The use of debrided human articular cartilage for autologous chondrocyte implantation: maintenance of chondrocyte differentiation and proliferation in type I collagen gels.

UNLABELLED: Autologous chondrocyte implantation (ACI) is the most promising surgical treatment for large full thickness knee joint articular cartilage (AC) defects where cells from healthy non-weight bearing area AC are multiplied in vitro and implanted into such defects. In the routine surgical procedure for symptomatic knee full thickness AC defects, damaged AC surrounding the edge and the base of such defects is usually debrided and discarded. The purpose of this study was to examine if chondrocytes from this 'debrided' AC can proliferate, synthesize a cartilage specific matrix and thus can be used for ACI. METHODS: Biopsies were retrieved from 12 patients (debrided articular cartilage: DAC, aged 35-61) and from two autopsies (normal articular cartilage: NAC, aged 21 and 25). Chondrocytes were isolated, seeded at low density in type I collagen gels and as monolayer cultures for 4 weeks without passage. RESULTS: After 4 weeks cultures in type I collagen gels, cell proliferation from DAC (18.34 +/- 1.95 fold) was similar to cells from NAC (11.24 +/- 1.02 fold). Syntheses of proteoglycan and collagen in DAC were also similar to NAC. Newly synthesized matrices in gel cultures consisted predominantly of type II collagen as shown by immuno-labelling and SDS-PAGE followed by fluorography. Chondrocytes from 'debrided human AC' cultured at low density in type I collagen gels may be used for the ACI procedure as they provide sufficient viable cell numbers for ACI and maintain their chondrocyte phenotype as they synthesize a cartilage-like matrix.

Effects of growth factors on cell proliferation and matrix synthesis of low-density, primary bovine chondrocytes cultured in collagen I gels.

Low cell density cell numbers and dedifferentiation are two major problems of human chondrocyte culture associated with articular cartilage repair. Bovine chondrocytes seeded at low density (3.5 x 10(4) cells/ml of gels) in three-dimensional collagen type I gels do proliferate and maintain their phenotype as shown by cell counts, morphology and matrix synthesis. The combination of three growth factors (3GFs; 10 ng/ml TGF-beta1 + 100 ng/ml IGF-I + 10 ng/ml b-FGF) added to serum-free culture medium in this culture system enhances the mitotic activity of bovine chondrocytes similar to 20% foetal calf serum (FCS). At day 21, cells proliferated by 41 fold in gels-FCS and 37 fold in gels-3GFs. Protein synthesis by gels-3GFs cultures was similar to 20% FCS when cultured for 3 weeks but much less proteoglycan was synthesized. The matrix deposition as observed by light and electron microscopy was quite different. More small diameter branching collagen fibrils and a denser matrix were presented in gels-FCS culture whilst loosely arranged larger diameter collagen fibrils were observed in gels-3GFs.

In vitro evaluation of voriconazole against clinical isolates of yeasts, moulds and dermatophytes in comparison with itraconazole, ketoconazole, amphotericin B and griseofulvin.

The in vitro activity of voriconazole (UK-109, 496), a new antifungal triazole derivative, against 650 clinical isolates of yeasts, moulds and dermatophytes was compared with that of itraconazole, ketoconazole, amphotericin B and griseofulvin. The geometric means of the minimum inhibitory concentrations (MICs) of voriconazole were 0.05 microgram ml-1 against yeasts (n = 187), 0.58 microgram ml-1 against moulds (n = 260) and 0.08 microgram ml-1 against dermatophytes (n = 203). The overall activity of voriconazole against yeasts and moulds was good, being similar to that of itraconazole, ketoconazole and amphotericin B. Voriconazole was highly effective against Aspergillus fumigatus (mean MIC 0.23 microgram ml-1) and other Aspergillus species and showed noteworthy activity (mean MICs 0.08-0.78 microgram ml-1) against emerging and less common clinical isolates of opportunistic moulds, such as Alternaria spp., Cladosporium spp., Acremonium spp., Chrysosporium spp. and Fusarium spp. On the other hand, voriconazole was less active in vitro than the comparative agents studied against various species of zygomycetes, such as Mucor spp., Rhizopus spp. and Absidia spp. Voriconazole and the other two azoles, itraconazole and ketoconazole, were more active than griseofulvin in vitro against most dermatophytes tested.

In vitro activity of voriconazole against yeasts, moulds and dermatophytes in comparison with fluconazole, amphotericin B and griseofulvin.

Voriconazole (CAS 137234-62-9, UK-109,496), a new antifungal triazole derivative, was studied in vitro against 650 clinical isolates, representing yeasts, moulds and dermatophytes, and was compared with fluconazole (CAS 86386-73-4), amphotericin B (CAS 1397-89-3), and griseofulvin (CAS 126-07-8). The mean minimum inhibitory concentrations (MICs) of voriconazole were 0.06 microgram/ml against yeasts (n = 187), 0.74 microgram/ml against moulds (n = 260) and 0.10 microgram/ml against dermatophytes (n = 203). Data from these in vitro studies showed that voriconazole was more potent than fluconazole against most species studied, but particularly against the isolates of moulds and dermatophytes. Overall, voriconazole and amphotericin B indicated comparably good activity against yeasts and moulds. Voriconazole was highly potent against 13 Aspergillus species studied (mean MIC 0.35 microgram/ml) and also showed noteworthy activity (mean MICs 0.08-0.78 microgram/ml) against emerging and less common clinical isolates of opportunistic moulds such as of Alternaria spp., Cladosporium spp., Acremonium spp., Chrysosporium spp., and Fusarium spp. In addition, voriconazole was more active in vitro than griseofulvin against most dermatophytes tested. The in vitro results confirmed that voriconazole has indeed a broad antifungal spectrum and could also be effective against a wide range of fungal infections in patients.