本文選題：3D打印 + 鈦合金�。� 參考：《第二軍醫(yī)大學(xué)》2017年博士論文

【摘要】：第一部分3D打印鈦合金表面多孔涂層及其機(jī)械性能測(cè)試目的:探討利用3D打印技術(shù)制作具有多孔涂層鈦合金假體的可行性及其機(jī)械性能的檢測(cè)。方法:使用計(jì)算機(jī)輔助設(shè)計(jì)建模,導(dǎo)入3D打印機(jī)后打印鈦合金假體部件;掃描電鏡對(duì)其進(jìn)行表征,測(cè)量孔徑孔隙率;使用電子萬(wàn)能試驗(yàn)機(jī)測(cè)試打印假體部件的抗壓、抗剪切性能。結(jié)果:利用3D打印技術(shù)制造出4組具有不同孔徑、孔隙率多孔涂層的鈦合金假體部件,經(jīng)掃描電鏡表征分別為:A組218.7μm,61.2%;B組262μm,68.1%;C組558μm,79.2%;D組753μm,89.0%。4組鈦合金多孔涂層的壓縮強(qiáng)度和剪切強(qiáng)度分別為A組78.02MPa,44.28 MPa;B組20.62MPa,23.17MPa;C組8.32MPa,12.64MPa;D組4.26MPa,8.53MPa。結(jié)論:3D打印技術(shù)可精確構(gòu)建鈦合金表面多孔涂層,具有良好的機(jī)械性能;隨著孔徑的逐漸增大,鈦合金多孔涂層抗壓、抗剪切性能急劇下降,其中孔徑262μm,孔隙率68.1%是相對(duì)理想內(nèi)植物微孔參數(shù),可為將來(lái)假體設(shè)計(jì)提供參考。第二部分3D打印鈦合金表面多孔涂層對(duì)MG-63細(xì)胞生物學(xué)行為影響目的:研究3D打印鈦合金多孔涂層的生物相容性及不同微孔參數(shù)對(duì)MG-63細(xì)胞的影響。方法:以各組鈦合金多孔涂層為支架培養(yǎng)MG-63細(xì)胞24h、48h、72h后,掃描電鏡觀察細(xì)胞形態(tài);DAPI染色后熒光顯微鏡下細(xì)胞計(jì)數(shù);CCK8檢測(cè)各組細(xì)胞成活率;用ALP試劑盒和免疫印跡技術(shù)檢測(cè)各組細(xì)胞堿性磷酸酶、骨鈣蛋白的表達(dá)情況。結(jié)果:各時(shí)間點(diǎn)均為A組和B組細(xì)胞伸展形態(tài)較好,伸出更多的偽足。培養(yǎng)24h細(xì)胞在A組和B組材料上粘附明顯多于C組和D組,差異有統(tǒng)計(jì)學(xué)意義,培養(yǎng)48h后,B組多于其他3組,差異有統(tǒng)計(jì)學(xué)意義,培養(yǎng)72h時(shí)4組細(xì)胞計(jì)數(shù)無(wú)統(tǒng)計(jì)學(xué)差異。培養(yǎng)24h細(xì)胞成活率A組低于空白組(無(wú)多孔涂層)、B組、C組、D組,差異均有統(tǒng)計(jì)學(xué)差異,B組與空白組無(wú)統(tǒng)計(jì)學(xué)差異,低于C組、D組,差異均有統(tǒng)計(jì)學(xué)差異,C組與空白組無(wú)統(tǒng)計(jì)學(xué)差異,但低于D組,差異有統(tǒng)計(jì)學(xué)差異,D組高于空白組,差異有統(tǒng)計(jì)學(xué)差異;培養(yǎng)48h細(xì)胞成活率A組低于空白組、B組、C組、D組,差異均有統(tǒng)計(jì)學(xué)差異,B組與C組無(wú)統(tǒng)計(jì)學(xué)差異,但高于A組,低于空白組,D組,差異均有統(tǒng)計(jì)學(xué)差異,C組與空白組無(wú)統(tǒng)計(jì)學(xué)差異,低于D組,差異均有統(tǒng)計(jì)學(xué)差異,D組高于空白組,差異有統(tǒng)計(jì)學(xué)差異;培養(yǎng)72h細(xì)胞成活率A組低于空白組、B組、C組、D組,差異均有統(tǒng)計(jì)學(xué)差異,B組低于空白組、C組、D組,差異均有統(tǒng)計(jì)學(xué)差異,C組低于空白組、D組,差異均有統(tǒng)計(jì)學(xué)差異,D組高于空白組,差異有統(tǒng)計(jì)學(xué)差異。各時(shí)間點(diǎn)各組間細(xì)胞堿性磷酸酶和骨鈣蛋白表達(dá)均無(wú)統(tǒng)計(jì)學(xué)差異。結(jié)論:3D打印鈦合金多孔涂層可以明顯影響MG-63細(xì)胞的生物學(xué)行為,相對(duì)較小的微孔(218.7μm,61.2%;262μm,68.1%)有利于細(xì)胞粘附和分化,較大的微孔(753μm,89.0%)有利于細(xì)胞增殖;較小孔徑的三維多孔涂層培養(yǎng)細(xì)胞收集可能存在誤差,較低的細(xì)胞增值率是否界定為材料毒性反應(yīng)或抑制增殖還需進(jìn)一步研究驗(yàn)證。第三部分3D打印鈦金屬多孔涂層促進(jìn)骨整合及其機(jī)制研究目的:研究3D打印不同微孔參數(shù)的鈦合金多孔涂層對(duì)骨整合及成骨基因表達(dá)的影響。方法:3D打印機(jī)打印出3組具有不同微孔參數(shù)多孔涂層的鈦合金假體,分別為(A組262μm,68.1%;B組558μm,79.2%;C組753μm,89.0%)。選擇27只新西蘭大白兔,隨機(jī)分為3組,將3種規(guī)格假體分別植入各組兔子雙側(cè)股骨髓腔,術(shù)后4周、8周、12周每組各處死3只兔子,進(jìn)行X線、Micro-CT掃描并測(cè)量骨體積分?jǐn)?shù)和組織礦物密度,制作硬組織切片進(jìn)行Goldner's染色和熒光顯微鏡觀察,行拔出實(shí)驗(yàn)測(cè)量最大拔出力,q PCR檢測(cè)Runx2和Osterix的表達(dá)。結(jié)果:X線和Micro-CT掃描均顯示4周時(shí)各組均無(wú)明顯骨質(zhì)沉積,隨著時(shí)間增加,新骨形成逐漸增多,12周時(shí)最多,且A組多于B組、C組。熒光顯微鏡觀察術(shù)后4周各組均熒光較弱,條帶較窄,隨時(shí)間增加熒光強(qiáng)度增強(qiáng),條帶變寬,黃色與綠色間隙增大,12周時(shí)最優(yōu),且A組優(yōu)于B組、C組。Goldner's染色示術(shù)后4周各組均以細(xì)胞核為主,隨時(shí)間增加,類骨質(zhì)逐漸增多,12周時(shí)達(dá)到頂峰,且A組優(yōu)于B組、C組。術(shù)后4周、8周時(shí),3組間骨體積分?jǐn)?shù)和組織礦物密度差異均無(wú)統(tǒng)計(jì)學(xué)意義;術(shù)后12周時(shí)A組骨體積分?jǐn)?shù)和組織礦物密度大于B組、C組,差異均有統(tǒng)計(jì)學(xué)意義,B組和C組差異無(wú)統(tǒng)計(jì)學(xué)意義。