CD93 promotes β1 integrin activation and fibronectin fibrillogenesis during tumor angiogenesis
Roberta Lugano, … , Elisabetta Dejana, Anna Dimberg
Roberta Lugano, … , Elisabetta Dejana, Anna Dimberg
Published May 15, 2018
Citation Information: J Clin Invest. 2018;128(8):3280-3297. https://doi.org/10.1172/JCI97459.
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Research Article Oncology Vascular biology

CD93 promotes β1 integrin activation and fibronectin fibrillogenesis during tumor angiogenesis

Abstract

Tumor angiogenesis occurs through regulation of genes that orchestrate endothelial sprouting and vessel maturation, including deposition of a vessel-associated extracellular matrix. CD93 is a transmembrane receptor that is upregulated in tumor vessels in many cancers, including high-grade glioma. Here, we demonstrate that CD93 regulates β1 integrin signaling and organization of fibronectin fibrillogenesis during tumor vascularization. In endothelial cells and mouse retina, CD93 was found to be expressed in endothelial filopodia and to promote filopodia formation. The CD93 localization to endothelial filopodia was stabilized by interaction with multimerin-2 (MMRN2), which inhibited its proteolytic cleavage. The CD93-MMRN2 complex was required for activation of β1 integrin, phosphorylation of focal adhesion kinase (FAK), and fibronectin fibrillogenesis in endothelial cells. Consequently, tumor vessels in gliomas implanted orthotopically in CD93-deficient mice showed diminished activation of β1 integrin and lacked organization of fibronectin into fibrillar structures. These findings demonstrate a key role of CD93 in vascular maturation and organization of the extracellular matrix in tumors, identifying it as a potential target for therapy.

Authors

Roberta Lugano, Kalyani Vemuri, Di Yu, Michael Bergqvist, Anja Smits, Magnus Essand, Staffan Johansson, Elisabetta Dejana, Anna Dimberg

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Figure 5

CD93 and MMRN2 colocalize in the tip cells during sprouting angiogenesis and are required for endothelial sprouting.

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CD93 and MMRN2 colocalize in the tip cells during sprouting angiogenesis...
(A) Immunofluorescent staining of CD93 and MMRN2 in the sprouting front (indicated by dotted line) of vessels formed in embryoid bodies (EBs) cultured in 2D on glass slides upon VEGF treatment. Differentiated endothelial cells were visualized by CD31 staining. Scale bars: 20 μm. Specific localization of CD93 and MMRN2 in CD31-positive sprouts and filopodia is shown in high-magnification images. (B) Immunofluorescent staining of CD93 and MMRN2 in EBs cultured in a 3D collagen gel. Scale bars: 200 μm. High-magnification images show a specific localization of CD93 in the tip cells and in filopodia. MMRN2 staining indicates its colocalization with CD31-positive sprouts and with CD93 in the tip cells. (C) CD31 immunofluorescent staining of shCD93- and shMMRN2-transfected EBs cultured in 2D glass slide. Untransfected (Ctrl) or empty vector–transfected (shCtrl) EBs were used as controls. Scale bars: 500 μm. (D) Quantification of CD31 sprouting expansion. Values represent mean ± SEM (at least n = 5 EBs per condition). ***P < 0.001; 1-way ANOVA with Dunnett’s multiple-comparisons test. (E) CD31 immunofluorescent staining of shCD93- and shMMRN2-transfected or control EBs (Ctrl and shCtrl) cultured in 3D collagen gel. Scale bars: 500 μm. (F) Quantification of CD31-positive sprouts. Values represent mean ± SEM (at least n = 10 EBs per condition). **P < 0.01, ***P < 0.001; 1-way ANOVA with Dunnett’s multiple-comparisons test. (G) Coculture of 2 individual EBs of each condition in 2D glass slide. (H) Coculture of control EBs (Ctrl or shCtrl) with shMMRN2 or shCD93 EBs. (I) Quantification of the CD31 sprouting expansion rescued upon coculture of control EBs with shMMRN2 or shCD93 EBs. *P < 0.05 vs. Ctrl; #P < 0.05 vs. shCtrl; 1-way ANOVA with Dunnett’s multiple-comparisons test.