Solidification behavior of PLLA/nHA nanocomposites

As part of a broader effort to establish processing-structure–property relationships in PLLA/nHA, which is currently under consideration for bioresorbable scaffolds for bone repair, hot stage optical microscopy and differential scanning calorimetry have been used to investigate the solidification behavior of a series of medical grade PLLA/nHA nanocomposites. The presence of the nHA resulted in an increase in the number of spherulites per unit volume during isothermal crystallization, but there was a substantial decrease in the spherulite growth rate with increasing nHA content in the temperature range 100–130 °C, argued to be associated with a significant increase in the melt viscosity in the presence of the nHA. The consequences for the global solidification rates and the phase structure of the PLLA/nHA nanocomposites are discussed.     

相分离法构建聚乳酸/纳米羟基磷灰石复合纳米纤维支架及性能

通过液.液相分离法构建纳米纤维聚左旋乳酸/蚋米羟基磷灰石(NF-PLLA/nHA)仿生复合支架,利用扫描电镜、压缩测试、微量二喹啉甲酸(BCA)法、X射线衍射及差示扫描量热等手段对其进行表征.结果显示,nHA均匀馕嵌在PLLA纳米纤维间隙中,不影响其纳米纤维结构且明显提高力学性能.同时,nHA的引入还能增加对牛血清白蛋...     

Efficient and toxicity-free surface immobilization of nano-hydroxyapatite for bone-regenerative composite scaffolds by grafting polyvinyl pyrrolidone

An effective approach to fabricate porous bone regenerative composite scaffolds with surface-immobilized nano-hydroxyapatite (nHA) is developed in this research. In the typical preparation process, surface-repellent stable colloidal nHA with surface carbonyl functionality was fabricated through in-situ polyvinyl pyrrolidone (PVP)-grafting synthesis without any organic solvents and potentially harmful additives. Followed by freezing and lyophilizing homogenous PVP-grafted nHA and chitosan mixtures, three dimensional nHA and chitosan composite scaffolds were then obtained. The TEM images and XPS analysis show that the discrete nHA was anchored at nano-level on the pore surface of chitosan scaffold by mediating PVP chemical-linkage. The PVP-grafting offers an efficient and safe approach to prepare nHA with the surface reactivity and is promising to be widely used in the fabrication of novel composite scaffolds for bone tissue engineering.     

Preparation of poly(3-hydroxybutyrate)/nano-hydroxyapatite composite scaffolds for bone tissue engineering

Poly(3-hydroxybutyrate)/nano-hydroxyapatite (PHB/nHA) composite scaffolds were fabricated via powder mixing, compression moulding, and particle leaching technique. The scaffolds had high porosity with interconnected porous architecture, a favorable structure for cell attachment and new bone tissue ingrowth. A homogeneous dispersion and a uniform distribution of HA nanoparticles in the polymer matrix were obtained. The scaffolds exhibited improved compressive modulus and compressive strength, which were all in the range of compressive modulus and compressive strength of cancellous bone. In addition, the use of toxic organic solvents was eliminated. Thus, the fabricated PHB/nHA composite scaffolds tend to be promising for application in bone tissue engineering.     

Characterization of poly(3-hydroxybutyrate)/nano-hydroxyapatite composite scaffolds fabricated without the use of organic solvents for bone tissue engineering applications

Poly(3-hydroxybutyrate)/nano-hydroxyapatite (PHB/nHA) composite scaffolds were fabricated without the use of organic solvents at different mass fractions of HA nanoparticles. HA nanoparticles were homogeneously dispersed as primary particles in the polymer matrix of the scaffolds at 10 and 15 wt.% nHA content. Agglomeration of HA nanoparticles occurred when the nHA content of the scaffolds reached 20 wt.%. All the scaffolds had high porosities with interconnected porous structure and optimized pore size ranges. Mechanical properties of all the scaffolds were in the range of mechanical properties of cancellous bone. Scaffolds were biocompatible to MG-63 cells in the indirect method of cytotoxicity evaluation. Also, the morphology of the attached MG-63 cells in direct contact with the scaffolds indicated the appropriate cell-scaffold interaction. Thus, the PHB/nHA composite scaffolds investigated in this study tend to be favorable for bone tissue engineering applications.     

