CREATION OF AN IN-VIVO CYTOSENSOR USING ENGINEERED MESANGIAL CELLS - AUTOMATIC SENSING OF GLOMERULAR INFLAMMATION CONTROLS TRANSGENE ACTIVITY

Automatic control over exogenous gene expression in response to the ac tivity of disease is a crucial hurdle for gene transfer-based therapie s. Towards achieving this goal, we created a ''cytosensor'' that perce ives local inflammatory states and subsequently regulates foreign gene expression, alpha-Smooth muscle actin is known to be expressed in glo merular mesangial cells exclusively in pathologic situations. CArG box element, the crucial regulatory sequence of the alpha-smooth muscle a ctin promoter, was used as a sensor for glomerular inflammation. Rat m esangial cells were stably transfected with an expression plasmid that introduces a beta-galactosidase gene under the control of CArG box el ements. In vitro, the established cells expressed beta-galactosidase e xclusively after stimulation with serum, To examine whether the cells are able to automatically control transgene activity in vivo, serum-st imulated or unstimulated cells were transferred into normal rat glomer uli or glomeruli subjected to anti-Thy 1 glomerulonephritis. When stim ulated cells were transferred into the normal glomeruli, beta-galactos idase expression was switched off in vivo within 3 d, In contrast, whe n unstimulated cells were transferred into the nephritic glomeruli, tr ansgene expression was substantially induced. These data indicate the feasibility of using the CArG box element as a molecular sensor for gl omerular injury. In the context of advanced forms of gene therapy, thi s approach provides a novel concept for automatic regulation of local transgene expression where the transgene is required to be activated d uring inflammation and deactivated when the inflammation has subsided.