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Transduction of axotomized retinal ganglion cells by adenoviral vector administration at the optic nerve stump: an in vivo model system for the inhibition of neuronal apoptotic cell death

Gene Therapy volume 6, pages 1759–1767 (1999)Cite this article

Abstract

Axotomy of the rat optic nerve leads to apoptotic cell death of retinal ganglion cells (RGCs). We have used adenoviral vectors to transduce RGCs from the cut optic nerve stump, a paradigm in which only those neurons are transduced which are directly affected by the axonal lesion. Transgenes encoded by the vectors were p35 and CrmA, which are potent intracellular anti-apoptotic proteins. We found that p35, but not CrmA exerted significant rescue effects on RGCs 14 days after axotomy. Expression of the transgenes was driven by the murine CMV (MCMV) promoter. The respective mRNAs were detectable 7 days but not 14 days after transduction. Since surviving RGCs were present beyond the time-point of detectable transcription of the p35 transgene, we conclude that apoptosis has been efficiently inhibited. In addition, we observed that transduction with two control vectors without a transgene in E1 also resulted in a minor but significant RGC rescue, implicating neuroprotective effects due to adenoviral transduction itself. This system will be useful in dissecting the pathways leading to neuronal cell death after axonal lesions and in the evaluation of the important question whether the cellular suicide program can be reverted to survival by therapeutic gene delivery.

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References

  1. Bergeron L, Yuan J . Sealing one’s fate: control of cell death in neurons Curr Opin Neurobiol 1998 8: 55–63

    Article  CAS  PubMed  Google Scholar 

  2. Jacobson MD, Weil M, Raff MC . Programmed cell death in animal development Cell 1997 88: 347–354

    Article  CAS  PubMed  Google Scholar 

  3. Strasser A, Huang DC, Vaux DL . The role of the bcl-2/ced-9 gene family in cancer and general implications of defects in cell death control for tumourigenesis and resistance to chemotherapy Biochim Biophys Acta 1997 1333: 151–178

    Google Scholar 

  4. Namura S et al. Activation and cleavage of caspase-3 in apoptosis induced by experimental cerebral ischemia J Neurosci 1998 18: 3659–3668

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Barinaga M . Stroke-damaged neurons may commit cellularsuicide Science 1998 281: 1302–1303

    Article  CAS  PubMed  Google Scholar 

  6. Guglielmo MA et al. The temporal profile and morphologic features of neuronal death in human stroke resemble those observed in experimental forebrain ischemia: the potential role of apoptosis Neurol Res 1998 20: 283–296

    Article  CAS  PubMed  Google Scholar 

  7. Iwahashi H et al. Synergistic anti-apoptotic activity between Bcl-2 and SMN implicated in spinal muscular atrophy Nature 1997 390: 413–417

    Article  CAS  PubMed  Google Scholar 

  8. Kim TW et al. Alternative cleavage of Alzheimer-associated presenilins during apoptosis by a caspase-3 family protease Science 1997 277: 373–376

    Article  CAS  PubMed  Google Scholar 

  9. Roperch JP et al. Inhibition of presenilin 1 expression is promoted by p53 and p21(WAF-1) and results in apoptosis and tumor suppression Nature Med 1998 4: 835–838

    Article  CAS  PubMed  Google Scholar 

  10. Conti AC, Raghupathi R, Trojanowski JQ, McIntosh TK . Experimental brain injury induces regionally distinct apoptosis during the acute and delayed post-traumatic period J Neurosci 1998 18: 5663–5672

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Yakovlev AG et al. Activation of CPP32-like caspases contributes to neuronal apoptosis and neurological dysfunction after traumatic brain injury J Neurosci 1997 17: 7415–7424

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Cohen GM . Caspases: the executioners of apoptosis Biochem J 1997 326: 1–16

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Ashkenazi A, Dixit VM . Death receptors: signaling and modulation Science 1998 281: 1305–1308

    Article  CAS  PubMed  Google Scholar 

  14. Frade JM, Barde YA . Microglia-derived nerve growth factor causes cell death in the developing retina Neuron 1998 20: 35–41

