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Assessment of Cell Death: Apoptosis, Necrosis, or In Between

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Animal Models of Acute Neurological Injuries II

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Abstract

Cell death and cell survival are the fundamental events in development and pathological conditions associated with virtually all disorders in the central nervous system (CNS) and peripheral nervous system (PNS). Assessment of cell death and identification of the type of cell death are critical for understanding the mechanism of the injury and for development of therapeutic treatment of diseases. Recent development in cellular and molecular biological discoveries has allowed better characterization of different cell death including necrosis, apoptosis, and autophagic death and development of numerous cell death assays. This chapter intends to introduce some common characteristics of different types of cell death and the most basic principle of assessments. Emerging evidence also show that sometimes it is difficult to separate different cell death mechanisms, and mixed cell death often is observed under in vivo situation. We hope the content of this chapter will help readers to better understand and develop novel ideas of cell death in diseases.

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References

  1. Garnier P, Prigent-Tessier A, Van Hoecke M, Bertrand N, Demougeot C, Sordet O, Swanson RA, Marie C, Beley A (2004) Hypoxia induces caspase-9 and caspase-3 activation without neuronal death in gerbil brains. Eur J Neurosci 20:937–946

    Article  PubMed  Google Scholar 

  2. Zhivotovsky B (2004) Apoptosis, necrosis and between. Cell Cycle 3:64–66

    Article  PubMed  CAS  Google Scholar 

  3. Xiao AY, Wei L, Xia S, Rothman S, Yu SP (2002) Ionic mechanism of ouabain-induced concurrent apoptosis and necrosis in individual cultured cortical neurons. J Neurosci 22:1350–1362

    PubMed  CAS  Google Scholar 

  4. Wei L, Ying DJ, Cui L, Langsdorf J, Yu SP (2004) Necrosis, apoptosis and hybrid death in the cortex and thalamus after barrel cortex ischemia in rats. Brain Res 1022:54–61

    Article  PubMed  CAS  Google Scholar 

  5. Wang XQ, Xiao AY, Yang A, LaRose L, Wei L, Yu SP (2003) Block of Na+, K+−ATPase and induction of hybrid death by 4-aminopyridine in cultured cortical neurons. J Pharmacol Exp Ther 305:502–506

    Article  PubMed  CAS  Google Scholar 

  6. Wei L, Han BH, Li Y, Keogh CL, Holtzman DM, Yu SP (2006) Cell death mechanism and protective effect of erythropoietin after focal ischemia in the whisker-barrel cortex of neonatal rats. J Pharmacol Exp Ther 317:109–116

    Article  PubMed  CAS  Google Scholar 

  7. Yakovlev AG, Faden AI (2004) Mechanisms of neural cell death: implications for development of neuroprotective treatment strategies. NeuroRx 1:5–16

    Article  PubMed  Google Scholar 

  8. Nicotera P, Melino G (2004) Regulation of the apoptosis-necrosis switch. Oncogene 23:2757–2765

    Article  PubMed  CAS  Google Scholar 

  9. Zeiss CJ (2003) The apoptosis-necrosis continuum: insights from genetically altered mice. Vet Pathol 40:481–495

    Article  PubMed  CAS  Google Scholar 

  10. Formigli L, Papucci L, Tani A, Schiavone N, Tempestini A, Orlandini GE, Capaccioli S, Orlandini SZ (2000) Aponecrosis: morphological and biochemical exploration of a syncretic process of cell death sharing apoptosis and necrosis. J Cell Physiol 182:41–49

    Article  PubMed  CAS  Google Scholar 

  11. Lemasters JJ (1999) V. Necrapoptosis and the mitochondrial permeability transition: shared pathways to necrosis and apoptosis. Am J Physiol 276:G1–G6

    PubMed  CAS  Google Scholar 

  12. Hu X, Xuan Y (2008) Bypassing cancer drug resistance by activating multiple death pathways–a proposal from the study of circumventing cancer drug resistance by induction of necroptosis. Cancer Lett 259:127–137

    Article  PubMed  CAS  Google Scholar 

  13. Moquin D, Chan FK (2010) The molecular regulation of programmed necrotic cell injury. Trends Biochem Sci 35(8):434–441

    Article  PubMed  CAS  Google Scholar 

  14. Degterev A, Huang Z, Boyce M, Li Y, Jagtap P, Mizushima N, Cuny GD, Mitchison TJ, Moskowitz MA, Yuan J (2005) Chemical inhibitor of nonapoptotic cell death with therapeutic potential for ischemic brain injury. Nat Chem Biol 1:112–119

