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Biomaterials respond to sterilization methods differently. Steam sterilization might decrease the performance of thermoplastic polyether-urethane (TPU); however, the effect of different gamma-radiation doses on this polymer is contradictory in present literature. The purpose of this study was to investigate the differences between irradiative doses in comparison with steam sterilization on a porous TPU scaffold produced by a new processing method. No significant differences in the surface chemical structure were found with attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) analysis when comparing with the sterilization methods. The molecular weight (M(w)) had a net increase from 11.5 +/- 0.039 to 13.2 +/- 0.072 kDa by gamma-sterilization from 10 to 60 kGy. The samples that were irradiated (>60 kGy) had also an increase in polydispersity index (PDI; 1.45 +/- 0.007) in comparison with the nonsterile ones (1.31 +/- 0.017), which indicate branching. Liquid chromatography/mass spectroscopy (LC/MS) analysis showed that there was a correlation between the concentration of the breakdown product, methyl dianiline, and cytotoxicity. The concentration of this compound was found to be four times higher in steam-sterilized sample (1.3 +/- 0.01 ppb) compared with that of the polymer sample gamma-sterilized at 10 kGy (0.3 +/- 0.01 ppb). The cytotoxicity of TPU was found to decrease with higher radiation doses, and was significantly higher for the steam-sterilized samples. It is recommended that TPU produced with the described foaming method should be sterilized by gamma-irradiation at 25 kGy or higher doses., ((c) 2006 Wiley Periodicals, Inc.)

A large-scale scaffold processing method with injection molding has been successfully developed. Water was used as a foaming agent for the new technique. NaCl was used as a porogen to achieve an open-cell structure. Organic solvents, which are common foaming agents for polyurethane, where not used. Toxic remains in the polymer were therefore prevented. Biocompatibility tested gave a mean optical density of 81% from WST-1 proliferation assay. In comparison to the previously study processing method, hot pressing (Haugen H, Ried V, Brunner M, Will J, Wintermantel E. J Mater Sci: Mater Med2004;15:343-346), the current scaffolds had an increase of 20% of the mean optical density. Cell seeding showed that human fibroblasts adhered to the surface and proliferated. The spread of the adhered fibroblasts was uniform on the surface. A quantitative MTT analysis proved that there was a significant (p < 0.01) increase in the OD level after 7 and 14 days of incubation. This cell layer thickened with increased incubation time from 7 to 14 days (p < 0.05) and had typical fibroblast morphology.

In this study we have used lectin histochemistry and scanning electron microscopy (SEM) to assess the growth and characterise the differentiation of human respiratory epithelial cells (REC) cultured on two biomaterial scaffolds. The first scaffold, based on a hyaluronic acid derivative, was observed to be non-adhesive for REC. This lack of adhesion was found to be unrelated to the presence of the hyaluronic acid binding domain on the surface of isolated REC. The other scaffold, consisting of equine collagen. was observed to encourage REC spreading and adhesion. Positive Ulex Europaeus agglutinin (UEA) lectin staining of this preparation indicated the presence of ciliated REC on the scaffold surface. However, the marked decrease in peanut agglutinin (PNA) positive staining, relative to that of control cultures and native tissue, indicates a dedifferentiation of the secretory cells of the REC monolayer. SEM analysis of REC cultured on the collagen scaffold confirmed the presence of ciliated cells thereby validating the UEA positive staining. The presence of both established and developing cilia was also verified. This study indicates that collagen biomaterials are appropriate for the tissue engineering of REC. Furthermore, that UEA and PNA staining is a useful tool in the characterisation of cells cultured on biomaterials, therefore helpful in identifying biomaterials that are suitable for specific tissue engineering purposes.

New cell culture techniques raise the possibility of creating cartilage in vitro with the help of tissue engineering. In this study, we compared two resorbable nonwoven cell scaffolds, a polyglycolic acid/poly-L-lactic acid (PGA/PLLA) (90/10) copolymer (Ethisorb) and pure PLLA (V 7-2), with different degradation characteristics in their aptitude for cartilage reconstruction. Chondrocytes were isolated enzymatically from human septal cartilage. The single cells were resuspended in agarose and transferred into the polymer scaffolds to create mechanical stability and retain the chondrocyte-specific phenotype. The cell-polymer constructs were then kept in perfusion culture for 1 week prior to subcutaneous transplantation into thymusaplastic nude mice. After 6, 12, and 24 weeks, the specimens were explanted and analyzed histochemically on the presence of collagen (azan staining), proteoglycans (Alcian blue staining), and calcification areas (von Kossa staining). Furthermore, different collagen types (collagen type I, which is found in most tissues, but not in hyaline cartilage matrix; and collagen type II, which is cartilage specific) were differentiated immunohistochemically by the indirect immunoperoxidase technique. Vascular ingrowth was investigated by a factor VIII antibody, which is a endothelial marker. Quantification of several matrix components was performed using the software Photoshop. Significant differences were found between both nonwoven structures concerning matrix synthesis and matrix quality as well as vascular ingrowth. Ethisorb, with a degradation time of approximately 3 weeks in vitro, showed no significant differences from normal human septal cartilage in the amount of collagen types I and II 24 weeks after transplantation. Thin fibrous tissue layers containing blood vessels encapsulated the transplants. V 7-2 constructs, which did not show strong signs of degradation even 24 weeks after transplantation, contained remarkably smaller amounts of cartilage-specific matrix components. At the same time, there was vascular ingrowth even in central parts of the transplants. In conclusion, polymer scaffolds with a short degradation time are suitable materials for the development of cartilage matrix products, while longer stability seems to inhibit matrix synthesis. Thus, in vitro engineering of human cartilage can result in a cartilage-like tissue when appropriate nonwovens are used. Therefore, this method could be the ideal cartilage replacement method without the risk of infection and with the possibility of reconstructing large defects with different configurations.

The aim of this study was to investigate the possibility of using the benzyl ester of hyaluronic acid (HYAFF 11), a recently developed semisynthetic resorbable material, as a scaffold for the culture of human nasoseptal chondrocytes in tissue-engineering procedures of cartilage reconstruction. Different techniques such as immunohistochemistry, scanning electron microscopy, and confocal laser scanning microscopy were used to study the behavior, morphology, and phenotype expression of the chondrocytes, which were initially expanded and then seeded on the material. The nonwoven cell carrier allowed good viability and adhesivity of the cells without any surface treatment with additional substances. Furthermore, the cultured cells expressed cartilage-specific collagen type II, indicating that they were able to redifferentiate within the scaffold of HYAFF 11 and were able to retain a chondrocyte phenotype even after a long period of in vitro conditions. Nevertheless, the expression of collagen type I, which was produced by dedifferentiated or incompletely redifferentiated chondrocytes, was noticeable. Additional data were obtained by subcutaneous implantation of samples seeded with human cells in the in vivo model of the athymic nude mouse. The results after 1 month revealed the development of tissue similar to hyaline cartilage. This study is promising for the use of this scaffold for tissue engineering of cartilage replacements.