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. 1998 May 1;101(9):1992–1999. doi: 10.1172/JCI2169

Caspase inhibitor affords neuroprotection with delayed administration in a rat model of neonatal hypoxic-ischemic brain injury.

Y Cheng 1, M Deshmukh 1, A D'Costa 1, J A Demaro 1, J M Gidday 1, A Shah 1, Y Sun 1, M F Jacquin 1, E M Johnson 1, D M Holtzman 1
PMCID: PMC508786  PMID: 9576764

Abstract

Programmed cell death (apoptosis) is a normal process in the developing nervous system. Recent data suggest that certain features seen in the process of programmed cell death may be favored in the developing versus the adult brain in response to different brain injuries. In a well characterized model of neonatal hypoxia-ischemia, we demonstrate marked but delayed cell death in which there is prominent DNA laddering, TUNEL-labeling, and nuclei with condensed chromatin. Caspase activation, which is required in many cases of apoptotic cell death, also followed a delayed time course after hypoxia-ischemia. Administration of boc-aspartyl(OMe)-fluoromethylketone, a pan-caspase inhibitor, was significantly neuroprotective when given by intracerebroventricular injection 3 h after cerebral hypoxia-ischemia. In addition, systemic injections of boc-aspartyl(OMe)-fluoromethylketone also given in a delayed fashion, resulted in significant neuroprotection. These findings suggest that caspase inhibitors may be able to provide benefit over a prolonged therapeutic window after hypoxic-ischemic events in the developing brain, a major contributor to static encephalopathy and cerebral palsy.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Armstrong R. C., Aja T. J., Hoang K. D., Gaur S., Bai X., Alnemri E. S., Litwack G., Karanewsky D. S., Fritz L. C., Tomaselli K. J. Activation of the CED3/ICE-related protease CPP32 in cerebellar granule neurons undergoing apoptosis but not necrosis. J Neurosci. 1997 Jan 15;17(2):553–562. doi: 10.1523/JNEUROSCI.17-02-00553.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brown A. W., Brierley J. B. Anoxic-ischaemic cell change in rat brain light microscopic and fine-structural observations. J Neurol Sci. 1972 May;16(1):59–84. doi: 10.1016/0022-510x(72)90102-5. [DOI] [PubMed] [Google Scholar]
  3. Cheng Y., Gidday J. M., Yan Q., Shah A. R., Holtzman D. M. Marked age-dependent neuroprotection by brain-derived neurotrophic factor against neonatal hypoxic-ischemic brain injury. Ann Neurol. 1997 Apr;41(4):521–529. doi: 10.1002/ana.410410416. [DOI] [PubMed] [Google Scholar]
  4. Collins R. J., Harmon B. V., Gobé G. C., Kerr J. F. Internucleosomal DNA cleavage should not be the sole criterion for identifying apoptosis. Int J Radiat Biol. 1992 Apr;61(4):451–453. doi: 10.1080/09553009214551201. [DOI] [PubMed] [Google Scholar]
  5. Deckwerth T. L., Johnson E. M., Jr Temporal analysis of events associated with programmed cell death (apoptosis) of sympathetic neurons deprived of nerve growth factor. J Cell Biol. 1993 Dec;123(5):1207–1222. doi: 10.1083/jcb.123.5.1207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Deshmukh M., Johnson E. M., Jr Programmed cell death in neurons: focus on the pathway of nerve growth factor deprivation-induced death of sympathetic neurons. Mol Pharmacol. 1997 Jun;51(6):897–906. doi: 10.1124/mol.51.6.897. [DOI] [PubMed] [Google Scholar]
