3D打印β-磷酸三钙支架复合淫羊藿苷微粒修复兔股骨头坏死
|
摘要
背景:前期研究制备了3D打印β-磷酸三钙复合淫羊藿苷支架。目的:观察3D打印β-磷酸三钙复合淫羊藿苷支架在兔股骨头坏死模型中的修复作用。方法:取新西兰大白兔(南京青龙山实验动物中心提供),制作激素型股骨头坏死模型,将造模成功的27只兔进行髓芯减压及清理死骨,随机分3组,分别植入自体骨(自体骨组)、β-磷酸三钙支架(磷酸三钙组)及3D打印β-磷酸三钙复合淫羊藿苷支架(复合支架组)。植入后第4,8,12周,进行股骨头micro-CT扫描及病理组织观察。结果与结论:①micro-CT显示,植入第4周,3组植入物周围及内部均有骨小梁生成;自体骨组植入后第8周植骨区有大量骨小梁,植入后第12周骨小梁结构致密;磷酸三钙组、复合支架组支架与骨界面整合良好,复合支架组植入第4周时支架附近有一定量的骨小梁生成,至第8周时支架上有骨小梁长入,磷酸三钙组植入第4周时仅可见微量薄弱骨小梁,第8周时才可见支架周围广泛骨小梁生成,第12周时骨小梁仍较其他两组稀疏;②植入第12周苏木精-伊红染色显示,自体骨、复合支架组成骨细胞分化成熟,软骨基质较少,新生骨组织与植入物整合良好,磷酸三钙组仍存在较多的软骨基质,新生骨与植入物尚未完成整合;③植入第12周Masson染色显示,复合支架组成骨细胞增殖低于自体骨组(P <0.05),但高于磷酸三钙组(P <0.05);④植入第12周TRAP染色显示,复合支架组破骨细胞数少于磷酸三钙组(P <0.05),与自体骨组比较无差异(P> 0.05);⑤植入第12周免疫组织化学染色显示,复合支架组血管内皮生长因子阳性率高于磷酸三钙组(P <0.05),但低于自体骨组(P <0.05);⑥结果表明,3D打印β-磷酸三钙复合淫羊藿苷支架植入股骨头坏死模型兔体内,可促进成骨细胞增殖分化、抑制破骨细胞活性,同时促进新生血管生成,具有促进兔股骨头坏死修复的作用。
BACKGROUND: Preliminary study has prepared three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin. OBJECTIVE: To investigate the role of three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin in the repair of rabbit models of osteonecrosis of the femoral head. METHODS: New Zealand white rabbits(provided by Qinglongshan Laboratory Animal Center of Nanjing) were selected to establish the steroid-induced osteonecrosis of the femoral head. The 27 model rabbits underwent core decompression and debridement, were randomly divided into three groups, and then implanted with autologous bone, β-tricalcium phosphate scaffold, three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin(composite scaffold group), respectively. The micro-CT scanning and pathological observation were performed at 4, 8, and 12 weeks after implantation. RESULTS AND CONCLUSION:(1) Micro-CT showed that at 4 weeks after implantation, trabecular bone was observed around and in implants in each group. In the autologous bone group, there were a large number of trabecular bones in the grafting area at 8 weeks, and the trabecular bone structure was dense at 12 weeks after implantation. In the tricalcium phosphate and composite scaffold groups, the scaffolds were well integrated with the bone interface. At 4 weeks after implantation, there was a certain amount of trabecular bone surrounding the scaffold, and trabecular grew into the scaffold until 8 weeks in the composite scaffold group. At 4 weeks after implantation, few thin trabecular bone was visible, and extensive trabecular bone formation was observed around the scaffold at 8 weeks in the tricalcium phosphate group.(2) Hematoxylin-eosin staining results showed that there were many mature osteoblasts, and few cartilage matrix, newly born bones integrated well to the implants at 12 weeks in the autologous bone and tricalcium phosphate groups. In the composite scaffold group, there were many cartilage matrixes, and newly born bones integrated poorly to the implants.(3) Masson staining showed that at 12 weeks after implantation, the osteogenic capacity in the composite scaffold group was lower than that in the autologous bone group(P < 0.05), but higher than that in the tricalcium phosphate group(P < 0.05).(4) TRAP staining results at 12 weeks after implantation revealed that the amount of osteoclast in composite scaffold group was less than that in the tricalcium phosphate group(P < 0.05), and was not significantly different from the autologous bone group(P > 0.05).(5) Immunohistochemical staining at 12 weeks after implantation revealed that the positive rate of vascular endothelial growth factor in the composite scaffold group was higher than that in the tricalcium phosphate group(P < 0.05), and lower than that in the autologous bone group(P < 0.05).(6) In summary, three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin implanted into the rabbit model of osteonecrosis of the femoral head can promote the proliferation and differentiation of osteoblasts, inhibit the viability of osteoclasts, promote the angiogenesis, and contribute to the repair of osteonecrosis of the femoral head in rabbits.
