Coordinated expression of Ets-1, pERK1/2, and VEGF in retina of streptozotocin-induced diabetic rats.
ABSTRACT To investigate the role played by E26 transformation-specific-1 (Ets-1), a transcription factor, and extracellular signal-regulated kinase 1/2 (ERK1/2) in the expression of vascular endothelial growth factor (VEGF), and the interaction of Ets-1 and ERK1/2 in the retina of diabetic rats.
Diabetes was induced in rats by an intraperitoneal injection of streptozotocin (STZ). To follow the time course in the expression of Ets-1, phosphorylated ERK1/2 (pERK1/2), and VEGF, rats were killed at 1, 2, 4, and 8 weeks after the injection of STZ, and total proteins were extracted from the isolated retinas. An adenovirus vector encoding dominant-negative Ets-1 and an inhibitor of PD98059 was injected intravitreally to investigate the effects of Ets-1 blockade and ERK1/2 inhibition on the expression of VEGF. Four weeks after the first intravitreal injection, total proteins and total RNA were extracted from the retinas for Western blot and Northern blot analyses.
The expression of Ets-1, pERK1/2, and VEGF in the retina increased in a time-dependent manner after STZ injection. The phosphorylation of ERK1/2 and protein level of VEGF were significantly reduced following intravitreal Ets-1. Inhibition of ERK1/2 phosphorylation resulted in a significant reduction in the expression of Ets-1 and the level of VEGF protein.
These results indicate that in the retina of STZ-induced diabetic rats: (1) the alterations of Ets-1, pERK1/2, and VEGF are approximately synchronized; (2) the phosphorylation of ERK1/2 is regulated by the expression of Ets-1; (3) the production of Ets-1 protein is dependent on the ERK1/2 pathway, and (4) the protein level of VEGF is regulated by both Ets-1 expression and ERK1/2 phosphorylation. We propose that VEGF, Ets-1, and ERK1 act synergistically in the development of diabetic retinopathy.
Article: Apoptosis induction by oxidized glycated LDL in human retinal capillary pericytes is independent of activation of MAPK signaling pathways.[show abstract] [hide abstract]
ABSTRACT: Pericyte loss is a cardinal feature of early diabetic retinopathy. We previously reported that highly oxidized-glycated low density lipoprotein (HOG-LDL) induces pericyte apoptosis in vitro. In this study, we investigated the role of the mitogen-activated protein kinase (MAPK) signaling pathways in HOG-LDL-induced apoptosis in human pericytes. Human retinal capillary pericytes (HRCP) were exposed to native LDL (N-LDL) and HOG-LDL, and apoptosis was measured using flow cytometry. Time- and dose-dependent responses of extracellular signal-regulated kinase (ERK), p38, and Jun N-terminal kinase (JNK) following exposure to N-LDL or HOG-LDL were determined using western blotting. U0126 (ERK inhibitor), SB203580 (p38 inhibitor), and SP600125 (JNK inhibitor) were used to determine the role of MAPK signaling in HOG-LDL-induced apoptosis. HOG-LDL induced apoptosis in HRCP in a dose-dependent manner at concentrations from 5 to 50 mg/l, with a constant effect from 50 to 200 mg/l. When compared to serum-free medium (SFM), this effect of HOG-LDL was found to be significant at all doses above 10 mg/l. In contrast, N-LDL at 200 mg/l did not induce apoptosis compared with SFM. Exposure to N-LDL versus HOG-LDL induced similar phosphorylation of ERK, p38, and JNK, peaking at 5 min, with similar dose-dependent responses up to 25 mg/l that were constant from 25 to 100 mg/l. Blocking of the ERK, p38, and JNK pathways did not inhibit pericyte apoptosis induced by HOG-LDL. Our data suggest that apoptosis induced by HOG-LDL in HRCP is independent of the activation of MAPK signaling pathways.Molecular vision 02/2009; 15:135-45. · 2.20 Impact Factor