Search

We investigated the local electrical properties of Pt Schottky contacts to a-plane n-type GaN using conductive atomic force microscopy (C-AFM). Current-voltage characteristics obtained by C-AFM showed rectifying properties, indicating nano-scale Schottky junction formation. Two-dimensional current maps revealed that the surface microstructures of GaN influenced transport properties of the junctions.

Poly(lactic-co-glycolic acid)(PLGA)/biphasic calcium phosphate (BCP) composite nanofibers with different BCP to PLGA ratios were fabricated using the electrospinning technique. The scanning electron microscopy (SEM) images showed a similar morphology and fibers in all groups. The incorporated BCP was dispersed homogenously throughout the nanofibers, and the surface roughness was affected by the input amount of BCP. The increase in amount of BCP incorporated was confirmed by several methods. BCP incorporation into the PLGA nanofibers did not affect the initial adhesion of osteoblasts and their adherent morphology. However, the proliferation of the cells cultured on the composite nanofibers for 10 days with larger amounts of BCP was delayed, suggesting that incorporated BCP may facilitate the switch from proliferation to differentiation of the osteoblasts. The incorporation of BCP enhanced the expression of osteogenic genes, as well as induced calcium deposition by the osteoblasts in the extracellular matrix(ECM) after 21 days of culture on the PLGA/BCP composite nanofibers. Overall, these results can provide evidence of the potential of BCP incorporation into the biomaterials for effective bone regeneration.

RGD-conjugated Tetronic–tyramine (RGD–Tet–TA) hydrogels were formed in situ, and cell attachment and spreading on the hydrogels were evaluated for an injectable tissue-regenerative scaffold. The hydrogels were prepared by simple mixing GRGDGGGGGY (RGD–Y) in a tyramine-conjugated Tetronic (Tet–TA) polymer solution in the presence of horseradish peroxidase (HRP) and hydrogen peroxide (H2O2). The physico-chemical properties such as mechanical properties and swelling ratio were evaluated and the conjugated RGD concentration and distribution were measured using fluoresamine assay and fluorescence observation. An in vitro cell study was investigated using osteoblasts (MC3T3–E1) on the hydrogel surfaces for 24 h, in which the viability and morphology of the adhered cells were observed using live/dead and F-actin assays. The results demonstrate that the in situ conjugation of RGD peptides to tyrosine residues enhanced the attachment and spreading of osteoblasts in a surface density-dependent manner. Therefore, this study suggests that in situ conjugation of the RGD peptide to injectable hydrogels via an enzymatic reaction may be an efficient tool to prepare in situ forming bio-mimetic hydrogels for tissue regenerative medicine.

In this study, novel fibrous matrices were developed as a depot to store and liberate growth factors in a controlled manner. Specifically, heparin was covalently conjugated onto the surface of fibrous matrices (composites of poly[caprolactone] and gelatin crosslinked with genipin), and basic fibroblast growth factor (bFGF) was then reversibly immobilized. The immobilization of bFGF was controlled as a function of the amount of conjugated heparin. The sustained release of bFGF from the fibrous matrices was successfully achieved over 4 weeks whereas physical adsorption of bFGF released quickly. The bFGF released from the fibrous matrices significantly enhanced in vitro proliferation of human umbilical vein endothelial cells. From the in vivo study, the group implanted with a higher amount of immobilized bFGF significantly facilitated neo-blood vessel formation as compared with other implantation groups. These results indicate that the sustained release of bFGF is important for the formation of blood vessels and that our fibrous matrices could be useful for regulation of tissue damage requiring angiogenesis. Further, our system can be combined with other growth factors with heparin binding domains, representing a facile depot for spatiotemporal control over the delivery of bioactive molecules in regenerative medicine.

In this study, we fabricated non-woven matrices using blends of polycaprolactone and gelatin with various spinning volumes to control the immobilized heparin content, which was ultimately intended to increase the immobilization efficiency of bFGF. The amount of bFGF on the heparin conjugated fibrous matrices depended on the thicknesses of the swollen matrices ranging from 35.4 ± 6.5 to 162.3 ± 14.0 ng and ≈90% of the bFGF was gradually released over a period of up to 56 d. The released bFGF enhanced the proliferation of human umbilical vein endothelial cells and human mesenchymal stem cells. In conclusion, our heparin-conjugated fibrous matrices have the potential to be used as a growth factor delivery system in tissue engineering applications.

Regulation of cell-material interactions is an important factor for modulating the cell function in many tissue engineering applications. A more attractive strategy for enhancing the cell-material interactions is to mimic the physical and chemical features of the native extracellular matrix (ECM). The main goal of this study was to develop ECM-like substrates that can control the cell-material interactions including adhesion, spreading, proliferation and differentiation. Poly(L-lactide-co-ɛ-caprolactone) (PLCL) fibrous meshes were fabricated using electrospinning. The meshes were functionalized with acrylic acid (AAc) using γ-ray irradiation, and Arg-Gly-Asp (RGD)-containing peptide was immobilized on the resulting mesh as a cell adhesive ligand. The adhesion and proliferation of the MC3T3-E1 pre-osteoblastic cells grown on the RGD-AAc-PLCL fibrous meshes were greater than those of the cells grown on the other fibrous meshes for up to 7 days. In addition, mature formation of F-actin stress fibers and focal adhesion (co-localized with vinculin) was only observed on the RGD-AAc-PLCL meshes. Moreover, the ALP activity and calcium content on the RGD-AAc-PLCL meshes were approximately 7.5 and 6.7 times higher than those on the other meshes, respectively. In addition, the expression of selected osteogenic genes, Cbfa1, ALP, and OCN, was significantly up-regulated (at least 5 to 9.7 times greater) on the RGD-AAc-PLCL meshes. This suggests that peptide-modified fibrous meshes eliciting desirable cellular responses may provide a useful tool for many tissue engineering applications.