Morphology, mechanical properties, and mineralization of rigid thermoplastic polyurethane/hydroxyapatite scaffolds for bone tissue applications: effects of fabrication approaches and hydroxyapatite size

Rigid thermoplastic polyurethane (TPU)/hydroxyapatite (HA) scaffolds were prepared with micro HA (mHA) and nano HA (nHA) particles, respectively, via the thermally induced phase separation method. The effects of solvent and co-solvent, addition of sodium chloride (NaCl) porogen, and HA particle size were studied together with the morphology, compressive properties, and mineralization behavior of the scaffolds. Depending on the solvent, co-solvent, or porogen used, different porous structures were produced. In particular, a ladder-like morphology was obtained when dioxane (Di) was used as the solvent, whereas an interconnected porous structure was obtained by using dioxane and deionized water (DiW) as co-solvents. Rectangular pores with interconnected channels on the pore walls were achieved by using NaCl crystals as porogens. The TPU/nHA scaffolds showed stronger compressive properties than the TPU/mHA scaffolds and the pure TPU scaffolds. The scaffolds prepared using dioxane and water as co-solvents exhibit the greatest compressive modulus. Furthermore, TPU scaffolds with nHA particles had the ability to form bone apatite when soaked in simulated body fluid (SBF). After being soaked in SBF for 3 weeks, the weight percentage of formed apatite in the TPU/nHA-DiW scaffold was 9.2 %wt of the initial TPU content. Preliminary cytotoxicity tests were conducted using NIH 3T3 fibroblast cells. The high survival rate of these cells and the mineralization behavior suggest biocompatibility and high potential of these composites being used in bone tissue engineering applications.     

Needle-like nano hydroxyapatite/poly(l-lactide acid) composite scaffold for bone tissue engineering application

In this paper, a new nano-hydroxyapatite / poly (l-lactide acid) (nHAP/PLLA) composite scaffold comprising needle-like nHAP particles was prepared. In the first step, the identification and morphology of chemically synthesized HAP particles were determined by XRD, EDX, FTIR and SEM analyses. The needle-like nHAP particles with an average size of approximately 30–60 nm in width and 100–400 nm in length were found similar to needle-like bone nano apatites in terms of chemical composition and morphology. In the second step, nHAP and micro-sized HAP (mHAP) particles were used to fabricate HAP filled PLLA (HAP/PLLA) composites scaffolds using solid–liquid phase separation method. The porosity of scaffolds was up to 85%, and their average macropore diameter was in the range of 64–175 µm. FTIR and XRD analyses showed the presence of molecular interactions and chemical linkages between HAP particles and PLLA matrix. The compressive strength of nanocomposite scaffolds could high up to 8.46 MPa while those of pure PLLA and microcomposite scaffolds were 1.79 and 4.61 MPa, respectively. The cell affinity and cytocompatibility of the nanocomposite scaffold were found to be higher than those of pure PLLA and microcomposite scaffolds. Based on the results, the newly developed nHAP/PLLA composite scaffold is comparable with cancellous bone in terms of microstructure and mechanical strength, so it may be a suitable alternative for bone tissue engineering applications.     

