Ukr.Biochem.J. 2017; Volume 89, Issue 2, Mar-Apr, pp. 43-54


Plasminogen and its fragments in rat brain: a plausible role for astrocytes in angiostatin generation

A. A. Tykhomyrov1, V. S. Nedzvetsky2,3, C. A. Ağca3,
V. V. Korsa1, T. V. Grinenko1

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Dnipropetrovsk National University, Dnipro, Ukraine;
3Bingöl University, Bingöl, Turkey;

The purpose of the present study was to examine the plasminogen localization and to detect levels of its fragments (angiostatins) in various regions of rat brain as well as to establish whether rat brain astrocytes could be involved in angiostatin production. It was shown immunohistochemically that plasminogen is distributed broadly in the various brain regions, with predominant expression in meningeal layer and IV, V, and VI layers or cerebral cortex, dentate gyrus, meningeal and Purkinje cells, molecular and granular layers of cerebellum, as well as vessel walls. Angiostatin polypeptides were detected by Western blot analysis mostly in the cerebral cortex and were represented by 50 and 40-30 kDa polypeptides. In the whole cell lysates from primary cultures of rat astrocytes, immunoreactive polypeptides with Mm ~ 92, 84, 65-60, 50, 40, 38-30 kDa, correspon­ding to native plasminogen and a variety of its truncated products, including angiostatin polypeptides, were revealed. Incubation of astrocytes with exogenous plasminogen resulted in gradual increasing levels of some plasminogen fragments, particularly 30 kDa protein. Moreover, this polypeptide appeared to be the single angiostatin released by astrocytes in vitro. We report here for the first time that astrocytes are one of the cell types in CNS that could be responsible for angiostatin formation and releasing.

