Role of PI3K/Akt and MEK/ERK in mediating hypoxia-induced expression of HIF-1alpha and VEGF in laser-induced rat choroidal neovascularization.
OBJECTIVE
The transcription factor hypoxia-inducible factor (HIF)-1 plays a central physiological role in oxygen and energy homeostasis and is activated during hypoxia by stabilization of the subunit HIF-1alpha. Hypoxia plays an important role in the development of choroidal neovascularization (CNV). Expression of HIF-1alpha has been demonstrated in CNV. Vascular endothelial growth factor (VEGF) is one of the most well-characterized angiogenic factors in CNV, which is under the regulation of HIF-1. The aim of the present study was to explore the upstream signaling pathways involved in regulating hypoxia-induced expression of HIF-1alpha and VEGF in laser-induced rat CNV.


METHODS
A well-established rat model of CNV and cultured human retinal pigment epithelium (hRPE) was used to investigate the role of PI3K/Akt and MEK/ERK pathways in regulating HIF-1alpha and VEGF expression in CNV in rat and hRPE under hypoxia by immunohistochemistry, Western blot analysis, real-time PCR, and ELISA.


RESULTS
pAkt, pERK, HIF-1alpha, and VEGF were upregulated in vivo and in vitro. PI3K inhibitor (Ly294002) significantly decreased pAkt activity and HIF-1alpha and VEGF expression in vivo and in vitro, whereas MEK inhibitor (PD98059) reduced ERK phosphorylation and the expression of VEGF but had no effect on HIF-1alpha. LY294002 and PD98059 severely inhibited the formation of CNV.


CONCLUSIONS
The PI3K/Akt pathway was required for hypoxia-induced expression of HIF-1alpha and VEGF, whereas the MEK/ERK pathway was required only for VEGF in laser-induced rat CNV.
Yang
Xiu-Mei
XM
Department of Ophthalmology, Eye Institute of Ophthalmology of Chinese PLA, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi Province, China.
Wang
Yu-Sheng
YS
Zhang
Jian
J
Li
Yan
Y
Xu
Jian-Feng
JF
Zhu
Jie
J
Zhao
Wei
W
Chu
Da-Ke
DK
Wiedemann
Peter
P
D000818:Animals; D015153:Blotting, Western; D020256:Choroidal Neovascularization / Q000378:metabolism* / Q000473:pathology; D004195:Disease Models, Animal; D004791:Enzyme Inhibitors / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D005451:Fluorescein Angiography; D000860:Hypoxia / Q000378:metabolism*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism*; D007124:Immunoenzyme Techniques; D017075:Laser Coagulation; D020930:MAP Kinase Kinase Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000502:physiology*; D051381:Rats; D011914:Rats, Inbred BN; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology; D015854:Up-Regulation; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
D004791:Enzyme Inhibitors; D048049:Extracellular Signal-Regulated MAP Kinases; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D020930:MAP Kinase Kinase Kinases; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt; D042461:Vascular Endothelial Growth Factor A; C497444:Hif1a protein, rat; C467498:vascular endothelial growth factor A, rat