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Background: Angiotensin II (Ang II) plays an important role in extracellular matrix deposition and tissue scarring in the kidney and the heart. The mechanism for extracellular matrix stimulation by Ang II is currently hypothetical, with one possibility pointing to a direct effect on cell synthesis of specific collagens., Methods: We studied the molecular mechanism for activation of type III collagen synthesis by Ang II in an in vitro cell model of myofibroblasts by evaluating (1) alpha1(III) collagen mRNA expression; (2) alpha1(III) collagen promoter activity; (3) DNA/protein binding with characterization of binding sites; (4) expression of transcription factors; and (5) the role of a short DNA segment as Ang II responsive element., Results: We found a specific dose-dependent stimulation of alpha1(III) collagen mRNA expression and a parallel effect on alpha1(III) collagen promoter activity. Transfection of constructs containing alpha1(III) collagen promoter fragments of different lengths localized the site of activation within the shortest 178 bp construct. By gel-retardation experiments, we observed the formation of a DNA-protein complex with crude extracts from Ang II-stimulated cells and an oligonucleotide spanning the 3 to 20 sequence. This complex was due to a sequence-specific interaction and was abolished by a 3 bp substitution mutation. The introduction of this mutation into the 178 bp construct abolished the stimulatory effect of Ang II., Conclusions: These results demonstrate that Ang II stimulates the expression of alpha1(III) collagen mRNA in myofibroblasts in vitro by activating the alpha1(III) collagen promoter at the level of a factor recognition site localized immediately downstream of the transcription start site. This mechanism could be involved in Ang II-induced renal and heart fibrosis.

The presence of isoalbumins with a less anionic charge than the normal protein (pI = 4.7) is the hallmark of proteinuria in minimal change nephropathy (MCN). Steroid-induced restoration of near normal levels of proteinuria is characterized by the appearance in urines of isoalbumins with a pI still more anionic than the normal. In our search for an explanation for the pI changes, we used preparative isoelectric focusing in granulated gels to split the microheterogeneous bands obtained from nine MCN-affected children into four fractions (A1, A2, A3, A4) with decreasing pI from 5.8 to 4.0 and we have determined their fatty acid content. The least anionic fraction, A1, was the most defatted, followed by A2, A3 and A4 in which fatty acid content progressively increased, A4 being the most fatted fraction. Accordingly, the mean content of fatty acids in urinary albumin in proteinuric children was lower than in both the remission phase and in normal children (2.17 +/- 0.03 vs. 20.91 +/- 0.38 and 20.94 +/- 0.39, respectively) and was lower by a factor of 4 compared to serum albumin in the same phase of the disease (2.17 +/- 0.03 vs. 8.59 +/- 1.64). Among medium and long-chain fatty acids, the ratio between serum and urinary albumin was the highest for linoleic acid (approximately 7), followed by that of oleic acid, palmitic acid and lauric acid. At variance in five other patients affected by non-MCN nephrotic syndrome this ratio was for practically all FAs about 1.(ABSTRACT TRUNCATED AT 250 WORDS)