Solvent Casting And Particulate Leaching Research Articles

Characterization of non-solvent- and thermal-induced phase separation applied polycaprolactone/demineralized bone matrix scaffold for bone tissue engineering

ObjectivePolycaprolactone (PCL) is a widely applied biomaterial in bone tissue engineering (BTE) due to its superior mechanical properties and biodegradability. However, the high hydrophobicity and low cell adhesion properties of PCL show limited cell interactions. Herein, we prepared the porous PCL/DBP composites with improved cell adhesion through the addition of demineralized bone powder (DBP). Three-dimensional scaffolds were fabricated by mixing various concentrations of DBP with PCL and applying non-solvent-induced phase separation (NIPS) and thermal-induced phase separation (TIPS) (N-TIPS) and solvent casting and particulate leaching (SCPL) to impart porosity.MethodsA characteristic evaluation was performed through X-ray diffraction (XRD), morphological analysis, physicochemical analysis, bioactivity test, and mechanical test. Upon culture with mouse bone marrow stem cells (mBMSCs), proliferation and osteogenic differentiation of mBMSC were evaluated using quantitative dsDNA analysis and alkaline phosphatase (ALP) activity, respectively.ResultsThe addition of DBP improved the physicochemical and mechanical properties of the scaffold and formed a large amount of hydroxyapatite (HAp). Also, cell proliferation and differentiation were increased by enhancing cell adhesion.ConclusionThe porous PCL/DBP scaffolds could provide a favorable microenvironment for cell adhesion and be a promising biomaterial model for bone tissue engineering.Graphical abstract

Macro- and microporous polycaprolactone/duck’s feet collagen scaffold fabricated by combining facile phase separation and particulate leaching techniques to enhance osteogenesis for bone tissue engineering

Herein, a facile macro- and microporous polycaprolactone/duck’s feet collagen scaffold (PCL/DC) was fabricated and characterized to confirm its applicability in bone tissue engineering. A biomimetic scaffold for bone tissue engineering and regeneration for bone defects is an important element. PCL is a widely applied biomaterial for bone tissue engineering due to its biocompatibility and biodegradability. However, the high hydrophobicity and low cell attachment site properties of PCL lead to an insufficient microenvironment in designing a scaffold. Collagen is a nature-derived biomaterial that is widely used in tissue engineering and has excellent biocompatibility, mechanical properties, and cell attachment moieties. Among the resources from which collagen can be obtained, DC contains a high amount of collagen type I (COL1), is biocompatible, and is cost-effective. In this study, the scaffolds were fabricated by blending DC with PCL in various ratios and applied non-solvent-induced phase separation (NIPS) and thermal-induced phase separation (TIPS) (N-TIPS), solvent casting and particulate leaching (SCPL), and gas foaming method to fabricate macro- and microporous structure. The characterization of the fabricated scaffolds was carried out by morphological analysis, bioactivity test, physicochemical analysis, and mechanical test. In vitro study was carried out by viability test, morphology observation, and gene expression. The results showed that the incorporation of DC enhances the physicochemical and mechanical properties of the scaffolds. Also, a large amount of bone mimetic apatite was formed according to the DC content in the bioactivity test. The in vitro study showed that the PCL/DC scaffold is biocompatible and the existence of apatite and DC formed a favorable microenvironment for cell proliferation and differentiation. Overall, the novel porous PCL/DC scaffold can be a promising biomaterial model for bone tissue engineering and regeneration.

Effect of sodium chloride as a porogen agent in mechanical properties of PLGA/HA nanocomposite scaffolds

In this study, the novel poly (lactic-co-glycolic acid)-Hydroxyapatite nanoparticles (PLGA/HA) nanocomposite scaffolds were fabricated with solvent casting and particulate leaching (SCPL) method. The role of sodium chloride (NaCl) particles with diameters of 250-400 μm as porogen agent in the mechanical strength of the produced scaffolds was evaluated. The prepared scaffolds were characterized using Fourier transform infrared (FT-IR), X‐ray diffraction (XRD), scanning electron microscopy (SEM), thermal gravimetric analysis (TGA) and compressive tests. The results showed the high compressive strength and homogenous porous structures for PLGA/HA nanocomposite scaffolds compared to pure PLGA due to the presence of HA nanoparticles in nanocomposites. Furthermore, the compressive strength of nanocomposite scaffolds increased by varying the weight ratio of hydroxyapatite nanoparticles to polymer (0, 20, 40 wt%) at constant salt ratio and decreased by increasing the weight ratio of salt particles to polymer from 1 to 3 wt% due to more porosity in nanocomposite scaffolds. These results confirmed that not only the nanocomposite scaffolds exhibited high mechanical properties, homogenous structures, as well as good porosity but also, they could be useful for bone tissue application.

