Calcium phosphate cements for bone engineering and their biological properties

Calcium phosphate cements （CPCs） are frequently used to repair bone defects. Since their discovery in the 1980s, extensive research has been conducted to improve their properties, and emerging evidence supports their increased application in bone tissue engineering. Much effort has been made to enhance the biological performance of CPCs, including their biocompatibility, osteoconductivity, osteoinductivity, biodegradability, bioactivity, and interactions with cells. This review article focuses on the major recent developments in CPCs, including 3D printing, injectability, stem cell delivery, growth factor and drug delivery, and pre- vascularization of CPC scaffolds via co-culture and tri-culture techniques to enhance angiogenesis and osteogenesis.

REINFORCEMENT OF CALCIUM PHOSPHATE CEMENTS WITH PHOSPHORYLATED CHITIN

Phosphorylated chitins (P-chitins) as the additives of calcium phosphate cements (CPCs) were prepared by the phosphorylation of chitin with phosphorus pentoxide in methanesulfonic acid. Their physical properties and effects on CPCs from monocalcium phosphate monohydrate (MCPM) and calcium oxide (CaO) or dicalcium phosphate dihydrate (DCPD) and calcium hydroxide [Ca(OH)(2)] were investigated. Addition of P-chitin (M-w = 2.60 x 10(4); degree of substitution, DS = 0.68) to the liquid phase in amounts up to 3 wt.% for MCPM and CaO cements or 1.5 wt.% for DCPD and Ca(OH)(2) cements could enhance the mechanical strength considerably, while little influence on the setting time was observed. However, further addition of P-chitin will cause no setting.

Effect of Additives on the Morphology of the Hydrated Product and Physical Properties of a Calcium Phosphate Cement

The morphology of a hydrated calcium phosphate cement （CPC） doped with several normally used additives was investigated by scanning electron microscopy （SEM） and the compressive strength of the cement was determined in this study. The hydrated products of CPC without additives was rod-like hydroxyapatite （HA） grains with around 2-5 μm in length and 100 nm in width. The addition of Sr obviously decreased the crystal size of the rod-like grains. CPCs containing carbonate, collagen and gelatin showed flake-like crystal morphology. Crylic acid-containing CPC presented flocculus-like structure. And malic acid-containing CPC exhibited oriented flake-like structure. The X-ray diffraction （XRD） analysis showed that the additives used in this study did not alter the hydration products of the cement. The compressive strength tests indicated that the compressive strength of the cement with rod-like morphology HA crystals was much higher than that of the cement with flake-like morphology HA crystals, and the cement with oriented flake-like morphology HA crystals exhibited the poorest compressive strength.

Development and characterization of α-tricalcium phosphate/monocalcium aluminate composite bone cement

Calcium phosphate cements have received much attention in recent decades owing to their biocompatibility, in situ handling, and shaping abilities. However, their low initial mechanical strength is still a major limitation. On the other hand, calcium aluminate cements (CACs) set fast and have a high initial strength and good corrosion resistance in contact with body fluids, making them excellent dental restorative materials. Therefore, the chemical, mechanical and biological properties of new-TCP/CA cement after aging in simulated body fluid (SBF) were investigated. The results indicated that the composites have setting times not appropriated for immediate applications and have degradation rates higher than those of the traditional CPCs. Moreover, the compressive strength of composite was lower than 5MPa and did not increase with SBF immersion. However, the α-TCP/CA composites showed a higher bioactivity at early stages and were not only more biocompatible but also more noncytotoxic.

Effect of carbon fiber on calcium phosphate bone cement

The calcium phosphate cement (α-TCP/TTCP) was reinforced with oxidation-t reated carbon fibers. The effect of aspect ratio and content of carbon fiber on the compression strength and bending strength of the hardened body was discussed . The results show that the reinforcing effect is optimal as the aspect ratio is 375 and the additive amount is 0.3%(mass fraction). Under this condition,the c ompressive strength is increased by 55% (maximum 63.46 MPa),and the bending st rength is nearly increased by 100% (maximum 11.95 MPa),respectively. However,if the additive quantity and aspect ratio are too high,the effect of the carbon fibers is limited because it can not be dispersed uniformly in the hardened bod y. The biological evaluation indicates that the calcium phosphate cement reinfor ced by carbon fibers has good biocompatibility.

