In the present study, we have examined a potential mechanism by which
sympathetic nerves regulate PEDF and whether its down regulation may be responsible for increased
capillary density observed after
sympathectomy. Six weeks post-sympathectomy,
eyes were removed from female
Sprague-Dawley rats for
Western blot analysis,
RNA isolation,
real-time PCR, and
immunohistochemistry for measurement of PEDF expression. The
contralateral or left
eye was used as an intra-animal control. In addition,
retinal pigment epithelial cells were grown in culture and treated with
norepinephrine and
propranolol. An
ELISA assay was used to determine the amount of PEDF
secreted into the RPE media. Quantitative results of
Western blot analysis and
real-time PCR confirm that both steady-state
gene expression and protein levels of PEDF are significantly decreased in the sympathectomized
retina (P<0.05) when compared to the
contralateral retina. Qualitative results of
immunohistochemistry verify that PEDF is located predominantly in the RPE cell layer of the
retina, and levels are decreased in the sympathectomized
retina.
ELISA results illustrate that
norepinephrine significantly increases PEDF
secretion by RPE cells and
propranolol slightly decreases PEDF
secretion into RPE cell medium. In conclusion, down regulation of PEDF may contribute to the increased
capillary density of the
outer plexiform layer in the
retina noted after
sympathectomy. Furthermore, expression of PEDF was significantly increased after treatment of
norepinephrine in RPE medium demonstrating a role of beta-adrenergic regulation of PEDF. Since
sympathetic nerves are damaged in
diabetes and PEDF appears to be regulated by
beta-adrenergic receptors, these results suggest a role for
sympathetic nerves in
diabetic retinopathy. This knowledge, in turn, may be used for future treatment and prevention of
diabetic retinopathy and other
ocular diseases.
