JOURNALOF MATERIALS SCIENCE: MATERIALS IN MEDICINE 14 (2003) 713±716 Engineered articular cartilage: in¯uence of the scaffold on cell phenotype and proliferation ANTONIO GIGANTE*, CLAUDIA BEVILACQUA, MASSIMO CAPPELLA, SANDRA MANZOTTI, FRANCESCO GRECO Department of Orthopaedics, University of Ancona, Ancona, Italy E-mail: agigante@iol.it Articular cartilage defects do not heal. Biodegradable scaffolds have been studied for cartilage engineering in order to implant autologous chondrocytes and help cartilage repair. We tested some new collagen matrices differing in collagen type, origin, structure and methods of extraction and puri®cation, and compared the behavior of human chondrocytes cultured on them. Human chondrocytes were grown for three weeks on four different equine type I collagen matrices, one type I, III porcine collagen matrix and one porcine type II collagen matrix. After 21 days, samples were subjected to histochemical, immunohistochemical and histomorphometric analysis to study phenotype expression and cell adhesion. At 7, 14 and 21 days cell proliferation was studied by incorporation of [3H]-thymidine. Our data evidence that the collagen type in¯uences cell morphology, adhesion and growth; indeed, cellularity and rate of proliferation were signi®cantly higher and cells were rounder on the collagen II matrix than on either of the collagen I matrices. Among the collagen I matrices, we observed a great variability in terms of cell adhesion and proliferation. The present study allowed us to identify one type I collagen matrix and one type II collagen matrix that could be usefully employed as a scaffold for chondrocyte transplantation. # 2003 Kluwer Academic Publishers Introduction Cartilage repair is a complex problem. Full-thickness defects do not heal because of absence of vascularity and low chondrocyte proliferation capacity [1]. Among the methods proposed to treat these defects [2], autologous chondrocyte implantation has yielded promising results [3]. However, the use of suspended cells and a periosteal ¯ap to hold the chondrocytes in place presents both theoretical and technical dif®culties [4,5]. To address these problems, cell-seeded matrices have been studied in cartilage engineering in order to facilitate in vivo implantation and promote cartilage repair [6]. A three-dimensional scaffold aids cell differentiation, allows for easier implantation and supports cell proliferation at the lesion site. Several natural and synthetic matrices have been studied in vitro and in vivo, among them ®brin gel [7, 8], hyaluronic acid [9], polyglycolic and polylactic acid [10], collagen gels, sponges and membranes [11±14], but results are not totally satisfactory. In the attempt to identify ever improved scaffolds for cartilage engineering, we tested some new collagen matrices differing in collagen type, origin, structure and methods of extraction and puri®cation, and compared the in vitro behavior of human chondrocytes cultured on them. Materials and methods Chondrocyte cultures Normal human articular cartilage was obtained from notch plasty during anterior cruciate ligament recon- structions. Cartilage slices were digested overnight in Dulbecco's modi®ed Eagle's medium (DMEM), 2% fetal bovine serum (FBS) (both from Gibco, Italy), 0.25% collagenase I-A and 0.01% DNAse I (both from Sigma, Italy). Cell viability was evaluated by Trypan blue stain. Cells were grown as monolayers in 75 cm 2 culture ¯asks (Corning Inc., NY) for two weeks, trypsinized (Sigma) and grown in 24-well plastic culture plates (Nunc A/S, Denmark) for three weeks on six different types of collagen membranes at an initial density of 1610 5 cells/cm 2 . The culture medium contained DMEM, 10% FBS (Gibco, Italy), 50 mg/ml ascorbic acid (Sigma), 1% penicillin-streptomycin and 1% fungizone (both from Gibco). Cultures were grown at *Author to whom all correspondence should be addressed: Clinica Ortopedica, Universita Á di Ancona, Via Conca, 60020, Torrette di Ancona, Ancona, Italia. 0957±4530 # 2003 Kluwer Academic Publishers 713