Rapid Fabrication of Anatomically-Shaped Bone Scaffolds Using Indirect 3D Printing and Perfusion Techniques.

Fused deposit modeling (FDM) 3D printing technology cannot generate scaffolds with high porosity while maintaining good integrity, anatomical-surface detail, or high surface area-to-volume ratio (S/V). Solvent casting and particulate leaching (SCPL) technique generates scaffolds with high porosity and high S/V. However, it is challenging to generate complex-shaped scaffolds; and solvent, particle and residual water removal are time consuming. Here we report techniques surmounting these problems, successfully generating a highly porous scaffold with the anatomical-shape characteristics of a human femur by polylactic acid polymer (PLA) and PLA-hydroxyapatite (HA) casting and salt leaching. The mold is water soluble and is easily removable. By perfusing with ethanol, water, and dry air sequentially, the solvent, salt, and residual water were removed 20 fold faster than utilizing conventional methods. The porosities are uniform throughout the femoral shaped scaffold generated with PLA or PLA-HA. Both scaffolds demonstrated good biocompatibility with the pre-osteoblasts (MC3T3-E1) fully attaching to the scaffold within 8 h. The cells demonstrated high viability and proliferation throughout the entire time course. The HA-incorporated scaffolds demonstrated significantly higher compressive strength, modulus and osteoinductivity as evidenced by higher levels of alkaline-phosphatase activity and calcium deposition. When 3D printing a 3D model at 95% porosity or above, our technology preserves integrity and surface detail when compared with FDM-generated scaffolds. Our technology can also generate scaffolds with a 31 fold larger S/V than FDM. We have developed a technology that is a versatile tool in creating personalized, patient-specific bone graft scaffolds efficiently with high porosity, good scaffold integrity, high anatomical-shaped surface detail and large S/V.

An oxygen plasma treated poly(dimethylsiloxane) bioscaffold coated with polydopamine for stem cell therapy.

In this study, 3D macroporous bioscaffolds were developed from poly(dimethylsiloxane) (PDMS) which is inert, biocompatible, non-biodegradable, retrievable and easily manufactured at low cost. PDMS bioscaffolds were synthesized using a solvent casting and particulate leaching (SCPL) technique and exhibited a macroporous interconnected architecture with 86 ± 3% porosity and 300 ± 100 µm pore size. As PDMS intrinsically has a hydrophobic surface, mainly due to the existence of methyl groups, its surface was modified by oxygen plasma treatment which, in turn, enabled us to apply a novel polydopamine coating onto the surface of the bioscaffold. The addition of a polydopamine coating to bioscaffolds was confirmed using composition analysis. Characterization of oxygen plasma treated-PDMS bioscaffolds coated with polydopamine (polydopamine coated-PDMS bioscaffolds) showed the presence of hydroxyl and secondary amines on their surface which resulted in a significant decrease in water contact angle when compared to uncoated-PDMS bioscaffolds (35 ± 3%, P < 0.05). Seeding adipose tissue-derived mesenchymal stem cells (AD-MSCs) into polydopamine coated-PDMS bioscaffolds resulted in cells demonstrating a 70 ± 6% increase in viability and 40 ± 5% increase in proliferation when compared to AD-MSCs seeded into uncoated-PDMS bioscaffolds (P < 0.05). In summary, this two-step method of oxygen plasma treatment followed by polydopamine coating improves the biocompatibility of PDMS bioscaffolds and only requires the use of simple reagents and mild reaction conditions. Hence, our novel polydopamine coated-PDMS bioscaffolds can represent an efficient and low-cost bioscaffold platform to support MSC therapies.

