In the aging heart, fibroblasts have a reduced ability to mature into myofibroblasts (expressing contractile actin a-SMA), which is necessary for scar contraction in wound healing. The mechanosensing pathway bridges the extracellular matrix (ECM) with the actin cytoskeleton and is altered during aging. We emphasize that changes in this pathway may affect myofibroblast activation. Using various methods (mass spectrometry, qPCR, flow, IF staining, and western), we found several abnormalities in the fibroblasts derived from old mouse hearts: 1) ECM deposited was altered with an increased quantity of collagens and the presence of fibronectin variant Anastellin that opposes myofibroblast maturation; 2) there was a 50% reduction of Kindlin-2, a protein that promotes integrin activation; 3) Polymerized (F-) to monomeric (G-) actin ratio was decreased by 65%, possibly due to a 5-fold increase in pERKs1/2 level, a negative regulator of F-actin; and pERKs inhibition (by 2.5μM, PD-0325901) increased by 2-fold F/G actin ratio; 4) we found an increased cytoplasmic retention of MRTF-A (by 44%), an actin-sensitive co-transcription factor (necessary for a-SMA) that shuttles into the nucleus in response to F-actin formation; 5) pERKs inhibition increased a-SMA transcription by 78%. Thus, the disrupted mechanosensing pathway contributes to the impaired myofibroblast maturation in the aging heart.
All experiments were performed in young (3-4 month-old) and old (24-30 month-old) hearts or primary cardiac fibroblasts isolated from male C57BL/6J mice. Unpaired Student’s t-test was used for statistical analysis, n=2 for mass spectrometry and n=4-10 for all other biological repeats.
Caffeine induces both short‐term and long‐term effects on gene expression and DNA methylation in the mouse heart (542.3)
