Congestive Heart Failure Effects on Atrial Fibroblast Phenotype: Differences between Freshly-Isolated and Cultured Cells

Introduction: Fibroblasts are important in the atrial fibrillation (AF) substrate resulting from congestive heart failure (CHF). We previously noted changes in in vivo indices of fibroblast function in a CHF dog model, but could not detect changes in isolated cells. This study assessed CHF-induced changes in the phenotype of fibroblasts freshly isolated from control versus CHF dogs, and examined effects of cell culture on these differences. Methods/Results: Left-atrial fibroblasts were isolated from control and CHF dogs (ventricular tachypacing 240 bpmx2 weeks). Freshly-isolated fibroblasts were compared to fibroblasts in primary culture. Extracellular-matrix (ECM) gene-expression was assessed by qPCR, protein by Western blot, fibroblast morphology with immunocytochemistry, and K+-current with patch-clamp. Freshly-isolated CHF fibroblasts had increased expression-levels of collagen-1 (10-fold), collagen-3 (5-fold), and fibronectin-1 (3-fold) vs. control, along with increased cell diameter (13.4 +/- 0.4 mu m vs control 8.4 +/- 0.3 mu m) and cell spreading (shape factor 0.81 +/- 0.02 vs. control 0.87 +/- 0.02), consistent with an activated phenotype. Freshly-isolated control fibroblasts displayed robust tetraethylammonium (TEA)-sensitive K+-currents that were strongly downregulated in CHF. The TEA-sensitive K+-current differences between control and CHF fibroblasts were attenuated after 2-day culture and eliminated after 7 days. Similarly, cell-culture eliminated the ECM protein-expression and shape differences between control and CHF fibroblasts. Conclusions: Freshly-isolated CHF and control atrial fibroblasts display distinct ECM-gene and morphological differences consistent with in vivo pathology. Culture for as little as 48 hours activates fibroblasts and obscures the effects of CHF. These results demonstrate potentially-important atrial-fibroblast phenotype changes in CHF and emphasize the need for caution in relating properties of cultured fibroblasts to in vivo systems. Citation: Dawson K, Wu C-T, Qi XY, Nattel S (2012) Congestive Heart Failure Effects on Atrial Fibroblast Phenotype: Differences between Freshly-Isolated and Cultured Cells. PLoS ONE 7(12): e52032. doi: 10.1371/journal.pone.0052032