本文選題：多孔支架 + 孔隙結構可控　； 參考：《西南交通大學》2016年博士論文

【摘要】：在骨組織工程支架中,多孔支架的物理結構特性是影響細胞行為、血管生長和骨形成的關鍵因素。它不但為細胞的生長和遷移提供模板,同時也為血管生長和骨形成提供充足的空間。雖然優(yōu)化多孔支架結構特性可有效促進骨組織形成,但是單純調控支架的物理結構促進骨再生的能力仍然是有限的。一些研究表面引入外源性生長因子和藥物可顯著增強骨組織再生,因此多孔支架與生物因子或藥物有效結合協(xié)同調控血管生長和骨形成受到越來越多的關注。在本研究中,通過調控支架的結構參數(shù)(孔徑、貫通性、孔徑分布和表面形貌)探索多孔羥基磷灰石(HA)支架結構特性與血管生長和異位骨形成的關系,進一步優(yōu)化支架的多孔結構。此外,通過將載藥高分子微球均勻分布在涂覆有海藻酸的多孔HA支架孔壁表面構建多孔支架緩釋體系,以增強多孔HA支架血管生長和骨形成能力。采用糖球造孔法通過去除糖球制備孔隙結構可控的3類多孔HA支架,控制糖球顆粒的粒徑和熱處理工藝精確調控多孔支架的宏孔孔徑和貫通孔尺寸。在本研究中,通過熱處理工藝將宏孔孔徑分別為500-650、700-950和1100-1250 μm的多孔HA支架的貫通孔尺寸/宏孔孔徑的比例(d/s比)統(tǒng)一調控約為0.26,確保3種孔徑支架具有相似的貫通性,以精確研究宏孔孔徑對異位骨形成和血管生長的影響。體內實驗結果顯示多孔支架的宏孔孔徑不但影響血管生長和骨形成的速度,同時影響新形成骨的分布。宏孔孔徑為700-950 μm的多孔HA支架比其他兩種孔徑多孔支架展現(xiàn)更高的血管數(shù)量、新骨生成量以及更為均勻的新骨分布。采用糖球造孔法和熱處理工藝技術制備宏孔孔徑為750-900μm、貫通孔徑尺寸分別為87、228和367μm (d/s比分別為0.09、0.26和0.45)的3種多孔HA支架,通過體內植入實驗研究多孔支架d/s比對異位誘導骨形成和血管生長的影響。結果顯示多孔支架的d/s比不但影響血管的生長,同時也影響骨組織的形成。植入4周后結果顯示新生血管的尺寸隨著支架貫通孔徑的增大而增大,體內植入12周后結果顯示d/s比為0.45的支架由于抗壓強度較低,在植入體內后發(fā)生脆裂導致新骨在支架內部不均勻的形成。d/s比為0.09的支架由于貫通孔徑較低限制了新生血管的數(shù)量和尺寸,導致骨形成能力的降低。而d/s比為0.26的支架則展現(xiàn)了最高的新骨生成量以及更均勻的新骨分布。通過采用梯度糖球模板制備2種孔徑分布相反的梯度分布多孔HA支架,即(1)內部孔徑為500-650μm、外周孔徑為1100-1250μm(HASL); (2)內部孔徑為1100-1250μm、外周孔徑為500-6500μm (HALS)。體內植入4周后結果顯示雖然HASL外周和中心區(qū)域的新生血管數(shù)量無顯著性差異,但是外周新生血管的尺寸明顯大于中心區(qū)域的新生血管。而HALS外周和中心區(qū)域的新生血管數(shù)量幾乎相同無顯著性差異,但是中心區(qū)域的新生血管數(shù)量小于外周區(qū)域的新生血管。HASL內總的新生血管數(shù)量和尺寸大于HALS的新生血管數(shù)。體內植入12周后結果顯示HASL在整個支架的外周及中心展現(xiàn)均勻的骨形成,而HALS只有外周有新骨的生成。HASL內總的新骨形成量明顯高于HALS。因此表明梯度多孔支架的孔徑分布不僅影響支架的血管化,還影響新形成骨的分布。外部大孔徑的梯度多孔支架更有利于再生新骨組織在多孔支架中的整體生長。采用一種新型的糖球顆粒模板濕化處理技術制備宏孔孔壁表面具有溝槽結構的多孔HA支架。結果發(fā)現(xiàn)濕化處理可調控糖球模板表面的水分含量,而孔壁表面溝槽的寬度又由糖球模板表面上的水分的含量所控制。細胞實驗顯示溝槽結構可誘導細胞沿溝槽方向定向排列。基因表達結果顯示溝槽結構有利于骨形成。通過一種新型的方法將載丹酚酸B (Sal B)的殼聚糖微球(CMs)均勻固定在涂覆有海藻酸的多孔HA支架表面構建支架緩釋體系。為增強CMs和HA支架的結合力,選用海藻酸作為多孔HA支架的涂覆材料。結果顯示通過靜電吸附的作用,微球穩(wěn)定地固定在涂有海藻酸的多孔HA支架表面。在吸附過程中,采用靜置和震蕩兩種組裝方式將Sal B/CMs組裝到多孔支架表面。與靜置組裝方式相比,采用震蕩組裝方式的多孔支架表面上的Sal B/CMs分布更為均勻,通過對比不同濃度的海藻酸研究載Sal B的CMs (Sal B/CMs)在多孔支架表面上的分布情況。結果顯示最優(yōu)的海藻酸濃度為1%,因為采用1%海藻酸作為涂覆層保障微球在支架表面均勻地分布并且不會影響多孔HA支架宏孔孔壁表面的多孔結構。細胞實驗結果顯示采用1%海藻酸作為涂覆層組裝Sal B/CMs的多孔HA支架在與細胞共培養(yǎng)3、7天后,明顯促進細胞的增殖,并且細胞均勻地粘附在支架的表面。
[Abstract]:In bone tissue engineering scaffolds, the physical structure of porous scaffolds is a key factor affecting cell behavior, vascular growth and bone formation. It not only provides a template for cell growth and migration, but also provides sufficient space for vascular growth and bone formation. Although optimizing the structural characteristics of porous scaffolds can effectively promote bone formation, However, the ability to regulate the physical structure of scaffolds to promote bone regeneration is still limited. The introduction of exogenous growth factors and drugs on some surfaces can significantly enhance bone tissue regeneration. Therefore, more and more attention has been paid to the synergistic regulation of vascular growth and bone formation by porous scaffolds with biological factors or drugs. The structural properties of the porous hydroxyapatite (HA) scaffold were investigated by regulating the structural parameters