BACKGROUND: Preliminary study has prepared three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin. OBJECTIVE: To investigate the role of three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin in the repair of rabbit models of osteonecrosis of the femoral head. METHODS: New Zealand white rabbits(provided by Qinglongshan Laboratory Animal Center of Nanjing) were selected to establish the steroid-induced osteonecrosis of the femoral head. The 27 model rabbits underwent core decompression and debridement, were randomly divided into three groups, and then implanted with autologous bone, β-tricalcium phosphate scaffold, three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin(composite scaffold group), respectively. The micro-CT scanning and pathological observation were performed at 4, 8, and 12 weeks after implantation. RESULTS AND CONCLUSION:(1) Micro-CT showed that at 4 weeks after implantation, trabecular bone was observed around and in implants in each group. In the autologous bone group, there were a large number of trabecular bones in the grafting area at 8 weeks, and the trabecular bone structure was dense at 12 weeks after implantation. In the tricalcium phosphate and composite scaffold groups, the scaffolds were well integrated with the bone interface. At 4 weeks after implantation, there was a certain amount of trabecular bone surrounding the scaffold, and trabecular grew into the scaffold until 8 weeks in the composite scaffold group. At 4 weeks after implantation, few thin trabecular bone was visible, and extensive trabecular bone formation was observed around the scaffold at 8 weeks in the tricalcium phosphate group.(2) Hematoxylin-eosin staining results showed that there were many mature osteoblasts, and few cartilage matrix, newly born bones integrated well to the implants at 12 weeks in the autologous bone and tricalcium phosphate groups. In the composite scaffold group, there were many cartilage matrixes, and newly born bones integrated poorly to the implants.(3) Masson staining showed that at 12 weeks after implantation, the osteogenic capacity in the composite scaffold group was lower than that in the autologous bone group(P < 0.05), but higher than that in the tricalcium phosphate group(P < 0.05).(4) TRAP staining results at 12 weeks after implantation revealed that the amount of osteoclast in composite scaffold group was less than that in the tricalcium phosphate group(P < 0.05), and was not significantly different from the autologous bone group(P > 0.05).(5) Immunohistochemical staining at 12 weeks after implantation revealed that the positive rate of vascular endothelial growth factor in the composite scaffold group was higher than that in the tricalcium phosphate group(P < 0.05), and lower than that in the autologous bone group(P < 0.05).(6) In summary, three-dimensional printing β-tricalcium phosphate scaffold loaded with icariin implanted into the rabbit model of osteonecrosis of the femoral head can promote the proliferation and differentiation of osteoblasts, inhibit the viability of osteoclasts, promote the angiogenesis, and contribute to the repair of osteonecrosis of the femoral head in rabbits.
引文
[1]Ye YH,Chen K,Jin KK,et al.Progress on surgical treatment for femoral head-preservering in the precollapse stage of femoral head necrosis.Zhongguo Gu Shang.2017;30(3):287-292.
[2]Utsunomiya T,Motomura G,Ikemura S,et al.Effects of sclerotic changes on stress concentration in early-stage osteonecrosis:A patient-specific,3D finite element analysis.J Orthop Res.2018;36(12):3169-3177.
[3]Issa K,Pivec R,Kapadia BH,et al.Osteonecrosis of the femoral head:the total hipreplacement solution.Bone Joint J.2013;95-B(11 Suppl A):46-50.
[4]朱旭日,杜斌,孙光权,等.髓芯减压打压植骨腓骨支撑术与头颈部开窗打压植骨术治疗早中期股骨头坏死疗效比较[J].中国骨与关节损伤杂志,2015,30(4):343-345.
[5]杜斌,孙鲁宁,袁滨,等.微创死骨清除打压植骨腓骨支撑配合中药补肾活血汤治疗早中期股骨头坏死的临床报道[J].中华关节外科杂志(电子版),2013,7(3):404-407.
[6]Korompilias AV,Beris AE,Lykissas MG,et al.Femoral head osteonecrosis:why choose free vascularized fibula grafting.Microsurgery.2011;31(3):223-228.
[7]党莹,李月,李瑞玉,等.骨组织工程支架材料在骨缺损修复及3D打印技术中的应用[J].中国组织工程研究,2017,21(14):2266-2273.
