生物活性玻璃强化骨质疏松椎弓根螺钉稳定性的生物力学及界面组织学研究
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摘要
目前椎弓根螺钉内固定技术已经成为脊柱外科的核心技术之一。椎弓根螺钉固定的稳定性取决于钉道对螺钉的把持力。在骨质疏松患者，由于骨小梁变薄、断裂，骨密度（bone mineral density，BMD）降低，钉道对螺钉的把持力明显下降，容易出现椎弓根螺钉松动、拔出等并发症从而导致内固定失败。如何提高骨质疏松条件下椎弓根螺钉内固定的稳定性成为骨科临床研究的重点和难点。目前国内外研究主要是将骨水泥类材料填充至钉道内，以强化螺钉固定。临床上常用聚甲基丙烯酸甲酯骨水泥（polymethylmethacry- late，PMMA)解决这一问题。PMMA虽然能显著增强椎弓根螺钉的稳定性，但它在体内无法吸收，在钉骨之间可形成二次界面，取钉极为困难，生物相容性差，放热反应等，临床医师对其应用持谨慎态度。45S5生物活性玻璃是美国Hench教授在1971年发现的一种新型生物活性材料，研究发现它能与骨形成骨键结合，具有很好的生物相容性和生物降解性，并能引导骨生长和诱导骨再生。生物活性玻璃已应用于骨修复领域，取得了很好的临床效果。国内外尚无学者将生物玻璃应用于强化螺钉固定的研究中。
研究目的：评价骨质疏松绵羊体内生物活性玻璃强化椎弓根螺钉稳定性的力学效果,并观察螺钉-骨质界面组织学变化。
研究方法:1.建立骨质疏松绵羊模型。8只成年雌性小尾寒羊,采用去势联合激素注射的方法建立骨质疏松动物模型,双能X线吸收法检测去势前和去势12月后绵羊腰椎骨密度,比较建模前后绵羊腰椎骨密度变化。2.取每只绵羊腰1-腰6双侧椎弓根为研究对象,将每只绵羊的12侧椎弓根随机分为3组:A组:空白组,对制备好的钉道不做任何处理,直接拧入椎弓根螺钉;B组:PMMA组,将配置好的骨水泥0.5ml注入钉道后拧入椎弓根螺钉;C组:BG组,将45S5生物玻璃约0.5g填入钉道后拧入椎弓根螺钉。分别饲养3个月和6个月后取材,轴向拔出实验评价3种方法强化螺钉稳定的力学效果。3、应用Micro-CT三维重建技术,荧光双标记方法和硬组织切片染色技术,观察BG组与空白组螺钉-骨界面骨生长、组织学等情况。
研究结果:
1、去势12个月后绵羊腰椎BMD(0.828±0.032g/cm~2),较去势前的绵羊腰椎BMD(1.015±0.037 g/cm~2)显著降低,平均下降22.70%,完成了建模。
2、生物力学结果显示:BG组在3个月和6个月时Fmax(分别为1083.09±86.48N、1175.20±134.84N)和E值(分别为1.8741±0.1724J、2.0375±0.2383J )显著高于空白组在相同时间点的Fmax (分别为871.76±79.03N、902.30±115.80N )和E值(分别为1.3058±0.1492J、1.4440±0.1759J),而显著低于PMMA组在相同时间点的Fmax(分别为1411.58±171.54N、1445.52±231.79N)和E值(分别为2.8418±0.2435J、3.0986±0.1599J),各组间差异有统计学意义(P<0.05)。不同时间点相同组间比较Fmax及E值差异无统计学意义(P>0.05)。
3、Micro-CT三维重建结果显示:BG组在3个月及6个月时其螺钉周围骨小梁数量较空白组显著增多,骨小梁结构更紧密,骨质情况优于空白组。骨计量学结果显示:3个月及6个月时BG组螺钉周围骨质参数中TMD、BVF、Tb.Th、Tb.N均显著高于空白组,而BS/BV、Tb.Sp显著低于空白组,差异均有统计学意义(P<0.05)。
4、荧光显微镜下观察结果显示:BG组在3个月时伴行的黄绿条带间的距离较宽,明显高于空白组及BG6个月组伴行的黄绿条带间距离。BG组在3个月时矿化沉积率(mineralization apposition rate,MAR)(11.94±1.26μm/d)显著高于空白组相同时间点的MAR(4.85±0.83μm/d),差异有统计学意义(P< 0.05 ),前者比后者增加了146.19%。而BG组在6个月时的MAR(4.86±1.12μm/d)与空白组相同时间点的MAR(4.74±0.72μm/d)无统计学差异(P>0.05)。荧光结果表明BG组在3月点时骨的生长矿化速度最快。
5、丽春红三色显微镜下观察结果显示:3个月时BG组螺钉周围骨量增多,骨小梁更密集,且对螺钉包裹紧密,螺钉-骨界面骨质情况明显优于空白组。6个月时BG组螺钉周围骨小梁结构更完善,骨小梁相对空白组增厚,结构密集,骨小梁对螺钉的包裹紧密,界面的骨质情况明显优于空白组。在两个时间点,均有少量BG存在螺钉周围骨质中,与骨结合紧密,6月组BG的量更少,降解的更完全。
结论:1、BG能增强OP条件下椎弓根螺钉固定的稳定性,且这种稳定性能长期维持。2、BG强化螺钉稳定性的力学效果不如PMMA。3、BG强化椎弓根螺钉稳定的机制是通过促进螺钉周围骨质生长,改善螺钉-骨界面骨质结构,提高界面对螺钉的把持作用而实现的。因此,BG可以成为临床上OP条件下提高骨螺钉界面强度进而提高椎弓根螺钉固定把持力的一种材料选择。
Currently pedicle screw fixation technique has already been one of the nucleus techniques in spinal column surgery. The pedicle screw fixing stationary stability is decided by the holding force of the screw pass vs. the screw. At osteoporosis patients, bone trabecula became thin and split, bone mineral density (BMD) stepped - down, the holding force of the screw pass vs. the screw obviously decreased, and the pedicle screw loosed and drawn out could easily appear, which would result in internal fixation failure. How to raise