ABSTRACT
Angiogenesis plays a critical role in many pathological processes, including irreversible blindness in eye diseases such as retinopathy of prematurity. Endothelial mitochondria are dynamic organelles that undergo constant fusion and fission and are critical signalling hubs that modulate angiogenesis by coordinating reactive oxygen species (ROS) production and calcium signalling and metabolism. In this study, we investigated the role of mitochondrial dynamics in pathological retinal angiogenesis. We showed that treatment with vascular endothelial growth factor (VEGF; 20 ng/ml) induced mitochondrial fission in HUVECs by promoting the phosphorylation of dynamin-related protein 1 (DRP1). DRP1 knockdown or pretreatment with the DRP1 inhibitor Mdivi-1 (5 µM) blocked VEGF-induced cell migration, proliferation, and tube formation in HUVECs. We demonstrated that VEGF treatment increased mitochondrial ROS production in HUVECs, which was necessary for HIF-1α-dependent glycolysis, as well as proliferation, migration, and tube formation, and the inhibition of mitochondrial fission prevented VEGF-induced mitochondrial ROS production. In an oxygen-induced retinopathy (OIR) mouse model, we found that active DRP1 was highly expressed in endothelial cells in neovascular tufts. The administration of Mdivi-1 (10 mg·kg-1·d-1, i.p.) for three days from postnatal day (P) 13 until P15 significantly alleviated pathological angiogenesis in the retina. Our results suggest that targeting mitochondrial fission may be a therapeutic strategy for proliferative retinopathies and other diseases that are dependent on pathological angiogenesis.
Subject(s)
Cell Movement , Dynamins , Human Umbilical Vein Endothelial Cells , Hypoxia-Inducible Factor 1, alpha Subunit , Mice, Inbred C57BL , Mitochondrial Dynamics , Quinazolinones , Reactive Oxygen Species , Retinal Neovascularization , Vascular Endothelial Growth Factor A , Mitochondrial Dynamics/drug effects , Animals , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Humans , Reactive Oxygen Species/metabolism , Dynamins/metabolism , Dynamins/antagonists & inhibitors , Vascular Endothelial Growth Factor A/metabolism , Quinazolinones/pharmacology , Retinal Neovascularization/metabolism , Retinal Neovascularization/pathology , Retinal Neovascularization/drug therapy , Cell Movement/drug effects , Mice , Cell Proliferation/drug effects , Mitochondria/metabolism , Mitochondria/drug effects , AngiogenesisABSTRACT
OBJECTIVE: To evaluate the curative effect of KGF-2 on the alkali-injured rabbit eye and to investigate the mechanism of KGF-2 accelerating corneal epithelial wound healing. METHODS: Alkali burn was produced in 24 corneas from 24 New Zealand rabbits. Four groups were randomly divided. Three groups (A, B, and C groups) were treated with KGF-2 solution (1, 50, 100 microg/ml, respectively), and one group (D group) was treated with phosphate-buffered saline (PBS) solution. The injured eyes were photographed after the fluorescence staining with a slit lamp and the pictures were analyzed with computer-aided picture analysis system to calculate the rate of corneal epithelial healing. Morphologic and immunohistological examinations (using P63, AE5 and EGFR antibodies) of the cornea were performed. RESULTS: KGF-2 at dosages ranging from 1 microg/ml to 100 microg/ml could enhance the cornea wound healing process. After 24 hours, epithelial healing rate of the 100 microg/ml KGF-2 group and the PBS treated group was 74% and 40%, respectively (P < 0.05). The corneal epithelial healing rate of each group was variable after four days and achieved complete healing after ten days. The P63 positive cells in KGF-2 groups appeared not only in the limbal area but also in the central area. For example, on the seventh day, in the limbal area, the P63 positive cells in the 100 microg/ml KGF-2 group, the PBS treated group and the normal group were 53.8 +/- 2.6, 29.5 +/- 2.2 and 17.0 +/- 2.1, respectively (P = 0.000). At the same time, the P63 positive cells in the non-limbal area in the 100 microg/ml KGF-2 group, the PBS treated group and the normal group were 69.5 +/- 2.8, 19.5 +/- 2.8 and 0, respectively (P = 0.000). CONCLUSIONS: These results suggested KGF-2 can stimulate the limbal epithelial stem cells to migrate to the central cornea. KGF-2 can accelerate the healing of alkali burned cornea.