術(shù)后4周、8周時(shí),A組Runx2和Osterix表達(dá)高于B組、C組,差異有統(tǒng)計(jì)學(xué)意義,B組和C組間無(wú)統(tǒng)計(jì)學(xué)差異;術(shù)后12周時(shí),C組Runx2表達(dá)低于A組、B組,差異有統(tǒng)計(jì)學(xué)意義,A組和B組間無(wú)統(tǒng)計(jì)學(xué)差異,Osterix表達(dá)3組間無(wú)統(tǒng)計(jì)學(xué)差異。術(shù)后4周時(shí),3組間最大拔出力差異均無(wú)統(tǒng)計(jì)學(xué)意義;術(shù)后8周時(shí),A組所需最大拔出力均大于B組、C組,差異均有統(tǒng)計(jì)學(xué)意義,B組和C組差異無(wú)統(tǒng)計(jì)學(xué)意義;術(shù)后12周時(shí),任意兩組最大拔出力比較,差異均有統(tǒng)計(jì)學(xué)意義,依次為A組B組C組。結(jié)論:3D打印鈦合金多孔涂層可以明顯促進(jìn)骨整合;隨著時(shí)間的延長(zhǎng),骨質(zhì)形成逐漸增多,沉積速率逐漸加快;孔徑262μm,孔隙率68.1%的多孔涂層最有利于促進(jìn)骨整合;微孔參數(shù)的變化對(duì)生物力學(xué)的影響大于對(duì)成骨效應(yīng)的影響。
[Abstract]:The first part of 3D printing titanium alloy surface porous coating and its mechanical properties test purpose: To explore the feasibility and mechanical properties of porous coated titanium alloy prosthesis by using 3D printing technology. Method: using computer aided design modeling, introducing 3D printer to print titanium alloy prosthesis components; scanning electron microscope Results: 4 groups of titanium alloy prosthesis components with different pore sizes and porous porous coatings were produced by 3D printing technology. The scanning electron microscope was characterized by A group 218.7 m, 61.2%; B group 262 mu m, 68.1%; C group 558 m, 79.2%; D group 753. The compressive strength and shear strength of the porous titanium alloy coating in group 89.0%.4 are A group 78.02MPa, 44.28 MPa, B group 20.62MPa, 23.17MPa, C group 8.32MPa, 12.64MPa, D group conclusion: it can accurately construct porous coating on the surface of titanium alloy, with good mechanical properties; with the increase of pore size, titanium alloy porous coating Layer compression and shear resistance dramatically decrease, the pore diameter is 262 mu m, and the porosity 68.1% is the relative ideal inner plant micropore parameters, which can provide reference for the future prosthesis design. Second part 3D printing titanium alloy surface porous coating on the biological behavior of MG-63 cells: the study of the biocompatibility and different microcompatibility of the porous coating of the 3D titanium alloy The effect of pore parameters on MG-63 cells. Methods: the cell morphology was observed by scanning electron microscope after MG-63 cells 24h, 48h and 72h were cultured with porous titanium alloy coating as scaffold. After DAPI staining, the cell counts were counted under fluorescence microscope; CCK8 was used to detect the survival rate of each cell, and the cell alkaline phosphatase, bone calcium egg was detected by ALP kit and immunoblotting technique. Results: at all time points, the cells in group A and group B extend well and extend more pseudo foot. The adhesion of 24h cells in A and B groups is more than that of C group and D group. The difference has statistical significance. After developing 48h, the B group is more than the other 3 groups. The difference has the significance of total count, and there is no statistical difference between the 4 groups when developing 72h. The