the stability of the pedicle screw fixation in OP condition becomes a key point and difficult point in orthopedics clinical research. Study mainly at home and abroad currently is filling the bone cement materials into the nail pass so as to enhance pedicle screw stability. In clinical we usually use the polymethylmethacrylate (PMMA) bone cement to resolve this problem. Although PMMA can obviously enhance the stability of pedicle screw fixation, it can not absorb in the body, forming two interfaces between screw and bone, taking out the screw extremely difficult, its biocompatibility is bad, exothermic reaction etc. clinical doctors had a careful attitude towards its application. 45S5 bioactive glass is a kind of new bioactive material discovered by Professor Hench from United States in 1971. Studies showed that BG could bond to the bone, had good biocompatibility and biodegradability, could lead bone growth and induce bone regeneration. Bioactive glass had already been used in bone repair realm, and obtained good clinical effect. At home and abroad there is nobody got bioactive glass into pedicle screw fixation study.
Objective: To evaluate the mechanical effect of pedicle screw augmentation using bioactive glass (BG) in osteoporosis sheep and observe the change of histology in the interface between bone and screw.
Methods: 1. An OP sheep model was established. Osteoporosis sheep models were established by ovariectomy combined with methylprednisolone injection in 8 adult female sheep, the BMD of the lumber of the sheep before and 12 months after OVX was detected by dual energy X-ray absorptiometry (DXA), and the BMD changes were compared. 2. Took L1-L6 bilateral pedicles of each as object of study, all 12 pedicles were randomly divided into 3 groups. Group A: control group, the pedicles were implanted with pedicle screws without any transactions; Group B: PMMA group, the pedicles were implanted with pedicle screws after the augmentation with PMMA (about 0.5ml); Group C: BG group, the pedicles were implanted with pedicle screws after the augmentation with 45S5 bioglass (about 0.5g). Got materials after 3 months or 6 months feeding respectively, and axial pull-out-test was taken to evaluate the mechanical effect of pedicle screw augmentation in 3 groups. 3. Apply a 3D reconstruction technique of Micro-CT, fluorescence double label method and sclerous tissue section dye technique, to observe the bone growth and the change of histology in the interface of bone and screw.
Result:
1. The bone mineral density (BMD) of the lumbar of the sheep 12 months after OVX (0.828±0.032g/cm~2) was significance decreased by 22.70% compared with that before OVX (1.015±0.037 g/cm~2), and the OP sheep model was completed.
