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Therapeutic effects of topical netrin-4 inhibits corneal neovascularization in alkali-burn rats.

Han Y, Shao Y, Liu T, Qu YL, Li W, Liu Z - PLoS ONE (2015)

Bottom Line: Netrin-4 and netrin-1 have been found to be either pro- or antiangiogenic factors.We found that netrin-4 functions similarly as netrin-1 in angiogenesis.These results indicate that netrin-4 shed new light on its potential roles in treatmenting for angiogenic diseases that affect the ocular surface, as well as other tissues.

View Article: PubMed Central - PubMed

Affiliation: Eye Institute of Xiamen University, Xiamen, Fujian, China; Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, Xiamen, Fujian, China.

ABSTRACT
Netrins are secreted molecules involved in axon guidance and angiogenesis. However, the role of netrins in the vasculature remains unclear. Netrin-4 and netrin-1 have been found to be either pro- or antiangiogenic factors. Previously, we found that netrin-1 acts as an anti-angiogenic factor in rats by inhibiting alkali burn-induced corneal neovascularization. Here, we further investigate the effects of netrin-4, another member of the same netrin family, on neovascularization in vitro and in vivo. We found that netrin-4 functions similarly as netrin-1 in angiogenesis. In vitro angiogenesis assay shows that netrin-4 affected human umbilical vein endothelial cell (HUVEC) tube formation, viability and proliferation, apoptosis, migration, and invasion in a dose-dependent manner. Netrin-4 was topically applied in vivo to alkali-burned rat corneas on day 0 (immediately after injury) and/or day 10 post-injury. Netrin-4 subsequently suppressed and reversed corneal neovascularization. Netrin-4 inhibited corneal epithelial and stromal cell apoptosis, inhibited vascular endothelial growth factor (VEGF), but promoted pigment epithelium-derived factor (PEDF) expression, decreased NK-KB p65 expression, and inhibits neutrophil and macrophage infiltration. These results indicate that netrin-4 shed new light on its potential roles in treatmenting for angiogenic diseases that affect the ocular surface, as well as other tissues.

No MeSH data available.


Related in: MedlinePlus

Effect of netrin-4 on HUVECs tube formation.Tube formation was assessed in tubulogenesis assay in vitro model, HUVECs were serum-starved with EBM2 medium (0.1% FBS, no growth factor) (Lonza) for 12 hours. HUVECs (1.5×104) were then cultured in a 24-well plate (Gibco) coated with 150 mL Matrigel Basement Membrane Matrix GFR (BD Biosciences). Different concentrations of netrin-4 (100, 500, 1000, and 5000 ng/mL) or PBS (control) were added, and the results were quantified 6 h later using Image J. (B) Representative images out of three independent experiments are shown. * p < 0.05; ** p < 0.01. Each experiment was performed three times and representative pictures are shown. Data are expressed as mean ± SD.
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pone.0122951.g003: Effect of netrin-4 on HUVECs tube formation.Tube formation was assessed in tubulogenesis assay in vitro model, HUVECs were serum-starved with EBM2 medium (0.1% FBS, no growth factor) (Lonza) for 12 hours. HUVECs (1.5×104) were then cultured in a 24-well plate (Gibco) coated with 150 mL Matrigel Basement Membrane Matrix GFR (BD Biosciences). Different concentrations of netrin-4 (100, 500, 1000, and 5000 ng/mL) or PBS (control) were added, and the results were quantified 6 h later using Image J. (B) Representative images out of three independent experiments are shown. * p < 0.05; ** p < 0.01. Each experiment was performed three times and representative pictures are shown. Data are expressed as mean ± SD.

Mentions: In vitro angiogenesis assay was performed according to a previously published protocol [26]. We first assessed the effects of different concentrations of netrin-4 (100, 500, 1000, and 5000 ng/mL) on HUVEC tube formation. The results show that 5000 ng/mL netrin-4 significantly inhibited HUVEC tube formation (Fig 3A and 3B). Therefore, 5000 ng/mL netrin-4 was used in the following experiments.


