Development of arginine-glycine-aspartate-immobilized 3D printed poly(propylene fumarate) scaffolds for cartilage tissue engineering

Abstract

Poly(propylene fumarate) (PPF) has known to be a good candidate material for cartilage tissue regeneration because of its excellent mechanical properties during its degradation processes. Here, we describe the potential application of PPF-based materials as 3D printing bioinks to create macroporous cell scaffolds using micro-stereolithography. To improve cell-matrix interaction of seeded human chondrocytes within the PPF-based 3D scaffolds, we immobilized arginine-glycine-aspartate (RGD) peptide onto the PPF scaffolds. We also evaluated various cellular behaviors of the seeded chondrocytes using MTS assay, microscopic and histological analyses. The results indicated that PPF-based biocompatible scaffolds with immobilized RGD peptide could effectively support initial adhesion and proliferation of human chondrocytes. Such a 3D bio-printable scaffold can offer an opportunity to promote cartilage tissue regeneration.     

Fabrication and characterization of poly(propylene fumarate) scaffolds with controlled pore structures using 3-dimensional printing and injection molding

Poly(propylene fumarate) (PPF) is an injectable, biodegradable polymer that has been used for fabricating preformed scaffolds in tissue engineering applications because of in situ crosslinking characteristics. Aiming for understanding the effects of pore structure parameters on bone tissue ingrowth, 3-dimensional (3D) PPF scaffolds with controlled pore architecture have been produced in this study from computer-aided design (CAD) models. We have created original scaffold models with 3 pore sizes (300, 600, and 900 microm) and randomly closed 0%, 10%, 20%, or 30% of total pores from the original models in 3 planes. PPF scaffolds were fabricated by a series steps involving 3D printing of support/build constructs, dissolving build materials, injecting PPF, and dissolving support materials. To investigate the effects of controlled pore size and interconnectivity on scaffolds, we compared the porosities between the models and PPF scaffolds fabricated thereby, examined pore morphologies in surface and cross-section using scanning electron microscopy, and measured permeability using the falling head conductivity test. The thermal properties of the resulting scaffolds as well as uncrosslinked PPF were determined by differential scanning calorimetry and thermogravimetric analysis. Average pore sizes and pore shapes of PPF scaffolds with 600- and 900-microm pores were similar to those of CAD models, but they depended on directions in those with 300-microm pores. Porosity and permeability of PPF scaffolds decreased as the number of closed pores in original models increased, particularly when the pore size was 300 microm as the result of low porosity and pore occlusion. These results show that 3D printing and injection molding technique can be applied to crosslinkable polymers to fabricate 3D porous scaffolds with controlled pore structures, porosity, and permeability using their CAD models.     

3D打印骨组织工程支架的制备技术

近年来,随着3D打印技术的飞速发展,人们开始通过3D打印技术去不断完善适合不同需求的定制骨组织工程支架。由于组织工程制造的支架是需要植入生物体内的,这就对支架有着极为严苛的要求。3D打印技术作为一种新兴制备骨组织工程支架的技术,其最大的优点是可以依照需求来定制个性化形状、结构,良好的宏微观结构、润湿性、机械强度和细胞反应的新型骨组织工程支架。本文回顾了2014―2019年间对骨组织工程支架的研究,对3D打印骨组织工程支架进行了总结,并且介绍了在多功能骨组织工程支架设计与制作中的理念与研究。     

Synthesis and properties of photocross-linked poly(propylene fumarate) scaffolds

The photocross-linking of poly(propylene fumarate) (PPF) to form porous scaffolds for bone tissue engineering applications was investigated. PPF was cross-linked using the photoinitiator bis(2,4,6-trimethylbenzoyl) phenylphosphine oxide (BAPO) and exposure to 30 min of long wavelength ultraviolet (UV) light. The porous photocross-linked PPF scaffolds (6.5 mm diameter cylinders) were synthesized by including a NaCl porogen (70, 80, and 90 wt% at cross-linking) prior to photocross-linking. After UV exposure, the samples were placed in water to remove the soluble porogen, revealing the porous PPF scaffold. As porogen leaching has not been used often with cross-linked polymers, and even more rarely with photoinitiated cross-linking, a study of the efficacy of this strategy and the properties of the resulting material was required. Results show that the inclusion of a porogen does not significantly alter the photoinitiation process and the resulting scaffolds are homogeneously cross-linked throughout their diameter. It was also shown that porosity can be generally controlled by porogen content and that scaffolds synthesized with at least 80 wt% porogen possess an interconnected pore structure. Compressive mechanical testing showed scaffold strength to decrease with increasing porogen content. The strongest scaffolds with interconnected pores had an elastic modulus of 2.3 ± 0.5 MPa and compressive strength at 1% yield of 0.11 ± 0.02 MPa. This work has shown that a photocross-linking/porogen leaching technique is a viable method to form porous scaffolds from photoinitiated materials.     

