Abstract 534: Adenoviral-mediated Knock-out of Focal Adhesion Kinase in Adult Mouse Cardiac Fibroblasts Promotes Cell Proliferation and Inhibits Cell Migration
Background: Focal adhesion kinase(FAK) is a critical protein for the regulation of integrin-mediated cellular function in many cell types. While prior studies have characterized FAK deficient embryonic derived fibroblasts, no data exists on FAK deficient adult cardiac fibroblasts(CFs). Thus, we studied how reduction of FAK in adult mouse CFs ex vivo changes phenotypes of these cells.
Methods and Results: CFs were isolated from homozygous FAK floxed mice(Beggs HE, et al, Neuron;40:501–14, 2003). The FAK knock-out(KO) CFs were created by infecting Cre-recombinase adenovirus(Ad-Cre). Ninety-six hours following Ad-Cre(200 moi) infection, FAK protein was significantly decreased, compared with lacZ virus-infected CFs control cells(65% reduction, p<0.05). The FAK KO CFs exhibited no significant differences in adhesion to fibronectin, independent of binding time(5, 15, 30 and 60 min), compared with control cells(p<0.05). Enhanced expression of paxillin and pyk2 was noted in FAK KO CFs, compared with controls. The proliferation activity of FAK KO CFs, determined by BrdU assay following PDGF-BB stimulation showed an 89% increase(p<0.05 vs. control group). Migration activity, examined by a modified Boyden chamber assay with PDGF-BB as a chemoattractant showed a 46% reduction in FAK KO CFs(p < 0.05 vs. control group). PDGF-BB-stimulated ERK1/2 kinase activity was significantly increased in FAK KO CFs, compared with controls, while p38, JNK and Akt kinase activities were not significantly different between the two groups.
Conclusion: Our results show that depletion of FAK in adult mouse CFs promotes cell proliferation and inhibits cell migration induced by PDGF-BB. Defective MAPK activation was found in the FAK KO cells suggesting that this signaling abnormality may play a role in the proliferative and migratory defects in these cells. Adhesion of FAK KO CFs was not impaired. Further study is required to elucidate the mechanism for our results.