本文選題：前交叉韌帶 + 脫細(xì)胞真皮基質(zhì)　； 參考：《第四軍醫(yī)大學(xué)》2013年碩士論文

【摘要】：目的：隨著對(duì)組織工程ACL構(gòu)建方法研究的不斷深入，發(fā)現(xiàn)多種細(xì)胞生長(zhǎng)因子在ACL愈合過(guò)程中起著關(guān)鍵的調(diào)控作用。其中TGF-β1和bFGF已成為研究ACL愈合的外源性生長(zhǎng)因子的熱點(diǎn)。ADM作為多種組織修復(fù)支架材料，其在臨床中修復(fù)組織缺損已得到廣泛應(yīng)用。本實(shí)驗(yàn)的目的在于探討特定濃度TGF-β1和bFGF對(duì)BMSCs在ADM支架上粘附，增值和分化的影響。并通過(guò)不同編織ADM重建兔ACL，比較重建ACL腱骨愈合，膠原含量分布和生物力學(xué)強(qiáng)度，從而為構(gòu)建組織工程ACL提供實(shí)驗(yàn)依據(jù)。 方法：1、將培養(yǎng)至第三代的兔BMSCs接種于ADM材料上，分別用普通培養(yǎng)液（對(duì)照組）及TGF-β1(濃度10ng/ml)和bFGF(濃度25ng/ml)培養(yǎng)液（實(shí)驗(yàn)組）培養(yǎng)，通過(guò)分別繪制第1、3、5、7、9、11天細(xì)胞增殖MTT曲線，第7天測(cè)定ALP含量，熒光顯微鏡及激光共聚焦掃描顯微鏡觀察，分析編織ADM支架基本力學(xué)性能，以及兩組培養(yǎng)液對(duì)細(xì)胞在ADM支架材料中的生長(zhǎng)增殖，分化和粘附的影響；2、通過(guò)編織4股束形支架（對(duì)照組）和4股柱形支架（實(shí)驗(yàn)組）重建兔ACL，分別取材第6、12周重建ACL，通過(guò)大體觀察關(guān)節(jié)功能和軟骨退變、腱-骨HE染色、重建ACL天狼猩紅染色及生物力學(xué)測(cè)試，分析兩組編織方法ADM支架的力學(xué)性能，及細(xì)胞在支架上生長(zhǎng)增值，，分泌I，III型膠原以及腱-骨愈合的差異。3、應(yīng)用SPSS12.0統(tǒng)計(jì)軟件對(duì)數(shù)據(jù)進(jìn)行統(tǒng)計(jì)學(xué)分析，P0.05為統(tǒng)計(jì)學(xué)差異標(biāo)準(zhǔn)。 結(jié)果：1、MTT曲線及熒光染色結(jié)果表明實(shí)驗(yàn)組培養(yǎng)液BMSCs在ADM支架上的粘附數(shù)量更多（P0.05），ALP染色證明TGF-β1(濃度10ng/ml)和bFGF(濃度25ng/ml)能夠促進(jìn)BMSCs細(xì)胞分泌更多的ALP(P0.05)，激光共聚焦掃描顯微鏡顯示實(shí)驗(yàn)組細(xì)胞粘附數(shù)量多，細(xì)胞形態(tài)更優(yōu)良。2、重建兔ACL術(shù)后6、12周腱-骨及ACL染色表明，實(shí)驗(yàn)組較對(duì)照組腱-骨愈合速度更快，兩組ACL分泌I、 III型膠原情況基本相當(dāng)。術(shù)后12周，生物力學(xué)測(cè)試結(jié)果實(shí)驗(yàn)組ACL的生物力學(xué)性能明顯優(yōu)于對(duì)照組(P0.05)。 結(jié)論：1、TGF-β1（濃度10ng/ml）及bFGF(濃度25ng/ml)培養(yǎng)液對(duì)BMSCs在ADM上的生長(zhǎng)，粘附及分化有明顯的刺激增強(qiáng)效果。2、編織ADM支架具備一定的生物力學(xué)強(qiáng)度，其中4股編織柱形ADM支架在兔體內(nèi)能夠促進(jìn)細(xì)胞分泌I,III型膠原，并提供較好的腱-骨愈合速度及生物力學(xué)強(qiáng)度。
[Abstract]:Aim: with the further study of ACL construction methods in tissue engineering, it was found that many cell growth factors play a key role in the healing of ACL. Among them, TGF- 尾 1 and bFGF have become the focus of exogenous growth factor in the study of ACL healing. ADM is used as a variety of scaffolds for tissue repair, and it has been widely used in clinical repair of tissue defects. The purpose of this study was to investigate the effects of specific concentrations of TGF- 尾 1 and bFGF on the adhesion, proliferation and differentiation of BMSCs on ADM scaffolds. The bone healing, collagen content distribution and biomechanical strength of reconstructed ACL tendon were compared by different braided ADM to provide experimental basis for the construction of tissue engineering ACL. Methods the third generation rabbit BMSCs was inoculated on the ADM material and cultured with normal culture medium (control group), TGF- 尾 1 (10 ng / ml) and bFGF- 尾 1 (25 ng / ml) culture medium (experimental group). The MTT curves of cell proliferation were plotted on the 7911 days of the first day, respectively. On the 7th day, the content of ALP was measured, the basic mechanical properties of braided ADM scaffolds were analyzed by fluorescence microscope and laser confocal scanning microscope, and the growth and proliferation of cells in ADM scaffold were also analyzed. ACLs were reconstructed by weaving 4 strands (control group) and 4 strands scaffolds (experimental group). ACLs were reconstructed at week 612, respectively. The articular function and cartilage degeneration were observed, and tendon-bone HE staining was performed. ACL sirius red staining and biomechanical test were performed to analyze the mechanical properties of ADM scaffolds and the growth of cells on the scaffolds. The secretion of type III collagen and tendon-bone union was different. The statistical analysis of the data by SPSS12.0 software was used as the standard of statistical difference (P0.05). Results the results of BMSCs and fluorescence staining showed that the adherent quantity of BMSCs on ADM scaffold was more than that of TGF- 尾 1 (10 ng / ml) and bFGF- 尾 1 (25 ng / ml). Laser confocal scanning microscope (confocal scanning microscope) showed that TGF- 尾 1 (10 ng / ml) and bFGF (25 ng / ml) could promote the secretion of ALP P0.05 by BMSCs cells. The results showed that the number of cell adhesion in experimental group was high, The morphology of the cells was better. The results of tendon bone and ACL staining at 612 weeks after ACL reconstruction showed that the healing rate of tendon-bone in the experimental group was faster than that in the control group. The level of I and III collagen secreted by ACL in the two groups was almost the same. 12 weeks after operation, the biomechanical properties of ACL in the experimental group were significantly better than those in the control group (P 0.05). Conclusion: TGF- 尾 _ 1 (10 ng / ml) and bFGF (25 ng / ml) can stimulate the growth, adhesion and differentiation of BMSCs on ADM significantly. The braided ADM scaffolds have a certain biomechanical strength. Among them, 4 strands of columnar ADM scaffolds could promote the secretion of type III collagen in rabbit cells, and provide a better rate of tendon-bone healing and biomechanical strength.
【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2013
【分類號(hào)】：R686.5

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本文編號(hào)：1969094