Synthesis and properties of photocross-linked poly(propylene fumarate) scaffolds

The photocross-linking of poly(propylene fumarate) (PPF) to form porous scaffolds for bone tissue engineering applications was investigated. PPF was cross-linked using the photoinitiator bis(2,4,6-trimethylbenzoyl) phenylphosphine oxide (BAPO) and exposure to 30 min of long wavelength ultraviolet (UV) light. The porous photocross-linked PPF scaffolds (6.5 mm diameter cylinders) were synthesized by including a NaCl porogen (70, 80, and 90 wt% at cross-linking) prior to photocross-linking. After UV exposure, the samples were placed in water to remove the soluble porogen, revealing the porous PPF scaffold. As porogen leaching has not been used often with cross-linked polymers, and even more rarely with photoinitiated cross-linking, a study of the efficacy of this strategy and the properties of the resulting material was required. Results show that the inclusion of a porogen does not significantly alter the photoinitiation process and the resulting scaffolds are homogeneously cross-linked throughout their diameter. It was also shown that porosity can be generally controlled by porogen content and that scaffolds synthesized with at least 80 wt% porogen possess an interconnected pore structure. Compressive mechanical testing showed scaffold strength to decrease with increasing porogen content. The strongest scaffolds with interconnected pores had an elastic modulus of 2.3+/-0.5 MPa and compressive strength at 1% yield of 0.11+/-0.02 MPa. This work has shown that a photocross-linking/porogen leaching technique is a viable method to form porous scaffolds from photoinitiated materials.     

骨髓间充质干细胞在淫羊藿苷/羟基磷灰石/聚乳酸-羟基乙酸共聚物支架上的成骨

文题释义：纳米结构：是尺寸介于分子和微米尺度间的物体结构。当纳米羟基磷灰石与高分子材料物理混合后,羟基磷灰石会发生自聚,从而在材料表面产生纳米结构。这种纳米结构有利于细胞(如骨髓充间质干细胞)的黏附,是骨修复材料表面细胞增殖和后期成骨分化的基础。成骨分化：当干细胞接受诱导时可以向成骨细胞转变。淫羊藿苷高分子复合支架与间充质干细胞共培养一段时间后,其骨分化标志物碱性磷酸酶和骨钙素的活性增高,同时成骨相关基因和蛋白(Runx-2、COLⅠ)表达水平上升,即细胞在淫羊藿苷诱导下发生了成骨分化。  摘要背景：近年来,骨组织工程技术为临床治疗骨缺损提供了全新的思路和模式。该研究首次将传统中药与组织工程支架的纳米结构结合,以期探索并构建一种可用于骨缺损治疗的新型骨组织替代材料。目的：研究淫羊藿苷(icariin,ICA)/羟基磷灰石(hydroxyapatite,HA)/聚乳酸-羟基乙酸共聚物(poly(lactic-co-glycolic acid),PLGA)复合支架的成骨活性。方法：将HA与PLGA通过物理共混的方式制成HA/PLGA复合支架,然后将其浸泡于不同浓度的ICA溶液中,从而得到ICA/HA/PLGA支架。利用兔骨髓间充质干细胞分别对复合支架的细胞黏附、增殖、成骨作用和细胞毒性进行评价。细胞黏附、细胞增殖和细胞毒性采用MTT法进行检测,碱性磷酸酶活性和骨钙素活性采用ELISA法进行检测,成骨相关基因和蛋白表达水平分别用荧光定量PCR和Western blot法进行检测。结果与结论：①PLGA中加入适量HA可以提高支架的力学强度,且在HA含量为10%时效果最佳,拉伸强度为(1.67±0.37) MPa;压缩模量为(4.17±1.62) MPa,且会在支架表面形成纳米结构;该微结构可以促进骨髓间充质干细胞在支架表面的黏附;②ICA不会影响骨髓间充质干细胞在复合支架上的增殖,且1.00 µmol/L ICA水溶液浸泡后的ICA/HA/PLGA复合支架具有最优的成骨分化功能,其碱性磷酸酶活性、骨钙素活性、成骨相关基因和蛋白(Runx-2和COLⅠ)的表达水平均最高;③ICA/HA/PLGA复合支架无细胞毒性;④结果表明,HA(10%)/ICA(1.00 µmol/L)/PLGA支架具有良好的机械性能、成骨作用和生物相容性,是一种具有良好应用潜力的骨组织工程支架。ORCID: 0000-0002-9770-9109(王德欣) 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