2. The biomechanics result showed that the Fmax and E of BG group in 3 month and 6 month (1083.09±86.48N, 1175.20±134.84N respectively and 1.8741±0.1724.K 2.0375±0.2383J respectively ) was much higher than the Fmax and E of the control group in the same time stamp (871.76±79.03N、902.30±115.80 respectively and 1.3058±0.1492K 1.4440±0.1759J respectively), and was much lower than the Fmax and E of the PMMA group in the same time stamp (1411.58±171.54N, 1445.52±231.79N respectively and 2.8418±0.2435K 3.0986+0.1599J respectively), and significant difference was found in each group (P<0.05). The Fmax and E in the same group of different time stamp had no significant difference (P>0.05).
3. The 3D reconstruction result of Micro-CT showed that the mass of the bone trabecular around the screw was much more and the structure was close-packed in BG group in 3 month and 3 month time stamp, and its bone substance surpassed the control group. The bone metrology result showed that the TMD, BVF, Tb.Th, Tb.N in BG group in 3 and 6 month stamp was much higher than that of the control group, and the BS/BV、Tb.Sp of the BG group was much lower than that of the control group, and significant difference was found (P< 0.05).
4. The fluorescence result under fluorescence microscope showed that the distance of the yellow and green fluorescence band of BG group in 3 month stamp was much bigger than that of the control group and BG group in 6 month stamp. The mineralization apposition rate (MAR) of BG group in 3 month time stamp (11.94±1.26um/d ) was much higher than the MAR of the control group in the same time stamp (4.85±0.83 um/d), significant different was found (P<0.05), and the former was increased by 146.19% compared whit the latter. The MAR of BG group in 6 month time stamp (4.86±1.12um/d) had no significant difference compared whit the control group in the same time stamp(4.74±0.72um/d) (P> 0.05). The fluorescence results indicated that the bone in the BG group in 3 month time stamp had a rapid growth rate.
5. Ponceau staining under microscope showed that the bone mass around the screw was much more and the bone trabecular was close-packed and wrapped the screws tightly in BG group of 3 month time stamp, the bone substance of the bone-screw interface surpassed the control group. In BG group of 6 month time stamp, the trabecular around the screw had a better bone structure and was thicker than that of the control group, the bone structure was close-packed and the trabecular wrapped the screws tightly, the bone substance of the interface surpassed the control group. In two time stamps, there were some BG found in the bone substance around the screw and combined with the bone well. In 6 month time stamp, the BG existed in the bone was much fewer, and had almost degradation completely.
Conclusion: 1. BG could enhance the stability of pedicle screw fixation in OP condition, and it could maintain the stability over a long term. 2. The mechanics effect of BG in pedicle screw augmentation was not equal to PMMA. 3. The mechanism of pedicle screw augmentation with BG was that the BG could induce growth of the bone around the screw, improve the bone structure of screw- bone interface, and enhance the holding force of the interface vs. the screw. Therefore, BG can be one of materials choosing in pedicle screw augmentation through improving bone-screw interface intensity and enhancing the hoding force of pedicle screw under OP condition.