Over the past decades, hydrogels have been widely studied as biomaterials for various biomedical applications like implants, drugs and cell delivery carriers because of their high biocompatibility, high water contents and excellent permeability for nutrients and metabolites. Especially, in situ forming hydrogel systems have received much attention because of their easy application based on minimal invasive techniques. Chemical cross-linking systems fabricated using enzymatic reactions have various advantages, such as high biocompatibility and easy control of reaction rates under mild condition. In this study, we report enzyme-triggered injectable and biodegradable hydrogels composed of Tetronic-tyramine conjugates. The Tetronic-tyramine conjugates were synthesized by first reacting Tetronic with succinic anhydride and subsequent conjugation with tyramine using DCC/NHS as coupling reagents. The chemical structure of Tetronic-succinic anhydride-tyramine (Tet-SA-TA) copolymer was characterized by (1)H NMR and FTIR. The hydrogels were prepared from a Tet-SA-TA solution above 3 wt % in the presence of horseradish peroxidase (HRP) and H(2)O(2) under physiological conditions. Their mechanical property, gelation time, swelling ratio and degradation time were evaluated at different polymer, HRP, and H(2)O(2) concentrations. In addition, a cyto-compatibility study was performed using the MC3T3-E1 cell line. In the cytotoxicity test, it was clear that the Tet-SA-TA hydrogel had no apparent cytotoxicity except for the hydrogel formed with 0.25 wt % H(2)O(2) due to the cytotoxicity of residual H(2)O(2). In conclusion, the obtained results demonstrated that the Tet-SA-TA hydrogel has great potential for use as an injectable scaffold for tissue engineering and as a drug carrier for controlled drug delivery systems.

We fabricated composite fibrous scaffolds from blends of poly(lactide-co-glycolide) (PLGA) and nano-sized hydroxyapatite (HA) via electrospinning. SEM-EDX and AFM analysis demonstrated that HA was homogeneously dispersed in the nanofibers, and the roughness increased along with the amount of incorporated HA. When hMSCs were cultured on these PLGA/HA composite nanofibers, we found that incorporation of HA on the nanofibers did not affect cell viability whereas increased ALP activity and expression of osteogenic genes as well as the calcium mineralization of hMSCs. Our results indicate that the composite nanofibers can be offered as a potential bone regenerative biomaterial for stem cell based therapies.

Composite nanofibers of poly(caprolactone) (PCL) and gelatin crosslinked with genipin are prepared. The contact angles and mechanical properties of crosslinked PCL-gelatin nanofibers decrease as the gelatin content increases. The proliferation of myoblasts is higher in the crosslinked PCL-gelatin nanofibers than in the PCL nanofibers, and the formation of myotubes is only observed on the crosslinked PCL-gelatin nanofibers. The expression level of myogenin, myosin heavy chain, and troponin T genes is increased as the gelatin content is increased. The results suggest that PCL-gelatin nanofibers crosslinked with genipin can be used as a substrate to modulate proliferation and differentiation of myoblasts, presenting potential applications in muscle tissue engineering.

Myotubes assemble with bundles of myofibers to form the structural units in skeletal muscle. Therefore, myotube formation plays an important role in restoring muscular functions, and substrates to promote the differentiation of myoblasts to myotubes need to be developed for muscle tissue engineering. In this study, we developed electrically conductive composite fibers of poly(L-lactide-co-epsilon-caprolactone) (PLCL) blended with polyaniline (PANi) using an electrospinning method, and then investigated the effect of these composite fibers on the differentiation of myoblasts. The prepared PLCL/PANi fibers showed no significant difference in fiber diameter or contact angle, regardless of the incorporation of PANi. The fibers containing 30% PANi (PLCL/PANi-30) maintained elastic properties of maximum elongation at break (160+/-14.4%). The composite fibers were cytocompatible, as the DNA content on each fiber was similar for up to 8 days of C2C12 myoblast culture. After 4 days of culture, the number of cells positive for sarcomeric myosin was 3.6-times greater on the electrically conductive fibers (21+/-1 and 19+/-2 for PLCL/PANi-15 and -30 fibers, respectively) than on the PLCL/PANi-0 fibers (6+/-2). Furthermore, the level of myogenin expression detected on day 8 of culture on PLCL/PANi-15 was approximately 1.6-fold greater than the PLCL/PANi-0 fibers. Similar results were observed for the expression of other genes including troponin T (2-fold greater) and the myosin heavy chain gene (3-fold greater). These results indicate that electrically conductive substrates can modulate the induction of myoblasts into myotube formation without additional electrical stimulation, suggesting that these fibers may have potential as a temporary substrate for skeletal tissue engineering.

Aims Cardiac tissue engineering has been proposed as an appropriate method to repair myocardial infarction (MI). Evidence suggests that a cell with scaffold combination was more effective than a cell-only implant. Nevertheless, to date, there has been no research into elastic biodegradable poly(lactide-co-e-caprolactone) (PLCL) scaffolds. The aim of this study was to investigate the effect of mesenchymal stem cells (MSCs) with elastic biodegradable PLCL scaffold transplants in a rat MI model. Methods and results Ten days after inducing MI through the cryoinjury method, a saline control, MSC, PLCL scaffold, or MSC-seeded PLCL scaffold was transplanted onto the hearts. Four weeks after transplantation, cardiac function and histology were evaluated. Transplanted MSCs survived and differentiated into cardiomyocytes in the injured region. Left ventricular ejection fraction in the MSC + PLCL group increased by 23% compared with that in the saline group; it was also higher in the MSC group. The infarct area in the MSC + PLCL group was decreased by 29% compared with that in the saline group; it was also reduced in the MSC group. Conclusion Mesenchymal stem cells plus PLCL should be an excellent combination for cardiac tissue engineering.

The effect of NBM incorporation in PLA nanofibers on their mechanical properties and the differentiation and mineralization of osteoblasts was studied. At 20% NBM, the Young's modulus of the nanofibers was 37.78 +/- 4.23, significantly larger than that of pure PLA nanofibers. MC3T3-E1 pre-osteoblasts attached to both types of nanofibers and developed full osteogenic phenotypes. A profound effect of NBM on the mineralization of MC3T3-E1 pre-osteoblasts was confirmed, suggesting that NBM/PLA composite nanofibers exhibit properties similar to those of the native collagen-rich mineralized bone matrix, and could therefore serve as a temporary substrate for facilitating the differentiation and mineralization of bone-forming cells.

In this work, electrically conductive polyaniline (PAni) doped with camphorsulfonic acid (CPSA) is blended with poly(L-lactide-co-epsilon-caprolactone) (PLCL), and then electrospun to prepare uniform nanofibers. The CPSA-PAni/PLCL nanofibers show a smooth fiber structure without coarse lumps or beads and consistent fiber diameters (which range from 100 to 700 nm) even with an increase in the amount of CPSA-PAni (from 0 to 30 wt.-%). However, the elongation at break decreases from 391.54 +/- 9.20% to 207.85 +/- 6.74% when 30% of CPSA-PAni is incorporated. Analysis of the surface of the nanofibers demonstrates the presence of homogeneously blended CPSA-PAni. Most importantly, a four-point probe analysis reveals that electrical properties are maintained in the nanofibers where the conductivity is significantly increased from 0.0015 to 0.0138 S x cm(-1) when the nanofibers are prepared with 30% CPSA-PAni. The cell adhesion tests using human dermal fibroblasts, NIH-3T3 fibroblasts, and C2C12 myoblasts demonstrate significantly higher adhesion on the CPSA-PAni/PLCL nanofibers than pure PLCL nanofibers. In addition, the growth of NIH-3T3 fibroblasts is enhanced under the stimulation of various direct current flows. The CPSA-PAni/PLCL nanofibers with electrically conductive properties may potentially be used as a platform substrate to study the effect of electrical signals on cell activities and to direct desirable cell function for tissue engineering applications.

