Aims 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.
References
[1]
Antonetti DA, Klein R, Gardner TW (2012) Diabetic retinopathy. N Engl J Med 366: 1227–1239. doi: 10.1056/nejmra1005073
[2]
Bressler NM, Beck RW, Ferris FL III (2011) Panretinal photocoagulation for proliferative diabetic retinopathy. N Engl J Med 365: 1520–1526. doi: 10.1056/nejmct0908432
[3]
Mason JO III, Colagross CT, Vail R (2006) Diabetic vitrectomy: risks, prognosis, future trends. Curr Opin Ophthalmol 17: 281–285. doi: 10.1097/01.icu.0000193098.28798.18
[4]
Stitt AW, Lois N, Medina RJ, Adamson P, Curtis TM (2013) Advances in our understanding of diabetic retinopathy. Clin Sci (Lond) 125: 1–17. doi: 10.1042/cs20120588
[5]
Chen Y, Hu Y, Lin M, Jenkins AJ, Keech AC, et al. (2013) Therapeutic effects of PPARalpha agonists on diabetic retinopathy in type 1 diabetes models. Diabetes 62: 261–272. doi: 10.2337/db11-0413
[6]
Hammes HP, Lin J, Renner O, Shani M, Lundqvist A, et al. (2002) Pericytes and the pathogenesis of diabetic retinopathy. Diabetes 51: 3107–3112. doi: 10.2337/diabetes.51.10.3107
[7]
Robinson R, Barathi VA, Chaurasia SS, Wong TY, Kern TS (2012) Update on animal models of diabetic retinopathy: from molecular approaches to mice and higher mammals. Dis Model Mech 5: 444–456.
[8]
Cheung N, Mitchell P, Wong TY (2010) Diabetic retinopathy. Lancet 376: 124–136. doi: 10.1016/s0140-6736(09)62124-3
[9]
Ceriello A, Esposito K, Ihnat M, Thorpe J, Giugliano D (2009) Long-term glycemic control influences the long-lasting effect of hyperglycemia on endothelial function in type 1 diabetes. J Clin Endocrinol Metab 94: 2751–2756. doi: 10.1210/jc.2009-0762
[10]
Kowluru RA (2003) Effect of reinstitution of good glycemic control on retinal oxidative stress and nitrative stress in diabetic rats. Diabetes 52: 818–823. doi: 10.2337/diabetes.52.3.818
[11]
Zhang L, Chen B, Tang L (2012) Metabolic memory: mechanisms and implications for diabetic retinopathy. Diabetes Res Clin Pract 96: 286–293. doi: 10.1016/j.diabres.2011.12.006
[12]
Kowluru RA, Chan PS (2007) Oxidative stress and diabetic retinopathy. Exp Diabetes Res 2007: 43603. doi: 10.1155/2007/43603
[13]
Wardlaw SL (2011) Hypothalamic proopiomelanocortin processing and the regulation of energy balance. Eur J Pharmacol 660: 213–219. doi: 10.1016/j.ejphar.2010.10.107
[14]
Yang Y (2011) Structure, function and regulation of the melanocortin receptors. Eur J Pharmacol 660: 125–130. doi: 10.1016/j.ejphar.2010.12.020
[15]
Zhang Y, Wu X, He Y, Kastin AJ, Hsuchou H, et al. (2009) Melanocortin potentiates leptin-induced STAT3 signaling via MAPK pathway. J Neurochem 110: 390–399. doi: 10.1111/j.1471-4159.2009.06144.x
[16]
Voisey J, Carroll L, van Daal A (2003) Melanocortins and their receptors and antagonists. Curr Drug Targets 4: 586–597. doi: 10.2174/1389450033490858
[17]
Nohara K, Zhang Y, Waraich RS, Laque A, Tiano JP, et al. (2011) Early-life exposure to testosterone programs the hypothalamic melanocortin system. Endocrinology 152: 1661–1669. doi: 10.1210/en.2010-1288
[18]
Baumgartl HJ, Standl E, Schmidt-Gayk H, Kolb HJ, Janka HU, et al. (1991) Changes of vitamin D3 serum concentrations at the onset of immune-mediated type 1 (insulin-dependent) diabetes mellitus. Diabetes Res 16: 145–148.
