自固化磷酸钙骨水泥的制备及其性能评估

Preparation and property evaluation of a self-setting calcium phosphate cement

背景:前期研究发现,乳酸-羟基乙酸共聚物(poly(lactic-co-glycolic acid),PLGA)可改善磷酸钙骨水泥的抗压强度与降解性能。目的:在前期研究的基础上制备具有更好力学性能、生物相容性及降解性的自固化磷酸钙骨水泥。方法:采用液相沉淀法制备乳酸-羟基乙酸共聚物/β-磷酸三钙/磷酸钙骨水泥,使复合骨水泥中β-磷酸三钙的含量分别为5%,10%,15%,20%,25%,30%,检测复合骨水泥及磷酸钙骨水泥的凝固时间、抗压强度、弹性模量及降解性能,筛选最佳β-磷酸三钙含量,进行细胞培养实验。分别以磷酸钙骨水泥浸提液(对照组)、PLGA/β-磷酸三钙/磷酸钙骨水泥浸提液(实验组)、含有体积分数10%胎牛血清及1%青霉素-链霉素双抗的α-MEM培养基(阴性对照组)、含体积分数6.4%苯酚的液体(阳性对照组)培养MC3T3-E1细胞,培养1,3,5 d,MTT法检测细胞增殖,同时观察细胞形态变化;培养1,3 d,检测培养液中乳酸脱氢酶活性。在PLGA/β-磷酸三钙/磷酸钙骨水泥与磷酸钙骨水泥表面接种MC3T3-E1细胞,3 d后进行扫描电镜观察。结果与结论:(1)随着β-磷酸三钙含量的增加,复合骨水泥的初凝时间、终凝时间均有所增加,与磷酸钙骨水泥初凝时间、终凝时间比较无差异(P>0.05);(2)复合骨水泥的抗压强度及弹性模量均高于磷酸钙骨水泥,其中以β-磷酸三钙含量为20%的复合骨水泥抗压强度最高,弹性模量也较好,因此实验选择β-磷酸三钙含量为20%的PLGA/β-磷酸三钙/磷酸钙骨水泥进行细胞实验;复合骨水泥的降解性能优于磷酸钙骨水泥;(3)随着培养时间的延长,实验组细胞吸光度、乳酸脱氢酶活性逐渐升高,不同时间点的吸光度值、乳酸脱氢酶活性与阴性对照组比较均无差异;实验组细胞生长良好;(4)在PLGA/β-磷酸三钙/磷酸钙骨水泥表面的MC3T3-E1细胞生长正常,能充分伸展,在材料表面伸出伪足,能够与材料紧密贴附;(5)结果表明,含20%β-磷酸三钙的PLGA/β-磷酸三钙/磷酸钙骨水泥复合材料具有较普通磷酸钙骨水泥更佳的抗压强度、弹性模量及降解性能,且细胞相容性好,无明显细胞毒性。

BACKGROUND： Preliminary studies have found that poly（lactic-co-glycolic acid） （PLGA） can improve the compressive strength and degradability of calcium phosphate cement. OBJECTIVE： To prepare a self-setting calcium phosphate cement which has better mechanical properties, biocompatibility and degradability on the basis of the previous findings. METHODS： Beta-tricalcium phosphate （β-TCP）, pure calcium phosphate cement （CPC） and PLGA powder were mixed at different mixing ratios for preparation of PLGA/β-TCP/CPC. Setting time, compressive strength, elastic modulus and degradation properties of the composite bone cement were evaluated to screen the optimal level of β-TCP. MC3T3-E1 cells were cultured in CPC extract （control）, PLGA/β-TCP/CPC extract （experimental）, α-MEM containing 10% fetal bovine serum and 1% penicillin-streptomycin double antibody （negative control）, and 6.4% phenol liquid （positive control）. MTT method was used to detect cell proliferation at 1, 3, 5 days after culture, and lactate dehydrogenase activity in culture media was detected at 1 and 3 days after culture. MC3T3-E1 cells were seeded on the surface of PLGA/β-TCP/CPC and pure CPC respectively, and were then observed by scanning electron microscopy after 3 days. RESULTS AND CONCLUSION： Initial setting time and final setting time among of the composite bone cement were increased with the increasing of β-TCP content, but had no significant difference from those of the CPC （P 〉 0.05）. The compressive strength and elastic modulus of the composite bone cement were higher than those of the CPC, and moreover, the composite bone cement with 20% β-TCP exhibited the highest compressive strength and higher elastic modulus as compared with the other groups. Therefore, PLGA/20% β-TCP/CPC was selected in the cell test. Moreover, the degradation properties of the composite bone cement were also better than those of the CPC. （3） With the growth of culture time, cell absorbance value and lactate dehydrogenase activity were gradually increased in the experimental group, and no difference existed between the experimental group and the negative control group. The cells in the experimental group also grew well. （4） MC3T3-E1 cells grew well and fully extended on the surface of PLGA/β-TCP/CPC, and cell pseudopodia on the material surface were tightly adhered to the material. To conclude, PLGA/20% β-TCP/CPC has better compressive strength, elastic modulus, degradation properties and cytocompatibility relative to the CPC, and moreover, the composite bone cement has no obvious cytotoxicity.