Increased myocardial collagen content in transgenic rats overexpressing cardiac angiotensin-converting enzyme is related to enhanced breakdown of N-acetyl-Ser-Asp-Lys-Pro and increased phosphorylation of Smad2/3.
BACKGROUND
Although increased activity of angiotensin-converting enzyme (ACE) has been associated with increased cardiac collagen, no studies to date have established a direct cause-and-effect relation between the two.


RESULTS
We used transgenic rats that overexpress human ACE selectively in the myocardium. Two independent heterozygous transgenic rat lines were studied, one expressing 2 to 3 copies (L1172) and the other expressing 5 to 10 copies (L1173) of the ACE transgene. These rats were normotensive but developed a proportionate increase in myocardial collagen depending on the ACE gene dose (up to 2.5-fold, P<0.01), but cardiac angiotensin II levels remained normal, whereas collagen content reversed to control levels on ACE inhibition. To explain these changes, we investigated N-acetyl-Ser-Asp-Lys-Pro (AcSDKP), an alternative substrate that is catabolized exclusively by ACE. Increased cardiac expression of ACE was paralleled by a reciprocal decrease in cardiac AcSDKP and a proportionate increase in phosphorylated Smad2 and Smad3, all of which normalized after both ACE inhibition and AcSDKP infusion. Furthermore, a functional link of this signaling cascade was demonstrated, because AcSDKP inhibited Smad3 phosphorylation in a dose-dependent manner in cultured cardiac fibroblasts and in vivo.


CONCLUSIONS
Our findings suggest that increased cardiac ACE activity can increase cardiac collagen content by degradation of AcSDKP, an inhibitor of the phosphorylation of transforming growth factor-beta signaling molecules Smad2 and Smad3. This implies that the antifibrotic effects of ACE inhibitors are mediated in part by increasing cardiac AcSDKP, with subsequent inhibition of Smad 2/3 phosphorylation.
Pokharel
Saraswati
S
Experimental and Molecular Cardiology, Cardiovascular Research Institute Maastricht (CARIM), University Hospital Maastricht, Maastricht, The Netherlands.
van Geel
Peter P
PP
Sharma
Umesh C
UC
Cleutjens
Jack P M
JP
Bohnemeier
Holger
H
Tian
Xiao-Li
XL
Schunkert
Heribert
H
Crijns
Harry J G M
HJ
Paul
Martin
M
Pinto
Yigal M
YM
D030201:Activin Receptors, Type I / Q000378:metabolism; D000804:Angiotensin II / Q000378:metabolism; D000818:Animals; D030801:Animals, Genetically Modified; D001667:Binding, Competitive; D003094:Collagen / Q000032:analysis*; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005355:Fibrosis; D006801:Humans; D017379:Hypertrophy, Left Ventricular / Q000517:prevention & control; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology; D009842:Oligopeptides / Q000378:metabolism / Q000494:pharmacology*; D007703:Peptidyl-Dipeptidase A / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases; D051381:Rats; D017207:Rats, Sprague-Dawley; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism; D051899:Smad2 Protein; D051900:Smad3 Protein; D013379:Substrate Specificity; D015534:Trans-Activators / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000378:metabolism; D053773:Transforming Growth Factor beta1
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D009842:Oligopeptides; D018125:Receptors, Transforming Growth Factor beta; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; D051899:Smad2 Protein; C497784:Smad2 protein, rat; D051900:Smad3 Protein; C497787:Smad3 protein, rat; C508960:TGFB1 protein, human; C508962:Tgfb1 protein, rat; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D000804:Angiotensin II; D003094:Collagen; D017346:Protein Serine-Threonine Kinases; D030201:Activin Receptors, Type I; D000077293:Receptor, Transforming Growth Factor-beta Type I; C000625843:Tgfbr1 protein, rat; D007703:Peptidyl-Dipeptidase A; C058504:goralatide