survival rate of 24h cells in A group was lower than that of blank group (no porous coating), group B, C group and D group, and there was no statistical difference between group B and blank group. There was no statistical difference between group B and D group. There was no statistical difference between group C and group D, but there was no statistical difference between the C group and the blank group, but the difference was statistically different from the D group, and the D group was higher than the blank group. There was statistical difference between the group and the group of D. The difference was statistically significant Difference: the survival rate of 48h cells in A group was lower than that of blank group, B group, C group and D group had statistical difference, but there was no statistical difference between group B and C group, but higher than that of group A, lower than that of blank group and D group, there was no statistical difference, there was no statistical difference between the C group and the blank group, and the difference was statistically different from that in the D group. The D group was higher than the blank group, and the difference was statistically significant. Difference: the survival rate of 72h cells in A group was lower than that of blank group, group B, C group and D group, and there were statistical differences in group B, group B, C group and D group, and C group was lower than that of blank group, D group, and D group was higher than that of blank group, the difference was statistically different. The cell alkaline phosphatase and bone in each group of time points were different. There is no statistical difference in the expression of calcium protein. Conclusion: 3D printing titanium alloy porous coating can obviously affect the biological behavior of MG-63 cells. Relatively small micropores (218.7 mu m, 61.2%; 262 mu m, 68.1%) are beneficial to cell adhesion and differentiation, larger micropores (753 mu m, 89%) are beneficial to cell proliferation; small pore size porous coating culture cells are beneficial to cell proliferation. To collect possible errors, whether the lower cell value added rate is defined as material toxicity or inhibition of proliferation needs further research. Third part 3D printing porous titanium porous coating to promote bone integration and its mechanism: the study of bone integration and osteogenesis gene expression of titanium alloy porous coating with different micropore parameters in 3D printing Methods: 3D printers print 3 groups of titanium alloy prosthesis with porous coating with different microporous parameters, which are (group A, 262, m, 68.1%; B 558 mu m, 79.2%; C group 753 m, 89%). Select 27 New Zealand white rabbits randomly into 3 groups. 3 kinds of prosthesis were implanted in each group of rabbit bilateral femur medullary cavity, 4 weeks, 8 weeks, 12 weeks in every group. 