Controlled adhesion and continuous growth of human mesenchymal stem cells (hMSCs) are essential for scaffold-based delivery of hMSCs in tissue engineering applications. The main goal of this study is to develop biofunctionalized synthetic substrates to actively control adhesion, spreading, and proliferation of hMSCs. gamma-Ray irradiation was employed to graft acrylic acid (AAc) to biodegeradable poly(L-lactide-co--caprolactone) (PLCL) films. Gelatin, a natural polymer, was then immobilized on the AAc grafted PLCL film (AAc-PLCL) to induce biomimetic interactions with the cells. The graft yield of AAc increased as the irradiation dose and AAc concentration increased, and the presence of gelatin (gelatin-AAc-PLCL) following immobilization was confirmed using ESCA. To investigate cell responses, hMSCs isolated from a human mandible were cultured on the various substrates and their adhesion, spreading, and proliferation were examined. After three days of culture, the DNA concentration from the cells cultured on gelatin-AAc-PLCL film was 2.9-fold greater than that on the PLCL film. Immunofluorescent staining of hMSCs cultured on the gelatin-AAc-PLCL films demonstrated homogeneous localization of F-Actin and vinculin in their cytoplasm, while mature adhesive structure was not observed from the cells cultured on other substrates. Furthermore, the ratio of projected area of adherent single cells on gelatin-AAc-PLCL films was significantly larger (116.80 +/- 12.78%) than that on the PLCL films (30.11 +/- 5.07%). Our results suggest that gelatin-immobilized PLCL substrates may be potentially used in tissue engineering, particularly as a stem cell delivery carrier for the regeneration of target tissue.

The production of nanofibrous PLA/HA composite scaffolds is described. The morphological, mechanical, surface, and thermal properties of the composites were extensively investigated. The results show that the mixture of PLA and HA formed smooth nanofibers without lumps. The incorporation of HA increased the mechanical strength of the nanofibers and changed the morphology, increasing the mean fiber diameter and pore size. Surface and internal properties confirmed that HA was homogeneously distributed inside the nanofibers and oriented towards their surface. The nanofiber composites allowed the adhesion and proliferation of pre-osteoblasts for up to 3 weeks.

Biodegradable and elastic poly(L-lactide-co-e-caprolactone) (PLCL) was electrospun to prepare nanofibers, andN-isopropylacrylamide (NIPAAm) was then grafted onto their surfaces under aqueous conditions using60Co-γ irradiation. The graft yield increased with increasing irradiation dose from 5 to 10 kGy and the nanofibers showed a greater graft yield compared with the films. SEM confirmed that the PLCL nanofibers maintained an interconnected pore structure after grafting with NIPAAm. However, overdoses of irradiation led to the excessive formation of homopolymer gels on the surface of the PLCL nanofibers. The equilibrium swelling and deswelling ratio of the PNIPAAm-g-PLCL nanofibers (prepared with 10 kGy) was the highest among the samples, which was consistent with the graft yield results. The phase-separation characteristics of PNIPAAm in aqueous conditions conferred a unique temperature-responsive swelling behavior of PNIPAAm-g-PLCL nanofibers, showing the ability to absorb a large amount of water at < 32 °C, and abrupt collapse when the temperature was increased to 40 °C. In accordance with the temperature-dependent changes in swelling behavior, the release rate of indomethacin and FITC-BSA loaded in PNIPAAm-g-PLCL nanofibers by a diffusion-mediated process was regulated by the change in temperature. Both model drugs demonstrated greater release rate at 40 °C relative to that at 25 °C. This approach of the temperature-controlled release of drugs from PNIPAAm-g-PLCL nanofibers using gamma-ray irradiation may be used to design drugs and protein delivery carriers in various biomedical applications.

Recently, much attention has been given to the fabrication of tissue-engineering scaffolds with nano-scaled structure to stimulate cell adhesion and proliferation in a microenvironment similar to the natural extracellular matrix milieu. In the present study, blends of gelatin and poly(L-lactide-co-epsilon-caprolactone) (PLCL) (blending ratio: 0, 30, 70 and 100 wt% gelatin to PLCL) were electrospun to prepare nano-structured non-woven fibers for the development of mechanically functional engineered skin grafts. The resulting nanofibers demonstrated the uniform and smooth fibers with mean diameters ranging from approx. 50 to 500 nm with interconnected pores, regardless of the composition. The contact angle decreased with increasing amount of gelatin in the blend and the water content of the nanofibers increased concurrently. PLCL nanofibers retained significant levels of recovery following application of uniaxial stress; GP-3 with 70% PLCL blend returned to the original length within less than 10% of deformation following 200% of uniaxial elongation. The overall tensile strength was inversely affected by increase in the gelatin content and degradation rates of the nanofibers were accelerated as the gelatin concentration increased. When seeded with human primary dermal fibroblasts and keratinocytes on the nanofibers, both initial cell adhesion and proliferation rate increased as a function of the gelatin content in the blend. Additionally, the total cell number was significantly greater on the nanofiber scaffolds than on polymer-coated glasses, indicating that nanofibrous structure facilitates cell proliferation. Taken together, gelatin/PLCL blend nanofiber scaffolds may serve as a promising artificial extracellular matrix for regeneration of mechanically functional skin tissue.

Mechanical stimulation is known to activate several cellular signal transduction pathways, leading to the induction of signaling molecules and extracellular matrix (ECM) proteins, thereby modulating cellular activities, such as proliferation and survival. In this study, primary human dermal fibroblasts (HDFs) were seeded onto chitosan-based scaffolds, and then cultured for 3 weeks in a bioreactor under a cyclic strain of 1 Hz frequency. Compared to control samples cultured under static conditions, the application of a cyclic strain stimulated the proliferation of HDFs in 1 week, and by week 3 the thickness of the cell/scaffold composites increased 1.56 fold. Moreover, immunohistochemical staining of the culture media obtained from the cell/scaffold samples subjected to the cyclic strain, revealed increases in the expression and secretion of ECM proteins, such as fibronectin and collagen. These results suggest that the preconditioning of cell/scaffold composites with a cyclic strain may enhance the proliferation of HDFs, and even facilitate integration of the engineered artificial dermal tissue into the host graft site.

