Development and physicochemical characterization of a porous calcium phosphate/bone matrix gelatin composite cement

doi:10.3969/j.issn.2095-4344.0640

Abstract

Abstract:

BACKGROUND: Calcium phosphate cement has good biocompatibility, osteoinductive activity and biodegradability, which meets some of the needs for repairing bone defects. However, single calcium phosphate cement is brittle and has poor biological activity without osteoinductive activity.

OBJECTIVE: To develop porous calcium phosphate/bone matrix gelatin composite cements and to explore the physicochemical characterization.

METHODS: Bone matrix gelatin was extracted from rabbits and poly(lactic-co-glycolic) acid (PLGA) microspheres were prepared. The new porous cement was developed using calcium phosphate cement composited with bone matrix gelatin and PLGA microspheres. The composited cement was then divided into three groups including blank group, 5% microsphere group and 10% microsphere group. Then, the physicochemical characterizations of the new porous composite bone cement in different groups were assessed by microstructure, setting time, injectability, resistance to collapsibility, in vitro degradation and porosity. Bone marrow mesenchymal stem cells were co-cultured with the extracts of these three kinds of composite cements, and cell proliferation rate was detected within 14 days of co-culture. The three kinds of composite cements were implanted into the lumbar vertebral bone defects in the rabbit model, and bone defect repair was analyzed by Micro-CT at 12 weeks after implantation.

RESULTS AND CONCLUSION: With the increasing ratio of the microsphere, the setting time was shortened (P < 0.05), the porosity was increased (P < 0.05), the compressive strength and elastic modulus were reduced (P < 0.05), the injectability and anti-diffusivity were weakened, and the in vitro degradation was fastened. Compared with the blank group, the cell proliferation was significantly increased in the 5% and 10% microsphere groups (P < 0.05). At 12 weeks after implantation, Micro-CT findings revealed new bone tissues formed at the bone defect site. Compared with the blank group, significantly increased bone mineral density, bone volume fraction, trabecular thickness, and trabecular number were observed in the 5% and 10% microsphere groups (P < 0.05), but the trabecular spacing was significantly lower than that in the blank group (P < 0.05). To conclude, the calcium phosphate/bone matrix gelatin composite cement with 5% PLGA microsphere has proper maneuverability, mechanical strength and cytocompatibility, degradability and osteoinductive activity.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

Key words: Calcium Phosphates, Porosity, Bone Matrix, Tissue Engineering

CLC Number:

R318

Wang Song, Yang Han, Yang Jian, Zhang Kaiquan, Kang Jianping, Wang Qing. Development and physicochemical characterization of a porous calcium phosphate/bone matrix gelatin composite cement[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(34): 5419-5425.

Figures/Tables 8

References

[1] O'Neill R,McCarthy HO,Montufar EB,et al.Critical review: Injectability of calcium phosphate pastes and cements.Acta Biomater.2016;9(16):30606-30607.

[2] Liao H,Félix Lanao RP,van den Beucken JJ,et al.Size matters: effects of PLGA-microsphere size in injectable CPC/PLGA on bone formation.J Tissue Eng Regen Med. 2016;10(8): 669-678.

[3] Urist MR,Lange RJ,Finerman GABone cell differentiation and growth factorsScience.1983; 220(4598):680-686.

[4] Vrancken NM,Claeys AD.Method for encapsulating water and compoundsin aqueous phase by extraction: US, 3526906[P].1970-08-11．

[5] 王松,杨函,杨剑,等.多孔磷酸钙/骨基质明胶复合骨水泥修复兔椎体骨缺损的研究[J].中国修复重建外科杂志, 2017,31(12): 1462-1467.

[6] 丁裕明.新型磷酸钙复合骨水泥的制备及性能研究[D].泸州:西南医科大学,2016:1-9.

[7] ISO.Cement-Test methods-Determination of setting time and soundness ISO 9597,2008.

[8] ASTM.Standard test method for time of setting of hydraulic-cement paste by Gillmore needles. ASTM C266–08, 2008.

[9] Habraken WJ,Wolke JG,Mikos AG,et al.PLGA microsphere/ calcium phosphate cement composites for tissue engineering: in vitro release and degradation characteristics.J Biomater Sci Polym Ed.2008;19(9):1171-1188.

[10] 马思遥,杨帅兵,岳晓华.Wistar大鼠骨髓间充质干细胞的培养与鉴定[J].临床医药文献杂志,2018,85(3):24-26.

[11] Khairoun I,Boltong MG,Driessens FC,et al.Effect of calcium carbonate on clinical compliance of apatitic calcium phosphate bone cement.J Biomed Mater Res. 1997;38(4): 356-360.

[12] Medri V,Mazzocchi M,Bellosi A.Doped calcium-aluminium- phosphate cements for biomedical applications.J Mater Sci Mater Med.2011;22(2):229-236.

[13] Shi M,Chen L,Wang Y,et al.Low-intensity pulsed ultrasound enhances antibiotic release of gentamicin-loaded, self-setting calcium phosphate cement.J Int Med Res. 2018;1: 300060518773023.

[14] Bohner M.Design of ceramic-based cements and putties for bone graft substitution.Eur Cell Mater.2010;20:1-12.