The Effect of Premixed Schedule on the Crystal Formation of Calcium Phosphate Cement-chitosan Composite with Added Tetracycline

In this study, calcium phosphate cements （CPC） were prepared by mixing cement powders of tetracalcium phosphate （TTCP） with a cement liquid of phosphate acid saline solution. Tetracycline （TTC）-CPC, chitosan-CPC and chitosan-TTC-CPC were investigated with different premixed schedule. It was demonstrate that both TTC and chitosan worked on the phase transition and crystal characteristics. TTCP mixed with phosphate acid saline solution had similar features of Fourier transform-infrared spectrometry （FT-IR） no matter it was mixed with chitosan or TTC or both. TTC premixed with cement liquid or powder had significant different features of FT-IR and 876 cm-1 seemed to be a special peak for TTC when TTC was premixed with cement liquid. This was also supported by XRD analysis, which showed that TTC premixed with cement liquid improved phase transition of TTCP to OCP. Chitosan, as organic additive, regulates the regular crystal formation and inhibits the phase transition of TTCP to OCP, except when it is mingled with cement liquid premixed with TTC in field scanning electron microscope. It was concluded that the premixed schedule influences the crystal formation and phase transition, which may be associated with its biocompatibility and bioactivities in vivo.

The In-situ Reinforcement of Calcium Phosphate Cement and Its Micro-structural Analysis

Carbon nanotubes （ CNTs ） and polyacrylic acid were employed to modify the setting process and hydration products of β-TCP/TTCP calcium phosphate cement. The micro-structure of hydration product and the fashion of how additives and hydration particles interconnected were investigated. With the modification effect of CNTs , the setting particles and CNTs got winded and interconnected and thus made the composite more compact and denser.

Effect of Substitutional Sr Ion on Mechanical Properties of Calcium Phosphate Bone Cement

A novel calcium phosphate cement was developed by adding different amount of SrO to tetracalcium phosphate during the fabrication process. The experimental results show that compressive strength of cements based on tetracalcium phosphate doped with SrO significantly increases with the increase of SrO content, approximately 60 MPa, whilst the mechanical behavior of CPC slightly decreases if 0.7wt% SrO is added. X-ray diffraction measurement confirms the setting reaction of doped cements is similar to that of pure calcium phosphate cement （CPC）. Low crystalline hydroxyapatite （HA） is found to be the main constituent of set cement. A mechanical reinforcement effect is resulted from the substitution of Sr ion to Ca2＋ in tetracalcium phosphate （TTCP）, accelerating HA crystal formation and a more cross-linked cement structure. In vitro bioactivity tests showed that CPC added with 0.5wt% SrO had a more rapid degradation compared with pure CPC.

Augmentation of Pedicle Screw Fixation with Calcium Phosphate Cement

To determine whether a biodegradable calcium phosphate cement(CPC) provides significant augmentation of pedicle screw fixation or not,an in vitro biomechanical study was carried out to evaluate the biomechanical effect of CPC in the restoration and augmentation of pedicle screw fixation.Axial pullout test and cyclic bending resistance test were employed in the experiment,and polymethylmethacrylate (PMMA) was chosen as control.The results demonstrate that the pullout strengths following CPC restoration and augmentation are 74% greater on an average than those of the control group,but less than those of PMMA restoration group and augmentation group respectively (increased by 126% versus control).In cyclic bending resistance test,the CPC augmented screws are found to withstand a greater number of cycles or greater loading with less displacement before loosening,but the augmentation effect of PMMA is greater than that of CPC.