研究目的：评价骨质疏松绵羊体内生物活性玻璃强化椎弓根螺钉稳定性的力学效果,并观察螺钉-骨质界面组织学变化。
研究方法:1.建立骨质疏松绵羊模型。8只成年雌性小尾寒羊,采用去势联合激素注射的方法建立骨质疏松动物模型,双能X线吸收法检测去势前和去势12月后绵羊腰椎骨密度,比较建模前后绵羊腰椎骨密度变化。2.取每只绵羊腰1-腰6双侧椎弓根为研究对象,将每只绵羊的12侧椎弓根随机分为3组:A组:空白组,对制备好的钉道不做任何处理,直接拧入椎弓根螺钉;B组:PMMA组,将配置好的骨水泥0.5ml注入钉道后拧入椎弓根螺钉;C组:BG组,将45S5生物玻璃约0.5g填入钉道后拧入椎弓根螺钉。分别饲养3个月和6个月后取材,轴向拔出实验评价3种方法强化螺钉稳定的力学效果。3、应用Micro-CT三维重建技术,荧光双标记方法和硬组织切片染色技术,观察BG组与空白组螺钉-骨界面骨生长、组织学等情况。
研究结果:
1、去势12个月后绵羊腰椎BMD(0.828±0.032g/cm~2),较去势前的绵羊腰椎BMD(1.015±0.037 g/cm~2)显著降低,平均下降22.70%,完成了建模。
2、生物力学结果显示:BG组在3个月和6个月时Fmax(分别为1083.09±86.48N、1175.20±134.84N)和E值(分别为1.8741±0.1724J、2.0375±0.2383J )显著高于空白组在相同时间点的Fmax (分别为871.76±79.03N、902.30±115.80N )和E值(分别为1.3058±0.1492J、1.4440±0.1759J),而显著低于PMMA组在相同时间点的Fmax(分别为1411.58±171.54N、1445.52±231.79N)和E值(分别为2.8418±0.2435J、3.0986±0.1599J),各组间差异有统计学意义(P<0.05)。不同时间点相同组间比较Fmax及E值差异无统计学意义(P>0.05)。
3、Micro-CT三维重建结果显示:BG组在3个月及6个月时其螺钉周围骨小梁数量较空白组显著增多,骨小梁结构更紧密,骨质情况优于空白组。骨计量学结果显示:3个月及6个月时BG组螺钉周围骨质参数中TMD、BVF、Tb.Th、Tb.N均显著高于空白组,而BS/BV、Tb.Sp显著低于空白组,差异均有统计学意义(P<0.05)。
4、荧光显微镜下观察结果显示:BG组在3个月时伴行的黄绿条带间的距离较宽,明显高于空白组及BG6个月组伴行的黄绿条带间距离。BG组在3个月时矿化沉积率(mineralization apposition rate,MAR)(11.94±1.26μm/d)显著高于空白组相同时间点的MAR(4.85±0.83μm/d),差异有统计学意义(P< 0.05 ),前者比后者增加了146.19%。而BG组在6个月时的MAR(4.86±1.12μm/d)与空白组相同时间点的MAR(4.74±0.72μm/d)无统计学差异(P>0.05)。荧光结果表明BG组在3月点时骨的生长矿化速度最快。
5、丽春红三色显微镜下观察结果显示:3个月时BG组螺钉周围骨量增多,骨小梁更密集,且对螺钉包裹紧密,螺钉-骨界面骨质情况明显优于空白组。6个月时BG组螺钉周围骨小梁结构更完善,骨小梁相对空白组增厚,结构密集,骨小梁对螺钉的包裹紧密,界面的骨质情况明显优于空白组。在两个时间点,均有少量BG存在螺钉周围骨质中,与骨结合紧密,6月组BG的量更少,降解的更完全。
结论:1、BG能增强OP条件下椎弓根螺钉固定的稳定性,且这种稳定性能长期维持。2、BG强化螺钉稳定性的力学效果不如PMMA。3、BG强化椎弓根螺钉稳定的机制是通过促进螺钉周围骨质生长,改善螺钉-骨界面骨质结构,提高界面对螺钉的把持作用而实现的。因此,BG可以成为临床上OP条件下提高骨螺钉界面强度进而提高椎弓根螺钉固定把持力的一种材料选择。
Currently pedicle screw fixation technique has already been one of the nucleus techniques in spinal column surgery. The pedicle screw fixing stationary stability is decided by the holding force of the screw pass vs. the screw. At osteoporosis patients, bone trabecula became thin and split, bone mineral density (BMD) stepped - down, the holding force of the screw pass vs. the screw obviously decreased, and the pedicle screw loosed and drawn out could easily appear, which would result in internal fixation failure. How to raise the stability of the pedicle screw fixation in OP condition becomes a key point and difficult point in orthopedics clinical research. Study mainly at home and abroad currently is filling the bone cement materials into the nail pass so as to enhance pedicle screw stability. In clinical we usually use the polymethylmethacrylate (PMMA) bone cement to resolve this problem. Although PMMA can obviously enhance the stability of pedicle screw fixation, it can not absorb in the body, forming two interfaces between screw and bone, taking out the screw extremely difficult, its biocompatibility is bad, exothermic reaction etc. clinical doctors had a careful attitude towards its application. 45S5 bioactive glass is a kind of new bioactive material discovered by Professor Hench from United States in 1971. Studies showed that BG could bond to the bone, had good biocompatibility and biodegradability, could lead bone growth and induce bone regeneration. Bioactive glass had already been used in bone repair realm, and obtained good clinical effect. At home and abroad there is nobody got bioactive glass into pedicle screw fixation study.
Objective: To evaluate the mechanical effect of pedicle screw augmentation using bioactive glass (BG) in osteoporosis sheep and observe the change of histology in the interface between bone and screw.