In vivo degradation of porous poly(propylene fumarate)/poly(DL-lactic-co-glycolic acid) composite scaffolds

This study investigated the in vivo degradation of poly(propylene fumarate) (PPF)/poly(DL-lactic-co-glycolic acid) (PLGA) composite scaffolds designed for controlled release of osteogenic factors. PPF/PLGA composites were implanted into 15.0mm segmental defects in the rabbit radius, harvested after 12 and 18 weeks, and analyzed using histological techniques to assess the extent of polymer degradation as well as the tissue response within the pores of the scaffolds. Polymer degradation was limited to micro-fragmentation of the scaffold at the ends and edges of the implant at both 12 and 18 weeks. The tissue within the pores of the scaffold consisted of fibrous tissue, blood vessels and some inflammatory cells. In areas where polymer breakdown was evident, an increased inflammatory response was observed. In contrast, areas of bone ingrowth into the polymer scaffold were characterized by minimal inflammatory response and polymer degradation. Our results show that minimal degradation of porous PPF occurs within 18 weeks of implantation in a rabbit model. Further, the in vivo degradation data of porous PPF/PLGA scaffolds are comparable with earlier obtained in vitro data.     

Preparation,in vitro degradability,cytotoxicity, and in vivo biocompatibility of porous hydroxyapatite whisker-reinforced poly(L-lactide) biocomposite scaffolds

Biodegradable and bioactive scaffolds with interconnected macroporous structures, suitable biodegradability, adequate mechanical property, and excellent biocompatibility have drawn increasing attention in bone tissue engineering. Hence, in this work, porous hydroxyapatite whisker-reinforced poly(L-lactide) (HA-w/PLLA) composite scaffolds with different ratios of HA and PLLA were successfully developed through compression molding and particle leaching. The microstructure, in vitro mineralization, cytocompatibility, hemocompatibility, and in vivo biocompatibility of the porous HA-w/PLLA were investigated for the first time. The SEM results revealed that these HA-w/PLLA scaffolds possessed interconnected pore structures. Compared with porous HA powder-reinforced PLLA (HA-p/PLLA) scaffolds, HA-w/PLLA scaffolds exhibited better mechanical property and in vitro bioactivity, as more formation of bone-like apatite layers were induced on these scaffolds after mineralization in SBF. Importantly, in vitro cytotoxicity displayed that porous HA-w/PLLA scaffold with HA/PLLA ratio of 1:1 (HA-w₁/PLLA₁) produced no deleterious effect on human mesenchymal stem cells (hMSCs), and cells performed elevated cell proliferation, indicating a good cytocompatibility. Simultaneously, well-behaved hemocompatibility and favorable in vivo biocompatibility determined from acute toxicity test and histological evaluation were also found in the porous HA-w₁/PLLA₁ scaffold. These findings may provide new prospects for utilizing the porous HA whisker-based biodegradable scaffolds in bone repair, replacement, and augmentation applications.     

