Biochemistry (Moscow)

, Volume 80, Issue 6, pp 790–799 | Cite as

Expression of neuronal and signaling proteins in penumbra around a photothrombotic infarction core in rat cerebral cortex

  • S. V. Demyanenko
  • S. N. Panchenko
  • A. B. UzdenskyEmail author


Photodynamic impact on animal cerebral cortex using water-soluble Bengal Rose as a photosensitizer, which does not cross the blood-brain barrier and remains in blood vessels, induces platelet aggregation, vessel occlusion, and brain tissue infarction. This reproduces ischemic stroke. Irreversible cell damage within the infarction core propagates to adjacent tissue and forms a transition zone — the penumbra. Tissue necrosis in the infarction core is too fast (minutes) to be prevented, but much slower penumbral injury (hours) can be limited. We studied the changes in morphology and protein expression profile in penumbra 1 h after local photothrombotic infarction induced by laser irradiation of the cerebral cortex after Bengal Rose administration. Morphological study using standard hematoxylin/eosin staining showed a 3-mm infarct core surrounded by 1.5–2.0 mm penumbra. Morphological changes in the penumbra were lesser and decreased towards its periphery. Antibody microarrays against 224 neuronal and signaling proteins were used for proteomic study. The observed upregulation of penumbra proteins involved in maintaining neurite integrity and guidance (NAV3, MAP1, CRMP2, PMP22); intercellular interactions (N-cadherin); synaptic transmission (glutamate decarboxylase, tryptophan hydroxylase, Munc-18-1, Munc-18-3, and synphilin-1); mitochondria quality control and mitophagy (PINK1 and Parkin); ubiquitin-mediated proteolysis and tissue clearance (UCHL1, PINK1, Parkin, synphilin-1); and signaling proteins (PKBα and ERK5) could be associated with tissue recovery. Downregulation of PKC, PKCβ1/2, and TDP-43 could also reduce tissue injury. These changes in expression of some neuronal proteins were directed mainly to protection and tissue recovery in the penumbra. Some upregulated proteins might serve as markers of protection processes in a penumbra.

