title: ERK1/2/COX-2/PGE2 signaling pathway mediates GPR91-dependent VEGF release in streptozotocin-induced diabetes.
abstract: OBJECTIVE
  Retinal vascular dysfunction caused by vascular endothelial growth factor (VEGF) is the major pathological change that occurs in diabetic retinopathy (DR). It has recently been demonstrated that G protein-coupled receptor 91 (GPR91) plays a major role in both vasculature development and retinal angiogenesis. In this study, we examined the signaling pathways involved in GPR91-dependent VEGF release during the early stages of retinal vascular change in streptozotocin-induced diabetes.
  
  
  METHODS
  Diabetic rats were assigned randomly to receive intravitreal injections of shRNA lentiviral particles targeting GPR91 (LV.shGPR91) or control particles (LV.shScrambled). Accumulation of succinate was assessed by gas chromatography-mass spectrometry (GC-MS). At 14 weeks, the ultrastructure and function of the retinal vessels of diabetic retinas with or without shRNA treatment were assessed using hematoxylin and eosin (HE) staining, transmission electron microscopy (TEM), and Evans blue dye permeability. The expression of GPR91, extracellular signal-regulated kinases 1 and 2 (ERK1/2) and cyclooxygenase-2 (COX-2) were measured using immunofluorescence and western blotting. COX-2 and VEGF mRNA were determined by quantitative RT-PCR. Prostaglandin E2 (PGE2) and VEGF secretion were detected using an enzyme-linked immunosorbent assay.
  
  
  RESULTS
  Succinate exhibited abundant accumulation in diabetic rat retinas. The retinal telangiectatic vessels, basement membrane thickness, and Evans blue dye permeability were attenuated by treatment with GPR91 shRNA. In diabetic rats, knockdown of GPR91 inhibited the activities of ERK1/2 and COX-2 as well as the expression of PGE2 and VEGF. Meanwhile, COX-2, PGE2, and VEGF expression was inhibited by ERK1/2 inhibitor U0126 and COX-2 inhibitor NS-398.
  
  
  CONCLUSIONS
  Our data suggest that hyperglycemia causes succinate accumulation and GPR91 activity in retinal ganglion cells, which mediate VEGF-induced retinal vascular change via the ERK1/2/COX-2/PGE2 pathway. This study highlights the signaling pathway as a potential target for intervention in DR.
authors.affiliation: Department of Ophthalmology, the Sixth People's Hospital, Shanghai Jiaotong University, Shanghai, China.
authors.forename: Tingting
authors.initials: T
authors.lastname: Li
authors.affiliation: Department of Ophthalmology, the Sixth People's Hospital, Shanghai Jiaotong University, Shanghai, China.
authors.forename: Jianyan
authors.initials: J
authors.lastname: Hu
authors.affiliation: Department of Ophthalmology, the Sixth People's Hospital, Shanghai Jiaotong University, Shanghai, China.
authors.forename: Shanshan
authors.initials: S
authors.lastname: Du
authors.affiliation: Department of Ophthalmology, the Sixth People's Hospital, Shanghai Jiaotong University, Shanghai, China.
authors.forename: Yongdong
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Ophthalmology, the Sixth People's Hospital, Shanghai Jiaotong University, Shanghai, China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Wang
authors.affiliation: Department of Ophthalmology, the Sixth People's Hospital, Shanghai Jiaotong University, Shanghai, China ; Shanghai Key Laboratory of Diabetes Mellitus, Shanghai, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Wu
mesh_terms: D000818:Animals; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D003921:Diabetes Mellitus, Experimental / Q000150:complications / Q000378:metabolism* / Q000473:pathology; D003930:Diabetic Retinopathy / Q000209:etiology / Q000378:metabolism* / Q000473:pathology; D015232:Dinoprostone / Q000378:metabolism*; D020935:MAP Kinase Signaling System; D008297:Male; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000008:administration & dosage / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D012165:Retinal Ganglion Cells / Q000378:metabolism / Q000473:pathology; D019802:Succinic Acid / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*
chemical_list: D012333:RNA, Messenger; D034741:RNA, Small Interfering; D043562:Receptors, G-Protein-Coupled; C523503:Sucnr1 protein, rat; D042461:Vascular Endothelial Growth Factor A; C467498:vascular endothelial growth factor A, rat; D019802:Succinic Acid; D051546:Cyclooxygenase 2; C489691:Ptgs2 protein, rat; D015232:Dinoprostone