    Article  PubMed  CAS  Google Scholar 

  15. Liu M, Dziennis S, Hurn PD, Alkayed NJ (2009) Mechanisms of gender-linked ischemic brain injury. Restor Neurol Neurosci 27:163–179

    PubMed  CAS  Google Scholar 

  16. Pearson-Fuhrhop KM, Kleim JA, Cramer SC (2009) Brain plasticity and genetic factors. Top Stroke Rehabil 16:282–299

    Article  PubMed  Google Scholar 

  17. Arnold M, Halpern M, Meier N, Fischer U, Haefeli T, Kappeler L, Brekenfeld C, Mattle HP, Nedeltchev K (2008) Age-dependent differences in demographics, risk factors, co-morbidity, etiology, management, and clinical outcome of acute ischemic stroke. J Neurol 255:1503–1507

    Article  PubMed  CAS  Google Scholar 

  18. Johansson BB (2004) Functional and cellular effects of environmental enrichment after experimental brain infarcts. Restor Neurol Neurosci 22:163–174

    PubMed  Google Scholar 

  19. Seidman C, Kirkham F, Pavlakis S (2007) Pediatric stroke: current developments. Curr Opin Pediatr 19:657–662

    Article  PubMed  Google Scholar 

  20. Raju TN, Nelson KB, Ferriero D, Lynch JK (2007) Ischemic perinatal stroke: summary of a workshop sponsored by the National Institute of Child Health and Human Development and the National Institute of Neurological Disorders and Stroke. Pediatrics 120:609–616

    Article  PubMed  Google Scholar 

  21. Machaalani R, Radford JL, Waters KA (2007) Tissue fixation effects on immunohistochemical staining of caspase-3 in brain tissue. Appl Immunohistochem Mol Morphol 15:463–470

    Article  PubMed  CAS  Google Scholar 

  22. Borst P, Rottenberg S (2004) Cancer cell death by programmed necrosis? Drug Resist Updat 7:321–324

    Article  PubMed  CAS  Google Scholar 

  23. Boujrad H, Gubkina O, Robert N, Krantic S, Susin SA (2007) AIF-mediated programmed necrosis: a highly regulated way to die. Cell Cycle 6:2612–2619

    Article  PubMed  CAS  Google Scholar 

  24. Galluzzi L, Kroemer G (2009) Shigella targets the mitochondrial checkpoint of programmed necrosis. Cell Host Microbe 5:107–109

    Article  PubMed  CAS  Google Scholar 

  25. Aigner T (2002) Apoptosis, necrosis, or whatever: how to find out what really happens? J Pathol 198:1–4

    Article  PubMed  CAS  Google Scholar 

  26. Galluzzi L, Aaronson SA, Abrams J, Alnemri ES, Andrews DW, Baehrecke EH, Bazan NG, Blagosklonny MV, Blomgren K, Borner C, Bredesen DE, Brenner C, Castedo M, Cidlowski JA, Ciechanover A, Cohen GM, De Laurenzi V, De Maria R, Deshmukh M, Dynlacht BD, El-Deiry WS, Flavell RA, Fulda S, Garrido C, Golstein P, Gougeon ML, Green DR, Gronemeyer H, Hajnoczky G, Hardwick JM, Hengartner MO, Ichijo H, Jaattela M, Kepp O, Kimchi A, Klionsky DJ, Knight RA, Kornbluth S, Kumar S, Levine B, Lipton SA, Lugli E, Madeo F, Malomi W, Marine JC, Martin SJ, Medema JP, Mehlen P, Melino G, Moll UM, Morselli E, Nagata S, Nicholson DW, Nicotera P, Nunez G, Oren M, Penninger J, Pervaiz S, Peter ME, Piacentini M, Prehn JH, Puthalakath H, Rabinovich GA, Rizzuto R, Rodrigues CM, Rubinsztein DC, Rudel T, Scorrano L, Simon HU, Steller H, Tschopp J, Tsujimoto Y, Vandenabeele P, Vitale I, Vousden KH, Youle RJ, Yuan J, Zhivotovsky B, Kroemer G (2009) Guidelines for the use and interpretation of assays for monitoring cell death in higher eukaryotes. Cell Death Differ 16:1093–1107

    Article  PubMed  CAS  Google Scholar 

  27. Broughton BR, Reutens DC, Sobey CG (2009) Apoptotic mechanisms after cerebral ischemia. Stroke 40:e331–e339

    Article  PubMed  Google Scholar 

  28. Ferrer I, Planas AM (2003) Signaling of cell death and cell survival following focal cerebral ischemia: life and death struggle in the penumbra. J Neuropathol Exp Neurol 62:329–339

