Comparative analysis of different collagen-based biomaterials as scaffolds for long-term culture of human fibroblasts.
Résumé
Biodegradable scaffolds, along with cells, are important components of most tissue-engineered constructs. In the study, there is a comparison of the behaviour of human fibroblasts cultured for up to six weeks in four different collagen-based three-dimensional matrices, in the form of sponges composed of pure native type I collagen (control), of collagen-GAG-chitosan (CGC) and of collagen cross-linked by two concentrations of diphenylphosphorylazide (DPPA-2 and DPPA-3). Variations in size and weight of the sponges, as well as fibroblast growth and migration, and total protein and collagen synthesis, are determined with time in culture. Owing to their low thermal stability, the partial denaturation and dissolution of the control sponges after incubation at 37 degrees C lead to considerable contraction and low cell proliferation. CGC sponges, stabilised by ionic interactions between the different components, show, after six weeks, limited contraction (20%) and weight increase (10% when seeded) and high cell growth (threefold increase). Similar results are obtained with weakly, cross-linked (DPPA-2) collagen sponges. Highly cross-linked (DPPA-3) sponges do not contract, whereas weight gain and cell proliferation are no different from those found with CGC and DPPA-2 sponges. Similar levels of total protein and collagen synthesis are shown for fibroblasts seeded in different matrices, with a slight general decrease (twofold) after three weeks, a much lower value than that observed with fibroblasts in culture within a contracted collagen gel (sixfold). Furthermore, the fraction of neo-synthesised collagen deposited in the sponges after six weeks represents more than 60% of the total, compared with only 10% obtained with fibroblasts in monolayer culture or 30% within a collagen gel. These results indicate that the matrices, particularly the CGC and DPPA-2 sponges, provide excellent supports for fibroblast growth and the formation of dermal and skin equivalents.Biodegradable scaffolds, along with cells, are important components of most tissue-engineered constructs. In the study, there is a comparison of the behaviour of human fibroblasts cultured for up to six weeks in four different collagen-based three-dimensional matrices, in the form of sponges composed of pure native type I collagen (control), of collagen-GAG-chitosan (CGC) and of collagen cross-linked by two concentrations of diphenylphosphorylazide (DPPA-2 and DPPA-3). Variations in size and weight of the sponges, as well as fibroblast growth and migration, and total protein and collagen synthesis, are determined with time in culture. Owing to their low thermal stability, the partial denaturation and dissolution of the control sponges after incubation at 37 degrees C lead to considerable contraction and low cell proliferation. CGC sponges, stabilised by ionic interactions between the different components, show, after six weeks, limited contraction (20%) and weight increase (10% when seeded) and high cell growth (threefold increase). Similar results are obtained with weakly, cross-linked (DPPA-2) collagen sponges. Highly cross-linked (DPPA-3) sponges do not contract, whereas weight gain and cell proliferation are no different from those found with CGC and DPPA-2 sponges. Similar levels of total protein and collagen synthesis are shown for fibroblasts seeded in different matrices, with a slight general decrease (twofold) after three weeks, a much lower value than that observed with fibroblasts in culture within a contracted collagen gel (sixfold). Furthermore, the fraction of neo-synthesised collagen deposited in the sponges after six weeks represents more than 60% of the total, compared with only 10% obtained with fibroblasts in monolayer culture or 30% within a collagen gel. These results indicate that the matrices, particularly the CGC and DPPA-2 sponges, provide excellent supports for fibroblast growth and the formation of dermal and skin equivalents.