    Article  CAS  PubMed  Google Scholar 

  15. Frade JM et al. Control of early cell death by BDNF in the chick retina Development 1997 124: 3313–3320

    CAS  PubMed  Google Scholar 

  16. Green DR, Reed JC . Mitochondria and apoptosis Science 1998 281: 1309–1312

    Article  CAS  PubMed  Google Scholar 

  17. Garcia-Valenzuela E, Gorczyca W, Darzynkiewicz Z, Sharma SC . Apoptosis in adult retinal ganglion cells after axotomy J Neurobiol 1994 25: 431–438

    Article  CAS  PubMed  Google Scholar 

  18. Rabacchi SA, Bonfanti L, Liu XH, Maffei L . Apoptotic cell death induced by optic nerve lesion in the neonatal rat J Neurosci 1994 14: 5292–5301

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Isenmann S et al. Up-regulation of Bax protein in degenerating retinal ganglion cells precedes apoptotic cell death after optic nerve lesion in the rat Eur J Neurosci 1997 9: 1763–1772

    Article  CAS  PubMed  Google Scholar 

  20. Klöcker N, Cellerino A, Bähr M . Free radical scavenging and inhibition of nitric oxide synthase potentiates the neurotrophic effects of brain-derived neurotrophic factor on axotomized retinal ganglion cells in vivo J Neurosci 1998 18: 1038–1046

    Article  PubMed  PubMed Central  Google Scholar 

  21. Kermer P, Klöcker N, Labes M, Bähr M . Inhibition of CPP32-like proteases rescues axotomized retinal ganglion cells from secondary cell death in vivo J Neurosci 1998 18: 4656–4662

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Finiels F et al. Specific and efficient gene transfer strategy offers new potentialities for the treatment of motor neurone diseases Neuroreport 1995 6: 2473–2478

    Article  CAS  PubMed  Google Scholar 

  23. Cayouette M, Gravel C . Adenovirus-mediated gene transfer of ciliary neurotrophic factor can prevent photoreceptor degeneration in the retinal degeneration (rd) mouse Hum Gene Ther 1997 8: 423–430

    Article  CAS  PubMed  Google Scholar 

  24. Bertin J et al. Apoptotic suppression by baculovirus P35 involves cleavage by and inhibition of a virus-induced CED-3/ICE-like protease J Virol 1996 70: 6251–6259

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Zhou Q et al. Interaction of the baculovirus anti-apoptotic protein p35 with caspases. Specificity, kinetics, and characterization of the caspase/p35 complex Biochemistry 1998 37: 10757–10765

    Article  CAS  PubMed  Google Scholar 

  26. Zhou Q et al. Target protease specificity of the viral serpin CrmA. Analysis of five caspases J Biol Chem 1997 272: 7797–7800

    Article  CAS  PubMed  Google Scholar 

  27. Addison CL, Hitt M, Kunsken D, Graham FL . Comparison of the human versus murine cytomegalovirus immediate early gene promoters for transgene expression by adenoviral vectors J Gen Virol 1997 7: 1653–1661

    Article  Google Scholar 

  28. Bilang-Bleuel A et al. Intrastriatal injection of an adenoviral vector expressing glial-cell-line-derived neurotrophic factor prevents dopaminergic neuron degeneration and behavioral impairment in a rat model of Parkinson disease Proc Natl Acad Sci USA 1997 94: 8818–8823

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Choi-Lundberg DL et al. Dopaminergic neurons protected from degeneration by GDNF gene therapy Science 1997 275: 838–841

    Article  CAS  PubMed  Google Scholar 

  30. Gravel C, Gotz R, Lorrain A, Sendtner M . Adenoviral gene transfer of ciliary neurotrophic factor and brain-derived neurotrophic factor leads to long-term survival of axotomized motor neurons Nature Med 1997 3: 765–770

    Article  CAS  PubMed  Google Scholar 

  31. Di-Polo A et al. Prolonged delivery of brain-derived neurotrophic factor by adenovirus-infected Muller cells temporarily rescues injured retinal ganglion cells Proc Natl Acad Sci USA 1998 95: 3978–3983

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Lawrence MS et al. Overexpression of Bcl-2 with herpes simplex virus vectors protects CNS neurons against neurological insults in vitro and in vivo J Neurosci 1996 16: 486–496

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Xu DG et al. Elevation of neuronal expression of NAIP reduces ischemic damage in the rat hippocampus Nature Med 1997 3: 997–1004

    Article  CAS  PubMed  Google Scholar 

  34. Niederkorn JY . Immune privilege and immune regulation in the eye Adv Immunol 1990 48: 191–226

    Article  CAS  PubMed  Google Scholar 

  35. Thornberry NA, Lazebnik Y . Caspases: enemies within Science 1998 281: 1312–1316

    Article  CAS  PubMed  Google Scholar 

  36. Kermer P et al. CPP-32 like proteases are major mediators of apoptosis in axotomized retinal ganglion cells in vivo Soc Neurosci 1998 24: 614.13 (Abstr.)