  7. Deshmukh M., Vasilakos J., Deckwerth T. L., Lampe P. A., Shivers B. D., Johnson E. M., Jr Genetic and metabolic status of NGF-deprived sympathetic neurons saved by an inhibitor of ICE family proteases. J Cell Biol. 1996 Dec;135(5):1341–1354. doi: 10.1083/jcb.135.5.1341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Deshpande J., Bergstedt K., Lindén T., Kalimo H., Wieloch T. Ultrastructural changes in the hippocampal CA1 region following transient cerebral ischemia: evidence against programmed cell death. Exp Brain Res. 1992;88(1):91–105. doi: 10.1007/BF02259131. [DOI] [PubMed] [Google Scholar]
  9. Ellis R. E., Yuan J. Y., Horvitz H. R. Mechanisms and functions of cell death. Annu Rev Cell Biol. 1991;7:663–698. doi: 10.1146/annurev.cb.07.110191.003311. [DOI] [PubMed] [Google Scholar]
  10. Ferrer I., Tortosa A., Macaya A., Sierra A., Moreno D., Munell F., Blanco R., Squier W. Evidence of nuclear DNA fragmentation following hypoxia-ischemia in the infant rat brain, and transient forebrain ischemia in the adult gerbil. Brain Pathol. 1994 Apr;4(2):115–122. doi: 10.1111/j.1750-3639.1994.tb00821.x. [DOI] [PubMed] [Google Scholar]
  11. Ferriero D. M., Arcavi L. J., Sagar S. M., McIntosh T. K., Simon R. P. Selective sparing of NADPH-diaphorase neurons in neonatal hypoxia-ischemia. Ann Neurol. 1988 Nov;24(5):670–676. doi: 10.1002/ana.410240512. [DOI] [PubMed] [Google Scholar]
  12. Ferriero D. M., Holtzman D. M., Black S. M., Sheldon R. A. Neonatal mice lacking neuronal nitric oxide synthase are less vulnerable to hypoxic-ischemic injury. Neurobiol Dis. 1996 Feb;3(1):64–71. doi: 10.1006/nbdi.1996.0006. [DOI] [PubMed] [Google Scholar]
  13. Gavrieli Y., Sherman Y., Ben-Sasson S. A. Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol. 1992 Nov;119(3):493–501. doi: 10.1083/jcb.119.3.493. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gidday J. M., Fitzgibbons J. C., Shah A. R., Park T. S. Neuroprotection from ischemic brain injury by hypoxic preconditioning in the neonatal rat. Neurosci Lett. 1994 Feb 28;168(1-2):221–224. doi: 10.1016/0304-3940(94)90455-3. [DOI] [PubMed] [Google Scholar]
  15. Golden J. P., Rana J. Z., Davis J., Zahm D. S., Jacquin M. F. Organization of the proximal, orbital segment of the infraorbital nerve at multiple intervals after axotomy at birth: a quantitative electron microscopic study in rat. J Comp Neurol. 1993 Dec 8;338(2):159–174. doi: 10.1002/cne.903380203. [DOI] [PubMed] [Google Scholar]
  16. Goto K., Ishige A., Sekiguchi K., Iizuka S., Sugimoto A., Yuzurihara M., Aburada M., Hosoya E., Kogure K. Effects of cycloheximide on delayed neuronal death in rat hippocampus. Brain Res. 1990 Nov 26;534(1-2):299–302. doi: 10.1016/0006-8993(90)90144-z. [DOI] [PubMed] [Google Scholar]
  17. Hagan P., Barks J. D., Yabut M., Davidson B. L., Roessler B., Silverstein F. S. Adenovirus-mediated over-expression of interleukin-1 receptor antagonist reduces susceptibility to excitotoxic brain injury in perinatal rats. Neuroscience. 1996 Dec;75(4):1033–1045. doi: 10.1016/0306-4522(96)00225-4. [DOI] [PubMed] [Google Scholar]
  18. Hara H., Friedlander R. M., Gagliardini V., Ayata C., Fink K., Huang Z., Shimizu-Sasamata M., Yuan J., Moskowitz M. A. Inhibition of interleukin 1beta converting enzyme family proteases reduces ischemic and excitotoxic neuronal damage. Proc Natl Acad Sci U S A. 1997 Mar 4;94(5):2007–2012. doi: 10.1073/pnas.94.5.2007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hill I. E., MacManus J. P., Rasquinha I., Tuor U. I. DNA fragmentation indicative of apoptosis following unilateral cerebral hypoxia-ischemia in the neonatal rat. Brain Res. 1995 Apr 10;676(2):398–403. doi: 10.1016/0006-8993(95)00145-g. [DOI] [PubMed] [Google Scholar]
