Amino-terminal fragments of laminin γ2 chain retract vascular endothelial cells and increase vascular permeability.
Bottom Line: Such effects of γ2pf led to prominent actin stress fiber formation in HUVECs, which was blocked by a ROCK inhibitor.As possible receptors, γ2pf interacted with heparan sulfate proteoglycans on the surface of HUVECs.Moreover, we localized the active site of γ2pf to the N-terminal epidermal growth factor-like repeat.
Affiliation: Department of Genome Science, Graduate School of Integrated Science and Nanobioscience, Yokohama City University, Yokohama, Japan; Division of Cell Biology, Kihara Institute for Biological Research, Yokohama City University, Yokohama, Japan.Show MeSH
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Mentions: The γ2pf-induced retraction of endothelial cells seemed to lead to their loss of barrier integrity. To confirm this possibility, we examined the activity of γ2pf on endothelial permeability in vitro (Fig. 4a). When the monolayers of HUVECs on the culture inserts were treated with purified γ2pf, the endothelial cell permeability, as measured by the diffusion of FITC-dextran through the HUVEC sheet, significantly increased compared to the untreated control cultures. In addition, enhanced permeability was observed with the full-length γ2 chain and its N-terminal domain V (γ2dV) (Fig. 4b, see also Fig. 1a). The order of the permeability activity was γ2dV > γ2pf > full-length γ2. Furthermore, we examined the effect of Lmγ2 on vascular permeability in vivo by Miles permeability assay with mice (Fig. 5). The intradermal injection of γ2pf increased the leakage of Evans blue dye two-fold compared to the PBS injection as control (Fig. 5a). Purified γ2dV also increased vascular permeability two-fold (Fig. 5b), but domain III of Lmγ2 did not show any significant activity (Figs 5c, S2, see also Fig. 1a). These results suggest that the N-terminal fragments of Lmγ2 chain function as vascular permeability-promoting factors in pathological conditions.
Affiliation: Department of Genome Science, Graduate School of Integrated Science and Nanobioscience, Yokohama City University, Yokohama, Japan; Division of Cell Biology, Kihara Institute for Biological Research, Yokohama City University, Yokohama, Japan.