Microstructure and Mechanical Properties of Calcium Phosphate Cement/Gelatine Composite Scaffold with Oriented Pore Structure for Bone Tissue Engineering

The macroporous calcium phosphate（CPC） cement with oriented pore structure was prepared by freeze casting. SEM observation showed that the macropores in the porous calcium phosphate cement were interconnected aligned along the ice growth direction. The porosity of the as-prepared porous CPC was measured to be 87.6% by Archimede＇s principle. XRD patterns of specimens showed that poorly crystallized hydroxyapatite was the main phase present in the hydrated porous calcium phosphate cement. To improve the mechanical properties of the CPC scaffold, the 15% gelatine solution was infiltrated into the pores under vacuum and then the samples were freeze dried to form the CPC/gelatine composite scaffolds. After reinforced with gelatine, the compressive strength of CPC/gelatine composite increased to 5.12 MPa, around fifty times greater than that of the unreinforced macroporous CPC scaffold, which was only 0.1 MPa. And the toughness of the scaffold has been greatly improved via the gelatine reinforcement with a much greater fracture strain. SEM examination of the specimens indicated good bonding between the cement and gelatine. Participating the external load by the deformable gelatine, patching the defects of the CPC pores wall, and crack deflection were supposed to be the reinforcement mechanisms. In conclusion, the calcium phosphate cement/gelatine composite with oriented rmre structure nrenared in this work might be a potential scaffold for bone tissue engineerinm

Evaluation of Calcium Phosphate Cement As a Root Canal Sealer Filling Material

Calcium phosphate cement for root end sealing was obtained by in mixing alpha-tricalcium phosphate and additives with an aqueous solation of citric. Powder and liquid were mixed at a ratio of 1.25g/mL. The biocompatibility of this material was investigated primarily by subcutaneous implantation tests. Then calcium phosphate cement was used to fill three adult dogs' root canal, both calcium hydroxide paste and hydroxyapatite paste as control. The animals were killed at 4, 12, 20 weeks postoperatively respectively. The effects of different materials on the apical closure, restoration of periapical tissues and adaptability to the dentinal surface were examined by optical and electronic microscope. The observation at 20 weeks shows that the calcium phosphate cement has the potentialities of being a root canal sealer filling material available for pulpless teeth with open-apex and destructive periapical tissue.

Preparation of a Novel Calcium Phosphate Cement Using N-methylene Phosphonic Chitosan as a Gelling Agent

A modified chitosan （ N-methylene phosphonic Chitosan, NMPC） was synthesized to improve solubility and ability to bind calcium ion. The properties of the raw material chitosan and its derivative NMPC were characterised using FTIR , ^1H- NMR . The aim of this study was to enhance the compressive CPC by reinforcing with NMPC. A formulation consisting of CPC powder , buffer solution and gelling agent was used for preparation of the CPC. CPC powder coasisted of tetracalcium phosphate（ TTCP ） and dicalcium phosphate anhydrous （ DCPA ）. NMPC which acted as the gelling ageut was dissohed into KH2PO4-Na2 HPO4 buffer solution. Each specimen in the mold was sandciched between two fritted glass sides and kept for 24 hours. Compressive strengths were determined, the setting product was identified using X-ray diffraction and scanning electron microscopy was used to investigate the hydroxyapatite particles size and porosity. The experimental results showed that the dominating influence on the compressive strengths of CPC-AMPC was the HA panicle size, its uniformity and appropriate porosity.

Preparation and Compressive Strength of Calcium Phosphate Bone Cement Containing N,O-carboxymethyl Chitosan

N, O-carboxymethyl chitosan （ CMCTS ）, a kind of biodegradable organic substance, was added to calcium phosphate bone cement （CPC） to prodnce a composite more similar in composition to human bone. The compressive strength of the new material was inereased by 10 times compared with conventional CPC.

Microstructure and Mechanical Properties of Calcium Phosphate Cement/gelatine Composite Scaffold with Oriented Pore Structure for Bone Tissue Engineering

In this study,the macroporous calcium phosphate cement with oriented pore structure was prepared by freeze casting.SEM observation showed that the macropores in the porous calcium phosphate cement were interconnected aligned along the ice growth direction.The porosity of the as-prepared porous CPC was measured to be 87.6% by Archimede's principle.XRD patterns of specimens showed that poorly crystallized hydroxyapatite was the main phase present in the hydrated porous calcium phosphate cement.To improve the mechanical properties of the CPC scaffold,the 15% gelatine solution was infiltrated into the pores under vacuum and then the samples were freeze dried to form the CPC/gelatine composite scaffolds.After reinforced with gelatine,the compressive strength of CPC/gelatin composite increased to 5.12 MPa,around 50 times greater than that of the unreinforced macroporous CPC scaffold,which was only 0.1 MPa.And the toughness of the scaffold has been greatly improved via the gelatine reinforcement with a much greater fracture strain.SEM examination of the specimens indicated good bonding between the cement and gelatine.In conclusion,the calcium phosphate cement/gelatine composite with oriented pore structure prepared in this study might be a potential scaffold for bone tissue engineering.