    PubMed  Google Scholar 

  29. Rami A (2008) Upregulation of Beclin 1 in the ischemic penumbra. Autophagy 4:227–229

    PubMed  CAS  Google Scholar 

  30. Zheng YQ, Liu JX, Li XZ, Xu L, Xu YG (2009) RNA interference-mediated downregulation of Beclin1 attenuates cerebral ischemic injury in rats. Acta Pharmacol Sin 30:919–927

    Article  PubMed  CAS  Google Scholar 

  31. Lai Y, Hickey RW, Chen Y, Bayir H, Sullivan ML, Chu CT, Kochanek PM, Dixon CE, Jenkins LW, Graham SH, Watkins SC, Clark RS (2008) Autophagy is increased after traumatic brain injury in mice and is partially inhibited by the antioxidant gamma-glutamylcysteinyl ethyl ester. J Cereb Blood Flow Metab 28:540–550

    Article  PubMed  CAS  Google Scholar 

  32. Lakhan SE, Kirchgessner A, Hofer M (2009) Inflammatory mechanisms in ischemic stroke: therapeutic approaches. J Transl Med 7:97

    Article  PubMed  Google Scholar 

  33. Yu SP, Yeh CH, Sensi SL, Gwag BJ, Canzoniero LM, Farhangrazi ZS, Ying HS, Tian M, Dugan LL, Choi DW (1997) Mediation of neuronal apoptosis by enhancement of outward potassium current. Science 278:114–117

    Article  PubMed  CAS  Google Scholar 

  34. Yu SP (2003) Regulation and critical role of potassium homeostasis in apoptosis. Prog Neurobiol 70:363–386

    Article  PubMed  CAS  Google Scholar 

  35. Yu SP, Choi DW (2000) Ions, cell volume, and apoptosis. Proc Natl Acad Sci USA 97:9360–9362

    Article  PubMed  CAS  Google Scholar 

  36. Koopman G, Reutelingsperger CP, Kuijten GA, Keehnen RM, Pals ST, van Oers MH (1994) Annexin V for flow cytometric detection of phosphatidylserine expression on B cells undergoing apoptosis. Blood 84:1415–1420

    PubMed  CAS  Google Scholar 

  37. Zhivotosky B, Orrenius S (2001) Assessment of apoptosis and necrosis by DNA fragmentation and morphological criteria. Curr Protoc Cell Biol Chapter 18:Unit 18.13

    Google Scholar 

  38. Burg ED, Remillard CV, Yuan JX (2006) K+ channels in apoptosis. J Membr Biol 209:3–20

    Article  PubMed  CAS  Google Scholar 

  39. Borrelli MJ, Carlini WG, Dewey WC, Ransom BR (1985) A simple method for making ion-selective microelectrodes suitable for intracellular recording in vertebrate cells. J Neurosci Methods 15:141–154

    Article  PubMed  CAS  Google Scholar 

  40. Ammann D, Chao PS, Simon W (1987) Valinomycin-based K+ selective microelectrodes with low electrical membrane resistance. Neurosci Lett 74:221–226

    Article  PubMed  CAS  Google Scholar 

  41. Nicholson C (1993) Ion-selective microelectrodes and diffusion measurements as tools to explore the brain cell microenvironment. J Neurosci Methods 48:199–213

    Article  PubMed  CAS  Google Scholar 

  42. Jezek P, Mahdi F, Garlid KD (1990) Reconstitution of the beef heart and rat liver mitochondrial K+/H+ (Na+/H+) antiporter. Quantitation of K+ transport with the novel fluorescent probe, PBFI. J Biol Chem 265:10522–10526

    PubMed  CAS  Google Scholar 

  43. Meuwis K, Boens N, De Schryver FC, Gallay J, Vincent M (1995) Photophysics of the fluorescent K+ indicator PBFI. Biophys J 68:2469–2473

    Article  PubMed  CAS  Google Scholar 

  44. Cook AL, Frydenberg M, Haynes JM (2002) Protein kinase G activation of K(ATP) channels in human-cultured prostatic stromal cells. Cell Signal 14:1023–1029

    Article  PubMed  CAS  Google Scholar 

  45. Liu D, Slevin JR, Lu C, Chan SL, Hansson M, Elmer E, Mattson MP (2003) Involvement of mitochondrial K+ release and cellular efflux in ischemic and apoptotic neuronal death. J Neurochem 86:966–979