[19]
Taylor AW, Lee D (2010) Applications of the Role of alpha-MSH in Ocular Immune Privilege. Adv Exp Med Biol 681: 143–149. doi: 10.1007/978-1-4419-6354-3_12
[20]
Shiratori K, Ohgami K, Ilieva IB, Koyama Y, Yoshida K, et al. (2004) Inhibition of endotoxin-induced uveitis and potentiation of cyclooxygenase-2 protein expression by alpha-melanocyte-stimulating hormone. Invest Ophthalmol Vis Sci 45: 159–164. doi: 10.1167/iovs.03-0492
[21]
Edling AE, Gomes D, Weeden T, Dzuris J, Stefano J, et al. (2011) Immunosuppressive activity of a novel peptide analog of alpha-melanocyte stimulating hormone (alpha-MSH) in experimental autoimmune uveitis. J Neuroimmunol 236: 1–9. doi: 10.1016/j.jneuroim.2011.04.015
[22]
Naveh N (2003) Melanocortins applied intravitreally delay retinal dystrophy in Royal College of Surgeons rats. Graefes Arch Clin Exp Ophthalmol 241: 1044–1050. doi: 10.1007/s00417-003-0781-y
[23]
Henri P, Beaumel S, Guezennec A, Poumes C, Stoebner PE, et al. (2011) MC1R expression in HaCaT keratinocytes inhibits UVA-induced ROS production via NADPH oxidase- and cAMP-dependent mechanisms. J Cell Physiol 227: 2578–2585. doi: 10.1002/jcp.22996
[24]
Kadekaro AL, Chen J, Yang J, Chen S, Jameson J, et al. (2012) Alpha-melanocyte-stimulating hormone suppresses oxidative stress through a p53-mediated signaling pathway in human melanocytes. Mol Cancer Res 10: 778–786. doi: 10.1158/1541-7786.mcr-11-0436
[25]
Kokot A, Metze D, Mouchet N, Galibert MD, Schiller M, et al. (2009) Alpha-melanocyte-stimulating hormone counteracts the suppressive effect of UVB on Nrf2 and Nrf-dependent gene expression in human skin. Endocrinology 150: 3197–3206. doi: 10.1210/en.2008-1315
[26]
Accili D, Arden KC (2004) FoxOs at the crossroads of cellular metabolism, differentiation, and transformation. Cell 117: 421–426. doi: 10.1016/s0092-8674(04)00452-0
[27]
Eijkelenboom A, Burgering BM (2013) FOXOs: signalling integrators for homeostasis maintenance. Nat Rev Mol Cell Biol 14: 83–97. doi: 10.1038/nrm3507
[28]
Ponugoti B, Dong G, Graves DT (2012) Role of forkhead transcription factors in diabetes-induced oxidative stress. Exp Diabetes Res 2012: 939751. doi: 10.1155/2012/939751
[29]
Tsuchiya K, Tanaka J, Shuiqing Y, Welch CL, DePinho RA, et al. (2012) FoxOs integrate pleiotropic actions of insulin in vascular endothelium to protect mice from atherosclerosis. Cell Metab 15: 372–381. doi: 10.1016/j.cmet.2012.01.018
[30]
Chuang PY, Yu Q, Fang W, Uribarri J, He JC (2007) Advanced glycation endproducts induce podocyte apoptosis by activation of the FOXO4 transcription factor. Kidney Int 72: 965–976. doi: 10.1038/sj.ki.5002456
[31]
Van Der Heide LP, Hoekman MF, Smidt MP (2004) The ins and outs of FoxO shuttling: mechanisms of FoxO translocation and transcriptional regulation. Biochem J 380: 297–309. doi: 10.1042/bj20040167
[32]
Urbich C, Knau A, Fichtlscherer S, Walter DH, Bruhl T, et al. (2005) FOXO-dependent expression of the proapoptotic protein Bim: pivotal role for apoptosis signaling in endothelial progenitor cells. FASEB J 19: 974–976. doi: 10.1096/fj.04-2727fje
Dietrich N, Hammes HP (2012) Retinal digest preparation: a method to study diabetic retinopathy. Methods Mol Biol 933: 291–302. doi: 10.1007/978-1-62703-068-7_19
[38]
Hu B, Zhang Y, Zeng Q, Han Q, Zhang L, et al. (2013) Intravitreal Injection of Ranibizumab and CTGF shRNA Improves Retinal Gene Expression and Microvessel Ultrastructure in a Rodent Model of Diabetes. Int J Mol Sci 15: 1606–1624. doi: 10.3390/ijms15011606