In vitro degradation of porous poly(propylene fumarate)/poly(DL-lactic-co-glycolic acid) composite scaffolds

This study investigated the in vitro degradation of porous poly(propylene fumarate) (PPF-based) composites incorporating microparticles of blends of poly(DL-lactic-co-glycolic acid) (PLGA) and poly(ethylene glycol) (PEG) during a 26-week period in pH 7.4 phosphate-buffered saline at 37 degrees C. Using a fractional factorial design, four formulations of composite scaffolds were fabricated with varying PEG content of the microparticles, microparticle mass fraction of the composite material, and initial leachable porogen content of the scaffold formulations. PPF scaffolds without microparticles were fabricated with varying leachable porogen content for use as controls. The effects of including PLGA/PEG microparticles in PPF scaffolds and the influence of alterations in the composite formulation on scaffold mass, geometry, water absorption, mechanical properties and porosity were examined for cylindrical specimens with lengths of 13 mm and diameters of 6.5 mm. The composite scaffold composition affected the extent of loss of polymer mass, scaffold length, and diameter, with the greatest loss of polymer mass equal to 15+/-5% over 26 weeks. No formulation, however, exhibited any variation in compressive modulus or peak compressive strength over time. Additionally, sample porosity, as determined by both mercury porosimetry and micro-computed tomography did not change during the period of this study. These results demonstrate that microparticle carriers can be incorporated into PPF scaffolds for localized delivery of bioactive molecules without altering scaffold mechanical or structural properties up to 26 weeks in vitro.     

Characterization of partially saturated poly(propylene fumarate) for orthopaedic application

A partially saturated linear polyester based on poly(propylene fumarate) (PPF) was synthesized for potential application in filling skeletal defects. The synthesis was carried out according to a two-step reaction scheme. Propylene glycol and fumaryl chloride were first combined to form an intermediate fumaric diester. The intermediate was then subjected to a transesterification to form the PPF-based polymer. This method allowed for production of a polymer with a number average molecular weight up to 1500 and a polydispersity index of 2.8 and below. The polymeric backbone structure was investigated through the use of FTIR and NMR. Kinetic studies of the transesterification allowed mapping of the molecular weight increase with reaction time. The final product was also characterized by thermal and solubility analysis.     

In vitro degradation of polymeric networks of poly(propylene fumarate) and the crosslinking macromer poly(propylene fumarate)-diacrylate

Polymeric networks of poly(propylene fumarate) (PPF) crosslinked with poly(propylene fumarate)-diacrylate (PPF-DA) are currently being investigated as an injectable, biodegradable bone cement. This study examined the effect of crosslinking density, medium pH, and the incorporation of a beta-tricalcium phosphate (beta-TCP) filler on the in vitro degradation of PPF/PPF-DA. Cylindrical specimens were submerged in buffered saline at 37 degrees C and the change in weight, geometry, and compressive mechanical properties were monitored over a 52-week period. All formulations showed an initial increase in modulus and yield strength over the first 12 weeks, achieving maxima of 1307+/-101 and 51+/-3MPa, respectively, for the beta-TCP composite. PPF/PPF-DA networks with the lower crosslinking density demonstrated the greatest degradation with a 17% mass loss. Samples in the lower buffer pH 5.0 compared to physiological pH 7.4 did not show any differences in mass loss, but exhibited a faster decrease in the compressive strength over time. The beta-TCP composites maintained their mechanical properties at the level following their initial increase. These results show that the degradation of PPF/PPF-DA networks can be controlled by the crosslinking density, accelerated at a lower pH, and prolonged with the incorporation of the beta-TCP filler.     

Bone formation in transforming growth factor beta-1-coated porous poly(propylene fumarate) scaffolds

This study determined the bone growth into pretreated poly(propylene fumarate) (PPF) scaffolds implanted into a subcritical size, rabbit cranial defect. PPF scaffolds were constructed by using a photocrosslinking-porogen leaching technique. These scaffolds were then either prewetted (PPF-Pw), treated with RF glow-discharge (PPF-Gd), coated with fibronectin (PPF-Fn), or coated with rhTGF-beta1 (PPF-TGF-beta1). One of each scaffold type was then placed into the cranium of nine rabbits. The rabbits were sacrificed after 8 weeks, and the scaffolds were retrieved for histological analysis. The most bone formation was present in the PPF-TGF-beta1 implants; the newly formed bone had a trabecular appearance together with bone marrow-like tissue. Little or no bone formation was observed in implants without rhTGF-beta1. These histological findings were confirmed by image analysis. Bone surface area, bone area percentage, pore fill percentage, and pore area percentage were significantly higher in the rhTGF-beta1-coated implants than in the noncoated implants. No statistical difference was seen between the PPF-Fn, PPF-Pw, or PPF-Gd scaffolds for these parameters. Quadruple fluorochrome labeling showed that in PPF-TGF-beta1 implants bone formation mainly started in the interior of a pore and proceeded toward the scaffold. We conclude that (a) PPF-TGF-beta1 scaffolds can indeed adequately induce bone formation in porous PPF, and (b) PPF scaffolds prepared by the photocrosslinking-porogen leaching technique are good candidates for the creation of bone graft substitutes.     

