Abstract 14371: HDAC9 Promotes Angiogenesis and Down-Regulates the Anti-Angiogenic MiR-17-92 Cluster in Endothelial Cells
Acetylation of histone proteins contributes to the regulation of gene expression by alteration of chromatin structure. Histone deacetylases (HDACs) act as modulators of gene expression by deacetylation of histone proteins as well as non-histone proteins. Previous studies identified the class IIa histone deacetylases HDAC5 as a negative regulator of angiogenesis. Furthermore, HDAC5 and HDAC9 knockout mice are prone to cardiac hypertrophic signals, indicating that HDAC5 and HDAC9 exhibit a redundant function in the heart. However, the role of HDAC9 in endothelial cells (EC) is unknown. SiRNA mediated knockdown of HDAC9 decreases EC sprouting from spheroids (46±8%) and tube formation (75±9%) in Matrigel assay without affecting cell viability. To confirm the role of HDAC9 in mammalian vessel formation, we performed a spheroid based matrigel plug assay in mice. Therefore, EC were transduced with shHDAC9 lentivirus or control virus. After selection of transduced cells, spheroids were generated, mixed with matrigel and subcutaneously injected into mice. Downregulation of HDAC9 significantly decreases vessel density (44±6%, p<0.05). Moreover, the average size (62±6%, p<0.05) and perfusion of vessels (19±6% control vs 1±1% shHDAC9, p<0.01) are reduced. Consistently, overexpression of HDAC9 wildtyp increases EC sprouting (139±10%). Furthermore, by using mutated constructs, we demonstrated that this pro-angiogenic effect requires the catalytic domain of HDAC9, nuclear localization and sumoylation of HDAC9. Downregulation of HDAC9 in EC increases the expression of the anti-angiogenic miR-17-92 cluster (214±7%, p<0.01). Consistently, single microRNAs of the cluster are upregulated after HDAC9 knockdown (namely miR-17a, -18a, -20 and -92a). Downregulation of HDAC9 significantly decreases the expression of the predicted miR17/20 target VEGF-A. In summary, we found that HDAC9, in contrast to HDAC5, promotes angiogenesis in vitro and in vivo as assessed by in vitro EC sprouting and in vivo vessel formation in mice. Mechanistically, the pro-angiogenic effect of HDAC9 in EC requires the catalytic domain, nuclear localization and sumoylation of HDAC9. Knockdown of HDAC9 in EC increases the expression of the anti-angiogenic miR-17-92 cluster.
- © 2010 by American Heart Association, Inc.