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

【摘要】：目的:本實(shí)驗(yàn)旨在觀察不同時(shí)間同種異體富血小板血漿聯(lián)合同種異體骨植入對(duì)骨缺損修復(fù)的影像學(xué)和組織學(xué)變化,以探討同種異體富血小板血漿對(duì)骨缺損植骨修復(fù)的療效,從而為臨床工作提供實(shí)驗(yàn)理論依據(jù)及指導(dǎo)意見(jiàn)。方法:1實(shí)驗(yàn)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(cè)髁建立骨缺損的模型后按分籠飼養(yǎng)編號(hào)隨機(jī)分為A、B兩組,A組(實(shí)驗(yàn)組,n=18),同種異體骨復(fù)合同種異體富血小板血漿植入;B組(對(duì)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(jìn)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(shí)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(fù)合型PRP凝膠。3血小板計(jì)數(shù):心臟取血后,使用血細(xì)胞分析儀,取少量血液進(jìn)行血小板計(jì)數(shù),記錄全血中血小板數(shù),參考Landesberg法通過(guò)兩次離心法制備PRP,同樣對(duì)PRP進(jìn)行血小板計(jì)數(shù),驗(yàn)證二次離心法制備出的PRP是否滿足實(shí)驗(yàn)要求。4觀察及評(píng)價(jià)指標(biāo):第4、8、12周時(shí),采用過(guò)量麻醉的方法處死各組實(shí)驗(yàn)動(dòng)物,首先設(shè)定特定的曝光條件攝股骨髁側(cè)位X光片(距離1m,條件220KV,30mA.曝光時(shí)間0.15s),觀察骨缺損植骨愈合情況。參照Yang氏放射學(xué)評(píng)分標(biāo)準(zhǔn)進(jìn)行評(píng)分,并通過(guò)X線平片測(cè)量記錄其骨缺損區(qū)上、中、下三點(diǎn)的阻射密度值,取平均值進(jìn)行統(tǒng)計(jì)分析。而后截取標(biāo)本對(duì)其進(jìn)行大體標(biāo)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(zhǎng)修復(fù)情況。制作病理組織切片及免疫組化染色,光學(xué)顯微鏡下觀察兔骨缺損再生與修復(fù)情況;并根據(jù)Lane氏組織學(xué)評(píng)分標(biāo)準(zhǔn)對(duì)每組動(dòng)物的骨修復(fù)情況進(jìn)行評(píng)分;應(yīng)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(jì)算其免疫組化切片的染色陽(yáng)性細(xì)胞平均光密度(Mean Optical Density,MOD)值,進(jìn)行統(tǒng)計(jì)學(xué)分析。結(jié)果:1一般情況及大體觀察:術(shù)后所有動(dòng)物切口愈合良好,從缺損邊界和骨痂形成情況,術(shù)后4周和8周時(shí),A組比B組缺損區(qū)組織修復(fù)骨缺損較平整,修復(fù)較好,術(shù)后12周,A組和B組骨組織缺損區(qū)域均得到完全修復(fù),邊界已無(wú)法分辨,部分可見(jiàn)異位骨化;2全血及PRP血小板計(jì)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(guò)兩次離心制備PRP的要求,滿足了目前實(shí)驗(yàn)需求;3影像學(xué)評(píng)分及觀察結(jié)果:術(shù)后4、8、12周各時(shí)間點(diǎn)A組與B組兩組放射學(xué)評(píng)分比較差異具有統(tǒng)計(jì)學(xué)意義(P0.05),A組評(píng)分顯著高于B組;而A組與B組兩組術(shù)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(jì)學(xué)意義(P0.05);4組織學(xué)評(píng)分及鏡下觀察情況:術(shù)后4、8、12周各時(shí)期A組和B組Lane組織學(xué)評(píng)分比較,實(shí)驗(yàn)組評(píng)分均高于對(duì)照組,且比較差異具有統(tǒng)計(jì)學(xué)意義(P0.05),光學(xué)顯微鏡下觀察結(jié)果顯示術(shù)后各時(shí)間點(diǎn)實(shí)驗(yàn)組成骨的面積均高于對(duì)照組,兩組比較差異具有統(tǒng)計(jì)學(xué)意義(P0.05);而免疫組化觀察結(jié)果顯示第4和8周時(shí)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(jì)學(xué)意義(P0.05),說(shuō)明第4和8周時(shí)PDGF、TGF-β蛋白的表達(dá)量高于對(duì)照組;術(shù)后第12周時(shí),PDGF、TGF-β蛋白的表達(dá)量均較第4和8周時(shí)下降,且實(shí)驗(yàn)組與對(duì)照組比較差異沒(méi)有統(tǒng)計(jì)學(xué)意義(P0.05)。結(jié)論:1同種異體富血小板血漿與異體骨組織復(fù)合后,能夠促進(jìn)骨組織生成,加快骨缺損修復(fù)。2血小板源性生長(zhǎng)因子(PDGF)和轉(zhuǎn)化生長(zhǎng)因子(TGF-β)于8周后顯著下降,是早期誘導(dǎo)骨組織再生的重要生長(zhǎng)因子。3術(shù)后12周時(shí)模型組的骨缺損處仍未完全修復(fù),證明實(shí)驗(yàn)?zāi)Ｐ统闪ⅰ?br/>[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(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R687.3

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