In-situ preparation of poly(propylene fumarate)--hydroxyapatite composite

In-situ precipitation of hydroxyapatite (HAp) in the presence of poly(propylene fumarate) (PPF) is investigated. Amorphous calcium phosphate (ACP) precipitates in the presence of the polymer and remains in the amorphous form for a relatively long time, e.g. even after 24 h of coexistence with the mother solution. Our observations suggest that PPF interacts with the surface of the ACP particles and prevents them from transformation to crystalline hydroxyapatite. The PPF polymer seems to be more efficient in hindering the ACP to HAp transformation at higher pH conditions. From spectroscopic observations we hypothesize that the C=O bond of the PPF molecules interact with the calcium ion of the ACP particles. In case of low molecular weight PPF this interaction may lead to the incorporation of the polymer within the growing ACP particles.     

Soft and hard tissue response to photocrosslinked poly(propylene fumarate) scaffolds in a rabbit model.

The treatment of large cranial defects may be greatly improved by the development of precisely formed bone tissue engineering scaffolds. Such scaffolds could be constructed by using UV laser stereolithography to photocrosslink a linear, biodegradable polymer into a three-dimensional implant. We have previously presented a method to photocrosslink the biodegradable polyester, poly(propylene fumarate) (PPF). To ensure the safety and effectiveness of this technique, the soft and hard tissue response to photocrosslinked PPF scaffolds of different pore morphologies was investigated. Four classes of photocrosslinked PPF scaffolds, constructed with differing porosities (57-75%) and pore sizes (300-500 or 600-800 microm), were implanted both subcutaneously and in 6.3-mm-diameter cranial defects in a rabbit model. The rabbits were sacrificed at 2 and 8 weeks, and the implants were analyzed by light microscopy, histological scoring analysis, and histomorphometric analysis. Results showed the PPF scaffolds elicit a mild tissue response in both soft and hard tissues. Inflammatory cells, vascularization, and connective tissue were observed at 2 weeks; a decrease in inflammatory cell density and a more organized connective tissue were observed at 8 weeks. Scaffold porosity and scaffold pore size were not found to significantly affect the observed tissue response. Evidence of scaffold surface degradation was noted both by histology and histomorphometric analysis. Bone ingrowth in PPF scaffolds implanted into cranial defects was <3% of the defect area. The results indicate that photocrosslinked PPF scaffolds are biocompatible in both soft and hard tissues and thus may be an attractive platform for bone tissue engineering.     

聚丙烯延胡索酸酯的合成、交联及生物力学、体外降解检测

目的 合成新型可注射性生物降解材料聚丙烯延胡索酸酯[poly(propylene fumarate), PPF],检测其交联温度、交联时间、生物力学性能和体外降解过程。方法 合成PPF,将PPF和N-乙烯基吡咯烷酮(N-vinyl pyrrolidinone,N-VP)在过氧化苯甲酰(benzoyl peroxide,BP)催化下交联,并且加入β-磷酸三钙(β-TCP)和氯化钠(NaCl),检测交联温度、时间、生物力学性能,和PMMA骨水泥进行比较。将PPF交联后浸泡于PBS。检测体外降解时质量和力学强度的变化。结果 交联温度在41.2±2.2℃和47.5±1.7℃间,交联时间从8.1±0.8min到63.7±4.4min,PPF的压应力为2.6±0.5MPa到12.0±2.3Mpa,压缩模量为26.6±8.7MPa到252.8±57.6Mpa,体外降解后4周PPF压应力为8.4±1.6Mpa。结论 PPF有合适的交联温度、交联时间和生物力学强度,体外降解过程中力学强度可以维持4周以上,是一个有应用前景的新型可注射生物高分子聚合材料。     

