本文選題：聚富馬酸丙二醇酯 + 增韌改性　； 參考：《浙江大學(xué)》2017年博士論文

【摘要】：第一部分聚富馬酸丙二醇酯/甲基丙烯酸羥乙酯彈性材料優(yōu)化制備與性能研究目的利用甲基丙烯酸羥乙酯作為共聚單體對聚富馬酸丙二醇酯進(jìn)行增韌改性,探索制備比傳統(tǒng)PPF韌性更好的材料的可行性,獲得制備彈性材料的甲基丙烯酸羥乙酯共聚單體最優(yōu)比例,在此基礎(chǔ)上調(diào)節(jié)HEMA共聚單體比例對PPF交聯(lián)密度、親水性及力學(xué)新能進(jìn)行精確調(diào)控,以期構(gòu)建出在結(jié)構(gòu)和力學(xué)性能上對天然瞼板組織有更高相似性的材料,為瞼板替代物的研制提供一個(gè)新的途徑。方法首先對兔瞼板組織微觀形貌、力學(xué)性能進(jìn)行分析,在此基礎(chǔ)上,采用具有良好生物相容性的PPF、HEMA進(jìn)行共聚,通過對HEMA共聚單體比例的調(diào)整,優(yōu)化共聚物各項(xiàng)參數(shù),制備基于PPF的彈性材料。傅里葉轉(zhuǎn)換紅外線光譜分析共聚物化學(xué)鍵及化學(xué)組成,差掃描量熱儀表征共聚物熱性能,掃描電鏡和透射電鏡檢測共聚物表面和斷面微觀結(jié)構(gòu),靜態(tài)水接觸角對共聚物的親疏水性進(jìn)行分析比較,萬能測試機(jī)檢測共聚物壓縮及拉伸力學(xué)強(qiáng)度,分析評價(jià)PPF/HEMA共聚材料作為瞼板替代材料的結(jié)構(gòu)與性能仿生可行性。結(jié)果兔瞼板平均楊氏模量為5.67±1.53 MPa。交聯(lián)固化后的PPF-HEMA共聚物能夠在3400 cm-1和1740cm-1處看到較強(qiáng)的羥基和羥基的伸縮振動峰,對比不同投料比的樣品中羥基和羰基振動峰的相對強(qiáng)度,隨著樣品中投料的HEMA量增加,羥基相對羰基含量增加。PPF-HEMA0.5、PPF-HEMA1、PPF-HEMA2的水接觸角隨著HEMA單體投料比的增加而降低;溶脹率分別為16%、20%和30%左右。溶脹前,除PPF-HEMA0.5樣品(14.3 ±2.6 MPa)以外,其他實(shí)驗(yàn)組和對照組壓縮模量均在19 MPa左右。拉伸模式下,PPF-HEMA 0.5樣品在溶脹前斷裂伸長率就能達(dá)到20%左右,PPF-HEMA 1和PPF-HEMA 2樣品的伸長率可以由溶脹前的不到10%提高到溶脹后的30～40%左右。在濕態(tài)循環(huán)加載條件下,各組分PPF-HEMA經(jīng)過50次循環(huán),樣品形貌保持完整,應(yīng)力-應(yīng)變曲線重合較好,應(yīng)力值分別為原始的90.1%,82.5%,and 85.9%,反應(yīng)材料具有較好的抗疲勞性。結(jié)論本部分研究選用HEMA為共聚單體,以1:0.5、1:1和1:2三種質(zhì)量置比投料,與PPF共聚制備了韌性材料。PPF-HEMA交聯(lián)體系中,通過增加HEMA共聚單體的比例改變固化后材料的化學(xué)組成,并在相當(dāng)程度上提高材料的溶脹性能,制備了一種具有良好的韌性和抗疲勞性的彈性材料,為模擬天然瞼板替代材料的研究和應(yīng)用奠定了實(shí)驗(yàn)基礎(chǔ)。第二部分聚富馬酸丙二醇酯/甲基丙烯酸羥乙酯組織工程多孔支架的制備及其理化性能和生物相容性研究目的利用生物相容性良好的明膠微粒作為致孔劑制備具有適度孔徑、高孔隙率和連通性好的多孔支架,在微觀層次探討PPF-HEMA共聚材料的化學(xué)組分、孔隙結(jié)構(gòu)對材料降解性能等理化性能和細(xì)胞黏附、增殖等生物活性的影響,進(jìn)一步在二維培養(yǎng)條件下驗(yàn)證該組織工程PPF-HEMA多孔支架對瞼板缺損修復(fù)修復(fù)研究中所涉及的目標(biāo)細(xì)胞的適用性。方法采用明膠粒子致孔法制備不同孔徑、不同共聚比例的多孔PPF-HEMA支架,掃描電鏡觀察支架的形貌和結(jié)構(gòu)并測定支架孔徑,排乙醇體積法測多孔支架孔隙率,根據(jù)1,2,3,6,9和12周支架的失重率分析體外降解性能。浸提液法用HDFs細(xì)胞考察材料的細(xì)胞毒性,并采用PrestoBlne試劑盒對人結(jié)膜基質(zhì)細(xì)胞和人結(jié)膜上皮細(xì)胞兩種不同目標(biāo)細(xì)胞在支架上的黏附與增殖能力進(jìn)行檢測,免疫熒光染色特異性標(biāo)記物觀察支架上細(xì)胞形態(tài),以評價(jià)組織工程PPF-HEMA多孔支架的生物相容性。結(jié)果明膠粒子致孔法180-280μm明膠微粒所制備的PPF-HEMA多孔支架具有規(guī)整的外形,而280-450μm、480-600μm明膠微粒致所制備的PPF-HEMA多孔支架因孔隙較大,280-450μm孔徑支架外形較為粗糙不規(guī),480-60μm孔徑支架甚至出現(xiàn)開裂。180-280μm明膠模板所制備的 PPF-HEMA 0.5、PPF-HEMA 1 和 PPF-HEMA 2多孔支架孔隙率分別在88.9±2.3%,82.3±1.6%和84.6±3.1%,與Image J軟件分析商業(yè)化的ADM孔隙率為91.5±3.2%相比無明顯差異。掃描電鏡可見多孔支架具有規(guī)整的外形,良好的孔徑分布和連通性。體外PBS孵育12周內(nèi),PPF-HEMA2組的支架質(zhì)量損失在第9周為10.9±1.3%,到12周質(zhì)量損失也僅達(dá)11.4±0.7%;而PPF-HEMA0.5和PPF-HEMA1到12周質(zhì)量損失分別達(dá)到15%和20%。MTT法檢測不同PPF-HEMA浸提液之間以及對比空白對照組,各時(shí)間點(diǎn)細(xì)胞增殖情況無明顯差異(P0.05)。在PPF-HEMA材料體系中,HEMA共聚單體的加入通過改變共聚體的親水性而調(diào)控支架的細(xì)胞相容性在PPF-HEMA材料體系中,PPF-HEMA 1能支持一定的CjECs增殖,但PPF-HEMA 0.5和PPF-HEMA 2多孔支架雖有一定細(xì)胞黏附,但并不有效地支持目標(biāo)細(xì)胞CjSCs生長。結(jié)論利用180-280μm明膠微粒致孔劑法可制備出具有適度孔徑、高孔隙率和連通性好的PPF-HEMA多孔支架,支架在體外降解12周質(zhì)量損失小于20%,具有良好的穩(wěn)定性,通過改變PPF-HEMA投料比可以通過調(diào)節(jié)共聚體的交聯(lián)密度可調(diào)控PPF-HEMA多孔支架體外降解行為。