antagomir-30d转染BMSCs联合CPC支架材料的异位成骨作用

目的:构建antagomir-30d/种子细胞骨髓间充质干细胞(BMSCs)/磷酸钙骨水泥(CPC)复合物,并将该复合物植入裸鼠皮下,研究antagomir-30d促进体内骨形成的作用。方法:将转染有150 nmol/L antagomir-30d、NC的BMSCs以及未经转染的空白BMSCs转移到CPC支架进行孵育,后植入到BALB/c-nu裸鼠皮下,以此研究裸鼠异位成骨。术后2、4、8周分别取出植入体。2周时,取出植入体,提取RNA进行RT-PCR检测,分析成骨基因碱性磷酸酶(ALP)、骨钙素(OC)以及Runt相关转录因子2(RUNX2)mRNA表达情况。此外,4周和8周的植入体分别进行苦味酸品红组织学染色和组织形态学分析新骨形成的情况。结果:植入2周RT-PCR结果显示,各成骨基因mRNA水平的表达量中转染有antagomir-30d组最高,并显著高于另外2组(P<0.05)。组织学染色结果表明,植入后4周,antagomir-30d组有少量新骨形成,而另外2组则少有新骨形成。组织形态学分析显示,新骨面积百分比antagomir-30d组为1.28%±0.19%。植入后8周,3组均可见明显新骨形成,但antagomir-30d组新骨形成量明显大于NC组及空白对照组(P<0.05),各组新骨面积百分比分别为17.79%±1.15%,1.82%±0.53%,2.33%±0.52%。结论:antagomir-30d可诱导裸鼠体内异位成骨,antagomir-30d/BMSCs/CPC复合物有希望作为组织工程复合物用于颌骨骨缺损及其他类型骨缺损的修复。

Bone tissue engineering via nanostructured calcium phosphate biomaterials and stem cells

Tissue engineering is promising to meet the increasing need for bone regeneration. Nanostructured calcium phosphate （CAP） biomaterials/scaffolds are of special interest as they share chemical/crystallographic similarities to inorganic components of bone. Three applications of nano-CaP are discussed in this review： nanostructured calcium phosphate cement （CPC）; nano-CaP composites; and nano-CaP coatings. The interactions between stem cells and nano-CaP are highlighted, including cell attachment, orientation/ morphology, differentiation and in vivo bone regeneration. Several trends can be seen： （i） nano-CaP biomaterials support stem cell attachment/proliferation and induce osteogenic differentiation, in some cases even without osteogenic supplements; （ii） the influence of nano-CaP surface patterns on cell alignment is not prominent due to non-uniform distribution of nano-crystals; （iii） nano-CaP can achieve better bone regeneration than conventional CaP biomaterials; （iv） combining stem cells with nano-CaP accelerates bone regeneration, the effect of which can be further enhanced by growth factors; and （v） cell microencapsulation in nano-CaP scaffolds is promising for bone tissue engineering. These understandings would help researchers to further uncover the underlying mechanisms and interactions in nano-CaP stem cell constructs in vitro and in vivo, tailor nano-CaP composite construct design and stem cell type selection to enhance cell function and bone regeneration, and translate laboratory findings to clinical treatments.

Calcium Phosphate Biomaterials: An Update

Current calcium phosphate （ CaP ） biomaterials for bone repair, substitution, augmentation and regeneration include hydroxyapatite （ HA ） from synthetic or biologic origin, beta-tricaicium phosphate （ β- TCP ）, biphasic calcium phosphate （BCP）, and are available as granules, porous blocks, components of compashes （CaP/pollymer） cements, and as coatings on orthopedic and dental implants. Experimental calcium phosphate biomaterials include CO3^- and F-substituted apatites, Mg-and Zn-substituted β-TCP, calcium phosphate glasses, This paper is a brief review of the different types of CaP biomaterials and their properties such as bioactivity , osteoconductivity , osteoinductivity.

