Thromboxane A2 regulates protein synthesis of cultured human mesangial cells

Cultured human glomerular mesangial cells express functional receptors for vasoconstrictor metabolites of arachidonate, such as thromboxane A2. Binding of thromboxane A2 analogs triggers phosphatidylinositol breakdown and a rise of intracellular free Ca2+, followed by complex effects on cell growth. We have evaluated the effects of the thromboxane A2 mimetic, U-46619, on the biosynthesis of human mesangial cell proteins. After 4 to 24 hours of incubation with U-46619, cells were metabolically labeled with tritiated leucine or sulfur 35-methionine and analyzed by one-dimensional and two-dimensional sodium dodecylsulfate polyacrylamide gel electrophoresis and fluorography. U-46619 modestly stimulated total protein synthesis in both quiescent and cycling cells to a maximum of 12% and 23% above control at 1 mumol/L after 24 hours, respectively. The eicosanoid selectively enhanced the labeling of crude membrane and cytosolic proteins of molecular weights of approximately 38 to 53 kd and 125 to 200 kd in the presence or absence of serum, respectively. Cellular tubulin and collagen type IV, but not actin, were markedly stimulated, as determined by immunoblotting of cell-associated proteins. On the contrary, U-46619 potently inhibited labeling of soluble proteins that were released into the media, to a maximum of 49% after 24 hours. Inhibition was confirmed by immunoblotting of collagen type IV. Intraglomerular accumulation of thromboxane A2 during chronic inflammation may modify the functional characteristics of the mesangium by regulation of the biosynthesis of structural proteins.