kn-62 and Cardiomegaly

Endothelin-1 (ET-1) induces cardiac hypertrophy. Because Ca(2+) is a major second messenger of ET-1, the role of Ca(2+) in ET-1-induced hypertrophic responses in cultured cardiac myocytes of neonatal rats was examined. ET-1 activated the promoter of the beta-type myosin heavy chain gene (beta-MHC) (-354 to +34 base pairs) by about 4-fold. This activation was inhibited by chelation of Ca(2+) and the blocking of protein kinase C activity. Similarly, the beta-MHC promoter was activated by Ca(2+) ionophores and a protein kinase C activator. beta-MHC promoter activation induced by ET-1 was suppressed by pretreatment with the calmodulin inhibitor, W7, the Ca(2+)/calmodulin-dependent kinase II (CaMKII) inhibitor, KN62, and the calcineurin inhibitor, cyclosporin A. beta-MHC promoter activation by ET-1 was also attenuated by overexpression of dominant-negative mutants of CaMKII and calcineurin. ET-1 increased the activity of CaMKII and calcineurin in cardiac myocytes. Pretreatment with KN62 and cyclosporin A strongly suppressed ET-1-induced increases in [(3)H]phenylalanine uptake and in cell size. These results suggest that Ca(2+) plays a critical role in ET-1-induced cardiomyocyte hypertrophy by activating CaMKII- and calcineurin-dependent pathways.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Animals; Animals, Newborn; Calcimycin; Calcineurin; Calcium; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases; Cardiomegaly; Cells, Cultured; Endothelin-1; Enzyme Inhibitors; Gene Expression Regulation; Heart; Ionomycin; Kinetics; Models, Cardiovascular; Myocardium; Myosin Heavy Chains; Promoter Regions, Genetic; Rats; Rats, Wistar; Sulfonamides; Tetradecanoylphorbol Acetate; Transfection

This study investigates whether leukemia inhibitory factor (LIF), a potent cardiac hypertrophic cytokine, affects the L-type Ca2+ current (I(Ca,L)) and intracellular Ca2+ concentrations ([Ca2+]i) in cardiomyocytes. I(Ca,L) was recorded using a whole cell patch clamp configuration in guinea pig cardiomyocytes, and the [Ca2+]i transient was detected by use of Fluo-3 in rat cardiomyocytes. Cells were preincubated with LIF (1000 U/ml) for 15 min before whole cell recording. LIF increased I(Ca,L) by 41.8%. LIF synergistically increased I(Ca,L) with isoproterenol. Preincubation with H89 did not inhibit the LIF-induced increase in I(Ca,L), indicating that this phenomenon is PKA-independent. PD98059 completely inhibited the increase in I(Ca,L), and this effect was dose-dependent (IC50=3.6 micromol/l). Other signal transduction inhibitors including AG490, SB203580, chelerythrine, genistein, and KN62 did not affect the LIF-induced increase in I(Ca,L). Perforated patch clamp recording revealed that LIF maximally increased the I(Ca,L) by 25% at 15 min. LIF also increased the peak [Ca2+]i transient level by 63% at 15 min. PD98059 fully inhibited the increase in the [Ca2+]i transient. In conclusion, LIF increased I(Ca,L) and the [Ca2+]i transient in cardiomyocytes, and the Raf-1/MEK/ERK pathway might be involved in the modulation of this activation.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Animals; Calcium; Calcium-Calmodulin-Dependent Protein Kinases; Cardiomegaly; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cytokines; Dose-Response Relationship, Drug; Enzyme Inhibitors; Flavonoids; Genistein; Growth Inhibitors; Guinea Pigs; Heart; Interleukin-6; Leukemia Inhibitory Factor; Lymphokines; Patch-Clamp Techniques; Rats; Rats, Sprague-Dawley; Signal Transduction; Time Factors