3 rabbits were killed, X-ray, Micro-CT scan, bone volume fraction and tissue mineral density were measured. Hard tissue sections were made by Goldner's staining and fluorescence microscopy. The maximum pulling force was drawn out, and the expression of Runx2 and Osterix was detected by Q PCR. Results: both X-ray and Micro-CT scans showed no obvious bone deposition at 4 weeks. With the increase of time, the formation of new bone gradually increased, at 12 weeks at most, and group A was more than group B, group C. The fluorescence microscope observed that the fluorescence intensity was weaker and the band was narrower in 4 weeks after the operation. The fluorescence intensity increased, the band widened, the yellow and green space increased, the best in the 12 week, and the A group was superior to the B group, and the C group.Goldner's staining showed that all groups after 4 weeks after the.Goldner's staining were all in all groups. With the nucleus based, the osteoid increased gradually with time, reaching the peak at 12 weeks, and the A group was superior to the B group. The difference of bone volume fraction and tissue mineral density between the 3 groups was not statistically significant at the time of 4 weeks and 8 weeks after the operation, and the bone volume fraction and tissue mineral density of group A were greater than that of group B and group C at 12 weeks after operation, and the difference was statistically significant, B group and C were significant. At 4 weeks and 8 weeks after operation, the expression of Runx2 and Osterix in group A was higher than that in group B, in group C, there was no statistical difference between group B and C group. At 12 weeks after operation, the Runx2 expression of group C was lower than that of A group and B group. There was no statistical difference between the A group and the group, and there was no statistical difference between the A group and the group. 4 weeks after the operation, there was no statistical difference between the group of A and the group. The maximum pulling force difference between the 3 groups was not statistically significant. At 8 weeks after the operation, the maximum pulling force needed in group A was greater than that of group B, and the difference was statistically significant in group C, and there was no statistical significance in group B and C group. The difference was statistically significant at 12 weeks after the operation, and the difference was statistically significant in group A B group C. Conclusion: 3D printing titanium alloy. Porous coating can obviously promote bone integration. As time prolongs, the formation of bone is increasing and the deposition rate is gradually accelerated; porous coating with a pore size of 262 mu m and 68.1% porosity is most conducive to promoting bone integration, and the changes in microporous parameters have greater effect on biomechanics than on the effect of osteogenesis.
【學(xué)位授予單位】：第二軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：TG174.4;R318.08

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