Development and characterisation of a collagen nano-hydroxyapatite composite scaffold for bone tissue engineering

Bone regeneration requires scaffolds that possess suitable mechanical and biological properties. This study sought to develop a novel collagen-nHA biocomposite scaffold via two new methods. Firstly a stable nHA suspension was produced and added to a collagen slurry (suspension method), and secondly, porous collagen scaffolds were immersed in nHA suspension after freeze-drying (immersion method). Significantly stronger constructs were produced using both methods compared to collagen only scaffolds, with a high porosity maintained (>98.9%). It was found that Coll-nHA composite scaffolds produced by the suspension method were up to 18 times stiffer than the collagen control (5.50 ± 1.70 kPa vs. 0.30 ± 0.09 kPa). The suspension method was also more reproducible, and the quantity of nHA incorporated could be varied with greater ease than with the immersion technique. In addition, Coll-nHA composites display excellent biological activity, demonstrating their potential as bone graft substitutes in orthopaedic regenerative medicine.     

Formation of bone-like apatite on poly(L-lactide) to improve osteoblast-like compatibility in vitro and in vivo

The biomimetic apatite coating process was adopted to modify poly(L-lactide) (PLLA) surfaces with osteoblasts-like cell compatibility. The apatite coating was formed on the pre-hydrolyzed PLLA film and scaffold surfaces by incubating in simulated body fluid (SBF). Scanning electron microscopy and energy dispersive X-ray analyzer were utilized to characterize the composition and the structure of the apatite coating. The cytocompatibility of the modified PLLA films was investigated by testing osteoblast-like attachment, proliferation, alkaline phosphatase (ALP) activity, and cell cycle. Subsequently, the modified PLLA scaffolds were co-cultured with the osteoblasts-like in vitro and subcutaneously implanted into nude mice. The experimental results showed that the formed apatite had a nano-sized particle and matrix configuration. The surface modification of PLLA with apatite coating significantly promoted osteoblast-like compatibility. After a four-week culture in vivo, no significant inflammatory signs were observed in the implanted regions and osteoblast-like congeries with bone-like structure began to form in the scaffolds. The positive results of this study suggest a good way to produce desirable PLLA biomaterials for bone tissue engineering.     

Human-like collagen/nano-hydroxyapatite scaffolds for the culture of chondrocytes

Three dimensional (3D) biodegradable porous scaffolds play a key role in cartilage tissue repair. Freeze-drying and cross-linking techniques were used to fabricate a 3D composite scaffold that combined the excellent biological characteristics of human-like collagen (HLC) and the outstanding mechanical properties of nano-hydroxyapatite (nHA). The scaffolds were characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and compression tests, using Relive® Artificial Bone (RAB) scaffolds as a control. HLC/nHA scaffolds displayed homogeneous interconnected macroporous structure and could withstand a compression stress of 2.67 ± 0.37 MPa, which was higher than that of the control group. Rabbit chondrocytes were seeded on the composite porous scaffolds and cultured for 21 days. Cell/scaffold constructs were examined using SEM, histological procedures, and biochemical assays for cell proliferation and the production of glycosaminoglycans (GAGs). The results indicated that HLC/nHA porous scaffolds were capable of encouraging cell adhesion, homogeneous distribution and abundant GAG synthesis, and maintaining natural chondrocyte morphology compared to RAB scaffolds. In conclusion, the presented data warrants the further exploration of HLC/nHA scaffolds as a potential biomimetic platform for chondrocytes in cartilage tissue engineering.     

Formation of bone-like apatite on poly(L-lactide) to improve osteoblast-like compatibility in vitro and in vivo

The biomimetic apatite coating process was adopted to modify poly(L-lactide) (PLLA) surfaces with osteoblasts-like cell compatibility. The apatite coating was formed on the pre-hydrolyzed PLLA film and scaffold surfaces by incubating in simulated body fluid (SBF). Scanning electron microscopy and energy dispersive X-ray analyzer were utilized to characterize the composition and the structure of the apatite coating. The cytocompatibility of the modified PLLA films was investigated by testing osteoblast-like attachment, proliferation, alkaline phosphatase (ALP) activity, and cell cycle. Subsequently, the modified PLLA scaffolds were co-cultured with the osteoblasts-like in vitro and subcutaneously implanted into nude mice. The experimental results showed that the formed apatite had a nano-sized particle and matrix configuration. The surface modification of PLLA with apatite coating significantly promoted osteoblast-like compatibility. After a four-week culture in vivo, no significant inflammatory signs were observed in the implanted regions and osteoblast-like congeries with bone-like structure began to form in the scaffolds. The positive results of this study suggest a good way to produce desirable PLLA biomaterials for bone tissue engineering.     

