Inhibitory effect of antisense oligodeoxynucleotide to p44/p42 MAPK on angiotensin II-induced hypertrophic response in cultured neonatal rat cardiac myocyte.
OBJECTIVE
To explore the inhibitory effect of antisense oligonucleotide (ODN) to mitogen activated protein kinase(MAPK) on cardiomyocyte hypertrophy induced by angiotensin II (Ang II).


METHODS
A 17-mer phosphorothioate-protected antisense ODN directed against the initiation of translation sites of the p42 and p44 MAPK isoforms by liposomal transfection was applied to inhibit the translation of p44/p42 MAPK mRNA. The sense and random ODNs to p44/p42MAPK were used as sequence controls. Neonatal cardiac myocytes were exposed to Ang II (10 nmol/L) for 5 min and then harvested in lysis buffer for the measurement of the activity and the phosphorylated protein content of p44/p42MAPK that were tested by P-81 phosphocellulose filter paper method and Western blotting, respectively. The rate of protein synthesis by [3H]leucine incorporation and the diameter of cell were measured after exposure to Ang II for 24 h and 72 h, respectively.


RESULTS
In cardiac myocyte Ang II increased p44/p42 MAPK activity and phosphorylated protein content by 140 % and 699 %, and also increased [3H]leucine incorporation and cell diameter by 40 % and 27 %. c-fos and c-myc mRNAs were induced significantly after exposure to Ang II. Antisense ODN to p44/p42MAPK (0.2 micromol/L) reduced Ang II-induced MAPK activity by 30 %, and phophorylated MAPK protein expression by 59 % in cardiac myocyte, and inhibited c-fos and c-myc mRNA expression induced by Ang II by 44 % and 43 %, respectively. The diameter and the rate of protein synthesis of cardiac myocyte induced by Ang II were decreased by 16 % and 22 % after pretreatment with antisense ODN to p44/p42MAPK.


CONCLUSIONS
Antisense ODN to p44/p42 MAPK inhibited the increase of rate of protein synthesis, and the augmentation of cell diameter and expression of c-fos and c-myc mRNA induced by Ang II in cultured cardiac myocytes. p44/p42 MAPK played a critical role in the hypertrophic response induced by Ang II in cultured neonatal rat cardiac myocytes.
Zhang
Shi-Qin
SQ
Laboratory of Molecular Pharmacology, Xiang Ya School of Medicine, Central South University, Changsha 410078, China. zhangshiqintj@hotmail.com
Ding
Bo
B
Guo
Zhao-Gui
ZG
Li
Yun-Xia
YX
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D006332:Cardiomegaly / Q000473:pathology; D002478:Cells, Cultured; D015870:Gene Expression / Q000187:drug effects; D016762:Genes, fos; D016259:Genes, myc; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar
D020319:Oligodeoxyribonucleotides, Antisense; D012333:RNA, Messenger; D000804:Angiotensin II; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases