MVL-PLA2, a Snake Venom Phospholipase A2, Inhibits Angiogenesis through an Increase in Microtubule Dynamics and Disorganization of Focal Adhesions

Abstract

Integrins are essential protagonists of the complex multi-step process of angiogenesis that has now become a major target for the development of anticancer therapies. We recently reported and characterized that MVL-PLA2, a novel phospholipase A2 from Macrovipera lebetina venom, exhibited anti-integrin activity. In this study, we show that MVL-PLA2 also displays potent anti-angiogenic properties. This phospholipase A2 inhibited adhesion and migration of human microvascular-endothelial cells (HMEC-1) in a dose-dependent manner without being cytotoxic. Using Matrigel™ and chick chorioallantoic membrane assays, we demonstrated that MVL-PLA2, as well as its catalytically inactivated form, significantly inhibited angiogenesis both in vitro and in vivo. We have also found that the actin cytoskeleton and the distribution of αvβ3 integrin, a critical regulator of angiogenesis and a major component of focal adhesions, were disturbed after MVL-PLA2 treatment. In order to further investigate the mechanism of action of this protein on endothelial cells, we analyzed the dynamic instability behavior of microtubules in living endothelial cells. Interestingly, we showed that MVL-PLA2 significantly increased microtubule dynamicity in HMEC-1 cells by 40%. We propose that the enhancement of microtubule dynamics may explain the alterations in the formation of focal adhesions, leading to inhibition of cell adhesion and migration.

Role of the Cytoskeleton in Formation and Maintenance of Angiogenic Sprouts

“…The vinblastine-induced collapse required Rho GTPases and was blocked with C3 exoenzyme and dominant negative forms of RhoA and RhoC, indicating microtubule depolymerization induced Rho GTPase activation and collapse of endothelial networks [153] . In separate studies, Luis and colleagues [173] recently reported that MVL-PLA2, a phospholipase A2 from snake venom, increased microtubule dynamics and blocked angiogenesis. MVL-PLA2 likewise disrupted focal adhesions, contributing to failed angiogenic responses [173] .…”

“…In separate studies, Luis and colleagues [173] recently reported that MVL-PLA2, a phospholipase A2 from snake venom, increased microtubule dynamics and blocked angiogenesis. MVL-PLA2 likewise disrupted focal adhesions, contributing to failed angiogenic responses [173] . In line with this observation, TAE226 inhibition of focal adhesion kinase blocked tube formation in vitro and microvessel density in subcutaneous tumors [174] .…”

Molecular Characterization of Lys49 and Asp49 Phospholipases A2 from Snake Venom and Their Antiviral Activities against Dengue virus

“…In vitro , the svPLA 2 s can also modulate cell adhesion and cell proliferation and have anti-angiogenic properties [9,10,11]. Furthermore, a number of studies describe the anti-microbial properties of several svPLA 2 s against Gram negative [12] and Gram positive bacteria [13].…”

Microtubule dynamics regulation contributes to endothelial morphogenesis

“…In addition, we did not observe a measurable growth rate increase in growth factor-induced protrusions in epithelial cells that appear morphologically quite similar to endothelial cells migrating along tension tracks 25 . In this regard it is interesting to note that low doses of microtubule perturbing drugs increase microtubule dynamics in endothelial cells, 21 , 22 , 26 an effect that also is unique to endothelial cells. Together, these observations suggest endothelial cell-specific mechanisms of microtubule regulation, and it will be interesting to see whether this can be therapeutically exploited.…”