PPF-HEMA 1能支持一定的CjECs增殖,但PPF-HEMA0.5PPF-HEMA2基本不能支持細(xì)胞增殖。第三部分聚富馬酸丙二醇酯/甲基丙烯酸羥乙酯多孔支架材料的體內(nèi)生物學(xué)效應(yīng)及眼瞼重建效果研究目的分析評價(jià)組織工程PPF-HEMA多孔支架的體內(nèi)生物相容性,同時(shí)觀測其原位缺損重建修復(fù)效果,為瞼板替代材料的構(gòu)建提供新的思路和合成方法,為眼瞼重建用組織工程瞼板替代材料的臨床應(yīng)用奠定前期實(shí)驗(yàn)基礎(chǔ)。方法構(gòu)建兔全層瞼板缺損動物模型,以去人脫細(xì)胞真皮基質(zhì)作為對照,植入PPF-HEMA多孔支架材料。通過觀察眼瞼缺損修復(fù)效果、組織切片HE染色和Masson染色觀察炎癥反應(yīng)和材料-組織反應(yīng)情況,評價(jià)PPF-HEMA多孔支架材料的可操作性和在體生物相容性。結(jié)果成功構(gòu)建兔全層瞼板缺損動物模型,術(shù)后8周ADM組和PPF-HEMA 2組眼瞼缺損完全修復(fù),未見任何成角畸形,PPF-HEMA 1組可見輕度成角畸形,對比陰性對照組可見明顯眼瞼成角畸形及疤痕形成。組織學(xué)切片HE染色可見術(shù)后1周各組炎癥反應(yīng)明顯,同一時(shí)間實(shí)驗(yàn)組與對照組炎癥反應(yīng)評分無統(tǒng)計(jì)學(xué)差異,隨植入時(shí)間增加各組炎癥反應(yīng)均明顯減輕,術(shù)后8周與術(shù)后1周炎癥反應(yīng)評分有統(tǒng)計(jì)學(xué)差異。組織學(xué)切片 Masson 染色可見PPF-HEMA多孔支架材料孔隙內(nèi)新生結(jié)締組織長入,支架周圍膠原纖維束包裹,ADM組、PPF-HEMA 1和PPF-HEMA 2組Image J 測量其厚度分別為 28.0 ± 9.3 μm,65.2 ± 16.8 μm 和 63.5 ± 17.3 μm。結(jié)論組織工程PPF-HEMA多孔支架具有良好的體內(nèi)生物相容性,在兔全層瞼板缺損動物模型中PPF-HEMA 2組表現(xiàn)出更好的修復(fù)效果,較低的炎癥反應(yīng)以及較慢的降解速率,相比于ADM其炎癥反應(yīng)無明顯差異,但長期穩(wěn)定性更好。本研究結(jié)果為仿天然瞼板結(jié)構(gòu)與性能高分子材料的研究奠定了扎實(shí)的實(shí)驗(yàn)基礎(chǔ),為臨床上眼表疾病的治療提供了一條新的途徑。
[Abstract]:The optimization of the preparation and properties of polyfumaric acid propanediol ester / hydroxyethyl methacrylate elastic material. Aim to toughen the polyfumaric acid propanediol ester with hydroxyethyl methacrylate as a copolymerized monomer, explore the feasibility of preparing the material better than the traditional PPF toughness, and obtain the methyl propylene for the preparation of the elastic material. The optimal proportion of hydroxyethyl ester copolymers is based on the ratio of HEMA copolymers to PPF crosslinking density, hydrophilicity and new mechanical energy, in order to construct a material with higher similarity in structure and mechanical properties to natural eyelid tissue, and to provide a new way for the development of eyelid substitutions. The microstructure and mechanical properties of the rabbit eyelid tissue were analyzed. On this basis, PPF and HEMA with good biocompatibility were used to copolymerized. By adjusting the proportion of HEMA copolymers, the parameters of the copolymers were optimized and the elastic materials based on PPF were prepared. The chemical bond and chemical composition of the copolymers were analyzed by Fourier transform infrared spectroscopy. The thermal performance of the copolymer was detected by differential scanning calorimeter. The microstructure of the surface and section of the copolymer was detected by scanning electron microscope and transmission electron microscope. The static water contact angle was used to analyze the hydrophobicity of the copolymer. The universal testing machine was used to detect the compressive strength of the copolymer and the tensile strength of the copolymer. The