  20. Holtzman D. M., Bayney R. M., Li Y. W., Khosrovi H., Berger C. N., Epstein C. J., Mobley W. C. Dysregulation of gene expression in mouse trisomy 16, an animal model of Down syndrome. EMBO J. 1992 Feb;11(2):619–627. doi: 10.1002/j.1460-2075.1992.tb05094.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Holtzman D. M., Deshmukh M. Caspases: a treatment target for neurodegenerative disease? Nat Med. 1997 Sep;3(9):954–955. doi: 10.1038/nm0997-954. [DOI] [PubMed] [Google Scholar]
  22. Holtzman D. M., Sheldon R. A., Jaffe W., Cheng Y., Ferriero D. M. Nerve growth factor protects the neonatal brain against hypoxic-ischemic injury. Ann Neurol. 1996 Jan;39(1):114–122. doi: 10.1002/ana.410390117. [DOI] [PubMed] [Google Scholar]
  23. Johnston M. V. Neurotransmitter alterations in a model of perinatal hypoxic-ischemic brain injury. Ann Neurol. 1983 May;13(5):511–518. doi: 10.1002/ana.410130507. [DOI] [PubMed] [Google Scholar]
  24. Kumar S., Lavin M. F. The ICE family of cysteine proteases as effectors of cell death. Cell Death Differ. 1996 Jul;3(3):255–267. [PubMed] [Google Scholar]
  25. Loddick S. A., MacKenzie A., Rothwell N. J. An ICE inhibitor, z-VAD-DCB attenuates ischaemic brain damage in the rat. Neuroreport. 1996 Jun 17;7(9):1465–1468. doi: 10.1097/00001756-199606170-00004. [DOI] [PubMed] [Google Scholar]
  26. Mobley W. C., Rutkowski J. L., Tennekoon G. I., Gemski J., Buchanan K., Johnston M. V. Nerve growth factor increases choline acetyltransferase activity in developing basal forebrain neurons. Brain Res. 1986 Jul;387(1):53–62. doi: 10.1016/0169-328x(86)90020-3. [DOI] [PubMed] [Google Scholar]
  27. Ni B., Wu X., Du Y., Su Y., Hamilton-Byrd E., Rockey P. K., Rosteck P., Jr, Poirier G. G., Paul S. M. Cloning and expression of a rat brain interleukin-1beta-converting enzyme (ICE)-related protease (IRP) and its possible role in apoptosis of cultured cerebellar granule neurons. J Neurosci. 1997 Mar 1;17(5):1561–1569. doi: 10.1523/JNEUROSCI.17-05-01561.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Nitatori T., Sato N., Waguri S., Karasawa Y., Araki H., Shibanai K., Kominami E., Uchiyama Y. Delayed neuronal death in the CA1 pyramidal cell layer of the gerbil hippocampus following transient ischemia is apoptosis. J Neurosci. 1995 Feb;15(2):1001–1011. doi: 10.1523/JNEUROSCI.15-02-01001.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Oppenheim R. W. Cell death during development of the nervous system. Annu Rev Neurosci. 1991;14:453–501. doi: 10.1146/annurev.ne.14.030191.002321. [DOI] [PubMed] [Google Scholar]
  30. Palmer C., Vannucci R. C. Potential new therapies for perinatal cerebral hypoxia-ischemia. Clin Perinatol. 1993 Jun;20(2):411–432. [PubMed] [Google Scholar]
  31. Portera-Cailliau C., Price D. L., Martin L. J. Excitotoxic neuronal death in the immature brain is an apoptosis-necrosis morphological continuum. J Comp Neurol. 1997 Feb 3;378(1):70–87. [PubMed] [Google Scholar]
  32. Raff M. C., Barres B. A., Burne J. F., Coles H. S., Ishizaki Y., Jacobson M. D. Programmed cell death and the control of cell survival: lessons from the nervous system. Science. 1993 Oct 29;262(5134):695–700. doi: 10.1126/science.8235590. [DOI] [PubMed] [Google Scholar]