of the scaffold (aperture, penetration, pore size distribution and surface morphology). The porous structure of the scaffold was further optimized. In addition, the high molecular weight microspheres were evenly distributed in the porous HA scaffold coated with alginate. The porous scaffold sustained-release system was constructed to enhance the vascular growth and bone formation ability of porous HA scaffolds. By using the sugar ball hole method, 3 kinds of porous HA scaffolds with controllable pore structure were prepared by removing sugar spheres. The size of the sugar sphere particles and the heat treatment process were controlled to accurately regulate the pore diameter and through hole size of porous scaffolds. In the heat treatment process, the penetration hole size / pore diameter ratio (d/s ratio) of the porous HA scaffold with the pore size of 500-650700-950 and 1100-1250 m, respectively, is regulated by about 0.26, ensuring the similar penetration of the 3 kinds of aperture scaffolds to accurately study the effect of the macropore diameter on the ectopic bone formation and the vascular growth. The results show that the pore diameter of the porous scaffold not only affects the speed of vascular growth and bone formation, but also affects the distribution of the newly formed bone. The porous HA scaffold with a macro pore diameter of 700-950 mu m exhibits higher blood vessel quantity, new bone formation and more uniform new bone distribution than the other two porous porous scaffolds. 3 kinds of porous HA scaffolds with macropore diameter of 750-900 m and 87228 and 367 mu m respectively (d/s ratio 0.09,0.26 and 0.45 respectively) were prepared by processing technology. The effect of d/s ratio of porous scaffold on ectopic bone formation and vascular growth was investigated by implantation in vivo. The results showed that the d/s ratio of porous scaffolds not only affected blood vessels. Growth, but also the formation of bone tissue. 4 weeks after implantation, the results showed that the size of the neovascularization increased with the increase of the perforation diameter of the stent. After 12 weeks in the body, the results showed that the d/s ratio 0.45 was low in compressive strength, and the embrittlement after implantation resulted in the uneven.D/s ratio of the new bone in the stent. The scaffold limited the number and size of the new blood vessels and reduced the bone formation ability. The d/s ratio of 0.26 to the scaffold showed the highest new bone formation and more uniform new bone distribution. The gradient distribution of porous HA scaffolds, that is, (1) internal pores, were prepared by using a gradient sugar sphere template. The diameter was 500-650 mu m, the circumference aperture was 1100-1250 mu m (HASL), and the inner aperture was 1100-1250 m and the peripheral aperture was 500-6500 mu m (HALS). The results showed that there was no significant difference in the number of neovascularization in the peripheral and central regions of the peripheral blood, but the size of the