PPF/HEMA copolymers were analyzed and evaluated as the junctions of the replacement material of the eyelid plate. The results show that the average young's modulus of the rabbit's eyelid is 5.67 + 1.53 MPa. after curing, and the PPF-HEMA copolymer can see the strong hydroxyl and hydroxyl expansion vibration peaks at 3400 cm-1 and 1740cm-1. The relative intensity of the hydroxyl and carbonyl vibration peaks in the samples with different feeding ratios is compared with the HEMA quantity of the samples in the sample. With the increase of the content of hydroxyl group, the relative carbonyl content of.PPF-HEMA0.5, PPF-HEMA1, PPF-HEMA2 decreased with the increase of the ratio of HEMA monomers, and the swelling rate was 16%, 20% and 30% respectively. Before the swelling, the compression modulus of the other experimental and control groups was around 19 MPa except the PPF-HEMA0.5 sample (14.3 + 2.6 MPa). Under tensile mode, PPF-HE The elongation at break of MA 0.5 can reach about 20% before swelling, and the elongation of PPF-HEMA 1 and PPF-HEMA 2 can be increased from less than 10% before swelling to about 30 to 40% after swelling. Under the condition of wet cyclic loading, each component PPF-HEMA passes through 50 cycles, the sample morphology is intact, stress strain curves reclose well, stress The value is 90.1%, 82.5%, and 85.9%, respectively, and the reaction material has good fatigue resistance. Conclusion this part of the study selected HEMA as a copolymerized monomer, 1:0.5,1:1 and 1:2 three kinds of mass ratio of material, and PPF copolymerization with PPF to prepare the.PPF-HEMA crosslinking system of ductile material and change the chemical properties of the solidified material by increasing the proportion of HEMA copolymers. An elastic material with good toughness and fatigue resistance was prepared to improve the swelling properties of the material to a considerable extent. The experimental foundation for the research and application of the natural replacement material for the natural eyelid plate was established. The preparation and theory of the second part polyfumaric acid propanediol ester / hydroxyethyl methacrylate porous scaffold Studies on chemical properties and biocompatibility with good biocompatible gelatin particles as porosities to prepare porous scaffolds with moderate pore size, high porosity and good connectivity. The chemical composition of PPF-HEMA copolymers, pore structure on material degradation properties, cell adhesion, proliferation and so on are discussed at the micro level. The effect of substance activity was further verified under two dimensional culture. The applicability of the tissue engineering PPF-HEMA porous scaffold to the target cells in the repair and repair of the eyelid defect was verified. Method the porous PPF-HEMA scaffold with different pore size, different