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N-cadherin acts upstream of VE-cadherin in controlling vascular morphogenesis.

Luo Y, Radice GL - J. Cell Biol. (2005)

Bottom Line: Contrary to previous studies, we found that N-cadherin localizes to endothelial cell-cell junctions in addition to its well-known diffusive membrane expression.Loss of N-cadherin in endothelial cells results in embryonic lethality at mid-gestation due to severe vascular defects.Intriguingly, loss of N-cadherin caused a significant decrease in VE-cadherin and its cytoplasmic binding partner, p120ctn.

View Article: PubMed Central - PubMed

Affiliation: Center for Research on Reproduction and Women's Health, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA.

ABSTRACT
Endothelial cells express two classic cadherins, VE-cadherin and N-cadherin. The importance of VE-cadherin in vascular development is well known; however, the function of N-cadherin in endothelial cells remains poorly understood. Contrary to previous studies, we found that N-cadherin localizes to endothelial cell-cell junctions in addition to its well-known diffusive membrane expression. To investigate the role of N-cadherin in vascular development, N-cadherin was specifically deleted from endothelial cells in mice. Loss of N-cadherin in endothelial cells results in embryonic lethality at mid-gestation due to severe vascular defects. Intriguingly, loss of N-cadherin caused a significant decrease in VE-cadherin and its cytoplasmic binding partner, p120ctn. The down-regulation of both VE-cadherin and p120ctn was confirmed in cultured endothelial cells using small interfering RNA to knockdown N-cadherin. We also show that N-cadherin is important for endothelial cell proliferation and motility. These findings provide a novel paradigm by which N-cadherin regulates angiogenesis, in part, by controlling VE-cadherin expression at the cell membrane.

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Cellular localization of cadherins in endothelial cells. Double immunostaining of N-cadherin and VE-cadherin in HUVEC (A–C) and HMVEC (D–F). N-cadherin exhibits both diffuse, nonjunctional as well as junctional staining (A and D). In contrast, VE-cadherin is restricted to regions of cell–cell contact (B and E) where it colocalizes with N-cadherin (C and F). The nuclei staining is due to nonspecific background. Bars, 50 μm.
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fig1: Cellular localization of cadherins in endothelial cells. Double immunostaining of N-cadherin and VE-cadherin in HUVEC (A–C) and HMVEC (D–F). N-cadherin exhibits both diffuse, nonjunctional as well as junctional staining (A and D). In contrast, VE-cadherin is restricted to regions of cell–cell contact (B and E) where it colocalizes with N-cadherin (C and F). The nuclei staining is due to nonspecific background. Bars, 50 μm.

Mentions: We report here that N-cadherin is also found at intercellular junctions colocalized with VE-cadherin in human umbilical vein endothelial cells (HUVEC) and human dermal microvascular endothelial cells (HMVEC), suggesting an unappreciated role for N-cadherin in EC–EC interactions (Fig. 1). To determine N-cadherin's function in ECs during vascular development, N-cadherin floxed mice (Kostetskii et al., 2005) were bred to Tie2-Cre transgenic mice (Kisanuki et al., 2001) to delete N-cadherin specifically from the EC lineage. Unexpectedly, N-cadherin loss in ECs caused recessive embryonic lethality at mid-gestation, before mural cell investment, raising the possibility of an important role for N-cadherin in EC–EC interactions. Mutant embryos were developmentally delayed compared with their wild-type littermates at E9.5, with lack of blood circulation accompanied by pericardial effusion (Fig. 2, A and B). By E10.5, mutant embryos were deteriorated (Fig. S1, available at http://www.jcb.org/cgi/content/full/jcb.200411127/DC1). Mutant embryos exhibited defective embryonic and yolk sac vasculature, including the lack of large vessels in the yolk sac. In contrast, wild-type yolk sac displayed great vessels at this stage (Fig. 2, C and D). Embryos also were examined by whole-mount staining with platelet endothelial cell adhesion molecule (PECAM) antibody to show the emerging vasculature. Mutant embryos exhibited an abnormal vascular plexus with smaller, less developed vessels in the brain and intersomitic region (Fig. 2 C; Fig. S2, available at http://www.jcb.org/cgi/content/full/jcb.200411127/DC1). Sectioning of the PECAM-stained mutant embryo showed smaller dorsal aorta and multiple endothelial vesicles instead of one large cardinal vein compared with the wild-type embryo (Fig. 2, D and E). Interestingly, endocardial cells in the heart tube appeared enlarged or swollen in the mutant embryo compared with the elongated morphology of the wild-type cells (Fig. 2, F and G). Although the Tie2-Cre transgene is not active in the myocardium, the myocardial layer appeared less compact in the mutant embryo compared with the wild-type embryo, suggesting cross-talk between the endocardium and myocardium involving cadherin (Fig. 2, F and G).


