Abstract
Background
Investigations into the response to implant debris tend to concentrate on how a population of cells proliferate in the presence of implant material, and how the regulation of cytokines change. For the problem of cobalt-chromium (CoCr) implants this has been done for osteoblasts and osteoclasts to understand how bone resorption, leading to aseptic loosening, is occurring. However, investigating the formation of the extracellular-matrix (ECM) may give a better indication of the mechanisms occurring. ECM is excreted from cells and is important for adhesion, structure, signaling and growth. Type I collagen is the most abundant protein in the ECM and is known to direct tissue development and is therefore a key part of understanding the mechanism behind aseptic loosening.
Methods
3T3-fibroblasts were seeded in Dulbecco's Modified Eagle Medium (DMEM) and supplemented with 100mM ascorbic acid. Every 48hours cells were fed with DMEM and doped with Co and Cr ions until fixation. Sirius Red dye was used to bind to the type I collagen, then removed using NaOH and analysed using UV absorption to show relative amounts of collagen. Type I collagen gel was formed in the presence of Co and Cr ions with and without DMEM and the fibers were imaged using AFM.
Results and Conclusions
Studies show a decrease in the collagen output of 3T3-fibroblasts with increasing concentration of Co ions, and low concentrations of Cr ions. Precipitation was also seen for Cr ions in the presence of DMEM which may explain the reduced effect of higher concentration of Cr ions in comparison to Co ions. Co and Cr ions do influence the ECM which will affect the bone re-modelling process. Further work will be performed to link these results to mineralisation mechanisms to quantitatively determine the impact of metal ion debris on the ECM and collagen fiber formation.
