Journal of Molecular Neuroscience

, Volume 54, Issue 3, pp 395–404

Davunetide (NAP) Protects the Retina Against Early Diabetic Injury by Reducing Apoptotic Death

  • Soraya Scuderi
  • Agata Grazia D’Amico
  • Alessandro Castorina
  • Concetta Federico
  • Giuseppina Marrazzo
  • Filippo Drago
  • Claudio Bucolo
  • Velia D’Agata


Davunetide (NAP) is an eight amino acid peptide that has been shown to provide potent neuroprotection. In the present study, we investigated the neuroprotective effect of NAP in diabetic retinopathy using an in vivo streptozotocin (STZ)-induced diabetic model. A single intraocular injection of NAP (100 μg/mL) or vehicle was administered 1 week after STZ injection. Three weeks after diabetes induction, we assessed the retinal expression and distribution of apoptosis markers, cleaved caspase-3, and Bcl2, by Western blot and immunofluorescent analysis. Furthermore, we evaluated the activation of mitogen-activated protein kinase/extracellular signal-regulated protein kinase (MAPK/ERK) and/or phosphatidylinositol-3 kinase/Akt pathways by measuring the protein levels of p-ERK and p-AKT with or without NAP treatment. Results demonstrated that NAP treatment reduced apoptotic event in diabetic retina, and it restored cleaved caspase-3 expression levels in the retina of STZ-injected rats as well as the decreased Bcl2. NAP treatment improved cellular survival through the activation of the MAPK/ERK pathway. Taken together, these findings suggested that NAP might be useful to treat retinal degenerative diseases.


Davunetide NAP Diabetic retinopathy Streptozotocin Retina Apoptosis MAPK/ERK pathway 


