Abstract 560: HDAC3 Unconventional Splicing Mediates Endothelial-mesenchymal Transition in Cardiac Fibrosis

Abstract

Introduction: Endothelial-mesenchymal transition (EndMT) makes a significant contribution to cardiac fibrosis. Histone deacetylase 3 (HDAC3) is essential in the maintenance of endothelial homeostasis. Our previous study indicates that mouse HDAC3 mRNA can undergo unconventional splicing, and that the HDAC3α spliced isoform promotes the EndMT process in human aortic endothelial cells (HAECs). This work aims to elucidate the role of HDAC3α in mediating EndMT and its part in the underlying mechanisms behind the development of pressure-overload-induced cardiac fibrosis. Methods and Results: Our previous study reveals that HDAC3α promotes EndMT in HAECs via the activation of transforming growth factor-beta 2, possibly through increased protease enzyme activity. HDAC3α was therefore overexpressed in HAECs via adenoviral gene transfer (Ad-HDAC3α). Protease array revealed that cells infected with Ad-HDAC3α have an elevated level of a disintegrin and metalloproteinase with thrombospondin motifs 1 (ADAMTS1), and this was further confirmed with increased mRNA and protein expression levels seen in Ad-HDAC3α-infected HAECs using RT-qPCR and Western blotting analysis respectively (both n = 4, P < 0.05). A mouse model of pressure overload-induced cardiac fibrosis was established by transverse aortic constriction (TAC), and cardiac fibrosis was confirmed by increased Col1a1 expression seen using RT-qPCR and picosinus red staining in heart tissue sections from mice 7 d post-TAC. Immunofluorescent staining detected CD31 + /α-SMA + (indicative of EndMT) and HDAC3α + /CD31 + cells in heart tissues from TAC mice but not in those from sham-operated mice. There were significant increases in mRNA expression of HDAC3α and ADAMTS1, together with EndMT transcription factors Snai1, Snai2 and Twist1 in heart tissues from 7 d post-TAC mice compared to those from 7 d post-sham mice (n = 7- 8, P < 0.05). Elevated HDAC3α and ADAMTS1 protein expression were also detected in the TAC heart tissues (n = 5, P < 0.05). Conclusion: These results indicate an association between the HDAC3 splicing isoform HDAC3α, ADAMTS1 and EndMT during TAC-induced cardiac fibrosis. Further investigation of the underlying mechanisms mediated by HDAC3α and ADAMTS1 in cardiac fibrosis is warranted.