[8]?poner P,Ku?era T,BrtkováJ,et al.Comparative Study on the Application of Mesenchymal Stromal Cells Combined with Tricalcium Phosphate Scaffold into Femoral Bone Defects.Cell Transplant.2018;27(10):1459-1468.
[9]Bonazza V,Hajistilly C,Pate lD,et al.Growth Factors Release From Concentrated Growth Factors:Effect ofβ-Tricalcium Phosphate Addition.J Craniofac Surg.2018;29(8):2291-2295.
[10]Wu JZ,Liu PC,Liu R.Icariin Restores Bone Structure and Strength in a Rat Model of Chronic High-Dose Alcohol-Induced Osteopenia.Cell Physiol Biochem.2018;46(4):1727-1736.
[11]Jing X,Yin W,Tian H,et al.Icariin doped bioactive glasses seeded with rat adipose-derived stem cells to promote bone repair via enhanced osteogenic and angiogenic activities.Life Sci.2018;202:52-60.
[12]薛鹏,杜斌,孙光权,等.可控释淫羊藿苷-β-磷酸三钙复合支架的制备[J].中国组织工程研究,2018,22(6):865-870.
[13]The Ministry of Science and Technology of the People’s Republic of China.Guidance Suggestions for the Care and Use of Laboratory Animals.2006-09-30.中华人民共和国科学技术部.关于善待实验动物的指导性意见.2006-09-30.
[14]曹良权,杜斌,孙光权,等.不同剂量糖皮质激素联合马血清构建激素诱导型股骨头缺血性坏死兔模型[J].中国组织工程研究,2017,21(8):1229-1235.
[15]Li D,Xie X,Kang P,et al.Percutaneously drilling through femoral head and neck fenestration combining with compacted autograft for early femoral head necrosis:A retrospective study.J Orthop Sci.2017;22(6):1060-1065.
[16]Zhu W,Zhao Y,Ma Q,et al.3D-printed porous titanium changed femoral head repair growth patterns:osteogenesis and vascularisation in porous titanium.J Mater Sci Mater Med.2017;28(4):62.
[17]Hu R,Lei P,Li B,et al.Real-time computerised tomography assisted porous tantalum implant in ARCO stage I-IInon-traumatic osteonecrosis of the femoral head:minimum five-year follow up.Int Orthop.2018;42(7):1535-1544.
[18]Cheng Q,Tang JL,Gu JJ,et al.Total hip arthroplasty following failure of tantalum rod implantation for osteonecrosis of the femoral head with 5-to 10-year follow-up.BMC Musculoskelet Disord.2018;19(1):289.
[19]Hsieh TP,Sheu SY,Sun JS,et al.Icariin isolated from Epimedium pubescens regulates osteoblasts anabolism through BMP-2,SMAD4,andCbfa1expression.Phytomedicine.2010;17(6):414-423.
[20]Shi W,Gao Y,Wang Y,et al.The flavonol glycoside icariin promotes bone formation in growing rats by activating the cAMP signaling pathway in primary cilia of osteoblasts.J Biol Chem.2017;292(51):20883-20896.
[21]Liu Y,Huang L,Hao B,et al.Use of an Osteoblast Overload Damage Model to Probe the Effect of Icariin on the Proliferation,Differentiation and Mineralization of MC3T3-E1 Cells through the Wnt/β-Catenin Signalling Pathway.Cell Physiol Biochem.2017;41(4):1605-1615.
[22]Ma XN,Ge BF,Chen KM,et al.Mechanisms of icariin in regulating bone formation of osteoblasts and bone resorption of osteoclasts.Zhongguo Yi Xue Ke Xue Yuan Xue Bao.2013;35(4):432-438.
[23]Chung BH,Kim JD,Kim CK,et al.Icariin stimulates angiogenesis by activating the MEK/ERK-and PI3K/Akt/eNOS-dependent signal pathways in human endothelial cells.Biochem Biophys Res Commun.2008;376(2):404-408.
[2]Utsunomiya T,Motomura G,Ikemura S,et al.Effects of sclerotic changes on stress concentration in early-stage osteonecrosis:A patient-specific,3D finite element analysis.J Orthop Res.2018;36(12):3169-3177.
[3]Issa K,Pivec R,Kapadia BH,et al.Osteonecrosis of the femoral head:the total hipreplacement solution.Bone Joint J.2013;95-B(11 Suppl A):46-50.
[4]朱旭日,杜斌,孙光权,等.髓芯减压打压植骨腓骨支撑术与头颈部开窗打压植骨术治疗早中期股骨头坏死疗效比较[J].中国骨与关节损伤杂志,2015,30(4):343-345.
