Objective: The vasoactive peptides arginine vasopressin (AVP) and angiotensin II (Ang II) induce similar second messengers in cultured glomerular mesangial cells, as shown by the rise in intracellular calcium and the activation of phospholipases C and A2. In contrast, AVP is a strong mitogen for cultured rat mesangial cells while Ang II is not. To elucidate the level of signal divergence, we examined the effects of AVP and Ang II on the expression of the immediate early genes c-fos, c-jun and Egr-1, which have been associated with cell growth. We also tested the effect of AVP and Ang II on the induction of the ornithine decarboxylase gene and enzyme in order to examine a process that is induced in cell activation, e.g. it has been associated with the G1 phase after mitogen-receptor binding.
Design and methods: Cellular effects of Ang II and AVP were studied in rat mesangial cells in conventional two-dimensional culture. Proliferation of the mesangial cells was measured by cell-counting and by [3H]-thymidine uptake. The presence of receptors for Ang II and AVP was assessed by measurement of prostaglandin E2 by a radioimmunoassay following stimulation with vasoactive peptides and a receptor-binding assay for Ang II. Messenger (m)RNA levels of c-fos, c-jun Egr-1 and ornithine decarboxylase were determined by Northern blot analysis. Ornithine decarboxylase activity was measured by an enzyme substrate assay.
Results: AVP (10(-7) mol/l) stimulated [3H]-thymidine uptake by 3.7-fold after 48 h and increased mesangial cell counts by 42% (P < 0.05), while Ang II was not mitogenic. Stimulation of resting mesangial cells with AVP (10(-9) to 10(-6) mol/l) caused maximal expression of the immediate early genes after 0.5-1 h, which disappeared after 2-4 h. The relative increases were: c-fos, 15-fold; c-jun, 12-fold; Egr-1, sixfold. Ang II (10(-9) to 10(-6) mol/l) did not induce these genes at any time. In contrast, ornithine decarboxylase mRNA and enzyme activity were induced by both AVP and Ang II.
Conclusions: The results demonstrate that AVP, but not Ang II, is a strong inducer of the immediate early genes c-fos, c-jun and Egr-1 in cultured mesangial cells. We conclude that signalling pathways activated by AVP and Ang II in mesangial cells diverge within 30 min after ligand-receptor binding and proximal to the transient expression of immediate early genes. While mitogenesis of cultured mesangial cells, as effected by AVP, involves the expression of immediate early genes, Ang II-induced non-mitogenic changes in the mesangial cell phenotype do not. The increase in ornithine decarboxylase mRNA and enzyme activity following stimulation with both AVP and Ang II indicates that its induction can occur independently of the immediate early gene expression and mitogenesis.