Modulation of macrophage phenotype using adenoviral transfection
The initial aim of this study was to examine the nature of the interaction between adenovirus and transfected macrophage, and to characterise the molecular mechanisms underlying macrophage response to adenoviral infection. Results showed that adenoviral transfection activated macrophages, promoting production of pro-inflammatory mediators and priming an enhanced response to other inflammatory stimuli. Activation was dependent on the nuclear factor kappa B (NFkB) signalling pathway, which was activated within two hours of transfection, and could be prevented using an inhibitory adenovirus which blocked NFkB signalling. The ultimate phenotype of the transfected macrophage was determined both by non-specific viral activation and by the properties of the transgene expressed. The second aim of this research was to investigate the effect of the local cytokine milieu on transgene expression in vitro. Results showed that transgene expression under the control of two different promoter constructs was subject to regulation by pro-inflammatory mediators, by mechanisms at least partly dependent on the NFkB signalling pathway. the results have important implications for the use of these promoters in gene therapy applications where the gene of interest is delivered into an inflammatory environment. The final stage of the project focused on the use of transfected macrophages in vivo, in a rat model of glomerulonephritis. Results showed that transfected macrophages expressing the anti-inflammatory cytokine IL-4 localised with enhanced efficiency to inflamed glomeruli after intra-renal artery injection of the mechanisms for this were examined. Better understanding of the mechanisms which promote localisation may ultimately permit the design of genetically-modified macrophages which selectively target sites of injury for delivery of anti-inflammatory cytokines.