Key words

stroke neurodegeneration penumbra proteomics 



collapsin response mediator protein 2


dual-specificity tyrosine-phosphorylated regulated kinase 1A


extracellular regulated kinase 2


γ-butyric acid


microtubule-associated protein 1


neuron navigator 3 protein


PTEN-induced mitochondrial protein kinase


protein kinase Bα


protein kinase C


protein kinase C isoform β1


peripheral myelin protein 22


photothrombotic infarction


NAD+-dependent deacetylase sirtuin-1


transactivation response DNA-binding protein


ubiquitin C-terminal hydrolase L1


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Meisel, A., Prass, K., Wolf, T., and Dirnagl, U. (2004) Stroke, in Neuroprotection: Models, Mechanisms and Therapies (Bahr, M., ed.) Wiley-Blackwell, Hoboken, NJ, pp. 9–43.Google Scholar
  2. 2.
    Iadecola, C., and Anrather, J. (2011) Stroke research at a crossroad: asking the brain for directions, Nat. Neurosci., 14, 1363–1368.PubMedCentralPubMedCrossRefGoogle Scholar
  3. 3.
    Moskowitz, M. A. (2010) Brain protection: maybe yes, maybe no, Stroke, 41, S85–S86.PubMedCrossRefGoogle Scholar
  4. 4.
    Zhiganshina, L. E., and Abakumova, T. R. (2013) Cerebrolysin in a treatment of acute ischemic stroke, Vestnik RAMN, 1, 21–29.Google Scholar
  5. 5.
    Watson, B. D., Dietrich, W. D., Busto, R., Wachtel, M. S., and Ginsberg, M. D. (1985) Induction of reproducible brain infarction by photochemically initiated thrombosis, Ann. Neurol., 17, 497–504.PubMedCrossRefGoogle Scholar
  6. 6.
    Dietrich, W. D., Watson, B. D., Busto, R., Ginsberg, M. D., and Bethea, J. R. (1987) Photochemically induced cerebral infarction. I. Early microvascular alterations, Acta Neuropathol., 72, 315–325.PubMedCrossRefGoogle Scholar
  7. 7.
    Pevsner, P. H., Eichenbaum, J. W., Miller, D. C., Pivawer, G., Eichenbaum, K. D., Stern, A., Zakian, K. L., and Koutcher, J. A. (2001) A photothrombotic model of small early ischemic infarcts in the rat brain with histologic and MRI correlation, J. Pharmacol. Toxicol. Methods, 45, 227–233.PubMedCrossRefGoogle Scholar
  8. 8.
    Shanina, E. V., Redecker, C., Reinecke, S., Schallert, T., and Witte, O. W. (2005) Long-term effects of sequential cortical infarcts on scar size, brain volume and cognitive function, Behav. Brain Res., 158, 69–77.PubMedCrossRefGoogle Scholar
  9. 9.
    Schmidt, A., Hoppen, M., Strecker, J. K., Diederich, K., Schabitz, W. R., Schilling, M., and Minnerup, J. (2012) Photochemically induced ischemic stroke in rats, Exp. Transl. Stroke Med., 4, 13.PubMedCentralPubMedCrossRefGoogle Scholar
  10. 10.
    Romanova, G. A., Barskov, I. V., Ostrovskaya, R. U., Gudasheva, T. A., and Viktorov, I. V. (1998) Behavioral and morphological changes induced by bilateral photoinduced thrombosis of cerebral vessels in the rat frontal cortex, Pathol. Physiol. Exp. Ther., No. 2, 8–10.Google Scholar
  11. 11.
    Romanova, G. A., Shakova, F. M., Barskov, I. V., Stelmashuk, E. V., Genrihs, E. E., Cheremnyh, A. M., Kalinina, T. I., and Yurin, V. L. (2014) Neuroprotective and anti-amnestic effect of erythropoietin derivatives at experimental ischemic damage to brain cortex, Bull. Exp. Biol. Med., 158, 299–302.Google Scholar
  12. 12.
    Uzdensky, A. B. (2010) Cellular and Molecular Mechanisms of Photodynamic Therapy [in Russian], Nauka, St. Petersburg.Google Scholar
  13. 13.
    Brundel, M., de Bresser, J., van Dillen, J. J., Kappelle, L. J., and Biessels, G. J. (2012) Cerebral microinfarcts: a systematic review of neuropathological studies, J. Cereb. Blood Flow Metab., 32, 425–436.PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Pantoni, L. (2010) Cerebral small vessel disease: from pathogenesis and clinical characteristics to therapeutic challenges, Lancet Neurol., 9, 689–701.PubMedCrossRefGoogle Scholar
  15. 15.
    Del Zoppo, G. J., and Mabuchi, T. (2003) Cerebral microvessel responses to focal ischemia, J. Cereb. Blood Flow Metab., 23, 879–894.PubMedCrossRefGoogle Scholar
  16. 16.
    Spisak, S., Tulassay, Z., Molnar, B., and Guttman, A. (2007) Protein microchips in biomedicine and biomarker discovery, Electrophoresis, 28, 4261–4273.PubMedCrossRefGoogle Scholar
  17. 17.
    Wingren, C., and Borrebaeck, C. A. (2009) Antibody-based microarrays, Methods Mol. Biol., 509, 57–84.PubMedGoogle Scholar