  1. Arimura A, Somogyvari-Vigh A, Weill C et al (1994) PACAP functions as a neurotrophic factor. Ann N Y Acad Sci 739:228–243PubMedCrossRefGoogle Scholar
  2. Arqués O, Chicote I, Tenbaum S, Puig I, Palmer Héctor G (2012) Standardized relative quantification of immunofluorescence tissue staining. Protocol Exchang. doi:10.1038/protex.2012.008 Google Scholar
  3. Ashur-Fabian O, Segal-Ruder Y, Skutelsky E et al (2003) The neuroprotective peptide NAP inhibits the aggregation of the beta-amyloid peptide. Peptides 24(9):1413–1423PubMedCrossRefGoogle Scholar
  4. Barber AJ, Lieth E, Khin SA, Antonetti DA, Buchanan AG, Gardner TW (1998) Neural apoptosis in the retina during experimental and human diabetes. Early onset and effect of insulin. J Clin Invest 102:783–791PubMedCentralPubMedCrossRefGoogle Scholar
  5. Barber AJ, Gardner TW, Abcouwer SF (2011) The significance of vascular and neural apoptosis to the pathology of diabetic retinopathy. Invest Ophthalmol Vis Sci 52:1156–1163PubMedCentralPubMedCrossRefGoogle Scholar
  6. Bassan M, Zamostiano R, Davidson A et al (1999) Complete sequence of a novel protein containing a femtomolar-activity-dependent neuroprotective peptide. J Neurochem 72(3):1283–1293PubMedCrossRefGoogle Scholar
  7. Belokopytov M, Shulman S, Dubinsky G, Gozes I, Belkin M, Rosner M (2011) Ameliorative effect of NAP on laser-induced retinal damage. Acta Ophthalmol 89(2):e126–e131PubMedCrossRefGoogle Scholar
  8. Beni-Adani L, Gozes I, Cohen Y et al (2001) A peptide derived from activity dependent neuroprotective protein (ADNP) ameliorates injury response in closed head injured mice. J Pharmacol Exp Ther 296:57–63PubMedGoogle Scholar
  9. Brenneman DE, Gozes I (1996) A femtomolar-acting neuroprotective peptide. J Clin Invest 97(10):2299–2307PubMedCentralPubMedCrossRefGoogle Scholar
  10. Bucolo C, Leggio GM, Drago F, Salomone S (2012) Eriodictyol prevents early retinal and plasma abnormalities in streptozotocin-induced diabetic rats. Biochem Pharmacol 84(1):88–92PubMedCrossRefGoogle Scholar
  11. Busciglio J, Pelsman A, Helguera P, Ashur-Fabian O, Pinhasov A, Brenneman DE, Gozes I (2007) NAP and ADNF-9 protect normal and Down’s syndrome cortical neurons from oxidative damage and apoptosis. Curr Pharm Des 13(11):1091–1098PubMedCrossRefGoogle Scholar
  12. Castorina A, Giunta S, Scuderi S, D’Agata V (2012) Involvement of PACAP/ADNP signaling in the resistance to cell death in malignant peripheral nerve sheath tumor (MPNST) cells. J Mol Neurosci 48(3):674–683PubMedCrossRefGoogle Scholar
  13. Cattano D, Valleggi S, Ma D et al (2008) Xenon induces transcription of ADNP in neonatal rat brain. Neurosci Lett 440(3):217–221PubMedCrossRefGoogle Scholar
  14. Curtis TM, Gardiner TA, Stitt AW (2009) Microvascular lesions of diabetic retinopathy: clues towards understanding pathogenesis. Eye 23:1496–1508PubMedCrossRefGoogle Scholar
  15. Frank RN (2004) Diabetic retinopathy. N Engl J Med 350:48–58PubMedCrossRefGoogle Scholar
  16. Gábriel R (2013) Neuropeptides and diabetic retinopathy. Br J Clin Pharmacol 75(5):1189–1201PubMedCentralPubMedCrossRefGoogle Scholar
  17. Giunta S, Castorina A, Bucolo C, Magro G, Drago F, D’Agata V (2012a) Early changes in pituitary adenylate cyclase-activating peptide, vasoactive intestinal peptide and related receptors expression in retina of streptozotocin-induced diabetic rats. Peptides 37(1):32–39PubMedCrossRefGoogle Scholar
  18. Giunta S, Castorina A, Scuderi S, Patti C, D’Agata V (2012b) Epidermal growth factor receptor (EGFR) and neuregulin (Neu) activation in human airway epithelial cells exposed to nickel acetate. Toxicol In Vitro 26(2):280–287PubMedCrossRefGoogle Scholar
  19. Gozes I, Divinski I (2004) The femtomolar-acting NAP interacts with microtubules: novel aspects of astrocyte protection. J Alzheimers Dis 6(6):S37–S41PubMedGoogle Scholar
  20. Gozes I, Giladi E, Pinhasov A, Bardea A, Brenneman DE (2000) Activity-dependent neurotrophic factor: intranasal administration of femtomolar-acting peptides improve performance in a water maze. J Pharmacol Exp Ther 293(3):1091–1098PubMedGoogle Scholar
  21. Gozes I, Steingart RA, Spier AD (2004) NAP mechanisms of neuroprotection. J Mol Neurosci 24(1):67–72PubMedCrossRefGoogle Scholar
  22. Gozes I, Zaltzman R, Hauser J, Brenneman DE, Shohami E, Hill JM (2005) The expression of activity-dependent neuroprotective protein (ADNP) is regulated by brain damage and treatment of mice with the ADNP derived peptide, NAP, reduces the severity of traumatic head injury. Curr Alzheimer Res 2(2):149–153PubMedCrossRefGoogle Scholar