Preparation, Characterization and Mechanical Assessment of Poly (Lactide-Co-Glycolide)/ Hyaluronic Acid/ Fibrin/ Bioactive Glass Nano-composite Scaffolds for Cartilage Tissue Engineering Applications

The purpose of this study was to evaluate the microstructural and mechanical properties of poly (Lactide-co-Glycolide) (PLGA) and PLGA/ Hyaluronic acid (Ha)/ Fibrin (F)/ Bioactive glass (BG) nanocomposite scaffolds for cartilage tissue engineering. Nanocomposite scaffolds composed of PLGA/ Ha/ F/ BG, containing different amounts of Ha, F, and BG nanoparticles were prepared via solvent casting and particulate leaching (SC/PL) techniques. Characterization techniques such as Fourier transform infrared spectroscopy (FT-IR), Scanning electron microscopy (SEM), Transmission electron microscopy (TEM) and X-ray diffraction (XRD), were performed. Furthermore, mechanical properties of the PLGA and nanocomposite scaffolds were determined. The results revealed that the scaffolds contain sufficient porosity with highly interconnected pore morphology. Desired distribution of BG nanoparticles with a few agglomerates was observed in the nanocomposite scaffolds by microscopic techniques. Increase the amount of fibrin enhanced compressive modulus and compressive strength of the nanocomposite scaffolds.

Characterization of controlled highly porous hyaluronan/gelatin cross-linking sponges for tissue engineering

This study examines the suitability of N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride for use as a cross-linker in the preparation of highly porous hyaluronan (HA)/gelatin sponges through the solvent casting and particulate leaching (SCPL) process. The modulus, water absorption rate, bonding, morphology, cell viability, and chondrocyte mRNA expression of the HA/gelatin sponges were measured and compared. Water uptake by the sponges resulted in pores with spherical diameters ranging from 100μm to 200μm. With higher porosity, the strength and modulus declined. Larger salt leaching amounts resulted in higher water uptake. The cross-linking bonds in the HA/gelatin sponges were mostly ester bonds. According to our study, the amount of salt in the SCPL process was the main factor that influenced strength, pore interconnectivity, and water uptake ability. The results also showed that the scaffold had a non-viability effect on human chondrocytes, but the mRNA expression level of aggrecan and type II collagen in the cartilage was significantly higher than that of the control group after 5d of culture. The sponges developed in the present study are potential candidates for chondrocyte proliferation and differentiation in cartilage regeneration.

Poly (d,l-lactide)/nano-hydroxyapatite composite scaffolds for bone tissue engineering and biocompatibility evaluation

Biodegradable polymer/bioceramic composite scaffolds can overcome the limitations of conventional ceramic bone substitutes such as brittleness and difficulty in shaping. However, conventional methods for fabricating polymer/bioceramic composite scaffolds often use organic solvents (e.g., the solvent casting and particulate leaching (SC/PL) method), which might be harmful to cells or tissues. In this study, Poly (D,L-lactide)/nano-hydroxyapatite (PDLLA/NHA) composites were prepared by in-situ polymerization, and highly porous scaffolds were fabricated using a novel method, supercritical CO2/salt-leaching method (SC CO2/SL). The materials and scaffolds were investigated by scanning electronic microscopy (SEM), transmission electronic microscopy (TEM) and gel permeation chromatography (GPC). GPC showed that the molecular weight of composites decreased with increase of NHA content. However, the water absorption and compressive strength increased dramatically. The SEM micrographs showed that the scaffolds with pore size about 250 microm were obtained by controlling parameters of SC CO2/SL. The biocompatibility of PDLLA/NHA porous scaffolds were evaluated in vitro and in vivo. The evaluation on the cytotoxicity were carried out by cell relative growth rate (RGR) method and cell direct contact method. The cytotoxicity of these scaffolds was in grade I according to ISO 10993-1. There was no toxicosis and death cases observed in acute systemic toxicity test. And histological observation of the tissue response (1 and 9 weeks after the implantation) showed that there are still some slight inflammation responses.

A poly(lactide‐co‐glycolide)/hydroxyapatite composite scaffold with enhanced osteoconductivity

Biodegradable polymer/ceramic scaffolds can overcome the limitations of conventional ceramic bone substitutes. However, the conventional methods of polymer/ceramic scaffold fabrication often use organic solvents, which might be harmful to cells or tissues. Moreover, scaffolds fabricated with the conventional methods have limited ceramic exposure on the scaffold surface since the polymer solution envelopes the ceramic particles during the fabrication process. In this study, we developed a novel fabrication method for the efficient exposure of ceramic onto the scaffold surface, which would enhance the osteoconductivity and wettability of the scaffold. Poly(D,L-lactide-co-glycolide)/nanohydroxyapatite (PLGA/HA) scaffolds were fabricated by the gas foaming and particulate leaching (GF/PL) method without the use of organic solvents. Selective staining of ceramic particles indicated that HA nanoparticles exposed to the scaffold surface were observed more abundantly in the GF/PL scaffold than in the conventional solvent casting and particulate leaching (SC/PL) scaffold. Both types of scaffolds were implanted to critical size defects in rat skulls for 8 weeks. The GF/PL scaffolds exhibited significantly enhanced bone regeneration when compared with the SC/PL scaffolds. Histological analyses and microcomputed tomography of the regenerated tissues showed that bone formation was more extensive on the GF/PL scaffolds than on the SC/PL scaffolds. Compared with the SC/PL scaffolds, the enhanced bone formation on the GF/PL scaffolds may result from the higher exposure of HA nanoparticles to the scaffold surface. These results show that the biodegradable polymer/ceramic composite scaffolds fabricated with the novel GF/PL method can enhance bone regeneration compared with those fabricated with the conventional SC/PL method.

