title: Formononetin, an active compound ofAstragalus membranaceus(Fisch) Bunge, inhibits hypoxia-induced retinal neovascularization via the HIF-1α/VEGF signaling pathway.
abstract: BACKGROUND
  It has been reported that formononetin (FMN), one of the main ingredients from famous traditional Chinese medicine "Huang-qi" (Astragalus membranaceus[Fisch] Bunge) for Qi-tonifying, exhibits the effects of immunomodulation and tumor growth inhibition via antiangiogenesis. Furthermore,A. membranaceusmay alleviate the retinal neovascularization (NV) of diabetic retinopathy. However, the information of FMN on retinal NV is limited so far. In the present study, we investigated the effects of FMN on the hypoxia-induced retinal NV and the possible related mechanisms.
  
  
  METHODS
  The VEGF secretion model of acute retinal pigment epithelial-19 (ARPE-19) cells under chemical hypoxia was established by the exposure of cells to 150 μM CoCl2and then cells were treated with 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1, a potent HIF-1α inhibitor, 1.0 μg/mL) or different concentrations of FMN (0.2 μg/mL, 1.0 μg/mL, and 5.0 μg/mL). The supernatants of cells were collected 48 hours later to measure the VEGF concentrations, following the manufacturer's instruction. The mRNA expressions of VEGF, HIF-1α, PHD-2, and β-actin were analyzed by quantitative reverse transcription polymerase chain reaction, and the protein expressions of HIF-1α and PHD-2 were determined by Western blot analysis. Furthermore, the rats with retinopathy were treated by intraperitoneal administration of conbercept injection (1.0 mg/kg) or FMN (5.0 mg/kg and 10.0 mg/kg) in an 80% oxygen atmosphere. The retinal avascular areas were assessed through visualization of the retinal vasculature by adenosine diphosphatase staining and hematoxylin and eosin staining.
  
  
  RESULTS
  FMN can indeed inhibit the VEGF secretion of ARPE-19 cells under hypoxia, downregulate the mRNA expression of VEGFA and PHD-2, and decrease the protein expression of VEGF, HIF-1α, and PHD-2 in vitro. Furthermore, FMN can prevent hypoxia-induced retinal NV in vivo.
  
  
  CONCLUSIONS
  FMN can ameliorate retinal NV via the HIF-1α/VEGF signaling pathway, and it may become a potential drug for the prevention and treatment of diabetic retinopathy.
authors.affiliation: Laboratory of Chinese Materia Medica, Department of Pharmacology, School of Pharmacy, Southwest Medical University, Luzhou; Post-Doctoral Research Station, Kanghong Pharmaceutical Group; Post-Doctoral Mobile Station, West China School of Pharmacy, Sichuan University, Chengdu, Sichuan, People's Republic of China.
authors.forename: Jianming
authors.initials: J
authors.lastname: Wu
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Xiao
authors.initials: X
authors.lastname: Ke
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Na
authors.initials: N
authors.lastname: Ma
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Wei
authors.initials: W
authors.lastname: Fu
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Hongcheng
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Manxi
authors.initials: M
authors.lastname: Zhao
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Xiaoping
authors.initials: X
authors.lastname: Gao
authors.affiliation: Post-Doctoral Research Station, Kanghong Pharmaceutical Group.
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Hao
authors.affiliation: Post-Doctoral Mobile Station, West China School of Pharmacy, Sichuan University, Chengdu, Sichuan, People's Republic of China.
authors.forename: Zhirong
authors.initials: Z
authors.lastname: Zhang
mesh_terms: D000818:Animals; D027885:Astragalus propinquus / Q000737:chemistry*; D015687:Cell Hypoxia / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis / Q000378:metabolism*; D007529:Isoflavones / Q000008:administration & dosage / Q000494:pharmacology*; D015394:Molecular Structure; D051381:Rats; D017207:Rats, Sprague-Dawley; D015861:Retinal Neovascularization / Q000188:drug therapy* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013329:Structure-Activity Relationship; D042442:Vascular Endothelial Growth Factors / Q000096:biosynthesis / Q000378:metabolism*
chemical_list: C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007529:Isoflavones; D042442:Vascular Endothelial Growth Factors; C007768:formononetin
keywords: angiogenesis; formononetin; hypoxia-inducible factor-1; oxygen-induced retinopathy; vascular endothelial growth factor