  23. Grant MB, Afzal A, Spoerri P, Pan H, Shaw LC, Mames RN (2004) The role of growth factors in the pathogenesis of diabetic retinopathy. Exp Opin Investig Drugs 13:1275–1293CrossRefGoogle Scholar
  24. Gressens P, Besse L, Robberecht P, Gozes I, Fridkin M, Evrard P (1999) Neuroprotection of the developing brain by systemic administration of vasoactive intestinal peptide derivatives. J Pharmacol Exp Ther 288(3):1207–1213PubMedGoogle Scholar
  25. Hu WK, Liu R, Pei H, Li B (2012) Endoplasmic reticulum stress-related factors protect against diabetic retinopathy. Exp Diabetes Res 507986Google Scholar
  26. Idan-Feldman A, Schirer Y, Polyzoidou E et al (2011) Davunetide (NAP) as a preventative treatment for central nervous system complications in a diabetes rat model. Neurobiol Dis 44(3):327–339PubMedCrossRefGoogle Scholar
  27. Idan-Feldman A, Ostritsky R, Gozes I (2012). Tau and caspase 3 as targets for neuroprotection. Int, J Alzheimers Dis. 2012:493670Google Scholar
  28. Jehle T, Dimitriu C, Auer S et al (2008) The neuropeptide NAP provides neuroprotection against retinal ganglion cell damage after retinal ischemia and optic nerve crush. Graefes Arch Clin Exp Ophthalmol 246(9):1255–1263PubMedCrossRefGoogle Scholar
  29. Jing G, Wang JJ, Zhang SX (2012) ER stress and apoptosis: a new mechanism for retinal cell death. Exp Diabetes Res 2012:589589PubMedCentralPubMedCrossRefGoogle Scholar
  30. Kim YH, Kim YS, Kang SS, Cho GJ, Choi WS (2010) Resveratrol inhibits neuronal apoptosis and elevated Ca2+/calmodulin-dependent protein kinase II activity in diabetic mouse retina. Diabetes 59(7):1825–1835PubMedCentralPubMedCrossRefGoogle Scholar
  31. Kowluru RA, Chakrabarti S, Chen S (2004) Re-institution of good metabolic control in diabetic rats and activation of caspase-3 and nuclear transcriptional factor (NF-kB) in the retina. Acta Diabetol 41:194–199PubMedCrossRefGoogle Scholar
  32. Kusner LL, Sarthy VP, Mohr S (2004) Nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase: a role in high glucose-induced apoptosis in retinal Muller cells. Invest Ophthalmol Vis Sci 45:1553–1561PubMedGoogle Scholar
  33. Leker RR, Teichner A, Grigoriadis N, Ovadia H, Brenneman DE, Fridkin M et al (2002) NAP, a femtomolar-acting peptide, protects the brain against ischemic injury by reducing apoptotic death. Stroke 33:1085–1092PubMedCrossRefGoogle Scholar
  34. Li Q, Zemel E, Miller B, Perlman I (2002) Early retinal damage in experimental diabetes: electroretinographical and morphological observations. Exp Eye Res 74:615–625PubMedCrossRefGoogle Scholar
  35. Liu Y, Tao L, Fu X, Zhao Y, Xu X (2013) BDNF protects retinal neurons from hyperglycemia through the TrkB/ERK/MAPK pathway. Mol Med Rep 7(6):1773–1778PubMedGoogle Scholar
  36. Mohr S, Xi X, Tang J, Kern TS (2002) Caspase activation in retinas of diabetic and galactosemic mice and diabetic patients. Diabetes 51:1172–1179PubMedCrossRefGoogle Scholar
  37. Nakamachi T, Li M, Shioda S, Arimura A (2006) Signaling involved in pituitary adenylate cyclase-activating polypeptide-stimulated ADNP expression. Peptides 27(7):1859–1864PubMedCrossRefGoogle Scholar
  38. Pan HZ, Zhang H, Chang D, Li H, Sui H (2008) The change of oxidative stress products in diabetes mellitus and diabetic retinopathy. Br J Ophthalmol 92:548–551PubMedCrossRefGoogle Scholar
  39. Paques M, Massin P, Gaudric A (1997) Growth factors and diabetic retinopathy. Diabetes Metab 23:125–130PubMedGoogle Scholar
  40. Park SH, Park WJ, Park SJ et al (2003) Apoptotic death of photoreceptors in the streptozotocin-induced diabetic rat retina. Diabetologia 46:1260–1268PubMedCrossRefGoogle Scholar
  41. Pascual M, Guerri C (2007) The peptide NAP promotes neuronal growth and differentiation through extracellular signal-regulated protein kinase and Akt pathways, and protects neurons co-cultured with astrocytes damaged by ethanol. J Neurochem 103(2):557–568PubMedCrossRefGoogle Scholar
  42. Poggi SH, Goodwin K, Hill JM et al (2003) The role of activity-dependent neuroprotective protein in a mouse model of fetal alcohol syndrome. Am J Obstet Gynecol 189(3):790–793PubMedCrossRefGoogle Scholar
  43. Qin X, Zhang Z, Xu H, Wu Y (2011) Notch signaling protects retina from nuclear factor-κB- and poly-ADP-ribose-polymerase-mediated apoptosis under high-glucose stimulation. Acta Biochim Biophys Sin (Shanghai) 43(9):703–711CrossRefGoogle Scholar