分级多孔壳聚糖/聚乳酸复合支架的制备及骨修复性能

为了仿生莲藕内部的贯穿大孔结构,以生物相容性好的壳聚糖(CS)作为基质材料,利用冰粒致孔、石蜡模具和冰模具成型3种成型方法制备了分级多孔CS支架材料,然后与力学强度较高的聚乳酸(PLLA)复合,制备网络互穿CS/PLLA复合支架。通过SEM、压缩强度测试和兔股骨髁骨缺损模型对CS/PLLA复合材料的形貌、力学强度和骨修复性能进行了表征。结果表明:利用冰模具制备的CS/PLLA复合支架能可控、批量制备,具有微米-毫米分级多孔结构,大孔孔径约为2mm,内部均匀分布着孔径约为60μm的贯穿微孔,并在微孔内形成密集的PLLA絮状网络结构。干态复合材料的压缩强度和模量分别比纯CS支架的提高了6倍和15倍。体内植入实验结果表明,CS/PLLA复合材料能够促进骨缺损的愈合,并随着新骨的形成,复合材料逐渐被降解吸收。     

Enhanced osteogenic differentiation of cord blood-derived unrestricted somatic stem cells on electrospun nanofibers

A new stem cell-scaffold construct based on poly-l-lactide (PLLA) nanofibers grafted with collagen (PLLA-COL) and cord blood-derived unrestricted somatic stem cells (USSC) were proposed to hold promising characteristics for bone tissue engineering. Fabricated nanofibers were characterized using SEM, ATR-FTIR, tensile and contact angle measurements. The capacity of PLLA, plasma-treated PLLA (PLLA-pl) and PLLA-COL scaffolds to support proliferation and osteogenic differentiation of USSC was evaluated using MTT assay and common osteogenic markers such as alkaline phosphatase (ALP) activity, calcium mineral deposition and bone-related genes. All three scaffolds showed nanofibrous and porous structure with suitable physical characteristics. Higher proliferation and viability of USSC was observed on PLLA-COL nanofibers compared to control surfaces. In osteogenic medium, ALP activity and calcium deposition exhibited the highest values on PLLA-COL scaffolds on days 7 and 14. These markers were also greater on PLLA and PLLA-pl compared to TCPS. Higher levels of collagen I, osteonectin and bone morphogenetic protein-2 were detected on PLLA-COL compared to PLLA and PLLA-pl. Runx2 and osteocalcin were also expressed continuously on all scaffolds during induction. These observations suggested the enhanced proliferation and osteogenic differentiation of USSC on PLLA-COL nanofiber scaffolds and introduced a new combination of stem cell-scaffold constructs with desired characteristics for application in bone tissue engineering.     

Vaterite deposition on biodegradable polymer foam scaffolds for inducing bone-like hydroxycarbonate apatite coatings

Hydroxycarbonate apatite (HCA) coatings on the surface of bioresorbable materials for bone tissue engineering scaffolds were produced using macroporous poly(DL-lactide) (PDLLA) foams impregnated by calcium carbonate in vaterite crystalline form. Stable and homogeneous vaterite deposition on PDLLA foams was achieved using a slurry dipping technique. In vitro studies in simulated body fluid (SBF) were performed to induce formation of (HCA) on the surface of vaterite/PDLLA composite foams. HCA was detected after immersion of foams in SBF for 7 days. Hence, depositing vaterite on materials followed by immersion in SBF is confirmed to induce HCA coatings on the surface of the material. The HCA coated, bioactive and resorbable PDLLA foams are intended for use as bone tissue engineering scaffolds.     

