Contribution of PI 3-kinase isoforms to angiotensin II- and alpha-adrenoceptor-mediated signalling pathways in cardiomyocytes.
OBJECTIVE
Angiotensin II stimulation increases the formation of reactive oxygen species (ROS), the phosphorylation of p38 mitogen-activated protein kinase (MAPK), and the expression of transforming growth factor beta (TGFbeta) in adult cardiomyocytes. The aim of this study was to determine the involvement of PI 3-kinase and to specify the participation of different isoforms in the angiotensin II-induced formation of ROS in comparison to the hypertrophic pathway triggered by alpha-adrenoceptor stimulation.


METHODS
Freshly isolated myocytes were used to examine formation of ROS via H(2)DCF fluorescence. p38 MAPK phosphorylation, p70(S6)-kinase phosphorylation, PI 3-kinase, and TGFbeta expression were measured by Western blotting. Sense and antisense oligonucleotides were used to down-regulate diverse PI 3-kinase isoforms. Hypertrophy was measured by (14)C-phenylalanine incorporation and cell volume.


RESULTS
Inhibition of PI 3-kinase by Ly294002 or wortmannin, two inhibitors, decreased formation of ROS, phosphorylation of p38 MAPK, and TGFbeta expression. Down-regulation of the p110beta isoform by antisense oligonucleotides inhibited the angiotensin II-induced signalling pathway but not the alpha-adrenoceptor-mediated hypertrophic growth of cardiomyocytes. In contrast, down-regulation of the p110alpha isoform decreased the alpha-adrenoceptor-mediated hypertrophic growth of cardiomyocytes but did not affect the angiotensin II-mediated signalling pathway.


CONCLUSIONS
Thus, our study identifies an involvement of PI 3-kinase in the angiotensin II-induced formation of ROS and provides a biochemical basis for ligand-specific responses for angiotensin II and alpha-adrenoceptor stimulation as relates to hypertrophy.
Wenzel
Sibylle
S
Physiologisches Institut, Justus-Liebig-Universität, Aulweg 129, D-35392 Giessen, Germany. sibylle.wenzel@phsiologie.med.uni-giessen.de
Abdallah
Yaser
Y
Helmig
Simone
S
Schäfer
Claudia
C
Piper
Hans Michael
HM
Schlüter
Klaus-Dieter
KD
D000804:Angiotensin II / Q000378:metabolism*; D000818:Animals; D006332:Cardiomegaly / Q000378:metabolism*; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008297:Male; D008856:Microscopy, Fluorescence; D009025:Morpholines / Q000494:pharmacology; D032383:Myocytes, Cardiac / Q000378:metabolism*; D010649:Phenylalanine / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000032:analysis / Q000378:metabolism; D011942:Receptors, Adrenergic, alpha / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism; D015398:Signal Transduction; D016212:Transforming Growth Factor beta / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D000804:Angiotensin II; D002867:Chromones; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D009025:Morpholines; D010649:Phenylalanine; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017382:Reactive Oxygen Species; D011942:Receptors, Adrenergic, alpha; D019893:Ribosomal Protein S6 Kinases; D016212:Transforming Growth Factor beta; D048051:p38 Mitogen-Activated Protein Kinases; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one