[15] Jang JH,Shin S,Kim HJ,et al.Improvement of physical properties of calcium phosphate cement by elastin-like polypeptide supplementation.Sci Rep.2018;8(1):5216.

[16] Sarkar SK,Lee BY,Padalhin AR,et al.Brushite-based calcium phosphate cement with multichannel hydroxyapatite granule loading for improved bone regeneration.J Biomater Appl.2016; 30(6):823-837.

[17] 张岩,郑欣,朱彦丞,等.微球在骨组织工程中的应用现状[J].中国矫形外科杂志,2016,24(16):1478-1481.

[18] Gunnella F,Kunisch E,Maenz S,et al.The GDF5 mutant BB-1 enhances the bone formation induced by an injectable, poly(l-lactide-co-glycolide) acid (PLGA) fiber-reinforced, brushite-forming cement in a sheep defect model of lumbar osteopenia.Spine J.2018;18(2):357-369.

[19] Gunnella F,Kunisch E,Bungartz M,et al.Low-dose BMP-2 is sufficient to enhance the bone formation induced by an injectable, PLGA fiber-reinforced, brushite-forming cement in a sheep defect model of lumbar osteopenia.Spine J. 2017; 17(11):1699-1711.

[20] Liu W,Zhang J,Weiss P,et al.The influence of different cellulose ethers on both the handling and mechanical properties of calcium phosphate cements for bone substitution. Acta Biomater.2013;3:5740-5750.

[21] Filipowska J,Pawlik J,Cholewa-Kowalska K,et al. Incorporation of sol-gel bioactive glass into PLGA improves mechanical properties and bioactivity of composite scaffolds and results in their osteoinductive properties.Biomed Mater. 2014;9(6):065001.

[22] Santos PS,Cestari TM,Paulin JB,et al.Osteoinductive porous biphasic calcium phosphate ceramic as an alternative to autogenous bone grafting in the treatment of mandibular bone critical-size defects.J Biomed Mater Res B Appl Biomater. 2018;106(4):1546-1557.

[23] 赵轶男,刘建,刘昌盛,等.不同孔径多孔CPC 材料修复兔大段骨缺损的实验研究[J].中国矫形外科杂志,2012,20(10):924-927.

[24] 江捍平,王大平,阮建明,等.纳米羟基磷灰石人工骨的生物相容性研究[J].中国现代医学杂志,2005, 15(10):1477-1480.

[25] 张红烨,张瑞生,夏永祥,等.磷酸钙骨水泥的生物相容性[J].中国组织工程研究与临床康复,2008,12(19):3713-3716.

[26] Song Y,Zhang C,Wang P,et al.Engineering bone regeneration with novel cell-laden hydrogel microfiber-injectable calcium phosphate scaffold.Mater Sci Eng C Mater Biol Appl. 2017;75: 895-905.

[27] Liu T,Li J,Shao Z,et al.Encapsulation of mesenchymal stem cells in chitosan/β-glycerophosphate hydrogel for seeding on a novel calcium phosphate cement scaffold.Med Eng Phys. 2018;56:9-15.

[28] Lamghari M,Berland S,Laurent A,et al.Bone reactions to nacre injected percutaneously into the vertebrae of sheep. Biomaterials.2001;22(6):555-562.

[29] Lamghari M,Antonietti P,Berland S,et al.Arthrodesis of lumbar spine transverse processes using nacre in rabbit.J Bone Miner Res.2001;16(6):2232-2237.

[30] Parker RM,Malham GM.Comparison of a calcium phosphate bone substitute with recombinant human bone morphogenetic protein-2: a prospective study of fusion rates, clinical outcomes and complications with 24-month follow-up.Eur Spine J.2017;26(3):754-763.

[31] Zheng YX,Wang J,Lin HT,et al.Reconstruction of orbital defect in rabbits with composite of calcium phosphate cement and recombinant human bone morphogenetic protein-2.Chin Med J (Engl). 2010;123(24):3658-3662.

[32] Perrier M,Lu Y,Nemke B,et al.Acceleration of second and fourth metatarsal fracture healing with recombinant human bone morphogenetic protein-2/calcium phosphate cement in horses.Vet Surg.2008;37(7):648-655.

[33] Palmer I,Nelson J,Schatton W,et al.Biocompatibility of calcium phosphate bone cement with optimised mechanical properties: an in vivo study.J Mater Sci Mater Med. 2016; 27(12):191.

[34] Lovasik BP,Holland CM,Howard BM,et al.Anterior Cervical Discectomy and Fusion: Comparison of Fusion, Dysphagia, and Complication Rates Between Recombinant Human Bone Morphogenetic Protein-2 and Beta-Tricalcium Phosphate. World Neurosurg.2017;97:674-683.

[35] Villavicencio AT,Burneikiene S.RhBMP-2-induced radiculitis in patients undergoing transforaminal lumbar interbody fusion: relationship to dose.Spine J.2016; 16(10):1208-1213.

[36] Toth JM,Wang M,Lawson J,et al.Radiographic, biomechanical, and histological evaluation of rhBMP-2 in a 3-level intertransverse process spine fusion: an ovine study.J Neurosurg Spine. 2016;25(6):733-739.