Previous in vitro studies showed that exchange protein directly activated by
cyclic AMP 1 (Epac1), which is a cAMP mediator, plays an important role in
maintenance of endothelial barrier function. Diabetic retinopathy is
characterized by impairment of retinal blood vessel integrity leading to
breakdown of blood retinal barrier, retinal hypoxia, and neuronal damage. Here,
we hypothesize that Epac1 regulates endothelial permeability and protects retina
from the retinal damage associated with diabetes. To test such hypothesis, we
first demonstrated that human retinal microvascular endothelial cells (HRMECs)
exposed to high glucose concentration at 25 mM or 35 mM showed the
decreased Epac1 expression level. Our preliminary data also showed that
Epac1-downstream activator, Rap1, a member of Ras GTPase, was also altered
by different glucose levels. In addition, retina from type 2 diabetic, db/db, mice
also showed the decreased Epac1 expression compared to that of non-diabetic,
db/m, mice. To further determine the role of Epac1 in diabetic retinopathy, we
made use of Epac1-deficient mice. The pathogenesis of diabetic retinopathy
share similar characteristics to that of ischemic retinopathy, such as neuronal cell
death, glial reactivity, and glutamate toxicity. Therefore, we used our previous
retinal ischemic model, i.e., transient middle cerebral artery occlusion (tMCAO).
Firstly, we determined the retinal morphology of Epac1-/- mice under normal
condition at 3wks. At 3 wks old, the Epac1-/- retinae showed a significantly
decreased thickness of outer plexiform layer (OPL) with a trend of increase in
inner nuclear layer (INL) thickness. Interestingly, there were obviously more
glutamine synthetase (GS)-positive M?ller cells and protein kinase C (PKC)-α
positive rod bipolar cells in INL. In addition, there were more IgG-positive
blood vessels in OPL. To further determine whether these phenotypes will lead
to more severe retinal damage, Epac1-/- mice were exposed to 2 hours of MCAO
followed by 22 hours of reperfusion, which we have previously shown to induce
retinal ischemia. There was no obvious difference in retinal thickness and
expressions of glial fibrillary acidic protein (GFAP) and GS in the contralateral
sides of Epac1+/+ and Epac1-/- retina after tMCAO suggesting that the
Epac1-deficiency may be compensated by either protein kinase A (PKA) or
Epac2. However, Epac2 level was not altered by Epac1-deficiency by Western
blot analysis. The ipsilateral sides of the retina of Epac1+/+ and Epac1-/- after
tMCAO also did not show obvious difference in swelling and cell death in inner
retina, GFAP, glutamate, GS, nitrotyrosine (NT), and peroxiredoxin 6 (Prx6),
suggesting that Epac1-deficiency may have been compensated by other cAMP
mediators, such as Epac2. However, Epac2 expression in the ipsilateral side of
Epac1+/+ and Epac1-/- retinae was not significantly different, although the
activities of Epac and PKA were not determined. Taken together, the
Epac1-deficient mice would serve as a useful model to determine the role of
Epac1 in retinal development, and to determine the detail mechanisms of
pathogenesis of diabetic and ischemic retinopathy. / published_or_final_version / Anatomy / Master / Master of Philosophy
| Identifer | oai:union.ndltd.org:HKU/oai:hub.hku.hk:10722/174370 |
| Date | January 2011 |
| Creators | Liu, Jin, 刘谨 |
| Contributors | Chung, SK |
| Publisher | The University of Hong Kong (Pokfulam, Hong Kong) |
| Source Sets | Hong Kong University Theses |
| Language | English |
| Detected Language | English |
| Type | PG_Thesis |
| Source | http://hub.hku.hk/bib/B47310157 |
| Rights | The author retains all proprietary rights, (such as patent rights) and the right to use in future works., Creative Commons: Attribution 3.0 Hong Kong License |
| Relation | HKU Theses Online (HKUTO) |
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