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Title: Effects of methotrexate on human bone cell responses to mechanical stimulation
Author: Elliot, Kerry Jane
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2004
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Mechanical forces influence human bone cell (HBC) metabolism and function. It has been shown that 0.33 Hz cyclical mechanical stimulation results in membrane hyperpolarisation of HBC. Membrane hyperpolarisation follows activation of a mechanotransduction pathway which involves the α5β1 integrin, tyrosine kinase activity, the actin cytoskeleton and interleukin-1β (IL-1β) secretion. IL-1β binds to IL-1 receptors and stimulates a signal cascade involving phospholipase C (PLC) and protein kinase C (PKC), leading to the activation of small conductance Ca2+ activated K+ (SK) channels. Experiments have been performed to further elucidate the mechanotransduction pathways activated following 0.33 Hz cyclical mechanical stimulation. It has been demonstrated that type 1 collagen mRNA levels are increased following 0.33 Hz cyclical mechanical stimulation, whereas bone morphogenic protein-4 (BMP-4) mRNA levels are decreased following 0.33 Hz cyclical mechanical stimulation. The effect of MTX on HBC responses to 0.33Hz cyclical mechanical stimulation was investigated. MTX dose-dependently inhibits the hyperpolarisation response following mechanical stimulation. MTX appears to block mechanically induced IL1-β release, rather than interfering with downstream IL1-β responses. Methylene blue cell adhesion assays have provided evidence to indicate that MTX may have effects on integrin receptors (in particular the β1-integrin subunit), although at present, the exact mechanisms involved are unknown. MTX does not affect mechanically induced changes in type 1 collagen and BMP-4 mRNA levels, which would indicate that the mechanotransduction pathways by which type 1 collagen and BMP-4 gene expression are regulated are not reliant on IL1-β function. MTX is known to affect purine biosynthesis resulting in an overproduction of adenosine. Studies have been performed to investigate the effects of adenosine on HBC responses to 0.33 Hz cyclical mechanical stimulation. It has been demonstrated that adenosine inhibits HBC hyperpolarisation following 0.33 Hz cyclical mechanical stimulation, and that the inhibitory effects of MTX on HBC responses to mechanical stimulation can be prevented by adenosine deaminase (ADA).
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available