Effects of PI3-k/Akt short hairpin RNA on proliferation, fibronectin production and synthesis of thrombospondin-1 and transforming growth factor-beta1 in glomerular mesangial cells induced by sublytic C5b-9 complexes.

Abstract

To explore proliferation of glomerular mesangial cells (GMC) and secretion of extracellular matrix (fibronectin induced by sublytic C5b-9 complexes), and then ascertain the role of phosphatidylinositol 3-kinase (PI3-k)/Akt signal pathway in these processes, by using small hairpin RNAs. The expression of cyclin D(2), (3)H-thymidine into DNA and production of fibronectin including thrombospondin-1 and transforming growth factor-beta(1) in the GMCs stimulated by sublytic C5b-9 or transfected with expression vectors of PI3-k and Akt short hairpin RNA or LY294002 (PI3-k inhibitor) were measured by Real-time quantitative polymerase chain reaction (PCR), Western blot, enzyme-linked immunosorbent assay (ELISA) and (3)H-thymidine incorporation ((3)H-TdR), respectively. The expression of cyclin D(2), (3)H-thymidine into DNA and fibronectin in the GMCs stimulated by sublytic C5b-9 could all be increased, and the elevations of these parameters mentioned above were also markedly reduced in the GMCs transfected with vectors of PI3-k and Akt short hairpin RNA or LY294002, respectively. These data indicate that sublytic C5b-9 can promote proliferation of GMCs and secretion of fibronectin as well as synthesis of thrombospondin-1 and transforming growth factor-beta(1). The PI3-k/Akt signal pathway in these reactions, mediated by sublytic C5b-9 complexes, may play at least a partial role.