    Article  PubMed  CAS  Google Scholar 

  46. Gogvadze V, Orrenius S, Zhivotovsky B (2003) Analysis of mitochondrial dysfunction during cell death. Curr Protoc Cell Biol Chapter 18:Unit 18.15

    Google Scholar 

  47. Kobayashi T, Kuroda S, Tada M, Houkin K, Iwasaki Y, Abe H (2003) Calcium-induced mitochondrial swelling and cytochrome c release in the brain: its biochemical characteristics and implication in ischemic neuronal injury. Brain Res 960:62–70

    Article  PubMed  CAS  Google Scholar 

  48. Bevers MB, Neumar RW (2008) Mechanistic role of calpains in postischemic neurodegeneration. J Cereb Blood Flow Metab 28:655–673

    Article  PubMed  CAS  Google Scholar 

  49. Chaitanya GV, Babu PP (2008) Activation of calpain, cathepsin-b and caspase-3 during transient focal cerebral ischemia in rat model. Neurochem Res 33:2178–2186

    Article  PubMed  CAS  Google Scholar 

  50. Jiang SX, Kappler J, Zurakowski B, Desbois A, Aylsworth A, Hou ST (2007) Calpain cleavage of collapsin response mediator proteins in ischemic mouse brain. Eur J Neurosci 26:801–809

    Article  PubMed  Google Scholar 

  51. Kaufmann SH, Kottke TJ, Martins LM, Henzing AJ, Earnshaw WC (2001) Analysis of caspase activation during apoptosis. Curr Protoc Cell Biol Chapter 18:Unit 18.12

    Google Scholar 

  52. Oo TF, Burke RE (2007) Histochemical methods for the detection of apoptosis in the nervous system. Curr Protoc Neurosci Chapter 1:Unit 1.15

    Google Scholar 

  53. McLaughlin B, Hartnett KA, Erhardt JA, Legos JJ, White RF, Barone FC, Aizenman E (2003) Caspase 3 activation is essential for neuroprotection in preconditioning. Proc Natl Acad Sci USA 100:715–720

    Article  PubMed  CAS  Google Scholar 

  54. Haince JF, Poirier GG, Kirkland JB (2004) Nonisotopic methods for determination of poly(ADP-ribose) levels and detection of poly(ADP-ribose) polymerase. Curr Protoc Cell Biol Chapter 18:Unit18.17

    Google Scholar 

  55. Hong SJ, Dawson TM, Dawson VL (2004) Nuclear and mitochondrial conversations in cell death: PARP-1 and AIF signaling. Trends Pharmacol Sci 25:259–264

    Article  PubMed  CAS  Google Scholar 

  56. Niizuma K, Endo H, Nito C, Myer DJ, Kim GS, Chan PH (2008) The PIDDosome mediates delayed death of hippocampal CA1 neurons after transient global cerebral ischemia in rats. Proc Natl Acad Sci USA 105:16368–16373

    Article  PubMed  CAS  Google Scholar 

  57. Chesselet MF, MacKenzie L, Hoang T (2001) Detection of DNA damage in tissue sections by in situ nick translation. Curr Protoc Neurosci Chapter 1:Unit 1.9

    Google Scholar 

  58. Yung YC, Kennedy G, Chun J (2009) Identification of neural programmed cell death through the detection of DNA fragmentation in situ and by PCR. Curr Protoc Neurosci Chapter 3:Unit 3.8

    Google Scholar 

  59. Rello S, Stockert JC, Moreno V, Gamez A, Pacheco M, Juarranz A, Canete M, Villanueva A (2005) Morphological criteria to distinguish cell death induced by apoptotic and necrotic treatments. Apoptosis 10:201–208

    Article  PubMed  CAS  Google Scholar 

  60. Blaschke AJ, Staley K, Chun J (1996) Widespread programmed cell death in proliferative and postmitotic regions of the fetal cerebral cortex. Development 122:1165–1174

    PubMed  CAS  Google Scholar 

  61. Chu CT, Plowey ED, Dagda RK, Hickey RW, Cherra SJ 3rd, Clark RS (2009) Autophagy in neurite injury and neurodegeneration: in vitro and in vivo models. Methods Enzymol 453:217–249

    Article  PubMed  CAS  Google Scholar 

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Genetta, T., Wei, L., Yu, S.P. (2012). Assessment of Cell Death: Apoptosis, Necrosis, or In Between. In: Chen, J., Xu, XM., Xu, Z., Zhang, J. (eds) Animal Models of Acute Neurological Injuries II. Springer Protocols Handbooks. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-576-3_2

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  • DOI: https://doi.org/10.1007/978-1-61779-576-3_2

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