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Queen Mary University of LondonQueen Mary University of London
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School of Engineering and Materials Science
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PhD Thesis: The in vitro response to simulated intra-articular environment associated with a cell-seeded ligament repair system

Author: PEARSON, Richard

Year: 2000

Supervisor(s): Dan Bader, David Lee

This thesis addresses biological issues associated with the development of a tissue engineering repair system (Bio-ACL). The proposed system consists of a biodegradable poly-L-lactide (PLLA) scaffold seeded with AH88 neonatal fibroblasts, for reconstruction of ruptured anterior cruciate ligaments (ACL).
The soluble PLLA degradation product was primarily composed of L-lactic acid. AH88 cells within monolayers sustained a 48 hour exposure to 28 mM L-lactic acid, with 59% remaining viable. Neutralisation of lactic acid to lactate markedly increased cellular tolerance to equimolar concentrations, with cells remaining viable at 222 mM. The effect of osmolality was also examined, significantly inhibiting proliferation and protein synthesis above 550 mOsmol. Cell seeded collagen lattices exposed to human synovial fluid (HuSF) collected post ACL trauma proliferated at a slower rate than DMEM + 10% FCS control cultures, identified by thymidine incorporation. However protein synthesis, determined by proline incorporation, was upregulated relative to control cultures, indicated by extensive rough endoplasmic reticulum. Maximum concentrations of cytokines, Fibroblastic Growth Factor 2 (FGF-2) and Transforming Growth Factor Beta 1 (TGF 1), were measured in HuSF immediately post-trauma. Conversely, HuSf hyaluronan increased in concentration during 18 months post-trauma. Monolayer cultures were exposed to these constituents at the concentrations measured. TGF 1 markedly increased proliferation and protein synthesis during a 24 hour culture period, whereas FGF-2 produced negligible effects compared to controls. Hyaluronan inhibited proliferation and protein synthesis, although a clear dose response correlation was not evident. These findings have been used to refine the design of the Bio-ACL within the overall programme.