Biofiltration for volatile organic compound (VOC) control in waste gas streams is best operated at steady contaminants loading. Variations in contaminant loading are, however, common in real applications. In order to provide long-term stable operation of a biofilter under adverse contaminant feeding conditions, a dual-fixed bed adsorption system involving a two-step cycle, i.e., adsorption and desorption, was proposed in this study. The beds of the adsorption unit were packed with granular activated carbon, and the unit was installed in-series before the biofilter. At transient toluene loadings averaging 46.9, 56.3, and 65.9 g/(m 3 h), a biofilter (total bed volume of 2.72 × 10 −3 m 3 ) with pre-cyclic adsorber/desorber unit (total volume of 2.06 × 10 −4 m 3 ) successfully reduced the waste gas toluene concentration below 5 mg/m 3 , while similar behavior was not achieved by a stand-alone biofilter (total volume of 2.72 × 10 −3 m 3 ). The net effect of the two-bed adsorption unit was to attenuate the peaks of toluene concentration, and to make the VOC-laden air stream amenable for biofiltration. It was concluded in this study that the integrated treatment scheme compared to a stand-alone biofilter attained the goal of reliably treating fluctuating toluene loading with high removal efficiency.

We have demonstrated synthesis and application of a water-soluble, folate-substituted poly(p-phenyleneethynylene) (PPE) as a fluorescent contrast agent to image cancer cells. This fluorescent polymer targets and images KB cancer cells in vitro with high selectivity. To deliver PPE to the cells, folate ligands have been attached to an amine-functionalized PPE via an amide coupling agent. The hydrolysis of the ester groups gave a water-soluble PPE, 5. The PPE 5 is minimally cytotoxic at concentrations of 1-10 microg/mL, which is sufficient to stain KB cancer cells efficiently. PPE 6, devoid of folate ligands, did not stain KB cells. As a low folic acid (-) receptor control group, NIH 3T3 fibroblast cells were incubated with 5 and did not show fluorescent labeling. The folate receptor-mediated endocytosis of KB cells was evidenced by laser scanning confocal microscopy and fluorescence microscopy. The photochemical stability and ability to sustain multivalency provide advantages of PPEs over other fluorescent contrast agents. Their minimal cytotoxicity makes the PPE superior to the cytotoxic CdSe quantum dots.

Success in tissue engineering requires an understanding of how cells integrate the signals presented from the microenvironment created by biomaterial scaffolds to alter their responses. Besides the presence of chemical stimuli, there is growing evidence that the spatial organization of cells and tissue within a 3-dimensional (3-D) extracellular matrix (ECM) context is a critical element in controlling cellular function. Therefore, in order to direct cells toward a desirable tissue structure, it is necessary to engineer biomaterials to have spatiotemporal control of the presentation of regulatory signals. Given that, micro-patterning techniques have profited by combining micro-fabrication technology with the chemical conjugation of biologically active molecules to provide new culture systems where cells can be cultured within a specific geometry. The micro-engineered environments have been developed as 2- and 3-D structures, which have proven greatly useful as versatile platforms to study cell, biomaterial, and ECM interactions on both macroscopic and microscopic levels. The main focus of this review is a brief summary of the use of micro-engineered substrates in the analysis of cell-biomaterial interactions with the aim to provide an introductory overview of practical applications available in the literature. In particular, topics regarding (1) the soft-lithography technique to prepare micro-patterned substrates for the spatial control of cell adhesion, (2) biomaterials stiffness-dependent cellular responses, and (3) the microarray techniques for analysis of cell/biomaterials interactions are discussed.

Current advances in biomaterial fabrication techniques have broadened their application in different realms of biomedical engineering, spanning from drug delivery to tissue engineering. The success of biomaterials depends highly on the ability to modulate cell and tissue responses, including cell adhesion, as well as induction of repair and immune processes. Thus, most recent approaches in the field have concentrated on functionalizing biomaterials with different biomolecules intended to evoke cell- and tissue-specific reactions. Marine mussels produce mussel adhesive proteins (MAPs), which help them strongly attach to different surfaces, even under wet conditions in the ocean. Inspired by mussel adhesiveness, scientists discovered that dopamine undergoes self-polymerization at alkaline conditions. This reaction provides a universal coating for metals, polymers, and ceramics, regardless of their chemical and physical properties. Furthermore, this polymerized layer is enriched with catechol groups that enable immobilization of primary amine or thiol-based biomolecules via a simple dipping process. Herein, this review explores the versatile surface modification techniques that have recently been exploited in tissue engineering and summarizes polydopamine polymerization mechanisms, coating process parameters, and effects on substrate properties. A brief discussion of polydopamine-based reactions in the context of engineering various tissue types, including bone, blood vessels, cartilage, nerves, and muscle, is also provided.

The immune system plays a crucial role in the success of tissue engineering strategies. Failure to consider the interactions between implantable scaffolds, usually containing cells and/or bioactive molecules, and the immune system can result in rejection of the implant and devastating clinical consequences. However, recent research into mesenchymal stem cells, which are commonly used in many tissue engineering applications, indicates that they may play a beneficial role modulating the immune system. Likewise, direct delivery of bioactive molecules involved in the inflammatory process can promote the success of tissue engineering constructs. In this article, we will review the various mechanisms in which modulation of the immune system is achieved through delivered bioactive molecules and cells and contextualize this information for future strategies in tissue engineering.

Chondrogenic commitments of mesenchymal stem cells (MSCs) require 3D cellular organization. Furthermore, recent progresses in bioreactor technology have contributed to the development of various biophysical stimulation platforms for efficient cartilage tissue formation. Here, an approach is reported to drive 3D cellular organization and enhance chondrogenic commitment of bone-marrow-derived human mesenchymal stem cells (BM-hMSCs) via magnetic nanoparticle (MNP)-mediated physical stimuli. MNPs isolated from Magnetospirillum sp. AMB-1 are endocytosed by the BM-hMSCs in a highly efficient manner. MNPs-incorporated BM-hMSCs are pelleted and then subjected to static magnetic field and/or magnet-derived shear stress. Magnetic-based stimuli enhance level of sulfated glycosaminoglycan (sGAG) and collagen synthesis, and facilitate the chondrogenic differentiation of BM-hMSCs. In addition, both static magnetic field and magnet-derived shear stress applied for the chondrogenic differentiation of BM-hMSCs do not show increament of hypertrophic differentiation. This MNP-mediated physical stimulation platform demonstrates a promising strategy for efficient cartilage tissue engineering.

