This study focuses on determining the effect of varying the composition and crosslinking of collagen-based films on their physical properties and interaction with myoblasts. Films composed of collagen or gelatin and crosslinked with a carbodiimide were assessed for their surface roughness and stiffness. These samples are significant because they allow variation of physical properties as well as offering different recognition motifs for cell binding. Cell reactivity was determined by the ability of myoblastic C2C12 and C2C12-α2+ cell lines (with different integrin expression) to adhere to and spread on the films. Significantly, crosslinking reduced the cell reactivity of all films, irrespective of their initial composition, stiffness or roughness. Crosslinking resulted in a dramatic increase in the stiffness of the collagen film and also tended to reduce the roughness of the films (Rq = 0.417 ± 0.035 μm, E = 31 ± 4.4 MPa). Gelatin films were generally smoother and more compliant than comparable collagen films (Rq = 7.9 ± 1.5 nm, E = 15 ± 3.1 MPa). The adhesion of α2-positive cells was enhanced relative to the parental C2C12 cells on collagen compared with gelatin films. These results indicate that the detrimental effect of crosslinking on cell response may be due to the altered physical properties of the films as well as a reduction in the number of available cell binding sites. Hence, although crosslinking can be used to enhance the mechanical stiffness and reduce the roughness of films, it reduces their capacity to support cell activity and could potentially limit the effectiveness of the collagen-based films and scaffolds.

No abstract

The structure of an ideal scaffold for tendon regeneration must be designed to provide a mechanical, structural and chemotactic microenvironment for native cellular activity to synthesize functional (i.e. load bearing) tissue. Collagen fibre scaffolds for this application have shown some promise to date, although the microstructural control required to mimic the native tendon environment has yet to be achieved allowing for minimal control of critical in vivo properties such as degradation rate and mass transport. In this report we describe the fabrication of a novel multi-fibre collagen fascicle structure, based on type-I collagen with failure stress of 25-49 MPa, approximating the strength and structure of native tendon tissue. We demonstrate a microscopic fabrication process based on the automated assembly of type-I collagen fibres with the ability to produce a controllable fascicle-like, structural motif allowing variable numbers of fibres per fascicle. We have confirmed that the resulting post-fabrication type-I collagen structure retains the essential phase behaviour, alignment and spectral characteristics of aligned native type-I collagen. We have also shown that both ovine tendon fibroblasts and human white blood cells in whole blood readily infiltrate the matrix on a macroscopic scale and that these cells adhere to the fibre surface after seven days in culture. The study has indicated that the synthetic collagen fascicle system may be a suitable biomaterial scaffold to provide a rationally designed implantable matrix material to mediate tendon repair and regeneration.

Tissue engineering is a promising technique for cartilage repair, but to optimize novel scaffolds before clinical trials, it is necessary to determine their characteristics for binding and release of growth factors. Toward this goal, a novel, porous collagen-glycosaminoglycan scaffold was loaded with a range of concentrations of insulinlike growth factor-1 (IGF-1) to evaluate its potential as a controlled delivery device. The kinetics of IGF-1 adsorption and release from the scaffold was demonstrated using radiolabeled IGF-1. Adsorption was rapid, and was approximately proportional to the loading concentration. Ionic bonding contributed to this interaction. IGF-1 release was studied over 14 days to compare the release profiles from different loading groups. Two distinct phases occurred: first, a burst release of up to 44% was noted within the first 24 h; then, a slow, sustained release (13%-16%) was observed from day 1 to 14. When the burst release was subtracted, the relative percentage of remaining IGF-1 released was similar for all loading groups and broadly followed t ½ kinetics until approximately day 6. Scaffold cross-linking using dehydrothermal treatment did not affect IGF-1 adsorption or release. Bioactivity of released IGF-1 was confirmed by seeding scaffolds (preadsorbed with unlabeled IGF-1) with human osteoarthritic chondrocytes and demonstrating increased proteoglycan production in vitro.

To deliver collagen implants as an open array of fibres allows optimal tendon-implant integration and good ingrowth of regenerated tissue. In the present study the resorption rate of both the examined implants was too low due to the high level of crosslinking. This led to only minor substitution of the implant with regenerated tissue, which in turn produced a low-strength implanted region. Further studies are needed to find the right balance between strength and resorption rate of collagen fibres.