Rationale: The mechanisms underlying lipotoxic/diabetic cardiomyopathy remain poorly understood. Saturated fatty acid (SFA) levels, elevated in obesity and type 2 diabetes, induce apoptosis in many cell types including cardiomyocytes. Signaling pathways, including the p38 alpha mitogen-activated kinase (MAPK)-dependent pathway, have been implicated in apoptosis due to a diverse range of insults. Objective: We tested the hypothesis that SFA-induced cardiomyocyte apoptosis is dependent on p38 alpha, activation. Methods and results: Human adult ventricular cardiomyocytes (AC16 cells) were exposed to high physiological levels of palmitate (PA), a SFA. The apoptotic response was measured using annexin-V by flow cytometry, and the p38 alpha-dependent pathway was evaluated using a p38 inhibitor PD169316, and by p38 alpha, small interfering RNA (siRNA) knockdown. PA exposure for 16 h dose-dependently increased apoptosis in AC16 cardiomyocytes (control: 2.6 +/- 0.6%, 150 mu M PA: 3.5 +/- 0.9%, 300 mu M PA: 11.5 +/- 1.6%, n = 4, p < 0.01). PA did not change total p38 alpha, protein levels, but increased p38 alpha, phosphorylation dose-dependently (n = 5, p <0.01). PD169316 tended to reduce PA-induced apoptosis (n = 4, p = 0.05). Specific p38 alpha siRNA markedly reduced the expression of p38 alpha but not p38 beta (n = 3, p < 0.0001), and dose-dependently attenuated PA-induced apoptosis (control siRNA: 7.7 +/- 1.0%, 300 mu M PA: 34.4 +/- 5.0%, 300 mu M PA + 30 pmol siRNA: 23.7 +/- 4.4%, 300 mu M PA + 60 pmol siRNA: 19.7 +/- 2.6%, 300 mu M PA + 120 pmol siRNA: 17.3 +/- 2.8%, n = 4, p < 0.0001). Conclusions: These results demonstrate that PA induces p38 alpha activation, and reducing p38 alpha expression attenuates PA-induced cardiomyocyte apoptosis. Our results support a potential mechanism by which high plasma SFA levels through p38 alpha activation may lead to the development of lipotoxic/diabetic cardiomyopathy. Published by Elsevier Inc.