Mechanical signals modulated vascular endothelial growth factor-A (VEGF-A) alternative splicing in osteoblastic cells through actin polymerisation

Abstract

To cite this version:Céline None of the authors has any conflicts of interest. A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT AbstractSince VEGF-A is involved in mechanically-induced bone gain and because vegf exists under 6 isoforms exerting various biological effects, we studied vegf isoforms expression and VEGF protein production in osteoblastic cells (rat Ros17/2.8 and human osteoblasts) submitted to 4 mechanical regimens. Mechanical regimens (1% stretch deformation)were designed with a fixed number of cycles (450) mRNA expressions of soluble VEGF isoforms (121,165) were specifically expressed under low frequency while matrix-bound VEGF isoforms (206, 189, 165, 145) were specifically expressed under high frequency in human osteoblasts. As F-actin stress fibers formation was significantly increased selectively in high frequencies conditions, we disrupted actin fibers in Ros17/2.8 and found that immobilisation of VEGF was abolished. Conversely, jasplakinolide treatment which increases stress fibers formation was able to mimic high frequency stretch-induced immobilisation of VEGF. Thus, we speculate that the stretchinduced increase in cell tension is responsible for matrix-bound vegf isoform production.Mechanically-induced selection of soluble or matrix-bound VEGF production may modify osteoblast and endothelial c ells crosstalk crucial during osteogenesis and fracture healing.

Fluid Flow-induced Soluble Vascular Endothelial Growth Factor Isoforms Regulate Actin Adaptation in Osteoblasts

“…Similar findings have been reported in a murine embryonic mesenchymal progenitor cell line and in bone marrow stromal cells that showed significantly increased VEGF mRNA expression in response to laminar or pulsatile fluid shear stress (28,29). Likewise, up-regulation of VEGF expression under cyclic strain has been shown in various cell types (30,31), including osteoblasts (14,32). Because VEGF expression in osteoblasts is induced by osteotropic factors such as prostaglandins, insulin-like growth factor 1, TGF-␤, PDGF, FGF2, bone morphogenetic proteins, and 1-␣,25-dihydroxyvitamin D 3 (5,33), it is possible that up-regulation of VEGF in response to PFSS results from the downstream signaling cas-FIGURE 5.…”

“…In rat and human osteoblasts, cyclic stretch has been shown to regulate the expression of VEGF isoforms that control actin polymerization in a frequency-dependent manner; soluble VEGF isoforms are more sensitive to low frequency cyclic stretch (0.05 Hz), whereas matrix-bound isoforms are more responsive to high frequency (5 Hz) (14). In addition, using latrunculin A (an inhibitor of actin polymeriza- tion) and jasplakinolide (a potent inducer of actin polymerization), Faure et al (14) demonstrated that not only actin polymerization but also cell contractility is key for the storage of matrix-bound VEGF.…”

“…Recent evidence using genetic manipulation revealed the important role of the HIFs/VEGF pathway in coupling angiogenesis and osteogenesis (41). In various cell types including osteoblasts, mechanical strain has been shown to induce both Hif1␣ and Vegf expression (14,42,43). However, in our previous cDNA microarray study Hif1␣ expression in MC3T3-E1 cells was not significantly altered after 5 h of exposure to PFSS (the National Center for Biotechnology Information Gene Expression Omnibus database, series GSE8117), indicating that flow-induced VEGF production and release are not directly regulated by HIFs.…”

“…At the molecular level, VEGF signaling has been shown to induce actin polymerization/depolymerization and to acti-vate focal adhesion kinases in both endothelial cells and osteoblasts (13)(14)(15)(16), in a manner very similar to changes observed when these cells are submitted to mechanical stimulation (17)(18)(19)(20). Based on these similarities and our recent findings indicating that Vegf expression in osteoblasts is upregulated by mechanical stimulation (2), we hypothesized that fluid shear stress regulates actin cytoskeleton adaptation during osteoblast mechanotransduction through VEGF release and autocrine signaling.…”

Characterizing the role of mechanical signals in gene regulatory networks using Long SAGE

“…In vivo and in vitro experiments showed that mechanical stress regulated the expression of FosB/ΔFosB (Inoue et al, 2004). The alternative splicing of vascular endothelial growth factor-A (Vegf-A) was regulated by mechanical stress in osteoblasts through actin polymerization (Faure et al, 2008). Although the mechanism of this process is still unclear, the up-regulation of splicing factors Hnrpk and Sf3b4 indicated the role of mechanical stress in regulating splicing process.…”