兼具显影与成骨功能多聚磷酸锶在磷酸钙骨水泥中的应用

背景:课题组前期研究发现,添加硫酸钡可以提高磷酸钙骨水泥的力学性能与显影性能,但伴随磷酸钙降解残存的显影剂难以降解,在植入部位产生的占位效应及破骨效应影响骨修复进程,因此需要开发一种可生物降解的新型显影材料。目的:探讨生物活性可降解材料多聚磷酸锶的显影能力及其对磷酸钙骨水泥理化性能与成骨效应的影响。方法:①分别制备磷酸钙骨水泥(CPC)、淀粉改性磷酸钙骨水泥(CPS)与含多聚磷酸锶(占骨水泥粉末的质量分数为20%)淀粉改性磷酸钙骨水泥(20%SrPP-CPN),表征3组骨水泥的理化性能。②将3组骨水泥浸提液分别与大鼠骨髓间充质干细胞共培养,检测细胞增殖、能量代谢与成骨分化能力。③在24只SD大鼠颅顶部两侧各制作直径5 mm的骨缺损,随机分为对照组(不进行任何干预)、CPC组、CPS组与20%SrPP-CPN组进行干预,每组6只。干预4,12周后取材进行相关检测。结果与结论:①与其他两组骨水泥比较,20%SrPP-CPN的显影能力增强、抗压强度与降解速率升高、固化时间延长,并且20%SrPP-CPN在降解过程中可稳定释放Sr^(2+)。②CCK-8实验显示20%SrPP-CPN不影响骨髓间充质干细胞的增殖;细胞饥饿实验(无血清培养)显示相较于其他两组骨水泥,20%SrPP-CPN可促进骨髓间充质干细胞的增殖;相较于其他两组骨水泥,20%SrPP-CPN可提升骨髓间充质干细胞内ATP浓度。碱性磷酸酶与茜素红染色显示,相较于其他两组骨水泥,20%SrPP-CPN可促进骨髓间充质干细胞的成骨分化。③在大鼠颅骨缺损实验中,Micro-CT扫描、组织学(苏木精-伊红、Masson染色)观察显示相较于CPC组、CPS组,20%SrPP-CPN组骨水泥降解明显,并且大量新生骨组织分散于降解的骨水泥中;免疫组化染色显示相较于CPC组、CPS组,20%SrPP-CPN组缺损处Runx2蛋白表达升高(P<0.01)。④结果表明,20%SrPP-CPN具有良好的显影性能与促成骨性能。

CPC/PMMA双向骨水泥性能及生物相容性的初步研究

目的将聚甲基丙烯酸甲酯骨水泥(PMMA)与磷酸钙骨水泥(CPC)制成一种注射型混合骨水泥,利用各自的特性,优势互补,从而使该混合骨水泥成为理想的椎体成形术及椎体后凸成形术填充材料。方法实验材料为注射型PMMA骨水泥和注射型CPC骨水泥。首先将PMMA与CPC按质量比1∶2、1∶1、2∶1均匀混合,制成不同比例的混合骨水泥,然后将PMMA、CPC、混合骨水泥(1∶2、1∶1、2∶1)制成标准试件(直径6mm、高12mm),最后对试件进行力学性能、凝固温度、凝固时间、电镜扫描、X线衍射、动物肌肉内植入实验,初步评估混合骨水泥的性能及生物相容性。结果 PMMA与CPC在1∶1比例混合时理化性能满意,其抗压强度较CPC提高约120%,凝固过程中的最高温度低于45℃,凝固时间约12～14 min,微观结构与CPC相似,孔径在100～300μm之间分布最多(约70%),大部分孔与孔之间有10～20μm小孔贯通。动物肌肉内植入实验显示混合骨水泥(1∶1)的组织相容性满意,但降解速度缓慢。结论 PMMA与CPC在1∶1比例混合能有效克服单一材料缺点,优势互补,从而使该混合骨水泥有望成为理想的椎体成形术及椎体后凸成形术填充材料。