copolymerization ratio was prepared by the gelatin particle hole method, and the morphology and junction of the scaffold were observed by scanning electron microscope. The pore size of the scaffold was measured and the porosity of the porous scaffold was measured by the method of ethanol volume. The degradation performance in vitro was analyzed according to the weight loss rate of the 1,2,3,6,9 and 12 weeks scaffold. The cytotoxicity of the material was examined by HDFs cells in the leaching solution, and the PrestoBlne Kit was used to treat the two different target cells of human conjunctival stromal cells and human nodal epithelial cells on the scaffold. The cell morphology on the scaffold was observed by the specific markers of immunofluorescence staining to evaluate the biocompatibility of the porous scaffolds of the tissue engineering PPF-HEMA. Results the PPF-HEMA porous scaffolds prepared by gelatin particles 180-280 um m gelatin particles have a regular shape, and 280-450 m, 480-600 mu m gelatin micro The porous PPF-HEMA scaffolds prepared by the particles are relatively rough, and the 280-450 - m diameter of the scaffold is rough and irregular. The 480-60 - M aperture bracket even appears PPF-HEMA 0.5 prepared by the cracking.180-280 - M gelatin template. The porosity of the porous scaffold for PPF-HEMA 1 and PPF-HEMA 2 is 88.9 + 2.3%, 82.3 + 1.6% and 84.6 +, respectively, and Image J software The porosity of the commercialized ADM was 91.5 + 3.2%. The scanning electron microscope showed that the porous scaffold had a regular shape, a good pore size distribution and connectivity. In the 12 week incubation of PBS, the mass loss of the PPF-HEMA2 group was 10.9 + 1.3%, and the mass loss was only 11.4 + 0.7% in the 12 week, while PPF-HEMA0.5 and PPF-HEMA1 to 1. The 2 weeks of mass loss reached 15% and 20%.MTT method to detect different PPF-HEMA extracts and contrast blank control group. There was no significant difference in cell proliferation at all time points (P0.05). In the PPF-HEMA material system, HEMA copolymers were added by changing the hydrophilicity of the copolymer to regulate the cell compatibility of the scaffold in the PPF-HEMA material body In the system, PPF-HEMA 1 can support a certain number of CjECs proliferation, but the porous scaffolds of PPF-HEMA 0.5 and PPF-HEMA 2 have certain cell adhesion, but they do not effectively support the CjSCs growth of the target cells. Conclusion the porous scaffold with moderate pore size, high pore rate and good connectivity can be prepared by using 180-280 micron gelatin microparticles to produce porous scaffolds with good pore size, high porosity and connectivity. The mass loss of 12 weeks in vitro is less than 20% and has good stability. By changing the ratio of PPF-HEMA feeding ratio, the degradation behavior of PPF-HEMA porous