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Cardiac myofibroblasts (CMF) play a key role in infarct repair and scar formation following myocardial infarction (MI) and are also an important source of proinflammatory cytokines. We postulated that interleukin-1[alpha] (IL-1[alpha]), a potential early trigger of acute inflammation post-MI, could stimulate human CMF to express additional proinflammatory cytokines. Furthermore, we hypothesized that these effects may be modulated by the anti-inflammatory cytokine interleukin-10 (IL-10). Human CMF were cultured from atrial biopsies from multiple patients. Interleukin-1[beta] (IL-1[beta]), tumor necrosis factor-[alpha] (TNF-[alpha]), interleukin-6 (IL-6), and cardiotrophin-I (CT-1) mRNA expression and secretion were measured using quantitative real-time RT-PCR and enzyme-linked immunosorbent assay. IL-1[alpha] (0.001-10 ng/ml, 0-6 h) stimulated IL-1[beta], TNF-[alpha], and IL-6 mRNA expression with distinct temporal and concentration profiles, resulting in increased cytokine secretion. The response to IL-1[alpha] was much greater than with TNF-[alpha]. Neither IL-1[alpha] nor TNF-[alpha] treatment modulated CT-1 mRNA expression. Immunoblotting with phosphospecific antibodies revealed that IL-1[alpha] stimulated the extracellular signal-regulated kinase (ERK)-1/2, p38 mitogen-activated protein kinase (MAPK), c-Jun N[H.sub.2]-terminal kinase (JNK), phosphatidylinositol 3-kinase (PI 3-kinase)/protein kinase B (Akt), and nuclear factor (NF)-[kappa]B signaling pathways. Pharmacological inhibitor studies indicated roles for PI 3-kinase/Akt and NF-[kappa]B pathways in mediating IL-1[beta] expression, and for NF-[kappa]B and p38 MAPK pathways in mediating TNF-[alpha] expression. IL-1[alpha]-induced IL-6 mRNA expression was reduced by p38 MAPK inhibition, but increased by ERK and JNK pathway inhibitors. IL-10 produced a consistent but modest reduction in IL-1[alpha]-induced IL-6 mRNA levels (not IL-1[beta] or TNF-[alpha]), but this was not reflected by reduced IL-6 protein secretion. In conclusion, IL-1[alpha] stimulates human CMF to express IL-1[beta], TNF-[alpha], and IL-6 via specific signaling pathways, responses that are unaffected by IL-10 exposure. cardiac fibroblasts; inflammation; signal transduction; cytokines; interleukin-10