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Proteolytic exposure of a cryptic site within collagen type IV is required for angiogenesis and tumor growth in vivo.

Xu J, Rodriguez D, Petitclerc E, Kim JJ, Hangai M, Moon YS, Davis GE, Brooks PC, Yuen SM - J. Cell Biol. (2001)

Bottom Line: Exposure of this cryptic site was associated with angiogenic, but not quiescent, blood vessels and was required for angiogenesis in vivo.A monoclonal antibody (HUIV26) directed to this site disrupts integrin-dependent endothelial cell interactions and potently inhibits angiogenesis and tumor growth.Together, these studies suggest a novel mechanism by which proteolysis contributes to angiogenesis by exposing hidden regulatory elements within matrix-immobilized collagen type IV.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, Kaplan Cancer Center, New York University School of Medicine, New York, NY 10016, USA.

ABSTRACT
Evidence is provided that proteolytic cleavage of collagen type IV results in the exposure of a functionally important cryptic site hidden within its triple helical structure. Exposure of this cryptic site was associated with angiogenic, but not quiescent, blood vessels and was required for angiogenesis in vivo. Exposure of the HUIV26 epitope was associated with a loss of alpha1beta1 integrin binding and the gain of alphavbeta3 binding. A monoclonal antibody (HUIV26) directed to this site disrupts integrin-dependent endothelial cell interactions and potently inhibits angiogenesis and tumor growth. Together, these studies suggest a novel mechanism by which proteolysis contributes to angiogenesis by exposing hidden regulatory elements within matrix-immobilized collagen type IV.

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Effects of purified Mab HUIV26 on angiogenesis in vivo. Rat corneal micropocket assays were performed to assess the effects of Mab HUIV26 on angiogenesis. (A) Representative corneas from rats implanted with hydron pellets containing bFGF (top), bFGF + Mab HUIV26 (middle) or bFGF + control Mab (bottom). Black arrows indicate angiogenic neovessels. Red arrows indicate preexisting limbal vessels. (B) Quantification of the area of neovascularization within rat corneas. Data bars represent the mean area of neovascularization from the limbus to hydron pellet. Experiments were performed at least twice with five to seven eyes per condition.
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fig4: Effects of purified Mab HUIV26 on angiogenesis in vivo. Rat corneal micropocket assays were performed to assess the effects of Mab HUIV26 on angiogenesis. (A) Representative corneas from rats implanted with hydron pellets containing bFGF (top), bFGF + Mab HUIV26 (middle) or bFGF + control Mab (bottom). Black arrows indicate angiogenic neovessels. Red arrows indicate preexisting limbal vessels. (B) Quantification of the area of neovascularization within rat corneas. Data bars represent the mean area of neovascularization from the limbus to hydron pellet. Experiments were performed at least twice with five to seven eyes per condition.

Mentions: Previous studies indicated that MMP-2–mediated proteolysis of collagen IV within the basement membrane preparation Matrigel resulted in enhanced endothelial cord formation (Schnaper et al., 1993). Moreover, our studies indicate that the HUIV26 cryptic site can be exposed after thermal denaturation of Matrigel (data not shown). Together, these findings suggest that cellular interactions with the HUIV26 cryptic epitope may facilitate angiogenesis. To examine this possibility, we analyzed the effects of Mab HUIV26 in the rat corneal micropocket assay (Dipietro et al., 1998). Hydron pellets containing either bFGF alone, bFGF plus HUIV26, or bFGF plus an isotype-matched control antibody were surgically implanted into the corneas of rats (Dipietro et al., 1998). After a 5-d incubation period, corneal angiogenesis was quantified (Koch et al., 1995). As shown in Fig. 4 A, bFGF induced a strong angiogenic response in the rat corneas (top). In contrast, little angiogenesis was detected in the corneas implanted with bFGF hydron pellets containing Mab HUIV26 (middle), as compared with either no treatment or treated with an isotype-matched control antibody (top and bottom). In fact, as shown in Fig. 4 B, rat corneal angiogenesis was inhibited by >70% as compared with controls (P < 0.001). Similar results were observed with bFGF- or VEGF-induced angiogenesis within the chick CAM model (data not shown). These findings provide evidence for a potent inhibitory activity for Mab HUIV26 on cytokine-induced angiogenesis in vivo.