  44. Sari Y, Gozes I (2006) Brain deficits associated with fetal alcohol exposure may be protected, in part, by peptides derived from activity-dependent neurotrophic factor and activity-dependent neuroprotective protein. Brain Res Rev 52(1):107–118PubMedCrossRefGoogle Scholar
  45. Schlingemann RO (2004) Role of growth factors and the wound healing response in age-related macular degeneration. Graefes Arch Clin Exp Ophthalmol 242:91–101PubMedCrossRefGoogle Scholar
  46. Schmittgen TD, Livak KJ (2008) Analyzing real-time PCR data by the comparative C(T) method. Nat Protoc 3:1101–1108PubMedCrossRefGoogle Scholar
  47. Scuderi S, D’Amico AG, Castorina A, Imbesi R, Carnazza ML, D’Agata V (2013) Ameliorative effect of PACAP and VIP against increased permeability in a model of outer blood retinal barrier dysfunction. Peptides 39:119–124PubMedCrossRefGoogle Scholar
  48. Snigdha S, Smith ED, Prieto GA, Cotman CW (2012) Caspase-3 activation as a bifurcation point between plasticity and cell death. Neurosci Bull 28(1):14–24PubMedCrossRefGoogle Scholar
  49. Spong CY, Abebe DT, Gozes I, Brenneman DE, Hill JM (2001) Prevention of fetal demise and growth restriction in a mouse model of fetal alcohol syndrome. J Pharmacol Exp Ther 297(2):774–779PubMedGoogle Scholar
  50. Steingart RA, Solomon B, Brenneman DE, Fridkin M, Gozes I (2000) VIP and peptides related to activity-dependent neurotrophic factor protect PC12 cells against oxidative stress. J Mol Neurosci 15(3):137–145PubMedCrossRefGoogle Scholar
  51. Toso L, Roberson R, Abebe D, Spong CY (2007) Neuroprotective peptides prevent some alcohol-induced alteration in gamma-aminobutyric acid A-beta3, which plays a role in cleft lip and palate and learning in fetal alcohol syndrome. Am J Obstet Gynecol 196:259PubMedCrossRefGoogle Scholar
  52. Van Dijk HW, Verbraak FD, Kok PH et al (2001) Inflammatory cytokines in vitreous fluid and serum of patients with diabetic vitreoretinopathy. J Diabetes Complicat 15:257–259CrossRefGoogle Scholar
  53. Villarroel M, Ciudin A, Hernández C, Simo R (2010) Neurodegeneration: an early event of diabetic retinopathy. World J Diabetes 15:57–64CrossRefGoogle Scholar
  54. Vulih-Shultzman I, Pinhasov A, Mandel S et al (2007) Activity-dependent neuroprotective protein snippet NAP reduces tau hyperphosphorylation and enhances learning in a novel transgenic mouse model. J Pharmacol Exp Ther 323(2):438–449PubMedCrossRefGoogle Scholar
  55. Xi X, Gao L, Hatala DA et al (2005) Chronically elevated glucose-induced apoptosis is mediated by inactivation of Akt in cultured Muller cells. Biochem Biophys Res Commun 326:548–553PubMedCrossRefGoogle Scholar
  56. Yamagishi S, Matsui T (2011) Advanced glycation end products (AGEs), oxidative stress and diabetic retinopathy. Curr Pharm Biotechnol 12:362–368PubMedCrossRefGoogle Scholar
  57. Yuuki T, Kanda T, Kimura Y et al (2001) Inflammatory cytokines in vitreous fluid and serum of patients with diabetic vitreoretinopathy. J Diabetes Complicat 15:257–259PubMedCrossRefGoogle Scholar
  58. Zaltzman R, Alexandrovich A, Beni SM, Trembovler V, Shohami E, Gozes (2004) Brain injury-dependent expression of activity-dependent neuroprotective protein. J Mol Neurosci 24(2):181–187PubMedCrossRefGoogle Scholar
  59. Zaltzman R, Alexandrovich A, Trembovler V, Shohami E, Gozes I (2005) The influence of the peptide NAP on Mac-1-deficient mice following closed head injury. Peptides 26(8):1520–1527PubMedCrossRefGoogle Scholar
  60. Zemlyak I, Sapolsky R, Gozes I (2009) NAP protects against cytochrome c release: inhibition of the initiation of apoptosis. Eur J Pharmacol 618(1–3):9–14PubMedCrossRefGoogle Scholar
  61. Zheng Y, Zeng H, She H, Liu H, Sun N (2010) Expression of peptide NAP in rat retinal Müller cells prevents hypoxia-induced retinal injuries and promotes retinal neurons growth. Biomed Pharmacother 64(6):417–423PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Soraya Scuderi
    • 1
  • Agata Grazia D’Amico
    • 1
  • Alessandro Castorina
    • 1
  • Concetta Federico
    • 2
  • Giuseppina Marrazzo
    • 3
  • Filippo Drago
    • 3
  • Claudio Bucolo
    • 3
  • Velia D’Agata
    • 1
  1. 1.Department of Bio-Medical Sciences, Section of Anatomy and HistologyUniversity of CataniaCataniaItaly
  2. 2.Department of Biological, Geological and Environmental Sciences, Section of Animal BiologyUniversity of CataniaCataniaItaly
  3. 3.Department of Clinical and Molecular Biomedicine, Section of Pharmacology and BiochemistryUniversity of CataniaCataniaItaly

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