Methods: 1. An OP sheep model was established. Osteoporosis sheep models were established by ovariectomy combined with methylprednisolone injection in 8 adult female sheep, the BMD of the lumber of the sheep before and 12 months after OVX was detected by dual energy X-ray absorptiometry (DXA), and the BMD changes were compared. 2. Took L1-L6 bilateral pedicles of each as object of study, all 12 pedicles were randomly divided into 3 groups. Group A: control group, the pedicles were implanted with pedicle screws without any transactions; Group B: PMMA group, the pedicles were implanted with pedicle screws after the augmentation with PMMA (about 0.5ml); Group C: BG group, the pedicles were implanted with pedicle screws after the augmentation with 45S5 bioglass (about 0.5g). Got materials after 3 months or 6 months feeding respectively, and axial pull-out-test was taken to evaluate the mechanical effect of pedicle screw augmentation in 3 groups. 3. Apply a 3D reconstruction technique of Micro-CT, fluorescence double label method and sclerous tissue section dye technique, to observe the bone growth and the change of histology in the interface of bone and screw.
Result:
1. The bone mineral density (BMD) of the lumbar of the sheep 12 months after OVX (0.828±0.032g/cm~2) was significance decreased by 22.70% compared with that before OVX (1.015±0.037 g/cm~2), and the OP sheep model was completed.
2. The biomechanics result showed that the Fmax and E of BG group in 3 month and 6 month (1083.09±86.48N, 1175.20±134.84N respectively and 1.8741±0.1724.K 2.0375±0.2383J respectively ) was much higher than the Fmax and E of the control group in the same time stamp (871.76±79.03N、902.30±115.80 respectively and 1.3058±0.1492K 1.4440±0.1759J respectively), and was much lower than the Fmax and E of the PMMA group in the same time stamp (1411.58±171.54N, 1445.52±231.79N respectively and 2.8418±0.2435K 3.0986+0.1599J respectively), and significant difference was found in each group (P<0.05). The Fmax and E in the same group of different time stamp had no significant difference (P>0.05).
3. The 3D reconstruction result of Micro-CT showed that the mass of the bone trabecular around the screw was much more and the structure was close-packed in BG group in 3 month and 3 month time stamp, and its bone substance surpassed the control group. The bone metrology result showed that the TMD, BVF, Tb.Th, Tb.N in BG group in 3 and 6 month stamp was much higher than that of the control group, and the BS/BV、Tb.Sp of the BG group was much lower than that of the control group, and significant difference was found (P< 0.05).
4. The fluorescence result under fluorescence microscope showed that the distance of the yellow and green fluorescence band of BG group in 3 month stamp was much bigger than that of the control group and BG group in 6 month stamp. The mineralization apposition rate (MAR) of BG group in 3 month time stamp (11.94±1.26um/d ) was much higher than the MAR of the control group in the same time stamp (4.85±0.83 um/d), significant different was found (P<0.05), and the former was increased by 146.19% compared whit the latter. The MAR of BG group in 6 month time stamp (4.86±1.12um/d) had no significant difference compared whit the control group in the same time stamp(4.74±0.72um/d) (P> 0.05). The fluorescence results indicated that the bone in the BG group in 3 month time stamp had a rapid growth rate.
5. Ponceau staining under microscope showed that the bone mass around the screw was much more and the bone trabecular was close-packed and wrapped the screws tightly in BG group of 3 month time stamp, the bone substance of the bone-screw interface surpassed the control group. In BG group of 6 month time stamp, the trabecular around the screw had a better bone structure and was thicker than that of the control group, the bone structure was close-packed and the trabecular wrapped the screws tightly, the bone substance of the interface surpassed the control group. In two time stamps, there were some BG found in the bone substance around the screw and combined with the bone well. In 6 month time stamp, the BG existed in the bone was much fewer, and had almost degradation completely.
Conclusion: 1. BG could enhance the stability of pedicle screw fixation in OP condition, and it could maintain the stability over a long term. 2. The mechanics effect of BG in pedicle screw augmentation was not equal to PMMA. 3. The mechanism of pedicle screw augmentation with BG was that the BG could induce growth of the bone around the screw, improve the bone structure of screw- bone interface, and enhance the holding force of the interface vs. the screw. Therefore, BG can be one of materials choosing in pedicle screw augmentation through improving bone-screw interface intensity and enhancing the hoding force of pedicle screw under OP condition.
引文
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