neovascularization in the peripheral blood was obviously greater than that of the neovascularization in the central region. And HALS There was no significant difference in the number of neovascularization in the peripheral and central regions, but the number of neovascularization in the central region was less than that of the new vascular.HASL in the peripheral region. The number and size of the new blood vessels in the new vascular.HASL were larger than that of the HALS. The results showed that HASL was uniform in the peripheral and center of the whole stent after the body implantation. Bone formation, and the formation of new bone in the HALS only with new bone in.HASL, is obviously higher than that of HALS.. Therefore, the pore size distribution of the gradient porous scaffold not only affects the vascularization of the scaffold, but also affects the distribution of the newly formed bone. The gradient porous scaffold with large external aperture is more beneficial to the overall growth of the new bone tissue in the porous scaffold. A porous HA scaffold with groove structure on the surface of the macroporous wall was prepared by a new type of sugar ball particle formwork. The results showed that the wetting treatment could regulate the water content of the surface of the sugar sphere template, and the width of the groove on the surface of the hole was controlled by the content of water on the surface of the sugar sphere template. Cell experiments showed the groove junction. The gene expression results show that the groove structure is beneficial to the formation of bone. The chitosan microsphere (CMs) of B (Sal B) is uniformly fixed to the porous HA scaffold coated with alginate on the surface of the scaffold to construct a scaffold release system. The binding force of CMs and HA scaffolds is selected. As the coating material of the porous HA scaffold, the alginic acid shows that the microspheres are immobilized on the surface of the porous HA scaffold coated with alginate by electrostatic adsorption. In the process of adsorption, the Sal B/CMs is assembled on the surface of the porous scaffold by two assembly methods of static and concussion. The distribution of Sal B/CMs on the surface of the porous scaffold is more uniform. The distribution of CMs (Sal B/CMs) on the porous scaffold on the surface of Sal B is studied by contrasting alginate with different concentrations. The results show that the optimal alginate concentration is 1%, because the use of 1% alginate as coating layer is evenly distributed on the surface of the scaffold and will not be used. The porous structure of the macroporous surface of the porous HA stent was affected. The results of cell experiment showed that the porous HA scaffold which used 1% alginate as the coating layer to assemble Sal B/CMs obviously promoted the cell proliferation after co culture with the cells, and the cells adhered to the surface of the scaffold evenly.
【學位授予單位】：西南交通大學
【學位級別】：博士
【學位授予年份】：2016
【分類號】：R318.08;TB383.4

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