Abstract
After trauma, articular cartilage often does not heal due to incomplete bonding of the fractured surfaces. In this study we investigated the ability of chemical cross-linkers to facilitate bonding of articular cartilage, either alone or in combination with a pre-treatment with surface-degrading agents. Articular cartilage blocks were harvested from the femoropatellar groove of bovine calves. Two cartilage blocks, either after pre-treatment or without, were assembled in a custom-designed chamber in partial apposition and subjected to cross-linking treatment. Subsequently, bonding of cartilage was measured as adhesive strength, that is, the maximum force at rupture of bonded cartilage blocks divided by the overlap area. In a first approach, bonding was investigated after treatment with cross-linking reagents only, employing glutaraldehyde, 1-ethyl-3-diaminopropyl-carbodiimide (EDC)/N-hydroxysuccinimide (NHS), genipin, or transglutaminase. Experiments were conducted with or without compression of the opposing surfaces. Compression during cross-linking strongly enhanced bonding, especially when applying EDC/NHS and glutaraldehyde. Therefore, all further experiments were performed under compressive conditions. Combinations of each of the four cross-linking agents with the degrading pre-treatments, pepsin, trypsin, and guanidine, led to distinct improvements in bonding compared to the use of cross-linkers alone. The highest values of adhesive strength were achieved employing combinations of pepsin or guanidine with EDC/NHS, and guanidine with glutaraldehyde. The release of extracellular matrix components, that is, glycosaminoglycans and total collagen, from cartilage blocks after pre-treatment was measured, but could not be directly correlated to the determined adhesive strength. Cytotoxicity was determined for all substances employed, that is, surface degrading agents and cross-linkers, using the resazurin assay. Taking the favourable cell vitality after treatment with pepsin and EDC/NHS and the cytotoxic effects of guanidine and glutaraldehyde into account, the combination of pepsin and EDC/NHS appeared to be the most advantageous treatment in this study. In conclusion, bonding of articular cartilage blocks was achieved by chemical fixation of their surface components using cross-linking reagents. Application of compressive forces and prior modulation of surface structures enhanced cartilage bonding significantly. Enzymatic treatment in combination with cross-linkers may represent a promising addition to current techniques for articular cartilage repair.
Highlights
After trauma, articular cartilage often does not heal due to incomplete bonding of the fractured surfaces
Adhesive strength of the bonding area At first, cartilage blocks of the two different geometries were fixed in partial apposition in the custom-made chamber
The cross-linking reagents EDC/NHS and glutaraldehyde led to measurable bonding of cartilage blocks in the absence of compressive strain, large increases in adhesive strength were achieved by additional compressive load during the bonding procedure
Summary
Articular cartilage often does not heal due to incomplete bonding of the fractured surfaces. Steroid hormone effect [5], and the synovial fluid flow, which might act at the interface in joint motion to keep the surfaces apart [7]. Based on these findings, therapeutic options for articular cartilage integration have been investigated. Collagen cross-linking has been stimulated over time in vitro [8] or articular cartilage surfaces have been degraded in order to stimulate repair in vitro [9,10,11,12] as well as in vivo [13]. Physical swelling of surface structures by guanidine was reported to stimulate the integrative repair process [10]
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