Photocrosslinking characteristics and mechanical properties of diethyl fumarate/poly(propylene fumarate) biomaterials

The development of tissue engineered materials for the treatment of large bone defects would provide attractive alternatives to the autografts, allografts, non-degradable polymers, ceramics, and metals that are currently used in clinical settings. To this end, poly(propylene fumarate) (PPF), a viscous polyester synthesized from diethyl fumarate (DEF), has been studied for use as an engineered bone graft. We have investigated the photocrosslinking of PPF dissolved in its precursor, DEF, using the photoinitiator bis(2,4,6-trimethylbenzoyl) phenylphosphine oxide (BAPO) and low levels of ultraviolet light exposure. A three factor, 2 x 2 x 4 factorial design was developed, studying the effects of PPF number average molecular weight, BAPO initiator content, and DEF content upon photocrosslinking characteristics and mechanical properties. Uncured DEF/PPF solution viscosity fell over three orders of magnitude as DEF content was increased from 0% to 75%. The exothermic photocrosslinking reaction released low levels of heat, with no more than 160J/g released from any formulation tested. As a result, the maximum photocrosslinking temperature remained below 47 degrees C for all samples. Both sol fraction and swelling degree generally increased with increasing DEF content. Compressive mechanical properties were within the range of trabecular bone, with the strongest samples possessing an elastic modulus of 195.3 +/- 17.5 MPa and a fracture strength of 68.8 +/- 9.4MPa. Finally, the results indicate that PPF crosslinking was facilitated at low DEF precursor concentrations, but hindered at higher precursor concentrations. These novel DEF/PPF solutions may be preferred over pure PPF as the basis for an engineered bone graft because they (1) exhibit reduced viscosity and thus are easily handled, (2) form polymer networks with compressive strength at fracture suitable for consideration for trabecular bone replacement, and (3) may be readily fabricated into solids with a wide range of structures.     

Tissue-engineered cartilage constructs using composite hyaluronic acid/collagen I hydrogels and designed poly(propylene fumarate) scaffolds

Our approach to cartilage tissue-engineering scaffolds combines image-based design and solid free-form (SFF) fabrication to create load-bearing constructs with user-defined parameters. In this study, 3-dimensional scaffolds with cubic and ellipsoidal pore architecture were fabricated using poly(propylene fumarate) (PPF). To increase seeding efficiency and cellular retention, hydrogels were used to deliver cells into the scaffolds. The first objective of this study was to evaluate the concentrations of composite hyaluronic acid (HyA) and collagen I hydrogels best able to stimulate proteoglycan synthesis in porcine chondrocytes in vitro and in vivo. The second objective was to evaluate the differences in extracellular matrix production due to pore geometry and scaffold design. For the in vitro assessment, chondrocytes were encapsulated in collagen I hydrogels with varying concentrations of HyA. Hydrogels were cultured for 1 and 2 weeks, and then the sulfated glycosaminoglycan (sGAG) content was quantified using a dimethyl-methylene blue assay. The concentration of HyA best able to increase ECM synthesis was 5% HyA/collagen I, or 0.23 mg/mL HyA. The results from the in vitro experiment were used as culture parameters for the in vivo analysis. Composite 5% HyA/collagen I or collagen I-only hydrogels were used to seed chondrocytes into SFF-fabricated scaffolds made of PPF with designed cubic or ellipsoidal pore geometry. The scaffolds were implanted subcutaneously in immunocompromised mice for 4 weeks. Histomorphometric analyses of sections stained with Safranin O were used to quantify the amount of ECM deposited by cells in the scaffolds. Scaffolds seeded with 5% HyA/collagen hydrogels had significantly greater areas of positive Safranin O staining (approximately 60%, compared with 30% for scaffolds with collagen I hydrogels only), indicating that greater numbers of chondrocytes retained their metabolic activity in the ectopic environment. These scaffolds also had greater stain intensities (corresponding to greater amounts of sGAG in the ECM) than their counterparts seeded with collagen I hydrogels alone. Significant differences in matrix production were not found between the scaffold pore designs. Overall, these results indicate that a combination of composite HyA hydrogels and designed SFF scaffolds could provide a functional tissue-engineered construct for cartilage repair with enhanced tissue regeneration in a load-bearing scaffold.     