    Google Scholar 

  37. Li P et al. Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade Cell 1997 91: 479–489

    Article  CAS  PubMed  Google Scholar 

  38. Pan G, Humke EW, Dixit VM . Activation of caspases triggered by cytochrome c in vitro FEBS Lett 1998 426: 151–154

    Article  CAS  PubMed  Google Scholar 

  39. Byrnes AP, Wood MJ, Charlton HM . Role of T cells in inflammation caused by adenovirus vectors in the brain Gene Therapy 1996 3: 644–651

    CAS  PubMed  Google Scholar 

  40. Byrnes AP, MacLaren RE, Charlton HM . Immunological instability of persistent adenovirus vectors in the brain: peripheral exposure to vector leads to renewed inflammation, reduced gene expression, and demyelination J Neurosci 1996 16: 3045–3055

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Oral HB et al. Ex vivo adenovirus-mediated gene transfer and immunomodulatory protein production in human cornea Gene Therapy 1997 4: 639–647

    Article  CAS  PubMed  Google Scholar 

  42. Palmer TD, Rosman GJ, Osborne WR, Miller AD . Genetically modified skin fibroblasts persist long after transplantation but gradually inactivate introduced genes Proc Natl Acad Sci USA 1991 88: 1330–1334

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Engel S et al . Limited transgene expression after adenoviral neurotrophin gene transfer in the rat retina is unrelated to host immune responses (submitted)

  44. Tiensiwakul P, Khoobyarian N . Adenovirus fiber protein produces synthesis of interferon in mouse spleen and macrophage cultures Intervirology 1983 20: 52–55

    Article  CAS  PubMed  Google Scholar 

  45. Reich N, Pine R, Levy D, Darnell JE Jr . Transcription of interferon-stimulated genes is induced by adenovirus particles but is suppressed by E1A gene products J Virol 1988 62: 114–119

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Harms JS, Splitter GA . Interferon-gamma inhibits transgene expression driven by SV40 or CMV promoters but augments expression driven by the mammalian MHC1 promoter Hum Gene Ther 1995 6: 1291–1297

    Article  CAS  PubMed  Google Scholar 

  47. Cayouette M, Gravel C . Adenovirus-mediated gene transfer to retinal ganglion cells Inv Opthal Vis Sci 1996 37: 2022–2028

    CAS  Google Scholar 

  48. Bett AJ, Haddara W, Prevec L, Graham FL . An efficient and flexible system for construction of adenovirus vectors with insertions or deletions in early regions 1 and 3 Proc Natl Acad Sci USA 1994 91: 8802–8806

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Dion LD, Fang J, Garver RI Jr . Supernatant rescue assay vs. polymerase chain reaction for detection of wild type adenovirus-contaminating recombinant adenovirus stocks J Virol Meth 1996 56: 99–107

    Article  CAS  Google Scholar 

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Acknowledgements

We are grateful to G Salvesen for the p35 cDNA and to J Yuan for the crmA cDNA. We thank Liane Meyn and Susanne Thomson for excellent technical assistance and Monika Labes for critical reading of the manuscript. This work was supported by BMBF grant 01 KS 9602.

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Authors and Affiliations

  1. Department of Neurology, University of Tuebingen, Medical School, Tuebingen, Germany

    S Kügler, N Klöcker, P Kermer, S Isenmann & M Bähr

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Kügler, S., Klöcker, N., Kermer, P. et al. Transduction of axotomized retinal ganglion cells by adenoviral vector administration at the optic nerve stump: an in vivo model system for the inhibition of neuronal apoptotic cell death. Gene Ther 6, 1759–1767 (1999). https://doi.org/10.1038/sj.gt.3301000

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