Related Experiment Video
Updated: Jun 1, 2026
Isolation, Culture, and Genetic Engineering of Mammalian Primary Pigment Epithelial Cells for Non-Viral Gene Therapy
Published on: February 26, 2021
Pigment epithelium-derived factor inhibits advanced glycation end-products-induced cytotoxicity in retinal pericytes
S Sheikpranbabu1, R Haribalaganesh, S Gurunathan
1Department of biotechnology, division of molecular and cellular biology, Kalasalingam university (Kalasalingam Academy of Research and Education), Krishnankoil-626126, Anand Nagar, Tamilnadu, India.
Aim:
This study investigated the effects of pigment epithelium-derived factor (PEDF) on advanced glycation end-product (AGE)-induced cytotoxicity in porcine retinal pericytes and the signalling mechanism involved.
Methods:
Retinal pericytes were isolated from porcine eyes and characterized by immunocytochemistry. The effect of AGEs and PEDF on cell proliferation was determined by bromodeoxyuridine (BrdU) assay. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity was analyzed by luminescence assay. Reactive oxygen species (ROS), nitric oxide (NO), superoxide dismutase (SOD) and glutathione peroxidase (GSH) were determined by biochemical assays. Induction of apoptosis was determined by caspase-3 colorimetric assay and DNA fragmentation analysis. Src activity was assessed by transient transfection analysis, and the status of Src phosphorylation at Y419 was analyzed by a competitive ELISA method.
Results:
AGEs significantly increased intracellular ROS generation in pericytes via NADPH oxidase and induced cell death via caspase-3 enzyme activation, whereas PEDF increased cell proliferation in a dose-dependent manner. In addition, PEDF inhibited AGE-induced ROS generation by increasing levels of SOD and GSH, and also blocked the activation of caspase-3. Furthermore, PEDF induced cell survival via the Src pathway by Src phosphorylation at Y419, as evidenced by a pharmacological inhibitor and Src mutants.
Conclusion:
These results suggest that PEDF abrogates AGE-induced oxidative stress and apoptosis in retinal pericytes via the Src pathway, thereby suggesting that PEDF is an effective therapeutic agent for the treatment of loss of pericytes in early diabetic retinopathy.