[39]
Jung DS, Li JJ, Kwak SJ, Lee SH, Park J, et al. (2008) FR167653 inhibits fibronectin expression and apoptosis in diabetic glomeruli and in high-glucose-stimulated mesangial cells. Am J Physiol Renal Physiol 295: F595–F604. doi: 10.1152/ajprenal.00624.2007
[40]
Xu L, Qu Z, Guo F, Pang M, Gao S, et al. (2013) Effects of ghrelin on gastric distention sensitive neurons in the arcuate nucleus of hypothalamus and gastric motility in diabetic rats. Peptides 48: 137–146. doi: 10.1016/j.peptides.2013.08.010
[41]
Hazra S, Rasheed A, Bhatwadekar A, Wang X, Shaw LC, et al. (2012) Liver X receptor modulates diabetic retinopathy outcome in a mouse model of streptozotocin-induced diabetes. Diabetes 61: 3270–3279. doi: 10.2337/db11-1596
[42]
Li Q, Verma A, Han PY, Nakagawa T, Johnson RJ, et al. (2010) Diabetic eNOS-knockout mice develop accelerated retinopathy. Invest Ophthalmol Vis Sci 51: 5240–5246. doi: 10.1167/iovs.09-5147
[43]
Stitt AW, Li YM, Gardiner TA, Bucala R, Archer DB, et al. (1997) Advanced glycation end products (AGEs) co-localize with AGE receptors in the retinal vasculature of diabetic and of AGE-infused rats. Am J Pathol 150: 523–531.
[44]
Hammes HP, Federoff HJ, Brownlee M (1995) Nerve growth factor prevents both neuroretinal programmed cell death and capillary pathology in experimental diabetes. Mol Med 1: 527–534.
[45]
Barber AJ, Lieth E, Khin SA, Antonetti DA, Buchanan AG, et al. (1998) Neural apoptosis in the retina during experimental and human diabetes. Early onset and effect of insulin. J Clin Invest 102: 783–791. doi: 10.1172/jci2425
[46]
Barber AJ, Gardner TW, Abcouwer SF (2011) The significance of vascular and neural apoptosis to the pathology of diabetic retinopathy. Invest Ophthalmol Vis Sci 52: 1156–1163. doi: 10.1167/iovs.10-6293
[47]
Zhang B, Hu Y, Ma JX (2009) Anti-inflammatory and antioxidant effects of SERPINA3K in the retina. Invest Ophthalmol Vis Sci 50: 3943–3952. doi: 10.1167/iovs.08-2954
[48]
Jarrett SG, Boulton ME (2012) Consequences of oxidative stress in age-related macular degeneration. Mol Aspects Med 33: 399–417. doi: 10.1016/j.mam.2012.03.009
[49]
Fujita K, Yamafuji M, Nakabeppu Y, Noda M (2012) Therapeutic approach to neurodegenerative diseases by medical gases: focusing on redox signaling and related antioxidant enzymes. Oxid Med Cell Longev 2012: 324256. doi: 10.1155/2012/324256
[50]
Schaible EV, Steinstrasser A, Jahn-Eimermacher A, Luh C, Sebastiani A, et al. (2013) Single Administration of Tripeptide alpha-MSH(11-13) Attenuates Brain Damage by Reduced Inflammation and Apoptosis after Experimental Traumatic Brain Injury in Mice. PLoS One 8: e71056. doi: 10.1371/journal.pone.0071056
[51]
Spaccapelo L, Bitto A, Galantucci M, Ottani A, Irrera N, et al. (2011) Melanocortin MC(4) receptor agonists counteract late inflammatory and apoptotic responses and improve neuronal functionality after cerebral ischemia. Eur J Pharmacol 670: 479–486. doi: 10.1016/j.ejphar.2011.09.015
[52]
Forslin AA, Spulber S, Oprica M, Winblad B, Post C, et al. (2007) Alpha-MSH rescues neurons from excitotoxic cell death. J Mol Neurosci 33: 239–251. doi: 10.1007/s12031-007-0019-2
[53]
Liu M, Zhang Y, Liu X, Zhang LJ, Li SL, et al. (2013) Protective effects of alpha-Melanocyte Stimulating Hormone on glutamate-induced retinal excitotoxicity. Chinese Journal of Experimental Ophthalmology 31: 440–445.