scaffold can be regulated by adjusting the crosslinking density of the copolymer,.PPF-HEMA 1 can support a certain CjECs proliferation, but the PPF-HEMA0.5PPF-HEMA2 base can not support cell proliferation. Third part of the poly (fumarate) The biological effects of propanediol / hydroxyethyl methacrylate porous scaffold materials and the effect of eyelid reconstruction are studied to evaluate the biocompatibility of tissue engineered PPF-HEMA porous scaffolds in vivo, and to observe the effect of reconstruction and repair of the defects in situ for the construction of palpebral replacement materials. Reconstruction of eyelid reconstruction with tissue engineering palpebral replacement material in the clinical application of the earlier experimental basis. Method to construct a rabbit full layer blepharo defect animal model, to remove the human acellular dermal matrix as the control, implanted PPF-HEMA porous scaffold material. By observing the effect of eyelid defect repair, HE staining and Masson staining of the tissue section were used to observe the inflammatory reaction. The maneuverability and biocompatibility of PPF-HEMA porous scaffold were evaluated with material tissue reaction. Results the rabbit model of full layer blepharo defect was successfully constructed. 8 weeks after operation, the eyelid defect was completely repaired in group ADM and PPF-HEMA 2, no angular deformity was found, and mild angle deformity in group PPF-HEMA 1 could be seen, compared with negative control group. HE staining showed that there was no significant difference in the inflammatory response between the experimental group and the control group at the same time 1 weeks after the operation. The inflammatory response in each group was significantly reduced with the time of implantation, and there was a statistically significant difference between the 8 weeks after the operation and the 1 weeks after the operation. Masson staining showed that the new connective tissue in the porous scaffold of PPF-HEMA was long into the pores, and the collagen fiber bundles were wrapped around the scaffold. The thickness of ADM group, PPF-HEMA 1 and PPF-HEMA 2 groups was 28 + 9.3 mu m, 65.2 + 16.8 micron and 63.5 + 17.3 micron M., respectively. In the animal model of full layer eyelid defect in rabbits, the PPF-HEMA 2 groups showed better repair effect, lower inflammatory response and slower degradation rate, and had no significant difference compared with ADM, but the long-term stability was better. The results of this study lay the foundation for the study of natural eyelid structure and properties of polymer materials A solid experimental foundation has provided a new way for the treatment of ocular surface diseases.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R318.08

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