Preparation and evaluation of porous poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) hydroxyapatite composite scaffolds

Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) and PHBHHx-hydroxyapatite (HAP) composite scaffolds have been prepared by phase separation and subsequent sublimation of the solvent for bone tissue engineering. Scanning electron microscopy (SEM), porosity measurement, mechanical tests, and thermogravimertric analysis (TGA) are used to analyze the physical properties of the scaffolds. The biocompatibility and osteoconductivity are assessed by examining the morphology, proliferation, and differentiation of MC3T3-E1 osteoprogenitor cells seeded on the scaffolds. The PHBHHx-HAP composite scaffolds show better mechanical properties, biocompatibility, and osteoconductivity than the PHBHHx scaffolds. The results suggest that PHBHHx-HAP composite scaffolds can be employed as a promising candidate for bone reconstruction.     

In vitro degradation and fracture toughness of multilayered porous poly(propylene fumarate)/beta-tricalcium phosphate scaffolds

This study investigated the in vitro degradation of poly(propylene fumarate)/beta-tricalcium phosphate (PPF/beta-TCP) scaffolds in pH 7.4 phosphate-buffered saline at 37 degrees C. Scaffold design consisted of three layers: two solid layers about a central layer of porous PPF foam. Solid PPF with molecular weights of 810 and 1450 Da was crosslinked under UV light. PPF foam was prepared by a photocrosslinking, porogen-leaching method with an initial porogen content of 80 wt % and two sizes, 150-300 and 300-500 microm. Comparison of initial and residual weights demonstrated a 14.3 +/- 2.0% loss of mass at 3 weeks and a 16.6 +/- 1.8% loss of mass at 6 weeks. Observed pH values for all constructs remained stable (7.15-7.40) throughout the 3 to 6 weeks. Scanning electron micrographs of these scaffolds revealed some loss of foam material between 3 and 6 weeks; however, foam microarchitecture was intact. Solid PPF fracture toughness was tested for high and low molecular weight PPF, 0.376 +/- 0.004 and 0.134 +/- 0.015 MPa(m)1/2, respectively. These values are roughly one magnitude less than human cortical bone.     

Injectable biodegradable polymer composites based on poly(propylene fumarate) crosslinked with poly(ethylene glycol)-dimethacrylate

New injectable, in situ crosslinkable biodegradable polymer composites were investigated consisting of poly(propylene fumarate) (PPF), poly(ethylene glycol)-dimethacrylate (PEG-DMA), and beta-tricalcium phosphate (beta-TCP). We examined the effects of the PEG-DMA/PPF double-bond ratio and beta-TCP content on the crosslinking characteristics of the composites including the maximum crosslinking temperature and the gel point, as well as the properties of the crosslinked composites such as the compressive strength and modulus, and the water-holding capacity. The maximum crosslinking temperature was constant averaging 39.7 degrees C for the composite formulations tested. The gel points varied from 8.0 +/- 1.0 to 12.6 +/- 2.5 min and were not affected by the relative amounts of PEG-DMA. The compressive strength at yield of PEG-DMA/PPF composites without beta-TCP increased from 5.9 +/- 1.0 to 11.2 +/- 2.2 MPa as the double-bond ratio of PEG-DMA/PPF increased from 0.38 to 1.88. An increase in compressive modulus was also observed from 30.2 +/- 3.5 to 58.4 +/- 6.2 MPa for the same range of the PEG-DMA/PPF double-bond ratio. Also, the addition of beta-TCP (33 wt%) enhanced the mechanical properties of all composites. The equilibrium water content of networks without beta-TCP increased from 21.7 +/- 0.2 to 30.6 +/- 0.2% for a double-bond ratio of PEG-DMA/PPF ranging from 0.38 to 1.88. However, the mechanical properties of the swollen composites under compression were smaller than the dry ones. These data demonstrate the feasibility of fabricating injectable biodegradable polymer composites with engineered mechanical properties for orthopedic tissue engineering.