Diabetic retinopathy remains a leading cause of blindness in the United States. Recent clinical trials have demonstrated that targeting vascular endothelial growth factor (VEGF) can effectively prevent progression of vision loss and for some patients restore visual acuity. These studies demonstrate that medical therapy for the retina can effectively treat diabetic retinopathy. However, not all patients respond to anti-VEGF therapies, which require repeat intra-ocular injections with the risk of endophthalmitis. In addition to VEGF, a number of inflammatory factors are elevated in patients with diabetic retinopathy that are believed to contribute to disease pathology including tumor necrosis factor, the chemokine (C-C motif) ligand 2 (CCL2), interleukin 6 and 8. Therefore, understanding the mechanisms by which growth factors and inflammatory cytokines alter the retinal vascular endothelium leading to vascular permeability and angiogenesis remains of high significance. Research from the previous funding period has identified two important signaling pathways that control permeability in response to VEGF. The first pathway involves conventional protein kinase C (PKC) activation and phosphorylation of the tight junction protein occludin and is required for VEGF induced vascular permeability. Published and preliminary data also reveal that occludin phosphorylation contributes to growth control and angiogenesis. Thus, occludin phosphorylation downstream of VEGF activation contributes to both vascular permeability and angiogenesis suggesting an important role in growth and blood-retinal barrier differentiation. Further, research over the previous funding period has identified a second signaling pathway involving atypical PKC activation as required for permeability response to both VEGF and inflammatory cytokines such as TNF and CCL2. Utilizing mass spectrometry phosphoproteomic analysis we have identified downstream targets linking this pathway to small G-protein regulation and control of permeability. Here we propose to define the role of occludin phosphorylation in permeability and angiogenesis in vivo while also elucidating the aPKC activation pathway to the control of vascular permeability. Collectively, these studies will provide novel insight into the mechanisms of VEGF induced permeability and angiogenesis specifically through the control of the tight junctions' complex and small G-protein regulation of the cytoskeleton. This research is expected to provide new insight into the nature of blood vessel growth and maturation and identify new targets for therapeutic intervention that are effective against both growth factors and inflammatory cytokines.

Public Health Relevance

This proposal provides innovative research to determine the mechanisms by which endothelial cells proliferate and mature contributing to the blood-retinal barrier and how diabetes alters these vessels leading to macular edema and pathologic angiogenesis. Importantly, this research explores required pathways that regulate vascular permeability and may lead to new therapies for macular edema.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY012021-18
Application #
8989102
Study Section
Special Emphasis Panel (ZRG1-CB-G (02))
Program Officer
Shen, Grace L
Project Start
1998-09-01
Project End
2019-12-31
Budget Start
2016-01-01
Budget End
2016-12-31
Support Year
18
Fiscal Year
2016
Total Cost
$386,360
Indirect Cost
$137,095
Name
University of Michigan Ann Arbor
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Muthusamy, Arivalagan; Lin, Cheng-Mao; Shanmugam, Sumathi et al. (2014) Ischemia-reperfusion injury induces occludin phosphorylation/ubiquitination and retinal vascular permeability in a VEGFR-2-dependent manner. J Cereb Blood Flow Metab 34:522-31
Keil, Jason M; Liu, Xuwen; Antonetti, David A (2013) Glucocorticoid induction of occludin expression and endothelial barrier requires transcription factor p54 NONO. Invest Ophthalmol Vis Sci 54:4007-15
Titchenell, Paul M; Hollis Showalter, H D; Pons, Jean-Francois et al. (2013) Synthesis and structure-activity relationships of 2-amino-3-carboxy-4-phenylthiophenes as novel atypical protein kinase C inhibitors. Bioorg Med Chem Lett 23:3034-8
Liu, Chunjian; Lin, James; Everlof, Gerry et al. (2013) Synthesis and evaluation of carbamoylmethylene linked prodrugs of BMS-582949, a clinical p38? inhibitor. Bioorg Med Chem Lett 23:3028-33
Abcouwer, Steven F; Lin, Cheng-Mao; Shanmugam, Sumathi et al. (2013) Minocycline prevents retinal inflammation and vascular permeability following ischemia-reperfusion injury. J Neuroinflammation 10:149
Titchenell, Paul M; Antonetti, David A (2013) Using the past to inform the future: anti-VEGF therapy as a road map to develop novel therapies for diabetic retinopathy. Diabetes 62:1808-15
Murakami, Tomoaki; Frey, Tiffany; Lin, Chengmao et al. (2012) Protein kinase c? phosphorylates occludin regulating tight junction trafficking in vascular endothelial growth factor-induced permeability in vivo. Diabetes 61:1573-83
Runkle, E Aaron; Sundstrom, Jeffrey M; Runkle, Kristin B et al. (2011) Occludin localizes to centrosomes and modifies mitotic entry. J Biol Chem 286:30847-58
Cai, Jun; Wu, Lin; Qi, Xiaoping et al. (2011) Placenta growth factor-1 exerts time-dependent stabilization of adherens junctions following VEGF-induced vascular permeability. PLoS One 6:e18076
Lopez-Quintero, Sandra V; Ji, Xin-Ying; Antonetti, David A et al. (2011) A three-pore model describes transport properties of bovine retinal endothelial cells in normal and elevated glucose. Invest Ophthalmol Vis Sci 52:1171-80

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