Abstract P1140: Acute Mechanosensitive Adaptation To Phenylephrine In Cardiomyocytes And Fibroblasts

Abstract

Mechanosensitive pathways are fine tuned to the physiological environment of the cell. Adrenergic stimulation induces cell type specific responses in the heart, including cardiomyocyte hypertrophy and fibroblast proliferation. We hypothesized acute induction of hypertrophy by phenylephrine (PHE) would activate mechanosensitive pathways in fibroblasts and myocytes, revealing novel molecular features of early hypertrophic adaptation. Male mice were subcutaneously injected with 20 mg/kg of PHE or PBS every other day for one week. Echocardiography revealed significant cardiac hypertrophy with increased ejection fraction (72.45% + 6.15 PHE v 56.32 + 5.32 PBS, p<0.0001) and LV wall thickness (2.25 mm + 0.29 PHE v 1.62 + 0.13 PBS, p<0.0001), along with diastolic impairment, as measured by elevated E/e’ ratio (37.25 + 7.29 PHE v 23.67 + 3.62 PBS, p<0.0001). Interestingly, histological analysis showed increased fibrotic deposition and cardiomyocyte cross-sectional area was preferential increased in fibrotic areas (265.5 um 2 + 67.5 PHE fibrotic v 195.1 + 47.46 PHE non-fibrotic v 165.6 + 46.96 PBS). While cardiac fibroblasts from PHE-treated animals did not show alpha smooth muscle actin staining, a traditional marker for activated myofibroblasts, fibroblasts from PHE-treated mice showed increased f-actin stress fibers and nuclear localization of Mrtfa, a mechanosensitive transcriptional coactivator. We performed RNA-seq on isolated cardiomyocytes, demonstrating activation of myocyte-specific responses to increased pressure ( Des, Nppa ), as well as activation of fibrotic genes ( Tgfbr1, Tgfbr2, Tgfb2 ) in myocytes . We sought to investigate mechanosensitive pathways initiated by PHE treatment in cardiomyocytes and identified transcriptional changes in Ankrd1, a Titin binding protein which translocates to the nucleus upon stress. We demonstrate a PHE induced increase in Ankrd1 protein expression and nuclear localization (nuclear area occupancy, as measured by microscopy, 17.04% + 8.76 PHE v 11.91% + 9.134 PBS, p=0.002) in cardiomyocytes. Together these findings demonstrate distinct acute phenotyptic adaptation to adrenergic stress in myocytes and fibroblasts, revealing cell type-specific mechanosensitive signaling mechanisms.