Development of stem cell delivery system with ability of control over mutilineage differentiation and improved engraft efficiency is imperative in regenerative medicine. We herein report transfer stamping of human mesenchymal stem cells (hMSCs) patches using thermally expandable hydrogels with tunable cell-adhesive properties. The hydrogels were prepared from functionalized four arm copolymer of Tetronic(®), and the cell adhesion on the hydrogel was modulated by incorporation of fibronectin (FN) or cell-adhesive peptide (RGD). The resulting hydrogels showed spontaneous expansion in size within 10min in response to the temperature reduction from 37 to 4°C. The adhesion and proliferation of hMSCs on FN-hydrogels were positively tunable in proportion to the amount of FN within hydrogels with complete monolayer of hMSCs (hMSC patch) being successfully achieved. The hMSC patch on the hydrogel was faced to the target substrate, which was then easily detached and re-attached to the target when the temperature was reduced from 37°C up to 4°C. We found that the transfer stamping of cell patch was facilitated at lower temperature of 4°C relative to 25°C, with the use of thinner hydrogels (0.5mm in thickness relatively to 1.0 or 1.5mm) and longer transfer time (>15min). Notably, the hMSC patch was simply transferred from the hydrogel to the subcutaneous mouse skin tissue within 15min with cold saline solution being dropped to the hydrogel. The hMSC patch following osteogenic or adipogenic commitment was also achieved with long-term culture of hMSCs on the hydrogel, which was successfully detached to the target surface. These results suggest that the hydrogels with thermally expandable and tunable cell-adhesive properties may serve as a universal substrate to harvest hMSC patch in a reliable and effective manner, which could potentially be utilized in many cell-sheet based therapeutic applications.

In this study, we investigated the effect of signaling peptides incorporated into oligo(poly(ethylene glycol) fumarate) (OPF) hydrogels on in vitro differentiation and mineralization of marrow stromal cells (MSCs) cultured in media without soluble osteogenic supplements (dexamethasone and beta-glycerol phosphate). When MSCs were cultured for 16 days on OPF hydrogels modified with Arg-Gly-Asp (RGD) containing peptides, the normalized cell number was dependent on the peptide concentration between days 0 and 5 and reached comparable values at day 10 regardless of the concentration. The alkaline phosphatase (ALP) activity of MSCs on the peptide-modified OPF hydrogels was also concentration-dependent: ALP activity showed peaks on day 10 or day 13 on OPF hydrogels modified with 2.0 and 1.0 micromol peptide/g, which were significantly greater than those on the OPF hydrogels modified with 0.1 micromol peptides/g or no peptide. A characteristic marker of osteoblastic differentiation, osteopontin (OPN), was detected for all the test groups. However, OPN secretion between days 0 and 10 was significantly higher on the peptide modified hydrogels compared to that on tissue culture-treated polystyrene. Taken together, the results indicate that the presence of signaling peptide allows for a favorable microenvironment for MSCs to differentiate into osteoblasts and produce mineralized matrix, although the soluble factors may further enhance calcium deposition. These findings further support the usefulness of OPF hydrogels as scaffolds for guided bone regeneration, and represent an initial step in exploring the complex relationship between soluble and insoluble factors in osteogenic differentiation on biodegradable materials.

We prepared oligo(poly(ethylene glycol) fumarate) (OPF) hydrogels modified with a rat osteopontin-derived peptide (ODP), Asp-Val-Asp-Val-Pro-Asp-Gly-Arg-Gly-Asp-Ser-Leu-Ala-Try-Gly (DVDVPDGRGDSLAYG), as well as Gly-Arg-Gly-Asp-Ser (GRGDS) and investigated the modulation of marrow stromal osteoblast function on the peptide-modified hydrogels. Osteoblast attachment was competitively inhibited by a soluble peptide suggesting that the interaction of osteoblasts with the hydrogel was ligand specific. The proliferation index of osteoblasts relative to the initial seeding density was similar on the hydrogels modified with ODP (1.18+/-0.13) and GRGDS (1.27+/-0.12). However, fibroblasts proliferated faster on GRGDS-modified hydrogels than on ODP-modified hydrogels as evidenced by the proliferation indices of 4.89+/-0.03 and 2.42+/-0.16, respectively. A megacolony migration assay conducted for 3 days with a seeding density of 53,000 cells/cm(2) showed that osteoblasts migrated to a longer distance on ODP-modified hydrogels (0.23+/-0.06 mm/day) than on hydrogels modified with GRGDS (0.15+/-0.02 mm/day). In addition, osteoblasts migrated faster than fibroblasts seeded at the same density on ODP-modified hydrogels (0.15+/-0.11 mm/day). The migration of osteoblasts on the peptide-modified hydrogels was dependent on the peptide concentration of the hydrogels resulting in an increased migration distance with increasing the peptide concentration for the concentrations tested. These results show that OPF-based biomimetic hydrogels hold promise for modulating cell proliferation and migration for specific applications by altering the specific ligand and its concentration in the hydrogels.

The development of biomaterials for tissue engineering applications has recently focused on the design of biomimetic materials that are capable of eliciting specific cellular responses and directing new tissue formation mediated by biomolecular recognition, which can be manipulated by altering design parameters of the material. Biomolecular recognition of materials by cells has been achieved by surface and bulk modification of biomaterials via chemical or physical methods with bioactive molecules such as a native long chain of extracellular matrix (ECM) proteins as well as short peptide sequences derived from intact ECM proteins that can incur specific interactions with cell receptors. The biomimetic materials potentially mimic many roles of ECM in tissues. For example, biomimetic scaffolds can provide biological cues for cell-matrix interactions to promote tissue growth, and the incorporation of peptide sequences into materials can also make the material degradable by specific protease enzymes. This review discusses the surface and bulk modification of biomaterials with cell recognition molecules to design biomimetic materials for tissue engineering. The criteria to design biomimetic materials such as the concentration and spatial distribution of modified bioactive molecules are addressed. Recent advances for the development of biomimetic materials in bone, nerve, and cardiovascular tissue engineering are also summarized.

This study evaluates the in vitro biocompatibility of an injectable and biodegradable polymeric network based on poly(propylene fumarate) (PPF) and the cross-linking agent PPF-diacrylate (PPF-DA). Using a methyl tetrazolium (MTT) assay, the effect of the concentrations of PPF and PPF-DA on the cytotoxicity of its unreacted macromers, cross-linked networks, and degradation products was examined. The influence of network structure properties on cell viability and attachment to the cross-linked material was also investigated. The unreacted macromers exhibited a time- and dose-dependent cytotoxic response that increased with more PPF-DA in the mixture. Conversely, the cross-linked networks formed with more PPF-DA did not demonstrate an adverse response because increases in conversion and cross-linking density prevented the extraction of toxic products. Fibroblast attachment was observed on the PPF/PPF-DA networks with the highest double bond conversions. The degradation products, obtained from the complete breakdown of the networks in basic conditions, displayed a dose-dependent cytotoxic response. These results show that there are concerns regarding the biocompatibility of injectable, biodegradable PPF/PPF-DA networks but also sheds light onto potential mechanisms to reduce the cytotoxic effects.

