Abstract 528: Cell Specific Signaling Regulated by PI3Kγ Modulates Myofibroblast Differentiation

Abstract

Phosphoinositide 3 Kinase γ (PI3Kγ) regulates anti-apoptotic Akt signaling. Previous studies have established a role for PI3Kγ in cardiac fibrosis, a key underlying cause of heart failure. However, less is known about the mechanism by which PI3Kγ regulates cardiac myofibroblast differentiation, hallmark of tissue fibrosis, characterized by smooth muscle α-actin (αSMA) overexpression. Measurement of αSMA abundance in cardiac lysates from PI3Kγ null mice (PI3Kγ -/- ) showed significant baseline and pressure overload induced upregulation compared to wildtype (WT), indicating that loss of PI3Kγ predisposes the hearts towards fibrosis. Furthermore, isolated cardiac fibroblasts (CF) from PI3Kγ -/- exhibited a myofibroblast phenotype with αSMA in stress fibers. Correspondingly, immunoblotting showed significantly higher αSMA in PI3Kγ -/- CF than WT. It has been previously shown that fibroblast growth factor mediated activation of the signaling pathway downregulates αSMA and that this inhibition of αSMA expression is through negative regulation by extracellular regulated kinase (ERK). To understand whether PI3Kγ regulates ERK signaling in a cell specific manner and thereby possibly αSMA, the phosphorylation of ERK by insulin stimulation were compared in CF isolated from WT and PI3Kγ -/- . Intriguingly, there was significant loss of ERK phosphorylation in CF from PI3Kγ -/- when compared to CF from WT. However, ERK phosphorylation was not altered in cardiomyocytes (CM) due to the absence of PI3Kγ. Confirming this differential regulation of ERK in CM and CF, there was no change in the association of ERK and protein phosphatase 2A (PP2A) in CM of WT and PI3Kγ -/- . However, there was increased association of PP2A with ERK in CF of PI3Kγ -/- when compared to CF from WT. This is because in the CM, ERK is regulated by Dual Specificity Phosphatase (DUSP8). However, in the CF, PP2A is the major regulator of ERK and thus PI3Kγ plays a major role in ERK signaling in CF. Consistent with previous observations, we found that association of DUSP8 with ERK was not observed in CF. Moreover, ERK-DUSP8 interaction was not dependent on the presence or absence of PI3Kγ. These data indicate that PI3Kγ regulates signaling pathways in a cell specific manner with respect to cardiac remodeling.