  33. Rice J. E., 3rd, Vannucci R. C., Brierley J. B. The influence of immaturity on hypoxic-ischemic brain damage in the rat. Ann Neurol. 1981 Feb;9(2):131–141. doi: 10.1002/ana.410090206. [DOI] [PubMed] [Google Scholar]
  34. Rink A., Fung K. M., Trojanowski J. Q., Lee V. M., Neugebauer E., McIntosh T. K. Evidence of apoptotic cell death after experimental traumatic brain injury in the rat. Am J Pathol. 1995 Dec;147(6):1575–1583. [PMC free article] [PubMed] [Google Scholar]
  35. Rothman S. M., Olney J. W. Glutamate and the pathophysiology of hypoxic--ischemic brain damage. Ann Neurol. 1986 Feb;19(2):105–111. doi: 10.1002/ana.410190202. [DOI] [PubMed] [Google Scholar]
  36. Schwartz L. M., Milligan C. E. Cold thoughts of death: the role of ICE proteases in neuronal cell death. Trends Neurosci. 1996 Dec;19(12):555–562. doi: 10.1016/s0166-2236(96)10067-9. [DOI] [PubMed] [Google Scholar]
  37. Schwartzman R. A., Cidlowski J. A. Apoptosis: the biochemistry and molecular biology of programmed cell death. Endocr Rev. 1993 Apr;14(2):133–151. doi: 10.1210/edrv-14-2-133. [DOI] [PubMed] [Google Scholar]
  38. Sidhu R. S., Tuor U. I., Del Bigio M. R. Nuclear condensation and fragmentation following cerebral hypoxia-ischemia occurs more frequently in immature than older rats. Neurosci Lett. 1997 Feb 21;223(2):129–132. doi: 10.1016/s0304-3940(97)13426-7. [DOI] [PubMed] [Google Scholar]
  39. Silverstein F. S., Barks J. D., Hagan P., Liu X. H., Ivacko J., Szaflarski J. Cytokines and perinatal brain injury. Neurochem Int. 1997 Apr-May;30(4-5):375–383. doi: 10.1016/s0197-0186(96)00072-1. [DOI] [PubMed] [Google Scholar]
  40. Silverstein F., Johnston M. V. Effects of hypoxia-ischemia on monoamine metabolism in the immature brain. Ann Neurol. 1984 Apr;15(4):342–347. doi: 10.1002/ana.410150407. [DOI] [PubMed] [Google Scholar]
  41. Sloviter R. S., Dean E., Neubort S. Electron microscopic analysis of adrenalectomy-induced hippocampal granule cell degeneration in the rat: apoptosis in the adult central nervous system. J Comp Neurol. 1993 Apr 15;330(3):337–351. doi: 10.1002/cne.903300305. [DOI] [PubMed] [Google Scholar]
  42. Tominaga T., Kure S., Narisawa K., Yoshimoto T. Endonuclease activation following focal ischemic injury in the rat brain. Brain Res. 1993 Apr 9;608(1):21–26. doi: 10.1016/0006-8993(93)90768-i. [DOI] [PubMed] [Google Scholar]
  43. Troy C. M., Stefanis L., Prochiantz A., Greene L. A., Shelanski M. L. The contrasting roles of ICE family proteases and interleukin-1beta in apoptosis induced by trophic factor withdrawal and by copper/zinc superoxide dismutase down-regulation. Proc Natl Acad Sci U S A. 1996 May 28;93(11):5635–5640. doi: 10.1073/pnas.93.11.5635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Vannucci R. C. Experimental biology of cerebral hypoxia-ischemia: relation to perinatal brain damage. Pediatr Res. 1990 Apr;27(4 Pt 1):317–326. doi: 10.1203/00006450-199004000-00001. [DOI] [PubMed] [Google Scholar]
  45. Wijsman J. H., Jonker R. R., Keijzer R., van de Velde C. J., Cornelisse C. J., van Dierendonck J. H. A new method to detect apoptosis in paraffin sections: in situ end-labeling of fragmented DNA. J Histochem Cytochem. 1993 Jan;41(1):7–12. doi: 10.1177/41.1.7678025. [DOI] [PubMed] [Google Scholar]
  46. Wyllie A. H., Kerr J. F., Currie A. R. Cell death: the significance of apoptosis. Int Rev Cytol. 1980;68:251–306. doi: 10.1016/s0074-7696(08)62312-8. [DOI] [PubMed] [Google Scholar]

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