N-cadherin acts upstream of VE-cadherin in controlling vascular morphogenesis.

Luo Y, Radice GL - J. Cell Biol. (2005)

Cellular localization of cadherins in endothelial cells. Double immunostaining of N-cadherin and VE-cadherin in HUVEC (A–C) and HMVEC (D–F). N-cadherin exhibits both diffuse, nonjunctional as well as junctional staining (A and D). In contrast, VE-cadherin is restricted to regions of cell–cell contact (B and E) where it colocalizes with N-cadherin (C and F). The nuclei staining is due to nonspecific background. Bars, 50 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171890&req=5

fig1: Cellular localization of cadherins in endothelial cells. Double immunostaining of N-cadherin and VE-cadherin in HUVEC (A–C) and HMVEC (D–F). N-cadherin exhibits both diffuse, nonjunctional as well as junctional staining (A and D). In contrast, VE-cadherin is restricted to regions of cell–cell contact (B and E) where it colocalizes with N-cadherin (C and F). The nuclei staining is due to nonspecific background. Bars, 50 μm.
Mentions: We report here that N-cadherin is also found at intercellular junctions colocalized with VE-cadherin in human umbilical vein endothelial cells (HUVEC) and human dermal microvascular endothelial cells (HMVEC), suggesting an unappreciated role for N-cadherin in EC–EC interactions (Fig. 1). To determine N-cadherin's function in ECs during vascular development, N-cadherin floxed mice (Kostetskii et al., 2005) were bred to Tie2-Cre transgenic mice (Kisanuki et al., 2001) to delete N-cadherin specifically from the EC lineage. Unexpectedly, N-cadherin loss in ECs caused recessive embryonic lethality at mid-gestation, before mural cell investment, raising the possibility of an important role for N-cadherin in EC–EC interactions. Mutant embryos were developmentally delayed compared with their wild-type littermates at E9.5, with lack of blood circulation accompanied by pericardial effusion (Fig. 2, A and B). By E10.5, mutant embryos were deteriorated (Fig. S1, available at http://www.jcb.org/cgi/content/full/jcb.200411127/DC1). Mutant embryos exhibited defective embryonic and yolk sac vasculature, including the lack of large vessels in the yolk sac. In contrast, wild-type yolk sac displayed great vessels at this stage (Fig. 2, C and D). Embryos also were examined by whole-mount staining with platelet endothelial cell adhesion molecule (PECAM) antibody to show the emerging vasculature. Mutant embryos exhibited an abnormal vascular plexus with smaller, less developed vessels in the brain and intersomitic region (Fig. 2 C; Fig. S2, available at http://www.jcb.org/cgi/content/full/jcb.200411127/DC1). Sectioning of the PECAM-stained mutant embryo showed smaller dorsal aorta and multiple endothelial vesicles instead of one large cardinal vein compared with the wild-type embryo (Fig. 2, D and E). Interestingly, endocardial cells in the heart tube appeared enlarged or swollen in the mutant embryo compared with the elongated morphology of the wild-type cells (Fig. 2, F and G). Although the Tie2-Cre transgene is not active in the myocardium, the myocardial layer appeared less compact in the mutant embryo compared with the wild-type embryo, suggesting cross-talk between the endocardium and myocardium involving cadherin (Fig. 2, F and G).

Bottom Line: Contrary to previous studies, we found that N-cadherin localizes to endothelial cell-cell junctions in addition to its well-known diffusive membrane expression.Loss of N-cadherin in endothelial cells results in embryonic lethality at mid-gestation due to severe vascular defects.Intriguingly, loss of N-cadherin caused a significant decrease in VE-cadherin and its cytoplasmic binding partner, p120ctn.

View Article: PubMed Central - PubMed

Affiliation: Center for Research on Reproduction and Women's Health, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA.

ABSTRACT
Endothelial cells express two classic cadherins, VE-cadherin and N-cadherin. The importance of VE-cadherin in vascular development is well known; however, the function of N-cadherin in endothelial cells remains poorly understood. Contrary to previous studies, we found that N-cadherin localizes to endothelial cell-cell junctions in addition to its well-known diffusive membrane expression. To investigate the role of N-cadherin in vascular development, N-cadherin was specifically deleted from endothelial cells in mice. Loss of N-cadherin in endothelial cells results in embryonic lethality at mid-gestation due to severe vascular defects. Intriguingly, loss of N-cadherin caused a significant decrease in VE-cadherin and its cytoplasmic binding partner, p120ctn. The down-regulation of both VE-cadherin and p120ctn was confirmed in cultured endothelial cells using small interfering RNA to knockdown N-cadherin. We also show that N-cadherin is important for endothelial cell proliferation and motility. These findings provide a novel paradigm by which N-cadherin regulates angiogenesis, in part, by controlling VE-cadherin expression at the cell membrane.

Show MeSH