Keywords: , , , , , ,


  1. Law RH, Abu-Ssaydeh D, Whisstock JC. New insights into the structure and function of the plasminogen/plasmin system. Curr Opin Struct Biol. 2013 Dec;23(6):836-41. Review. PubMed, CrossRef
  2. Melchor JP, Strickland S. Tissue plasminogen activator in central nervous system physiology and pathology. Thromb Haemost. 2005 Apr;93(4):655-60. Review. PubMed, PubMedCentral, CrossRef
  3. Niego B, Freeman R, Puschmann TB, Turnley AM, Medcalf RL. t-PA-specific modulation of a human blood-brain barrier model involves plasmin-mediated activation of the Rho kinase pathway in astrocytes. Blood. 2012 May 17;119(20):4752-61. PubMed, CrossRef
  4. Calabresi P, Napolitano M, Centonze D, Marfia GA, Gubellini P, Teule MA, Berretta N, Bernardi G, Frati L, Tolu M, Gulino A. Tissue plasminogen activator controls multiple forms of synaptic plasticity and memory. Eur J Neurosci. 2000 Mar;12(3):1002-12. PubMed, CrossRef
  5. Silverman MA, Johnson S, Gurkins D, Farmer M, Lochner JE, Rosa P, Scalettar BA. Mechanisms of transport and exocytosis of dense-core granules containing tissue plasminogen activator in developing hippocampal neurons. J Neurosci. 2005 Mar 23;25(12):3095-106. PubMed, CrossRef
  6. Soff GA. Angiostatin and angiostatin-related proteins. Cancer Metastasis Rev. 2000;19(1-2):97-107. Review. PubMed, CrossRef
  7. Wahl ML, Owen CS, Grant DS. Angiostatin induces intracellular acidosis and anoikis in endothelial cells at a tumor-like low pH. Endothelium. 2002;9(3):205-16. PubMed, CrossRef
  8. Radziwon-Balicka A, Moncada de la Rosa C, Zielnik B, Doroszko A, Jurasz P. Temporal and pharmacological characterization of angiostatin release and generation by human platelets: implications for endothelial cell migration. PLoS One. 2013;8(3):e59281.  PubMed, PubMedCentral, CrossRef
  9. van Tilborg AA, Sweep FC, Geurts-Moespot AJ, Wetzels AM, de Waal RM, Westphal JR, Massuger LF. Plasminogen activators are involved in angiostatin generation in vivo in benign and malignant ovarian tumor cyst fluids. Int J Oncol. 2014 Apr;44(4):1394-400. PubMed, CrossRef
  10. Böhm MR, Hodes F, Brockhaus K, Hummel S, Schlatt S, Melkonyan H, Thanos S, et al. Is Angiostatin Involved in Physiological Foveal Avascularity? Invest Ophthalmol Vis Sci. 2016 Sep 1;57(11):4536-52. PubMed, CrossRef
  11. Liang YZ, Zeng ZL, Hua LL, Li JF, Wang YL, Bi XZ. Expression and significance of angiostatin, vascular endothelial growth factor and matrix metalloproteinase-9 in brain tissue of diabetic rats with ischemia reperfusion. Asian Pac J Trop Med. 2016 Jun;9(6):587-91. PubMed, CrossRef
  12. Deininger MH, Winkler S, Kremsner PG, Meyermann R, Schluesener HJ. Angiogenic proteins in brains of patients who died with cerebral malaria. J Neuroimmunol. 2003 Sep;142(1-2):101-11. PubMed, CrossRef
  13. Taniguchi Y, Inoue N, Morita S, Nikaido Y, Nakashima T, Nagai N, Okada K, Matsuo O, Miyata S. Localization of plasminogen in mouse hippocampus, cerebral cortex, and hypothalamus. Cell Tissue Res. 2011 Feb;343(2):303-17. PubMed, CrossRef
  14. Zhang G, Jin G, Nie X, Mi R, Zhu G, Jia W, Liu F. Enhanced antitumor efficacy of an oncolytic herpes simplex virus expressing an endostatin-angiostatin fusion gene in human glioblastoma stem cell xenografts. PLoS One. 2014 Apr 22;9(4):e95872. PubMed, PubMedCentral, CrossRef
  15. Perri SR, Nalbantoglu J, Annabi B, Koty Z, Lejeune L, François M, Di Falco MR, Béliveau R, Galipeau J. Plasminogen kringle 5-engineered glioma cells block migration of tumor-associated macrophages and suppress tumor vascularization and progression. Cancer Res. 2005 Sep 15;65(18):8359-65. PubMed, CrossRef
  16. Pekny M, Pekna M, Messing A, Steinhäuser C, Lee JM, Parpura V, Hol EM, Sofroniew MV, Verkhratsky A. Astrocytes: a central element in neurological diseases. Acta Neuropathol. 2016 Mar;131(3):323-45. Review. PubMed, CrossRef
  17. Broux B, Gowing E, Prat A. Glial regulation of the blood-brain barrier in health and disease. Semin Immunopathol. 2015 Nov;37(6):577-90. Review. PubMed, CrossRef
  18. Argaw AT, Asp L, Zhang J, Navrazhina K, Pham T, Mariani JN, Mahase S, Dutta DJ, Seto J, Kramer EG, Ferrara N, Sofroniew MV, John GR. Astrocyte-derived VEGF-A drives blood-brain barrier disruption in CNS inflammatory disease. J Clin Invest. 2012 Jul;122(7):2454-68. PubMed, PubMedCentral, CrossRef
  19. Hajrasouliha AR, Jiang G, Lu Q, Lu H, Kaplan HJ, Zhang HG, Shao H. Exosomes from retinal astrocytes contain antiangiogenic components that inhibit laser-induced choroidal neovascularization. J Biol Chem. 2013 Sep 27;288(39):28058-67. PubMed, PubMedCentral, CrossRef