Currently, oligo[poly(ethylene glycol) fumarate] (OPF) hydrogels are being investigated as an injectable and biodegradable system for tissue engineering applications. In this study, cytotoxicity of each component of the OPF hydrogel formulation and the resulting cross-linked network was examined. Specifically, OPF synthesized with poly(ethylene glycol) (PEG) of different molecular weights (MW), the cross-linking agent [PEG-diacrylate (PEG-DA)], and the redox initiator pair [ammonium persulfate (APS) and ascorbic acid (AA)] were evaluated for cytotoxicity at 2 and 24 h using marrow stromal cells (MSCs) as model cells. The effect of leachable byproducts of OPF hydrogels on cytotoxicity was also investigated. Upon exposure to various concentrations of OPF for 2 h, greater than 50% of the MSCs were viable, regardless of OPF molecular weight or concentration in the media. After 24 h, the MSCs maintained more than 75% viability except for OPF concentrations higher than 25% (w/v). When examining the cross-linking agent, PEG-DA of higher MW (3400) demonstrated significantly higher viability compared to PEG-DA with MW 575 at all concentrations tested. Considering initiators, when MSCs were exposed to AA and APS, as well as the combination of AA and APS, higher viability was observed at lower concentrations. Once cross-linked, the leachable products from the OPF hydrogels had minimal adverse effects on the viability of MSCs (percentage of live cells was higher than 90% regardless of hydrogel types). The results suggest that, after optimization of cross-linking parameters, OPF-based hydrogels hold promise as novel injectable scaffolds or cell carriers in tissue engineering.

Although the coculture of multiple cell types has been widely employed in regenerative medicine, in vivo transplantation of cocultured cells while maintaining the hierarchical structure remains challenging. Here, a spatially assembled bilayer cell sheet of human mesenchymal stem cells and human umbilical vein endothelial cells on a thermally expandable hydrogel containing fibronectin is prepared and its effect on in vitro proangiogenic functions and in vivo ischemic injury is investigated. The expansion of hydrogels in response to a temperature change from 37 to 4 °C allows rapid harvest and delivery of the bilayer cell sheet to two different targets (an in vitro model glass surface and in vivo tissue). The in vitro study confirms that the bilayer sheet significantly increases proangiogenic functions such as the release of nitric oxide and expression of vascular endothelial cell genes. In addition, transplantation of the cell sheet from the hydrogels into a hindlimb ischemia mice model demonstrates significant retardation of necrosis particularly in the group transplated with the bilayer sheet. Collectively, the bilayer cell sheet is readily transferrable from the thermally expandable hydrogel and represents an alternative approach for recovery from ischemic injury, potentially via improved cell-cell communication.

Summary We fabricated composite fibrous matrices with various volumes, ultimately to control the amount of immobilized basic fibroblast growth factor (bFGF). The fibrous matrices appeared to have an open-porous structure and the amount of bFGF on fibrous matrices was increased with increasing the volumes of matrices. Controlled release of bFGF from the matrices was achieved for 56 days. During in vitro test, the released bFGF maintained its bioactivity and enhanced proliferation of human mesenchymal stem cells.

A novel oligomer based on alternating fumaric acid and poly(ethylene glycol) (PEG) units was synthesized and characterized. End group analysis by nuclear magnetic resonance (NMR) spectroscopy showed that the oligo (PEG fumarate) (OPF) had end PEG chains and multiple fumarate groups along its macromolecular chain. According to thermal characterization by differential scanning calorimetry (DSC), the crystallinity of OPF was found to be lower than PEG. The molecular weight and the degree of oligomerization of OPF determined by gel permeation chromatography (GPC) revealed that the conversion of OPF was substantially affected by the steric factors associated with large PEG chains. OPF was cross-linked using radical polymerization in the presence of either a chemical initiator or photoinitiator. As the OPF cross-linked, the unsaturated fumarate bonds disappeared and the PEG proton peak broadened in the NMR spectra. The cross-linked OPF gel showed swelling behavior that was dependent on the molecular weight of P...

A novel macromer, oligo(poly(ethylene glycol) fumarate) (OPF), was synthesized by the reaction between poly(ethylene glycol) (PEG) of molecular weight 1000 (PEG 1.0K) and fumaryl chloride. The oligo(PEG fumarate) (OPF 1.0K) was modified with a peptide known to modulate cellular functions, Gly-Arg-Gly-Asp (GRGD), after being activated with 4-nitrophenyl chloroformate (NPC). The determined yield of the GRGD modification in 0.1 M sodium bicarbonate buffer of pH 8.3 was 83% as determined by NMR measurements. The OPF 1.0K and the OPF 1.0K modified with GRGD were cross-linked with an unsaturated biodegradable polyester, poly(propylene fumarate) (PPF), by photopolymerization. The cross-linked PPF was characterized by Fourier transform infrared (FT-IR) spectroscopy and contact angle measurements. The equilibrium contact angle of water on the cross-linked PPF surface decreased with the incorporation of OPF 1.0K and the OPF 1.0K modified with GRGD. The results suggest that the OPF macromer can be used for the preparation of functionalized networks incorporating cell adhesion specific sequences.

Peripheral arterial disease is a common manifestation of systemic atherosclerosis, which results in more serious consequences of ischemic events in peripheral tissues such as the lower extremities. Cell therapy has been tested as a treatment for peripheral ischemia that functions by inducing angiogenesis in the ischemic region. However, the poor survival and engraftment of transplanted cells limit the efficacy of cell therapy. In order to overcome such challenges, we applied genetically engineered cell sheets using a cell-interactive and thermosensitive hydrogel and nonviral polymer nanoparticles. C2C12 myoblast sheets were formed on Tetronic-tyramine (Tet-TA)-RGD hydrogel prepared through a highly efficient and noncytotoxic enzymatic reaction. The myoblast sheets were then transfected with vascular endothelial growth factor (VEGF) plasmids using poly(β-amino ester) nanoparticles to increase the angiogenic potential of the sheets. The transfection increased the VEGF expression and secretion from the C2C12 sheets. The enhanced angiogenic effect of the VEGF-transfected C2C12 sheets was confirmed using an in vitro capillary formation assay. More importantly, the transplantation of the VEGF-transfected C2C12 sheets promoted the formation of capillaries and arterioles in ischemic muscles, attenuated the muscle necrosis and fibrosis progressed by ischemia, and eventually prevented ischemic limb loss. In conclusion, the combination of cell sheet engineering and genetic modification can provide more effective treatment for therapeutic angiogenesis.

