Activation of mesangial cell signaling cascades in response to mechanical strain

Background. Mesangial cells (MCs) are constantly exposed to pulsatile stret ch and relaxation in their role as architectural support for the glomerulus . There is no cell proliferation in normal glomeruli. In contrast, animal m odels of increased glomerular capillary pressure are characterized by resid ent glomerular cell proliferation and elaboration of extracellular matrix ( ECM) protein, resulting in glomerulosclerosis. This process can be ameliora ted by maneuvers, such as angiotensin converting enzyme inhibition, that re duce glomerular capillary pressure. MCs grown on ECM-coated plates and expo sed to cyclic stretch/relaxation proliferate and produce ECM protein, sugge sting that this may be a useful in vitro model for MC behavior in response to increased physical forces. Previous work has shown induction of c-fos in response to application of mechanical strain to MCs, which may induce incr eases in AP-1 transcription factor activity, which, in turn, may augment EC M protein and transforming growth factor beta transcription and cell prolif eration. Stimuli that lead to c-fos induction pass through mitogen-activate d protein kinase (MAPK) pathways. Three MAPK cascades have been characteriz ed in mammalian cells-p44/42 (classic MAPK), the stress-activated protein k inase/Jun terminal kinase (SAPK/JNK) pathway, and p38/HOG-and mechanical st rain activates p44/42 and SAPK/JNK in cardiac fibroblasts. However, in cont rast to MCs, these cells do not proliferate in response to physical force. Accordingly, we studied activation of the MAPK pathways in MCs exposed to m echanical strain. Methods. MCs (passages 5 to 10) cultured on type 1 collagen-coated, flexibl e-bottom plates were exposed to 30, 60, or 120 minutes of cyclic strain (60 cycles/min) by computer-driven generation of vacuums of -14 and -28 kPa, i nducing 20% and 29% elongations in the diameter of the surfaces, respective ly. Control MCs were grown on coated rigid bottom plates. Proliferation was assessed at 24 hours by H-3-thymidine incorporation. Protein levels (by We stern blot) and activity assays for all three kinase cascades were performe d at 30, 60, and 120 minutes. Results. Cyclic strain/relaxation lead to an approximate doubling of H-3-th ymidine incorporation at 24 hours (N = 3, P < 0.05) only in cultures stretc hed 29%, but not in cultures stretched 20%. At -29% elongation, the increas e in H-3-thymidine incorporation was preceded by early activation of MAPK s ignaling pathways. p44/42 activity increased to a maximum of eightfold grea ter than control at 60 minutes. p38/HOG activity was not measurable at base line but was increased markedly at 30 minutes, which was sustained through to 120 minutes. SAPK/JNK activity was present at a very low level in MCs an d was not changed by stretch. However, it was markedly increased by sorbito l. In MCs stretched to 20% elongation, lesser increases in p44/42 were seen with a similar time course, whereas no increases in p38/HOG or SAPK could be detected at the time points studied. No increase in any kinase pathway a ctivity was seen at any time in static cultures. Conclusions. High-pressure cyclic stretch leads to MC proliferation, preced ed by marked activation of p44/42 and p38/HOG MAPKs. Cell proliferation is not seen with low-pressure stretch, and there is only modest p44/42 MAPK ac tivation, suggesting that glomerular capillary hypertension may lead to cel l proliferation and injury partly through differential activation of kinase cascades.