[5]杜斌,孙鲁宁,袁滨,等.微创死骨清除打压植骨腓骨支撑配合中药补肾活血汤治疗早中期股骨头坏死的临床报道[J].中华关节外科杂志(电子版),2013,7(3):404-407.
[6]Korompilias AV,Beris AE,Lykissas MG,et al.Femoral head osteonecrosis:why choose free vascularized fibula grafting.Microsurgery.2011;31(3):223-228.
[7]党莹,李月,李瑞玉,等.骨组织工程支架材料在骨缺损修复及3D打印技术中的应用[J].中国组织工程研究,2017,21(14):2266-2273.
[8]?poner P,Ku?era T,BrtkováJ,et al.Comparative Study on the Application of Mesenchymal Stromal Cells Combined with Tricalcium Phosphate Scaffold into Femoral Bone Defects.Cell Transplant.2018;27(10):1459-1468.
[9]Bonazza V,Hajistilly C,Pate lD,et al.Growth Factors Release From Concentrated Growth Factors:Effect ofβ-Tricalcium Phosphate Addition.J Craniofac Surg.2018;29(8):2291-2295.
[10]Wu JZ,Liu PC,Liu R.Icariin Restores Bone Structure and Strength in a Rat Model of Chronic High-Dose Alcohol-Induced Osteopenia.Cell Physiol Biochem.2018;46(4):1727-1736.
[11]Jing X,Yin W,Tian H,et al.Icariin doped bioactive glasses seeded with rat adipose-derived stem cells to promote bone repair via enhanced osteogenic and angiogenic activities.Life Sci.2018;202:52-60.
[12]薛鹏,杜斌,孙光权,等.可控释淫羊藿苷-β-磷酸三钙复合支架的制备[J].中国组织工程研究,2018,22(6):865-870.
[13]The Ministry of Science and Technology of the People’s Republic of China.Guidance Suggestions for the Care and Use of Laboratory Animals.2006-09-30.中华人民共和国科学技术部.关于善待实验动物的指导性意见.2006-09-30.
[14]曹良权,杜斌,孙光权,等.不同剂量糖皮质激素联合马血清构建激素诱导型股骨头缺血性坏死兔模型[J].中国组织工程研究,2017,21(8):1229-1235.
[15]Li D,Xie X,Kang P,et al.Percutaneously drilling through femoral head and neck fenestration combining with compacted autograft for early femoral head necrosis:A retrospective study.J Orthop Sci.2017;22(6):1060-1065.
[16]Zhu W,Zhao Y,Ma Q,et al.3D-printed porous titanium changed femoral head repair growth patterns:osteogenesis and vascularisation in porous titanium.J Mater Sci Mater Med.2017;28(4):62.
[17]Hu R,Lei P,Li B,et al.Real-time computerised tomography assisted porous tantalum implant in ARCO stage I-IInon-traumatic osteonecrosis of the femoral head:minimum five-year follow up.Int Orthop.2018;42(7):1535-1544.
[18]Cheng Q,Tang JL,Gu JJ,et al.Total hip arthroplasty following failure of tantalum rod implantation for osteonecrosis of the femoral head with 5-to 10-year follow-up.BMC Musculoskelet Disord.2018;19(1):289.
[19]Hsieh TP,Sheu SY,Sun JS,et al.Icariin isolated from Epimedium pubescens regulates osteoblasts anabolism through BMP-2,SMAD4,andCbfa1expression.Phytomedicine.2010;17(6):414-423.
[20]Shi W,Gao Y,Wang Y,et al.The flavonol glycoside icariin promotes bone formation in growing rats by activating the cAMP signaling pathway in primary cilia of osteoblasts.J Biol Chem.2017;292(51):20883-20896.
[21]Liu Y,Huang L,Hao B,et al.Use of an Osteoblast Overload Damage Model to Probe the Effect of Icariin on the Proliferation,Differentiation and Mineralization of MC3T3-E1 Cells through the Wnt/β-Catenin Signalling Pathway.Cell Physiol Biochem.2017;41(4):1605-1615.
[22]Ma XN,Ge BF,Chen KM,et al.Mechanisms of icariin in regulating bone formation of osteoblasts and bone resorption of osteoclasts.Zhongguo Yi Xue Ke Xue Yuan Xue Bao.2013;35(4):432-438.
[23]Chung BH,Kim JD,Kim CK,et al.Icariin stimulates angiogenesis by activating the MEK/ERK-and PI3K/Akt/eNOS-dependent signal pathways in human endothelial cells.Biochem Biophys Res Commun.2008;376(2):404-408.