Proteolytic exposure of a cryptic site within collagen type IV is required for angiogenesis and tumor growth in vivo.

Xu J, Rodriguez D, Petitclerc E, Kim JJ, Hangai M, Moon YS, Davis GE, Brooks PC, Yuen SM - J. Cell Biol. (2001)

Effects of purified Mab HUIV26 on angiogenesis in vivo. Rat corneal micropocket assays were performed to assess the effects of Mab HUIV26 on angiogenesis. (A) Representative corneas from rats implanted with hydron pellets containing bFGF (top), bFGF + Mab HUIV26 (middle) or bFGF + control Mab (bottom). Black arrows indicate angiogenic neovessels. Red arrows indicate preexisting limbal vessels. (B) Quantification of the area of neovascularization within rat corneas. Data bars represent the mean area of neovascularization from the limbus to hydron pellet. Experiments were performed at least twice with five to seven eyes per condition.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196184&req=5

fig4: Effects of purified Mab HUIV26 on angiogenesis in vivo. Rat corneal micropocket assays were performed to assess the effects of Mab HUIV26 on angiogenesis. (A) Representative corneas from rats implanted with hydron pellets containing bFGF (top), bFGF + Mab HUIV26 (middle) or bFGF + control Mab (bottom). Black arrows indicate angiogenic neovessels. Red arrows indicate preexisting limbal vessels. (B) Quantification of the area of neovascularization within rat corneas. Data bars represent the mean area of neovascularization from the limbus to hydron pellet. Experiments were performed at least twice with five to seven eyes per condition.
Mentions: Previous studies indicated that MMP-2–mediated proteolysis of collagen IV within the basement membrane preparation Matrigel resulted in enhanced endothelial cord formation (Schnaper et al., 1993). Moreover, our studies indicate that the HUIV26 cryptic site can be exposed after thermal denaturation of Matrigel (data not shown). Together, these findings suggest that cellular interactions with the HUIV26 cryptic epitope may facilitate angiogenesis. To examine this possibility, we analyzed the effects of Mab HUIV26 in the rat corneal micropocket assay (Dipietro et al., 1998). Hydron pellets containing either bFGF alone, bFGF plus HUIV26, or bFGF plus an isotype-matched control antibody were surgically implanted into the corneas of rats (Dipietro et al., 1998). After a 5-d incubation period, corneal angiogenesis was quantified (Koch et al., 1995). As shown in Fig. 4 A, bFGF induced a strong angiogenic response in the rat corneas (top). In contrast, little angiogenesis was detected in the corneas implanted with bFGF hydron pellets containing Mab HUIV26 (middle), as compared with either no treatment or treated with an isotype-matched control antibody (top and bottom). In fact, as shown in Fig. 4 B, rat corneal angiogenesis was inhibited by >70% as compared with controls (P < 0.001). Similar results were observed with bFGF- or VEGF-induced angiogenesis within the chick CAM model (data not shown). These findings provide evidence for a potent inhibitory activity for Mab HUIV26 on cytokine-induced angiogenesis in vivo.

Bottom Line: Exposure of this cryptic site was associated with angiogenic, but not quiescent, blood vessels and was required for angiogenesis in vivo.A monoclonal antibody (HUIV26) directed to this site disrupts integrin-dependent endothelial cell interactions and potently inhibits angiogenesis and tumor growth.Together, these studies suggest a novel mechanism by which proteolysis contributes to angiogenesis by exposing hidden regulatory elements within matrix-immobilized collagen type IV.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, Kaplan Cancer Center, New York University School of Medicine, New York, NY 10016, USA.

ABSTRACT
Evidence is provided that proteolytic cleavage of collagen type IV results in the exposure of a functionally important cryptic site hidden within its triple helical structure. Exposure of this cryptic site was associated with angiogenic, but not quiescent, blood vessels and was required for angiogenesis in vivo. Exposure of the HUIV26 epitope was associated with a loss of alpha1beta1 integrin binding and the gain of alphavbeta3 binding. A monoclonal antibody (HUIV26) directed to this site disrupts integrin-dependent endothelial cell interactions and potently inhibits angiogenesis and tumor growth. Together, these studies suggest a novel mechanism by which proteolysis contributes to angiogenesis by exposing hidden regulatory elements within matrix-immobilized collagen type IV.

Show MeSH
Related in: MedlinePlus