  20. Le DM, Besson A, Fogg DK, Choi KS, Waisman DM, Goodyer CG, Rewcastle B, Yong VW. Exploitation of astrocytes by glioma cells to facilitate invasiveness: a mechanism involving matrix metalloproteinase-2 and the urokinase-type plasminogen activator-plasmin cascade. J Neurosci. 2003 May 15;23(10):4034-43. PubMed
  21. Bermúdez O, Parra A, Ramírez L. Purification and activation of caprine and canine plasminogens: comparison with human plasminogen. IATREIA. 2011;24(2):117-125.
  22. Stoscheck CM. Quantitation of protein. Methods Enzymol. 1990;182:50-68. PubMed, CrossRef
  23. Abraham S, de Vellis J, Vernadakis A, Haber B. eds. A dissection and tissue culture manual of the nervous system. New York: Alan R. Liss, Inc. 1989.
  24. Miles LA, Castellino FJ, Gong Y. Critical role for conversion of glu-plasminogen to Lys-plasminogen for optimal stimulation of plasminogen activation on cell surfaces. Trends Cardiovasc Med. 2003 Jan;13(1):21-30. Review. PubMed, CrossRef
  25. Bohmfalk JF, Fuller GM. Plaminogen is synthesized by primary cultures of rat hepatocytes. Science. 1980 Jul 18;209(4454):408-10. PubMed, CrossRef
  26. Basham ME, Seeds NW. Plasminogen expression in the neonatal and adult mouse brain. J Neurochem. 2001 Apr;77(1):318-25. PubMed, CrossRef
  27. Yepes M, Lawrence DA. New functions for an old enzyme: nonhemostatic roles for tissue-type plasminogen activator in the central nervous system. Exp Biol Med (Maywood). 2004 Dec;229(11):1097-104. Review. PubMed
  28. Pang PT, Teng HK, Zaitsev E, Woo NT, Sakata K, Zhen S, Teng KK, Yung WH, Hempstead BL, Lu B. Cleavage of proBDNF by tPA/plasmin is essential for long-term hippocampal plasticity. Science. 2004 Oct 15;306(5695):487-91. PubMed, CrossRef
  29. Wu F, Torre E, Cuellar-Giraldo D, Cheng L, Yi H, Bichler EK, García PS, Yepes M. Tissue-type plasminogen activator triggers the synaptic vesicle cycle in cerebral cortical neurons. J Cereb Blood Flow Metab. 2015 Dec;35(12):1966-76. PubMed, PubMedCentral, CrossRef
  30. Oray S, Majewska A, Sur M. Dendritic spine dynamics are regulated by monocular deprivation and extracellular matrix degradation. Neuron. 2004 Dec 16;44(6):1021-30. PubMed, CrossRef
  31. Li J, Yu L, Gu X, Ma Y, Pasqualini R, Arap W, Snyder EY, Sidman RL. Tissue plasminogen activator regulates Purkinje neuron development and survival. Proc Natl Acad Sci USA. 2013 Jun 25;110(26):E2410-9. PubMed, PubMedCentral, CrossRef
  32. Tsilibary E, Tzinia A, Radenovic L, Stamenkovic V, Lebitko T, Mucha M, Pawlak R, Frischknecht R, Kaczmarek L. Neural ECM proteases in learning and synaptic plasticity. Prog Brain Res. 2014;214:135-57. Review. PubMed, CrossRef
  33. Ma HI, Lin SZ, Chiang YH, Li J, Chen SL, Tsao YP, Xiao X. Intratumoral gene therapy of malignant brain tumor in a rat model with angiostatin delivered by adeno-associated viral (AAV) vector. Gene Ther. 2002 Jan;9(1):2-11. PubMed, CrossRef
  34. Ryu JK, Little JP, Klegeris A, Jantaratnotai N, McLarnon JG. Actions of the anti-angiogenic compound angiostatin in an animal model of Alzheimer’s disease. Curr Alzheimer Res. 2013 Mar;10(3):252-60. PubMed, CrossRef
  35. Guzyk MM, Tykhomyrov AA, Nedzvetsky VS, Prischepa IV, Grinenko TV, Yanitska LV, Kuchmerovska TM. Poly(ADP-Ribose) Polymerase-1 (PARP-1) Inhibitors Reduce Reactive Gliosis and Improve Angiostatin Levels in Retina of Diabetic Rats. Neurochem Res. 2016 Oct;41(10):2526-2537. PubMed, CrossRef
  36. Pearce JW, Janardhan KS, Caldwell S, Singh B. Angiostatin and integrin alphavbeta3 in the feline, bovine, canine, equine, porcine and murine retina and cornea. Vet Ophthalmol. 2007 Sep-Oct;10(5):313-9. PubMed, CrossRef
  37. Di Bari G, Gentile E, Latronico T, Corriero G, Fasano A, Nonnis Marzano C, Liuzzi GM. Inhibitory Effect of Aqueous Extracts from Marine Sponges on the Activity and Expression of Gelatinases A (MMP-2) and B (MMP-9) in Rat Astrocyte Cultures. PLoS One. 2015 Jun 8;10(6):e0129322. PubMed, PubMed, CrossRef
  38. Li F, Yang J, Liu X, He P, Ji S, Wang J, Han J, Chen N, Yao L. Human glioma cell BT325 expresses a proteinase that converts human plasminogen to kringle 1-5-containing fragments. Biochem Biophys Res Commun. 2000 Nov 30;278(3):821-5. PubMed, CrossRef
  39. Nakajima K, Tsuzaki N, Nagata K, Takemoto N, Kohsaka S. Production and secretion of plasminogen in cultured rat brain microglia. FEBS Lett. 1992 Aug 17;308(2):179-82. PubMed, CrossRef
  40. Nakanishi H. Microglial functions and proteases. Mol Neurobiol. 2003 Apr;27(2):163-76. Review. PubMed, CrossRef

Creative CommonsThis work is licensed under a Creative Commons Attribution 4.0 International License.