Composite poly-l-lactic acid/poly-(α,β)-dl-aspartic acid/collagen nanofibrous scaffolds for dermal tissue regeneration

Tissue engineering scaffolds for skin tissue regeneration is an ever expounding area of research, as the products that meet the necessary requirements are far and elite. The nanofibrous poly-l-lactic acid/poly-(α,β)-dl-aspartic acid/Collagen (PLLA/PAA/Col I&III) scaffolds were fabricated by electrospinning and characterized by SEM, contact angle and FTIR analysis for skin tissue regeneration. The cell-scaffold interactions were analyzed by cell proliferation and their morphology observed in SEM. The results showed that the cell proliferation was significantly increased (p ≤ 0.05) in PLLA/PAA/Col I&III scaffolds compared to PLLA and PLLA/PAA nanofibrous scaffolds. The abundance and accessibility of adipose derived stem cells (ADSCs) may prove to be novel cell therapeutics for dermal tissue regeneration. The differentiation of ADSCs was confirmed using collagen expression and their morphology by CMFDA dye extrusion technique. The current study focuses on the application of PLLA/PAA/Col I&III nanofibrous scaffolds for skin tissue engineering and their potential use as substrate for the culture and differentiation of ADSCs. The objective for inclusion of a novel cell binding moiety like PAA was to replace damaged extracellular matrix and to guide new cells directly into the wound bed with enhanced proliferation and overall organization. This combinatorial epitome of PLLA/PAA/Col I&III nanofibrous scaffold with stem cell therapy to induce the necessary paracrine signalling effect would favour faster regeneration of the damaged skin tissues.     

Biocompatibility,degradability, bioactivity and osteogenesis of mesoporous/macroporous scaffolds of mesoporous diopside/poly(l-lactide) composite

Bioactive mesoporous diopside (m-DP) and poly(l-lactide) (PLLA) composite scaffolds with mesoporous/macroporous structure were prepared by the solution-casting and particulate-leaching method. The results demonstrated that the degradability and bioactivity of the mesoporous/macroporous scaffolds were significantly improved by incorporating m-DP into PLLA, and that the improvement was m-DP content-dependent. In addition, the scaffolds containing m-DP showed the ability to neutralize acidic degradation products and prevent the pH from dropping in the solution during the soaking period. Moreover, the scaffolds containing m-DP enhanced attachment, proliferation and alkaline phosphatase activity of MC3T3-E1 cells, which were also m-DP content-dependent. Furthermore, the histological and immunohistochemical analysis results showed that the scaffolds with m-DP significantly promoted new bone formation and improved the materials degraded in vivo, indicating good biocompatibility. The results suggested that the mesoporous/macroporous scaffolds of the m-DP/PLLA composite with osteogenesis had a potential for bone regeneration.     

Poly(L-lactic acid) nanocylinders as nanofibrous structures for macroporous gelatin scaffolds

Electrospun Nanofiber sheets have been shown to mimic the structure of extracellular matrix (ECM). Although these nanofibers have shown great potential for use as tissue engineering scaffolds, it is difficult for the electrospun nanofiber based sheets to be shaped into the desired three-dimensional structure. In this study, poly(L-lactic acid) (PLLA), a biodegradable and biocompatible polyester, was electrospun to produce nanofibers that were treated with an amino group containing base in order to fabricate polymeric nanocylinders. The aspect ratio of the PLLA nanocylinders was tunable by varying the aminolysis time and density of the amino group containing base. The effects of changes in nanofibrous morphology of the PLLA nanocylinders/macro-porous gelatin scaffolds on cell adhesion and proliferation were evaluated. The results revealed different cell morphology, adhesion, and proliferation in the nanocylinders composite gelatin scaffold versus gelatin scaffold alone. Confocal laser scanning microscopy observation showed more spreading and a more flattened cell morphology after NIH3T3 cells were cultured on PLLA nanocylinders/gelatin scaffolds for 10 hours and 4 days. These results indicate that the gelatin/PLLA nanocylinder composite is a promising way to fabricate 3D nanofibrous scaffolds that accelerates cell adhesion and proliferation for tissue engineering.     