In this study, thermosensitive hydrogels incorporated with multiple cell-interactive factors were developed as a substrate to form monolayer of human umbilical vein endothelial cells (HUVECs) that can be detached and transferrable to target sites as a cell-sheet in response to temperature change. The cell adhesive peptide (RGD) and growth factor (bFGF) covalently incorporated within the hydrogel significantly enhanced adhesion and proliferation of HUVECs, allowing for the formation of their confluent monolayer. Meanwhile, the precisely controllable change in the size of the hydrogels was observed by a repeated increase and decrease in temperature from 37 to 4 °C. By exploiting this unique behavior, the detachment and transfer of HUVEC sheet confluently cultured at 37 °C was rapidly induced within 10 min by expansion of the hydrogels when the temperature was decreased to 4 °C. The transferred cell sheet was highly viable and maintained robust cell−cell junction. Finally, the process of cell sheet transfer was directly applied onto an ischemic injury in the hind limb of mice. The transplanted HUVECs as a sheet retarded tissue necrosis over 14 days in comparison with that of direct injection of the same number of cells. Our results suggest that the developed multifunctional Tetronic-tyramine hydrogels could serve as an ideal substrate to modulate the formation of an endothelial cell layer that could potentially be utilized to treat peripheral arterial disease.

Incomplete pressure equalization (PE) is practiced in a commercial oxygen concentrator for medical use by adopting simultaneous PE and feed-pressurization for pressurizing an adsorption bed. In such a cycle configuration, extent of equalization during PE affects process performance. In order to assess the effect, performance of pressure swing adsorption (PSA) process with incomplete PE was determined by both simulations and experiments. In simulations, an equilibrium model was used with the assumptions of multicomponent Langmuir isotherms, isothermal operation, and no pressure drop through a bed. The required parameters for simulations were measured in separate experiments. PSA experiments were performed for a two-bed cycle with PE. Two kinds of pressurization, feed and product, were examined. Effects of purge amount and extent of equalization on process performance were assessed in view of productivity and light-component recovery. From the obtained results performance contours were constructed. 95 oxygen mole percent production from air with zeolite 13× was considered as a case study. In both pressurizations, an optimal specific purge and an extent of equalization for the productivity and recovery were observed, but with a different level of equalization. For a maximum productivity feed-pressurization favored incomplete PE, while a maximum recovery occurred at complete PE for both pressurizations. The simulations depicted well existence of optimum conditions, though they showed quantitative disagreement with experiments.

Interpenetrating polymer networks (IPNs) composed of poly(vinyl alcohol) (PVA) and poly(acrylic acid) (PAAc) exhibited electrical-sensitive behavior. PAAc as an initial network was prepared inside a PVA solution using UV irradiation; then, PVA networks as a secondary network were formed by a repetitive freeze–thawing process. Their mechanical properties were influenced by the swelling ratio, crosslinking by UV radiation and a freeze–thawing process, and intermolecular force by hydrogen bonding. When a swollen PVA/PAAc IPN was placed between a pair of electrodes, the IPN exhibited bending behavior upon the applied electric field. The equilibrium bending angle (EBA) and the bending speed of the PVA/PAAc IPN increased with the applied voltage and the content of the PAAc network having negatively charged ionic groups within the IPN. The electroresponsive behavior of the present IPN was also affected by the electrolyte concentration of the external solution. Particularly, IPN37 showed a maximum EBA when the critical ionic strength was 0.1. Anisotropic deswelling of the IPN was observed in a direct contact with a pair of electrodes under aerobic conditions. The PVA/PAAc IPN also showed stepwise bending behavior depending on the electric stimulus. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 73: 1675–1683, 1999

The swelling behaviors of poly(vinyl alcohol)–poly(acrylic acid) (PVA–PAAc) interpenetrating networks (IPN) hydrogels in the presence of electrolytes were studied. The ionized carboxylic group within IPN hydrogels at pH 7 strongly interacted with electrolytes in the medium and caused anomalous swelling pattern. The permeabilities of 5 representative solutes were regulated as a function of temperature, pH, ionic strength, solute size, and ionic properties of solutes. The permeation of nonionic solutes followed the swelling behaviors dependent on external stimuli, including the above factors. However, the ionic solutes showed different trends in their permeation through IPN hydrogels. © 1998 John Wiley & Sons, Inc. J Appl Polm Sci 69: 479–486, 1998

A temperature- and pH-responsive drug delivery system was studied by using interpenetrating polymer network (IPN) hydrogels constructed with poly(acrylic acid) (PAAc) and poly(vinyl alcohol) (PVA). The release of indomethacin incorporated into these hydrogels showed pulsatile patterns in response to both pH and temperature. Indomethacin diffused from the polymer matrices through the swelling and deswelling mechanism. The release amount increased at higher temperature because of the swelling caused by the dissociation of hydrogen bonding. The drastic change of drug release was achieved by alternating pH of the buffer solution and was attributed to the change of states of ionic groups within IPN hydrogels. The free water contents were calculated by using differential scanning calorimetry (DSC), and were proved to be the main factor in the swelling. These results demonstrated that the drug release could be controlled by the swelling/deswelling degree of IPN hydrogels as functions of pH and/or temperature. © 1997 John Wiley & Sons, Inc. J Appl Polm Sci 65: 685–693

Natural vessel has three types of concentric cell layers that perform their specific functions. Here, the fabrication of vascular structure is reported by transfer printing of three different cell layers using thermosensitive hydrogels. Tetronic-tyramine and RGD peptide are co-crosslinked to prepare cell adhesive and thermosensitive hydrogels. The hydrogel increases its diameter by 1.26 times when the temperature reduces from 37 °C to 4 °C. At optimized seeding density, three types of cells form monolayers on the hydrogel, which is then transferred to the target surface within 3 min. Three monolayers are simultaneously transferred on one substrate with controlled shape and arrangement. The same approach is applied onto nanofiber scaffolds that are cultured for more than 5 d. Every type of monolayer shows proliferation and migration on nanofiber scaffolds, and the formation of robust cell-cell contact is revealed by CD31 staining in endothelial cell layer. A vascular structure with multicellular components is fabricated by transfer of three monolayers on nanofibers that are manually rolled with the diameter and length of the tube being approximately 3 mm and 12 mm, respectively. Collectively, it is concluded that the tissue transfer printing is a useful tool for constructing a vascular structure and mimicking natural structure of different types of tissues.

