Abstract

Background: Endothelial cells regulate angiogenesis and vessel wall homeostasis via bioactive factors, such as cysteine-rich 61 (Cyr61) and Integrin-_αv (Itg_αv). Pre-mRNA splicing leads to the expression of two Cyr61 mRNA splice variants (Cyr61_IS, Cyr61_IR) and three Itg_αv isoforms (Itg_αv_1, 2, 3). Splicing is important for the functional multiplicity and is regulated via the serine/arginine-rich (SR) protein kinases, Cdc2-like kinases (Clk) and DNA topoisomerase I (DNA topo I). Here, we study the impact of these SR protein kinases on the Cyr61 and Itg_αv isoform expression as well as on the endothelial cell proliferation and pro-angiogenic properties of human microvascular endothelial cells (HMEC-1).

Methods: We assessed the expression of Cyr61 and Itg_αv isoforms via RT-PCR and Western blotting. Cellular functions were determined by adequate assays (MTT proliferation assay, in vitro angiogenesis assay).

Results: We found Clks as well as DNA topo I to modulate the differential isoform expression of Cyr61 and Itg_αv, the protein expression and secretion in resting as well as in TNF-α-stimulated HMEC-1. Moreover, these processes affected the endothelial cell proliferation and pro-angiogenic tube formation by HMEC-1. Finally, treatment of cells with recombinant Cyr61_IS or siRNA-mediated silencing of Cyr61_IS and Cyr61_IR also modulated these functions.

Conclusions: This study indicates DNA topo I and Clks to regulate the differential isoform expression Cyr61 and Itg_αv and to affect regenerative endothelial functions under normal and pro-inflammatory conditions.

Keywords: Endothelial cells, inflammation, cytokines, kinase