Fabrication and characterization of electrospun osteon mimicking scaffolds for bone tissue engineering

Skeletal loss and bone deficiencies are a major worldwide problem with over 600,000 procedures performed in the US alone annually, making bone one of the most transplanted tissues, second to blood only. Bone is a composite tissue composed of organic matrix, inorganic bone mineral, and water. Structurally bone is organized into two distinct types: trabecular (or cancellous) and cortical (or compact) bones. Trabecular bone is characterized by an extensive interconnected network of pores. Cortical bone is composed of tightly packed units, called osteons, oriented parallel along to the axis of the bone. While the majority of scaffolds attempt to replicate the structure of the trabecular bone, fewer attempts have been made to create scaffolds to mimic the structure of cortical bone. The aim of this study was to develop a technique to fabricate scaffolds that mimic the organization of an osteon, the structural unit of cortical bone. We successfully built a rotating stage for PGA fibers and utilized it for collecting electrospun nanofibers and creating scaffolds. Resulting scaffolds consisted of concentric layers of electrospun PLLA or gelatin/PLLA nanofibers wrapped around PGA microfiber core with diameters that ranged from 200 to 600 μm. Scaffolds were mineralized by incubation in 10× simulated body fluid, and scaffolds composed of 10%gelatin/PLLA had significantly higher amounts of calcium phosphate. The electrospun scaffolds also supported cellular attachment and proliferation of MC3T3 cells over the period of 28 days.     

海藻酸钙水凝胶/聚乳酸复合材料的制备与性能

通过化学发泡-冷冻干燥-粒子滤出复合法制备聚乳酸(PLLA)大孔支架, 然后在大孔内以海藻酸钠(SA)、碳酸钙、葡萄糖酸内酯(GDL)为原料, 通过原位相转变制备海藻酸钙水凝胶/聚乳酸复合材料(CA/PLLA); 分别利用SEM、压缩强度测试和细胞培养对CA/PLLA支架的形貌、力学性能及生物相容性进行了研究。结果表明: PLLA具有直径小于2 mm、孔道相互连通的孔洞, 且在大孔中能够形成均匀的CA。CA/PLLA复合材料的压缩强度(2.74 MPa)远大于单一的海藻酸钙水凝胶的压缩强度(0.10 MPa)。在CA/PLLA复合支架中, 软骨细胞呈簇状圆形生长状态, 与其在天然软骨陷窝里生长状态一致。这种软硬结合、天然与合成高分子杂化的CA/PLLA复合材料的力学强度和生物相容性同时得到提高, 可进一步作为骨和软骨修复材料研究。     

Coating of collagen on a poly(l-lactic acid) sponge surface for tissue engineering

Porous scaffolds play important roles in tissue engineering. Biodegradable synthetic polymers, such as poly(l-lactic acid) (PLLA), frequently are used in the preparation of porous scaffolds. Pretreating the surface of a PLLA porous scaffold is required to increase its wettability for smooth cell seeding due to the hydrophobic property of the scaffold's surface. In this study, a simple coating method was used to modify the surface of the PLLA sponges. The coating method included three steps: filling the PLLA sponge pores with collagen aqueous solution, centrifuging to remove excess collagen, and, finally, freeze-drying. Compared with the uncoated PLLA sponge, the collagen-coated PLLA sponge demonstrated both improved wettability and high water absorption. Cells were smoothly seeded in the collagen-coated PLLA sponges by dropping a cell suspension solution onto the sponges. Cells adhered to the collagen-coated sponge and were distributed homogeneously throughout the collagen-coated PLLA sponge.