Peripheral arterial disease (PAD) is characterized by the altered structure and function of arteries caused by accumulated plaque. There have been many studies on treating this disease by the direct injection of various types of therapeutic cells, however, the low cell engraftment efficiency and diffusion of the transplanted cells have been major problems. In this study, we developed an approach (transfer printing) to deliver monolayer of cells to the hindlimb ischemic tissue using thermosensitive hydrogels, and investigated its efficacy in long term retention upon transplantation and therapeutic angiogenesis. We first investigated the in vitro maintenance of robust cell–cell contacts and stable expression of the ECM proteins in myoblast layer following transfer printing process. In order to confirm the therapeutic effect of the myoblasts in vivo, we cultured a monolayer of C2C12 myoblasts on thermosensitive hydrogels, which was then transferred to the hindlimb ischemia tissue of athymic mice directly from the hydrogel by conformal contact. The transferred myoblast layer was retained for a longer period of time than an intramuscularly injected cell suspension. In addition, the morphology of the mice and laser Doppler perfusion (28 days after treatment) supported that the myoblast layer enhanced the therapeutic effects on the ischemic tissue. In summary, the transplantation of the C2C12 myoblast layer using a tissue transfer printing method could represent a new approach for the treatment of PAD by therapeutic angiogenesis.

The ultimate goal of tissue engineering is to replace damaged tissues by applying engineering technology and the principles of life sciences. To successfully engineer a desirable tissue, three main elements of cells, scaffolds and growth factors need to be harmonized. Biomaterial-based scaffolds serve as a critical platform both to support cell adhesion and to deliver growth factors. Various methods of fabricating scaffolds have been investigated. One recently developed method that is growing in popularity is called electrospinning. Electrospinning is known for its capacity to make fibrous and porous structures that are similar to natural extracellular matrix (ECM). Other advantages to electrospinning include its ability to create relatively large surface to volume ratios, its ability to control fiber size from micro- to nano-scales and its versatility in material choice. Although early work with electrospun fibers has shown promise in the regeneration of certain types of tissues, further modification of their chemical, biological and mechanical properties would permit future advancements. In this paper, current approaches to the development of modular electrospun fibers as scaffolds for tissue engineering are discussed. Their chemical and physical characteristics can be tuned for the regeneration of specific target tissues by co-spinning of multiple materials and by post-modification of the surface of electrospun fibers. In addition, topology or structure can also be controlled to elicit specific responses from cells and tissues. The selection of proper polymers, suitable surface modification techniques and the control of the dimension and arrangement of the fibrous structure of electrospun fibers can offer versatility and tissue specificity, and therefore provide a blueprint for specific tissue engineering applications.

The development of ideal barrier membranes with appropriate porosity and bioactivity is essential for the guidance of new bone formation in orthopedic and craniomaxillofacial surgery. In this study, we developed bioactive electrospun fibers based on poly (lactide-co-glycolic acid) (PLGA) by immobilizing bone-forming peptide 1 (BFP1) derived from the immature region of bone morphogenetic protein 7 (BMP7). We exploited polydopamine chemistry for the immobilization of BFP1; polydopamine (PD) was coated on the electrospun PLGA fibers, on which BFP1 was subsequently immobilized under weakly basic conditions. The immobilization of BFP1 was verified by characterizing the surface chemical composition and quantitatively measured by fluorescamine assay. The immobilization of BPF1 on the electrospun fibers supported the compact distribution of collagen I and the spreading of human mesenchymal stem cells (hMSCs). SEM micrographs demonstrated the aggregation of globular mineral accretions, with significant increases in ALP activity and calcium deposition when hMSCs were cultured on fibers immobilized with BFP1 for 14 days. We then implanted the prepared fibers onto mouse calvarial defects and analyzed bone formation after 2 months. Semi-quantification of bone growth from representative X-ray images showed that the bone area was approximately 20% in the defect-only group, while the group implanted with PLGA fibers showed significant improvements of 44.27 ± 7.37% and 57.59 ± 15.24% in the groups implanted with PD-coated PLGA and with BFP1-coated PLGA, respectively. Based on these results, our approach may be a promising tool to develop clinically-applicable bioactive membranes for guided bone regeneration."

Adult stem cells have recently drawn considerable attention for potential cell therapy applications. However, critical details about their specific in vivo environments and cellular activities are unclear. Adipose tissue-derived mesenchymal stem cells (ASCs) are attractive candidates for treating bone defects, but most studies focus on delivery of in vitro-differentiated cells. We assessed various scaffolding materials for the ability to support osteogenic differentiation of undifferentiated human ASCs in vivo, in athymic nude rat calvaria. Twenty-four 9- to 10-week-old athymic nude Sprague-Dawley rats (250 g) were used in these experiments. Fat tissue from 3 patients was harvested from abdominal tissue discarded during reconstructive breast surgery by transverse rectus abdominis myocutaneous flap, performed at the Asan Medical Center after resection of breast cancer. Human ASCs were extracted from discarded adipose tissue and isolated based on standard International Society for Cellular Therapy protocols. Adipose tissue-derived mesenchymal stem cells were seeded on polylactic glycolic acid, atelocollagen, and hydroxyapatite scaffolds, and osteogenesis was evaluated using bone mineral densitometry, histology, immunohistochemistry, and reverse transcription polymerase chain reaction. The gross appearance of scaffolds seeded with ASCs was strikingly different from that of scaffolds alone. Bone mineral densitometry analysis revealed a 2- to 3-fold increase in mineral density in ASC-seeded scaffolds. In addition, undifferentiated ASCs seeded onto hydroxyapatite scaffolds, but not onto collagen or polylactic glycolic acid scaffolds, expressed human messenger RNA for osteogenic markers such as alkaline phosphatase, osteopontin, osteocalcin, and osteonectin. These results indicate that undifferentiated human ASCs can differentiate into osteocytes or osteoblasts in athymic nude rat calvaria, and the importance of appropriate scaffolding for in vivo ASC differentiation.

Gas separation of a binary gas mixture by various pressure swing adsorption (PSA) cycles was studied by a numerical simulation in order to provide a guidance in selecting PSA cycles. PSA cycles considered in this study are 3, 4-step cycles for production of only one component and a cycle with pressure equalization for production of a light component. 4 and 5-step cycles for simultaneous production of both components of a binary gas mixture are also considered. Separation of a CH4/CO2 gas mixture with zeolite 5A was chosen as a case study. Performances of cycles were examined and compared in view of purity, recovery and productivity. Their relative advantages were discussed. Inclusion of a purging step to a 3-step cycle for production of only one component improves a cycle performance. Further performance improvement of a cycle for production of a light component can be achieved by employing pressure equalization. Sircar's 4-step cycle with a recycle of effluent shows the best performance in view of purity and recovery among cycles for simultaneous production of both components.