Abstract 9739: MicroRNA26 Regulation of TRPC3 Subunits Underlies Profibrillatory Fibroblast Activation in a Canine Atrial Fibrillation Model
Background: Cardiac fibroblast (FB) proliferation and differentiation into extracellular matrix (ECM) secreting myofibroblasts contributes to AF-promoting fibrotic remodeling. We previously observed that Ca2+ entry through transient receptor potential canonical 3 (TRPC3) channels regulates cardiac FB proliferation/differentiation. Here, we examined the potential involvement of TRPC3 in a canine AF model.
Methods: Dogs kept continuously in AF for 1 week by pacing at 600 bpm (n=11) were compared to controls (CTL, n=11). Echocardiographic, hemodynamic and electrophysiological properties were studied in vivo. Left atrial (LA) FBs were isolated for nonselective cation current (INSC) recording (patch clamp), cell cycle analysis (flow cytometry), protein (immunoblot) and mRNA (qPCR) expression.
Results: AF increased LA diastolic area by 13%* (*p<0.05 vs CTL) and LA pressure by 150%* without LV dilation. AF also shortened atrial refractory periods from 98±4 to 79±5 ms* (BCL 150 ms) and prolonged spontaneously maintained AF duration from 26±8 to 937±209 s*. In freshly isolated FBs, INSC sensitive to a TRPC3 selective blocker, pyrazole3 (Pyr3, 3 μ M) increased in AF by 89%*, indicating increased TRPC3 current. In freshly cultured FBs, the cell number increase-rate, the G2/M cell content (an index of cell division) and α-smooth muscle actin protein expression increased in AF by 185*, 33* and 154* % respectively, indicating increased proliferation and differentiation; in vitro treatment with Pyr3 (3 μ M) decreased these profibrotic indices by 78%*, 12%* and 74%* vs vehicle respectively. In fresh FBs, collagen-1 and -3 gene expression increased in AF by 1210%* and 750%* respectively, indicating increased ECM production. TRPC3 protein expression in fresh FBs increased in AF by 64%*; however, TRPC3 mRNA decreased by 44%*, suggesting epigenetic regulation by microRNA (miR). AF reduced FB expression of miR-26a, predicted to target and suppress TRPC3 translation, by 65%*. In vitro miR-26a knockdown upregulated TRPC3 protein by 54%*, mimicking AF effects.
Conclusions: AF increases FB proliferation, differentiation and ECM gene expression by increasing TRPC3 subunit expression and current, likely by downregulating the TRPC3-targeting microRNA miR-26.
- © 2011 by American Heart Association, Inc.