The influence of architecture on degradation and tissue ingrowth into three-dimensional poly(lactic- co-glycolic acid) scaffolds in vitro and in vivo

The in vitro and in vivo degradation properties of poly(lactic- co-glycolic acid) (PLGA) scaffolds produced by two different technologies—thermally induced phase separation (TIPS), and solvent casting and particulate leaching (SCPL) were compared. Over 6 weeks, in vitro degradation produced changes in SCPL scaffold dimension, mass, internal architecture and mechanical properties. TIPS scaffolds produced far less changes in these parameters providing significant advantages over SCPL. In vivo results were based on a microsurgically created arteriovenous (AV) loop sandwiched between two TIPS scaffolds placed in a polycarbonate chamber under rat groin skin. Histologically, a predominant foreign body giant cell response and reduced vascularity was evident in tissue ingrowth between 2 and 8 weeks in TIPS scaffolds. Tissue death occurred at 8 weeks in the smallest pores. Morphometric comparison of TIPS and SCPL scaffolds indicated slightly better tissue ingrowth but greater loss of scaffold structure in SCPL scaffolds. Although advantageous in vitro, large surface area:volume ratios and varying pore sizes in PLGA TIPS scaffolds mean that effective in vivo (AV loop) utilization will only be achieved if the foreign body response can be significantly reduced so as to allow successful vascularisation, and hence sustained tissue growth, in pores less than 300 μm.

Poly(lactide- co-glycolide)/hydroxyapatite composite scaffolds for bone tissue engineering

Biodegradable polymer/bioceramic composite scaffolds can overcome the limitations of conventional ceramic bone substitutes such as brittleness and difficulty in shaping. However, conventional methods for fabricating polymer/bioceramic composite scaffolds often use organic solvents (e.g., the solvent casting and particulate leaching (SC/PL) method), which might be harmful to cells or tissues. Furthermore, the polymer solutions may coat the ceramics and hinder their exposure to the scaffold surface, which may decrease the likelihood that the seeded osteogenic cells will make contact with the bioactive ceramics. In this study, a novel method for fabricating a polymer/nano-bioceramic composite scaffold with high exposure of the bioceramics to the scaffold surface was developed for efficient bone tissue engineering. Poly( d, l-lactic- co-glycolic acid)/nano-hydroxyapatite (PLGA/HA) composite scaffolds were fabricated by the gas forming and particulate leaching (GF/PL) method without the use of organic solvents. The GF/PL method exposed HA nanoparticles at the scaffold surface significantly more than the conventional SC/PL method does. The GF/PL scaffolds showed interconnected porous structures without a skin layer and exhibited superior enhanced mechanical properties to those of scaffolds fabricated by the SC/PL method. Both types of scaffolds were seeded with rat calvarial osteoblasts and cultured in vitro or were subcutaneously implanted into athymic mice for eight weeks. The GF/PL scaffolds exhibited significantly higher cell growth, alkaline phosphatase activity, and mineralization compared to the SC/PL scaffolds in vitro. Histological analyses and calcium content quantification of the regenerated tissues five and eight weeks after implantation showed that bone formation was more extensive on the GF/PL scaffolds than on the SC/PL scaffolds. Compared to the SC/PL scaffolds, the enhanced bone formation on the GF/PL scaffolds may have resulted from the higher exposure of HA nanoparticles at the scaffold surface, which allowed for direct contact with the transplanted cells and stimulated the cell proliferation and osteogenic differentiation. These results show that the biodegradable polymer/bioceramic composite scaffolds fabricated by the novel GF/PL method enhance bone regeneration compared with those fabricated by the conventional SC/PL method.