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represents the main mechanism for the transport of molecules in physiological con-
ditions, the paracellular transport becomes a prominent way for plasma leakage in
inflammation, ischemia-reperfusion injury, adult respiratory distress syndrome,
tumor growth and metastatization, and several other pathological conditions
(Komarova and Malik 2010 ; Hu et al. 2008 ; Mehta and Malik 2006 ; Wallez and
Huber 2008 ). The endothelial barrier can be altered by various inflammatory and
vasoactive agents, which, consequently, increase endothelial permeability.
Experimental evidence suggests that CgA, at low concentrations, can preserve the
integrity of the endothelial barrier function when these cells are exposed to inflam-
matory stimuli, such as tumor necrosis factor-α (TNF) or vascular endothelial
growth factor (VEGF) (Ferrero et al. 2004 ). Structure-function studies showed that
an active site is located in the N-terminal domain. Indeed, the CgA1-78 fragment is
sufficient to protect vessels from TNF-induced vascular leakage in vivo and to
inhibit the paracellular flux of radio-labeled albumin through endothelial cell mono-
layers in vitro (Ferrero et al. 2004 ). Mechanistic studies showed that CgA1-78 can
inhibit the TNF-induced disassembly of VE-cadherin-dependent adherence junc-
tions and the conversion of peripheral actin bundles into stress fibers in endothelial
cell monolayers (Ferrero et al. 2004 ). The inhibitory effect of CgA and CgA1-78 on
TNF-induced alteration of endothelial barrier integrity is unlikely related to inhibi-
tion of TNF/TNF-receptor interactions, as CgA does not bind these molecules
(Ferrero et al. 2004 ). Experimental data obtained with bovine pulmonary arterial
endothelial cells showed that CgA1-78 can inhibit TNF-induced phosphorylation of
p38MAPK by a pertussis toxin-sensitive mechanism, implicating a role for CgA1-
78 in protecting the endothelial integrity via regulation of the stress-activated
MAPK pathway in a G-protein-dependent manner (Blois et al. 2006a). CgA1-78
can also inhibit TNF-induced intercellular cell adhesion (ICAM)-1 expression,
monocyte chemoactractant protein (MCP)-1 release and relocation of high-mobility
group box (HMGB)-1 in human microvascular endothelial cells, further supporting
an anti-inflammatory role of this CgA fragment (Di Comite et al. 2009a).
The receptors responsible for these biological effects are still unknown. It has
been reported that bovine aorta endothelial cells bind and internalize 1 nM
(^125) I-labeled CgA (Mandalà et al. 2000 ). Furthermore, fluorescein-isotiocyanate
(FITC)-labeled CgA1-78 is bound and internalized in endocytotic vesicles by
human umbilical vein endothelial cells (HUVECs) in culture (Ferrero et al. 2004 ).
In vitro binding studies performed with the Langmuir apparatus showed that CgA1-
78 can interact with membrane phospholipids, particularly with phosphatidylserine
(Dondossola et al. 2010 ; Blois et al. 2006b). The specific, enhanced fluidity exerted
by low nanomolar concentrations of CgA1-78 in monolayers of phosphatidylserine
may suggest that CgA1-78 can indirectly interact with, or perturb, specific receptors
in phosphatidylserine-rich microdomains in the outer leaflet of the plasma mem-
brane (Blois et al. 2006b). CgA1-78 can also induce a marked increase of caveolae-
dependent endocytosis in bovine aortic endothelial cells, which is significantly
reduced by heparinase III and by wortmannin, a specific phosphoinositide 3-kinase
(PI3K) inhibitor (Ramella et al. 2010 ). As these compounds also abolished the
CgA1-78-dependent phosphorylation of endothelial nitric oxide synthase (eNOS),
F. Curnis et al.