Calpain‐2 regulation of VEGF‐mediated angiogenesis

Abstract

Angiogenesis is a complex process involving endothelial cell migration, proliferation, and differentiation as well as tube formation. These processes are stimulated by a variety of growth factors such as vascular endothelial growth factor (VEGF). VEGF-induced cytoskeletal reorganization plays a crucial role in the angiogenic processes. In the present study, we evaluated the role of calpain in VEGF-induced angiogenesis in vitro and in vivo. Human pulmonary microvascular endothelial cells (PMEC) were incubated with VEGF (10-60 ng/ml) for 2-24 h, after which we measured calpain activities, protein contents of the calpain subunits and of calpastatin, endothelial monolayer wound repair, tube formation, and actin cytoskeleton changes. Incubation of PMEC with VEGF resulted in dose- and time-dependent increases in calpain activity and protein content of calpain-2. VEGF did not change the protein contents of calpain-1 and the small subunit or of calpastatin. Incubation of PMEC with a VEGF receptor blocker prevented the VEGF-induced increase in calpain activity. Inhibition of calpain activity by siRNA directed against calpain-2 and by overexpression of calpastatin prevented VEGF-induced increases in actin stress fibers in endothelial cells and angiogenesis. Overexpression of calpastatin also inhibits vessel formation in subcutaneous (s.c.) matrigel plugs in mice. These results indicate that calpain mediates VEGF-induced angiogenic effects by modulating actin cytoskeletal organization.

Protein Phosphotyrosine Phosphatase 1B (PTP1B) in Calpain-dependent Feedback Regulation of Vascular Endothelial Growth Factor Receptor (VEGFR2) in Endothelial Cells

“…We and others have previously shown evidences of a VEGF/ VEGFR2/calpain/PI3K/AMPK/Akt/eNOS/NO signaling pathway (1)(2)(3)(4). In this study, we set out to investigate any calpain dependent feedback regulation of VEGFR2.…”

“…We and others have previously shown that calpain activity is increased within 10 min of VEGF stimulation of endothelial cells, which can be inhibited by antagonist of the VEGF receptor (1)(2)(3)(4). In addition, we have demonstrated an important role of calpain in mediating VEGFinduced PI3K/AMPK/Akt activation and consequent eNOS phosphorylation and nitric oxide (NO) production (1).…”

“…and Akt-Our previous data showed robust calpain activation in VEGF-treated bovine aortic endothelial cells (BAECs), which can be abolished by pretreatment with calpain inhibitor calpeptin or ALLN (1) or VEGF receptor inhibitor 4-[(4Ј-chloro-2Ј-fluoro) phenylamino]-6,7-dimethoxyquinazoline (2). Knocking down calpain inhibited VEGF-induced NO production in endothelial cells (1).…”

Calpain mediates pulmonary vascular remodeling in rodent models of pulmonary hypertension, and its inhibition attenuates pathologic features of disease

“…Expression of calpain-1 and calpain-2 was silenced using siRNA technology (73). The target sequences for the mRNA of calpain-1 and calpain-2 were 5′-AAGCTAGTGTTCGTG-CACTCT-3′ and 5′-AAACCAGAGCTTCCAGGAAAA-3′, respectively.…”

Calpain as a Novel Regulator of Autophagosome Formation

“…1 Their most studied involvement is in the regulation of the dynamic rearrangements of cytoskeletal architecture during cell migration 2 : depending on the cellular context, a range of stemming effects have been shown to be controlled by calpains such as synaptic function/remodelling in neurons 3 or angiogenesis in endothelial cells. 4 m-calpain and m-calpain, encoded by Capn1 and Capn2, respectively, are ubiquitously expressed and require a common 28-kDa regulatory subunit encoded by the Capns1 gene for stability and activity. Targeted disruption of Capns1 is embryonic lethal at day 10-11, due to severe defects in vascular development.…”