Therapeutic effects of topical netrin-4 inhibits corneal neovascularization in alkali-burn rats.

Han Y, Shao Y, Liu T, Qu YL, Li W, Liu Z - PLoS ONE (2015)

Effect of netrin-4 on HUVECs tube formation.Tube formation was assessed in tubulogenesis assay in vitro model, HUVECs were serum-starved with EBM2 medium (0.1% FBS, no growth factor) (Lonza) for 12 hours. HUVECs (1.5×104) were then cultured in a 24-well plate (Gibco) coated with 150 mL Matrigel Basement Membrane Matrix GFR (BD Biosciences). Different concentrations of netrin-4 (100, 500, 1000, and 5000 ng/mL) or PBS (control) were added, and the results were quantified 6 h later using Image J. (B) Representative images out of three independent experiments are shown. * p < 0.05; ** p < 0.01. Each experiment was performed three times and representative pictures are shown. Data are expressed as mean ± SD.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4390284&req=5

pone.0122951.g003: Effect of netrin-4 on HUVECs tube formation.Tube formation was assessed in tubulogenesis assay in vitro model, HUVECs were serum-starved with EBM2 medium (0.1% FBS, no growth factor) (Lonza) for 12 hours. HUVECs (1.5×104) were then cultured in a 24-well plate (Gibco) coated with 150 mL Matrigel Basement Membrane Matrix GFR (BD Biosciences). Different concentrations of netrin-4 (100, 500, 1000, and 5000 ng/mL) or PBS (control) were added, and the results were quantified 6 h later using Image J. (B) Representative images out of three independent experiments are shown. * p < 0.05; ** p < 0.01. Each experiment was performed three times and representative pictures are shown. Data are expressed as mean ± SD.
Mentions: In vitro angiogenesis assay was performed according to a previously published protocol [26]. We first assessed the effects of different concentrations of netrin-4 (100, 500, 1000, and 5000 ng/mL) on HUVEC tube formation. The results show that 5000 ng/mL netrin-4 significantly inhibited HUVEC tube formation (Fig 3A and 3B). Therefore, 5000 ng/mL netrin-4 was used in the following experiments.

Bottom Line: Netrin-4 and netrin-1 have been found to be either pro- or antiangiogenic factors.We found that netrin-4 functions similarly as netrin-1 in angiogenesis.These results indicate that netrin-4 shed new light on its potential roles in treatmenting for angiogenic diseases that affect the ocular surface, as well as other tissues.

View Article: PubMed Central - PubMed

Affiliation: Eye Institute of Xiamen University, Xiamen, Fujian, China; Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, Xiamen, Fujian, China.

ABSTRACT
Netrins are secreted molecules involved in axon guidance and angiogenesis. However, the role of netrins in the vasculature remains unclear. Netrin-4 and netrin-1 have been found to be either pro- or antiangiogenic factors. Previously, we found that netrin-1 acts as an anti-angiogenic factor in rats by inhibiting alkali burn-induced corneal neovascularization. Here, we further investigate the effects of netrin-4, another member of the same netrin family, on neovascularization in vitro and in vivo. We found that netrin-4 functions similarly as netrin-1 in angiogenesis. In vitro angiogenesis assay shows that netrin-4 affected human umbilical vein endothelial cell (HUVEC) tube formation, viability and proliferation, apoptosis, migration, and invasion in a dose-dependent manner. Netrin-4 was topically applied in vivo to alkali-burned rat corneas on day 0 (immediately after injury) and/or day 10 post-injury. Netrin-4 subsequently suppressed and reversed corneal neovascularization. Netrin-4 inhibited corneal epithelial and stromal cell apoptosis, inhibited vascular endothelial growth factor (VEGF), but promoted pigment epithelium-derived factor (PEDF) expression, decreased NK-KB p65 expression, and inhibits neutrophil and macrophage infiltration. These results indicate that netrin-4 shed new light on its potential roles in treatmenting for angiogenic diseases that affect the ocular surface, as well as other tissues.

No MeSH data available.


Related in: MedlinePlus