  18. 18.
    Dayon, L., Turck, N., Garci-Berrocoso, T., Walter, N., Burkhard, P. R., Vilalta, A., Sahuquillo, J., Montaner, J., and Sanchez, J. C. (2011) Brain extracellular fluid protein changes in acute stroke patients, J. Proteome Res., 10, 1043–1051.PubMedCrossRefGoogle Scholar
  19. 19.
    Demyanenko, S. V., Uzdensky, A. B., Sharifulina, S. A., Lapteva, T. O., and Polyakova, L. P. (2014) PDT-induced epigenetic changes in the mouse cerebral cortex: a protein microarray study, Biochim. Biophys. Acta, 1840, 262–270.PubMedCrossRefGoogle Scholar
  20. 20.
    Zilles, K. (1985) The Cortex of the Rat: A Stereotaxis Atlas, Springer-Verlag, Berlin.CrossRefGoogle Scholar
  21. 21.
    Villa, R. F., Gorini, A., Ferrari, F., and Hoyer, S. (2013) Energy metabolism of cerebral mitochondria during aging, ischemia and post-ischemic recovery assessed by functional proteomics of enzymes, Neurochem. Int., 63, 765–781.PubMedCrossRefGoogle Scholar
  22. 22.
    Datta, A., Park, J. E., Li, X., Zhang, H., Ho, Z. S., Heese, K., Lim, S. K., Tam, J. P., and Sze, S. K. (2010) Phenotyping of an in vitro model of ischemic penumbra by iTRAQ-based shotgun quantitative proteomics, J. Proteom. Res., 9, 472–484.CrossRefGoogle Scholar
  23. 23.
    Bu, X., Zhang, N., Yang, X., Liu, Y., Du, J., Liang, J., Xu, Q., and Li, J. (2011) Proteomic analysis of PKCßII-interacting proteins involved in HPC-induced neuroprotection against cerebral ischemia of mice, J. Neurochem., 117, 346–356.PubMedCrossRefGoogle Scholar
  24. 24.
    Hara, H., Onodera, H., Yoshidomi, M., Matsuda, Y., and Kogure, K. (1990) Staurosporine, a novel protein kinase C inhibitor, prevents postischemic neuronal damage in the gerbil and rat, J. Cereb. Blood Flow Metab., 10, 646–653.PubMedCrossRefGoogle Scholar
  25. 25.
    Felipo, V., Minana, M. D., and Grisolia, S. (1993) Inhibitors of protein kinase C prevent the toxicity of glutamate in primary neuronal cultures, Brain Res., 604, 192–196.PubMedCrossRefGoogle Scholar
  26. 26.
    Bright, R., and Mochly-Rosen, D. (2005) The role of protein kinase C in cerebral ischemic and reperfusion injury, Stroke, 36, 2781–2790.PubMedCrossRefGoogle Scholar
  27. 27.
    Chou, W. H., and Messing, R. O. (2005) Protein kinase C isozymes in stroke, Trends Cardiovasc. Med., 15, 47–51.PubMedCrossRefGoogle Scholar
  28. 28.
    Lee, B. K., Yoon, J. S., Lee, M. G., and Jung, Y. S. (2014) Protein kinase C-β mediates neuronal activation of Na+/H+ exchanger-1 during glutamate excitotoxicity, Cell Signal., 26, 697–704.PubMedCrossRefGoogle Scholar
  29. 29.
    Wang, J., Bright, R., Mochly-Rosen, D., and Giffard, R. G. (2004) Cell-specific role for e-and βI-protein kinase C isozymes in protecting cortical neurons and astrocytes from ischemia-like injury, Neuropharmacology, 47, 136–145.PubMedCrossRefGoogle Scholar
  30. 30.
    Zhao, H., Sapolsky, R. M., and Steinberg, G. K. (2006) Phosphoinositide-3-kinase/akt survival signal pathways are implicated in neuronal survival after stroke, Mol. Neurobiol., 34, 249–270.PubMedCrossRefGoogle Scholar
  31. 31.
    Wang, R. M., Zhang, Q. G., Li, C. H., and Zhang, G. Y. (2005) Activation of extracellular signal-regulated kinase 5 may play a neuroprotective role in hippocampal CA3/DG region after cerebral ischemia, J. Neurosci. Res., 80, 391–399.PubMedCrossRefGoogle Scholar
  32. 32.
    Laguna, A., Aranda, S., Barallobre, M. J., Barhoum, R., Fernandez, E., Fotaki, V., Delabar, J. M., de la Luna, S., Villa, P., and Arbones, M. L. (2008) The protein kinase DYRK1A regulates caspase-9-mediated apoptosis during retina development, Dev. Cell, 15, 841–853.PubMedCrossRefGoogle Scholar
  33. 33.
    Choi, H. K., and Chung, K. C. (2011) DYRK1A positively stimulates ASK1-JNK signaling pathway during apoptotic cell death, Exp. Neurobiol., 20, 35–44.PubMedCentralPubMedCrossRefGoogle Scholar
  34. 34.
    Guo, X., Williams, J. G., Schug, T. T., and Li, X. (2010) DYRK1A and DYRK3 promote cell survival through phosphorylation and activation of SIRT1, J. Biol. Chem., 285, 3223–3232.Google Scholar
  35. 35.
    Trancikova, A., Tsika, E., and Moore, D. J. (2012) Mitochondrial dysfunction in genetic animal models of Parkinson’s disease, Antioxid. Redox Signal., 16, 896–919.PubMedCentralPubMedCrossRefGoogle Scholar