[54]
Aleksandrushkina NI, Vanyushin BF (2012) Endonucleases and apoptosis in animals. Biochemistry (Mosc) 77: 1436–1451. doi: 10.1134/s0006297912130032
[55]
Kay JG, Grinstein S (2013) Phosphatidylserine-mediated cellular signaling. Adv Exp Med Biol 991: 177–193. doi: 10.1007/978-94-007-6331-9_10
[56]
Chuang PY, Dai Y, Liu R, He H, Kretzler M, et al. (2011) Alteration of forkhead box O (foxo4) acetylation mediates apoptosis of podocytes in diabetes mellitus. PLoS One 6: e23566. doi: 10.1371/journal.pone.0023566
[57]
Behl Y, Krothapalli P, Desta T, Roy S, Graves DT (2009) FOXO1 plays an important role in enhanced microvascular cell apoptosis and microvascular cell loss in type 1 and type 2 diabetic rats. Diabetes 58: 917–925. doi: 10.2337/db08-0537
[58]
Wilson JF, Harry FM (1980) Release, distribution and half-life of alpha-melanotrophin in the rat. J Endocrinol 86: 61–67. doi: 10.1677/joe.0.0860061
[59]
Ugwu SO, Blanchard J, Dorr RT, Levine N, Brooks C, et al. (1997) Skin pigmentation and pharmacokinetics of melanotan-I in humans. Biopharm Drug Dispos 18: 259–269. doi: 10.1002/(sici)1099-081x(199704)18:3<259::aid-bdd20>3.0.co;2-x
[60]
Rudman D, Hollins BM, Kutner MH, Moffitt SD, Lynn MJ (1983) Three types of alpha-melanocyte-stimulating hormone: bioactivities and half-lives. Am J Physiol 245: E47–E54.
[61]
Lee DJ, Biros DJ, Taylor AW (2009) Injection of an alpha-melanocyte stimulating hormone expression plasmid is effective in suppressing experimental autoimmune uveitis. Int Immunopharmacol 9: 1079–1086. doi: 10.1016/j.intimp.2009.05.001
[62]
Rahmouni K, Haynes WG, Morgan DA, Mark AL (2003) Role of melanocortin-4 receptors in mediating renal sympathoactivation to leptin and insulin. J Neurosci 23: 5998–6004.
[63]
Haynes WG, Morgan DA, Djalali A, Sivitz WI, Mark AL (1999) Interactions between the melanocortin system and leptin in control of sympathetic nerve traffic. Hypertension 33: 542–547. doi: 10.1161/01.hyp.33.1.542
[64]
da Silva AA, do Carmo JM, Kanyicska B, Dubinion J, Brandon E, et al. (2008) Endogenous melanocortin system activity contributes to the elevated arterial pressure in spontaneously hypertensive rats. Hypertension 51: 884–890. doi: 10.1161/hypertensionaha.107.100636
