Mesangial cell signaling cascades in response to mechanical strain and glucose

Background. Elevated glucose levels and glomerular hypertension (Pgc) are c onsidered to contribute to the elaboration of matrix protein by mesangial c ells (MCs) in diabetic glomeruli. MCs grown in 30 mM of glucose produce exc essive matrix protein, as do MCs exposed to cyclic strain, and the combinat ion of the two exacerbates this. Tight glucose control or reduction in Pgc clinically delays progression of diabetic nephropathy. MC c-fos is induced in response to either application of strain or high ambient glucose, induci ng increases in activated protein-1 transactivational activity and extracel lular matrix production. Stimuli that lead to c-fos induction pass through the three mitogen-activated protein (MAP) kinase pathways: p44/42, SAPK/JNK , and p38/HOG. We studied MAP kinase activation in MCs exposed to mechanica l strain and a high glucose. Methods. MCs (passage 5 through 10) cultured for 96 hours on type 1 collage n-coated flexible-bottom plates in either 5.6 or 30 mM glucose were exposed to 5, 10, or 30 minutes of cyclic strain (60 cycles per min) by computer-d riven generation of vacuums of -14 kPa, inducing 20% elongation in the diam eter of the surface. Control MCs were grown on both coated rigid and flexib le-bottom plates. Protein levels (by Western blot) and activity assays for all three kinase cascades were performed at baseline and after 5, 10, and 3 0 minutes. All experiments were performed in triplicate. Results. MAP kinase signaling was seen in response to stretch, and high amb ient glucose affected the pattern of activation. Both p44/42 and p38HOG kin ase activities showed small increases to a maximum of 2.5- to 3.5-fold grea ter than static MCs at 10 minutes. Activity in both kinase cascades was sli ghtly suppressed by 30 mM glucose. In contrast, SAPK/JNK activity was prese nt at a very low level in static MCs and increased markedly by 10 minutes o f stretch. Thirty micromolars of glucose augmented this effect by a factor of six over MCs cultured in 5.6 mM glucose after 10 minutes of stretch. Nei ther glucose concentration nor mechanical strain had any effect on the prot ein expression of any of the kinases by Western blot. Conclusions. MAP kinase cascade signaling is seen when physical force is ap plied to MCs, and glucose affects the pattern of activity. Thirty micromola rs of glucose markedly increase the level of SAPK/JNK activation. This may have implications in diabetic signal transduction and matrix protein produc tion.