  36. 36.
    De Vries, R. L., and Przedborski, S. (2013) Mitophagy and Parkinson’s disease: be eaten to stay healthy, Mol. Cell Neurosci., 55, 37–43.PubMedCrossRefGoogle Scholar
  37. 37.
    Caldeira, M. V., Salazar, I. L., Curcio, M., Canzoniero, L. M., and Duarte, C. B. (2014) Role of the ubiquitin-proteasome system in brain ischemia: friend or foe? Prog. Neurobiol., 112, 50–69.PubMedCrossRefGoogle Scholar
  38. 38.
    Yamauchi, T., Sakurai, M., Abe, K., Matsumiya, G., and Sawa, Y. (2008) Ubiquitin-mediated stress response in the spinal cord after transient ischemia, Stroke, 39, 1883–1889.PubMedCrossRefGoogle Scholar
  39. 39.
    Kruger, R. (2004) The role of synphilin-1 in synaptic function and protein degradation, Cell Tissue Res., 318, 195–199.PubMedCrossRefGoogle Scholar
  40. 40.
    Maes, T., Barcelo, A., and Buesa, C. (2002) Neuron navigator: a human gene family with homology to unc-53, a cell guidance gene from Caenorhabditis elegans, Genomics, 80, 21–30.PubMedCrossRefGoogle Scholar
  41. 41.
    Halpain, S., and Dehmelt, L. (2006) The MAP1 family of microtubule-associated proteins, Genome Biol., 7, 2–4.CrossRefGoogle Scholar
  42. 42.
    Chen, A., Liao, W. P., Lu, Q., Wong, W. S., and Wong, P. T. (2007) Up-regulation of dihydropyrimidinase-related protein 2, spectrin alpha II chain, heat shock cognate protein 70 pseudogene 1 and tropomodulin 2 after focal cerebral ischemia in rats — a proteomics approach, Neurochem. Int., 50, 1078–1086.PubMedCrossRefGoogle Scholar
  43. 43.
    Hou, S. T., Jiang, S. X., Aylsworth, A., Ferguson, G., Slinn, J., Hu, H., Leung, T., Kappler, J., and Kaibuchi, K. (2009) CaMKII phosphorylates collapsin response mediator protein 2 and modulates axonal damage during glutamate excitotoxicity, J. Neurochem., 111, 870–881.PubMedCrossRefGoogle Scholar
  44. 44.
    Quarles, R. H. (2002) Myelin sheaths: glycoproteins involved in their formation, maintenance and degeneration, Cell. Mol. Life Sci., 59, 1851–1871.PubMedCrossRefGoogle Scholar
  45. 45.
    Gallwitz, D., and Jahn, R. (2003) The riddle of the Sec1/Munc-18 proteins — new twists added to their interactions with SNAREs, Trends Biochem. Sci., 28, 113–116.PubMedCrossRefGoogle Scholar
  46. 46.
    Lee, E. B., Lee, V. M., and Trojanowski, J. Q. (2011) Gains or losses: molecular mechanisms of TDP43-mediated neurodegeneration, Nat. Rev. Neurosci., 13, 38–50.PubMedCentralPubMedGoogle Scholar
  47. 47.
    Kanazawa, M., Kakita, A., Igarashi, H., Takahashi, T., Kawamura, K., Takahashi, H., Nakada, T., Nishizawa, M., and Shimohata, T. (2011) Biochemical and histopathological alterations in TAR DNA-binding protein-43 after acute ischemic stroke in rats, J. Neurochem., 116, 957–965.PubMedCrossRefGoogle Scholar
  48. 48.
    Zechariah, A. E., Ali, A., Hagemann, N., Jin, F., Doeppner, T. R., Helfrich, I., Mies, G., and Hermann, D. M. (2013) Hyperlipidemia attenuates vascular endothelial growth factor-induced angiogenesis, impairs cerebral blood flow, and disturbs stroke recovery via decreased pericyte coverage of brain endothelial cells, Arterioscler. Thromb. Vasc. Biol., 33, 1561–1567.PubMedCrossRefGoogle Scholar
  49. 49.
    Back, T., Ginsberg, M. D., Dietrich, W. D., and Watson, B. D. (1996) Induction of spreading depression in the ischemic hemisphere following experimental middle cerebral artery occlusion: effect on infarct morphology, J. Cereb. Blood Flow Metab., 16, 202–213.PubMedCrossRefGoogle Scholar
  50. 50.
    Puyal, J., Ginet, V., and Clarke, P. G. (2013) Multiple interacting cell death mechanisms in the mediation of excitotoxicity and ischemic brain damage: a challenge for neuroprotection, Prog. Neurobiol., 105, 24–48.PubMedCrossRefGoogle Scholar
  51. 51.
    Sims, N. R., and Anderson, M. F. (2002) Mitochondrial contributions to tissue damage in stroke, Neurochem. Int., 40, 511–526.PubMedCrossRefGoogle Scholar

Copyright information

© Pleiades Publishing, Ltd. 2015

Authors and Affiliations

  • S. V. Demyanenko
    • 1
  • S. N. Panchenko
    • 2
  • A. B. Uzdensky
    • 1
    Email author
  1. 1.Academy of Biology and BiotechnologySouthern Federal UniversityRostov-on-DonRussia
  2. 2.Rostov State Medical UniversityRostov-on-DonRussia

Personalised recommendations