α-Melanocyte-stimulating hormone protects retinal vascular endothelial cells from oxidative stress and apoptosis in a rat model of diabetes.
OBJECTIVE
Oxidative stress and apoptosis are among the earliest lesions of diabetic retinopathy. This study sought to examine the anti-oxidative and anti-apoptotic effects of α-melanocyte-stimulating hormone (α-MSH) in early diabetic retinas and to explore the underlying mechanisms in retinal vascular endothelial cells.


METHODS
Sprague-Dawley rats were injected intravenously with streptozocin to induce diabetes. The diabetic rats were injected intravitreally with α-MSH or saline. At week 5 after diabetes, the retinas were analyzed for reactive oxygen species (ROS) and gene expression. One week later, the retinas were processed for terminal deoxynucleotidyl transferase dUTP nick-end labeling staining and transmission electron microscopy. Retinal vascular endothelial cells were stimulated by high glucose (HG) with or without α-MSH. The expression of Forkhead box O genes (Foxos) was examined through real-time PCR. The Foxo4 gene was overexpressed in endothelial cells by transient transfection prior to α-MSH or HG treatment, and oxidative stress and apoptosis were analyzed through CM-H2DCFDA and annexin-V assays, respectively.


RESULTS
In diabetic retinas, the levels of H2O2 and ROS and the total anti-oxidant capacity were normalized, the apoptotic cell number was reduced, and the ultrastructural injuries were ameliorated by α-MSH. Treatment with α-MSH also corrected the aberrant changes in eNOS, iNOS, ICAM-1, and TNF-α expression levels in diabetic retinas. Furthermore, α-MSH inhibited Foxo4 up-regulation in diabetic retinas and in endothelial cells exposed to HG, whereas Foxo4 overexpression abrogated the anti-oxidative and anti-apoptotic effects of α-MSH in HG-stimulated retinal vascular endothelial cells.


CONCLUSIONS
α-MSH normalized oxidative stress, reduced apoptosis and ultrastructural injuries, and corrected gene expression levels in early diabetic retinas. The protective effects of α-MSH in retinal vascular endothelial cells may be mediated through the inhibition of Foxo4 up-regulation induced by HG. This study suggests an α-MSH-mediated potential intervention approach to early diabetic retinopathy and a novel regulatory mechanism involving Foxo4.
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Lijuan
L
Zhang
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Lijie
L
Dong
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Xun
X
Liu
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Yuanfeng
Y
Jiang
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Lingjun
L
Zhang
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Xiaomin
X
Zhang
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Xiaorong
X
Li
Tianjin Medical University Eye Hospital/Eye Institute, Tianjin, China.
Yan
Y
Zhang
D000818:Animals; D017209:Apoptosis / Q000235:genetics / Q000502:physiology*; D003921:Diabetes Mellitus, Experimental / Q000235:genetics / Q000378:metabolism*; D003930:Diabetic Retinopathy / Q000235:genetics / Q000378:metabolism; D042783:Endothelial Cells / Q000378:metabolism*; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000235:genetics; D006861:Hydrogen Peroxide / Q000378:metabolism; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism; D008297:Male; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress / Q000235:genetics / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D012160:Retina / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000235:genetics / Q000502:physiology; D000521:alpha-MSH / Q000235:genetics / Q000378:metabolism*
C524737:FOXO4 protein, rat; D051858:Forkhead Transcription Factors; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D000521:alpha-MSH; D006861:Hydrogen Peroxide; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496318:Nos2 protein, rat; C496316:Nos3 protein, rat