏡下觀察情況:術后4、8、12周各時期A組和B組Lane組織學評分比較,實驗組評分均高于對照組,且比較差異具有統(tǒng)計學意義(P0.05),光學顯微鏡下觀察結果顯示術后各時間點實驗組成骨的面積均高于對照組,兩組比較差異具有統(tǒng)計學意義(P0.05);而免疫組化觀察結果顯示第4和8周時A組PDGF、TGF-β免疫組化切片平均光密度(MOD)明顯高于B組,比較差異具有統(tǒng)計學意義(P0.05),說明第4和8周時PDGF、TGF-β蛋白的表達量高于對照組;術后第12周時,PDGF、TGF-β蛋白的表達量均較第4和8周時下降,且實驗組與對照組比較差異沒有統(tǒng)計學意義(P0.05)。結論:1同種異體富血小板血漿與異體骨組織復合后,能夠促進骨組織生成,加快骨缺損修復。2血小板源性生長因子(PDGF)和轉化生長因子(TGF-β)于8周后顯著下降,是早期誘導骨組織再生的重要生長因子。3術后12周時模型組的骨缺損處仍未完全修復,證明實驗模型成立。
[Abstract]:Objective: To study the change of this experiment is to observe the different time of allogeneic platelet rich plasma combined with bone allograft implantation for repairing bone defect imaging and tissue, to investigate the effect of allogeneic platelet rich plasma clinical effects of bone grafting on the repair of bone defect, so as to provide the experimental basis and theoretical guidance for clinical work. Methods: 1 experimental groups: 36 New Zealand rabbits were selected in rabbit limb medial femoral condyle to establish bone defect model by feeding numbers were randomly divided into A, B two groups, A group (experimental group, n=18), allogeneic bone allograft combined with platelet rich plasma implantation group (control group, B; n=18), simple implantation of allograft bone, another 6 only to establish bone defect model, defect not treated as model group.2 allogenic PRP preparation: heart blood 50ml, were transferred to 10ml centrifuge tube two centrifugal, first to 200g centrifugal Centrifugal force, 10min, second times the same with 200g centrifugal force, the time above, the preparation of PRP about 1ml, PRP and allograft bone mixed according to the proportion of 2:1 after activation, made of composite PRP gel.3 platelet count: blood from the heart, blood cell analyzer, take a small amount of blood in the platelet count, recording whole blood platelet count, Landesberg method two PRP were prepared by centrifugal method, also the platelet count of PRP, verify whether the prepared two PRP centrifugal method can meet the requirement of.4 observation and evaluation index: 4,8,12 weeks, using the method of excessive anesthesia sacrificed animal, first set the exposure condition specific perturbation the lateral femoral condyle X ray (1m 220KV, 30mA. distance, exposure time 0.15s), to observe the bone defect bone graft healing. According to Yang's radiological score standard for evaluation, and by measuring the bone defect records of X Xianping In the area, resistance, density three point value, average value of statistical analysis. Then the interception were gross observation on the repair of growth observed surface changes of implant materials and bone defect area of bone. Making histological and immunohistochemical staining, observe the regeneration of bone defect in rabbits with the restoration of optical microscope; and score according to the standard for evaluation of Lane's bone repair in each animal; the application of image analysis system (HMIAS-2000) and the professional image analysis software (Image-Pro Plus 6) positive cells to calculate the average optical density of immunohistochemical slices (Mean Optical Density, MOD) the value for statistical analysis. Results: 1 the general situation and general observation: after all animal wound healing well from the defect boundary and callus formation, after 4 and 8 weeks, A group than in B group of defects Tissue repair bone defect repair is better than the flat, 12 weeks after operation, A group and B group of bone defects were completely repaired, the boundary has been unable to distinguish, partially visible heterotopic ossification; 2 PRP and blood platelet count: the number of platelets in the blood of PRP (4.30 + 0.28) times, according to Landesberg by the method of two centrifugal PRP preparation requirements, meet the experimental requirements; score and observation results of 3 imaging: statistically significant 4,8,12 weeks each time point in A group and B group two radiology group after the score difference (P0.05), A group was significantly higher than B group; compared their X-ray the density of A group and B group after operation in two groups, A group resistance, density value is better than that of B group, the difference was statistically significant (P0.05); 4 histological grading and microscopic observation: 4,8,12 weeks after surgery in each period of Lane group A group and B group was compared with the experimental group were higher than those in the control group and more, The difference was statistically significant (P0.05), optical microscope observation showed that the experimental group at each time point after operation of bone area were higher than the control group, with significant differences between the two groups (P0.05); and immunohistochemistry results showed that fourth and 8 weeks in group A, PDGF, TGF- P immunohistochemistry average light density (MOD) was significantly higher than that of B group, the difference was statistically significant (P0.05), fourth and 8 weeks of PDGF, the expression of TGF- protein was higher than that in the control group; twelfth weeks after the operation, PDGF, the expression of TGF- beta protein were fourth and 8 weeks of the fall, and the comparison between the experimental group and the the control group was not statistically significant difference (P0.05). Conclusion: 1 allogeneic platelet rich plasma and bone allograft composite tissue, can promote bone formation and accelerate bone repair.2 platelet-derived growth factor (PDGF) and transforming growth factor (TGF-) in 8 weeks decreased significantly early The bone defect of the model group was still not completely repaired at 12 weeks after the important growth factor.3, which proved that the experimental model was established.

【學位授予單位】：河北醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R687.3

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