Cell Death during Developmental Processes


Embryonic development and differentiation to adult form depends on orchestration of cell division and death. In embryos, programmed death sculpts form, opens lumens, separates or splits tissue layers, allows tissue layers to fuse and removes vestigial organs. Both the central nervous and immune systems overproduce cells and destroy those that do not form successful synapses or produce unusable antibodies. Cell death is first seen in mammalian embryos when the blastocyst expands, but elsewhere, the first deaths are not seen before the maternal–zygotic transition. Abnormal timing, amount or localisation of cell death leads to abnormalities or death of embryos.

Several signalling pathways trigger cell death. Usually, the signals activate caspases (first discovered in embryonic cell death in nematodes) and lead to apoptosis, although apoptosis is not the only form of cell death. The signalling mechanisms that control cell death in embryos are not well understood, but should be if we hope to understand normal and teratological development.

Key Concepts

  • Cell death can be seen in both embryonic development and normal growth of adult tissue.
  • The embryonic cell deaths are highly programmed in that they are predictable in location, time and amount. In the simplest instances, such as in nematodes, control of cell death is under direct control of a small number of genes.
  • Most but not all of the embryonic deaths are apoptotic.
  • Embryonic cell to sculpt the embryo and define the boundaries of tissues and organs. In the central nervous system and the immune system, overgrowth (production of excessive cells) and subsequent pruning by cell death generate the high specificity that characterises these systems.
  • Deregulation of apoptosis can produce many embryonic abnormalities and teratologies and, later in life, produces cancers, autoimmune disease or neurodegenerative disease.
  • There are many means to study cell death, but only a few are directly applicable to the study of cell death in embryos. Nevertheless, further study is needed to understand the signalling mechanisms that decide the death of cells in specific locations and times.
  • Autophagy is increasingly recognised as a factor influencing the likelihood of onset of apoptosis and cell death.
  • New ultra‐resolution fluorescence microscopic techniques, capable of simultaneously analysing energy flux, autophagy and apoptosis, may lead to new insights into these questions.
  • Learning more about cell death in embryos will help us understand how it is controlled in adults.

Keywords: apoptosis; autophagy; caspase; cell death receptor; embryo; embryonic development; gene expression; programmed cell death; techniques

Figure 1. Schematic overview of the major cell death (apoptotic) pathways. Two major pathways lead to apoptosis: external (type I or ligand‐mediated) and internal (type II or metabolically mediated). When apoptosis is initiated through the external pathway, signalling occurs through the interaction between extracellular death ligands such as FasL or tumour necrosis factor α (TNFα) and death receptors such as type I receptor for tumour necrosis factor (TNFR1) and TNF‐related apoptosis‐inducing ligand (TRAIL) receptors followed by activation of the death domain of intracellular part of receptor. Then these components form the death‐inducing signalling complex (DISC), which recruits and activates procaspase‐8. Activated caspase‐8 is an initiator caspase that subsequently cleaves and activates downstream effector caspases such as caspase‐3 and caspase‐7 and finally results in apoptotic cell death. The intrinsic cell death pathway is triggered by a metabolic problem, the nature of which is unknown for embryonic as opposed to experimental models. The signal initiates from within the cell and is mediated by mitochondria, which releases proapoptotic factors from the mitochondria. This release is tightly controlled by the proteins Bak and Bax and BH3‐only protein. The released factors, including cytochrome c and smac/Diablo, lead to the formation of apoptosome in association with Apaf‐1 and procaspase‐9. Activation of caspase‐9 leads to activation of caspase‐3 and apoptotic cell death. The apoptosis‐inducing factor (AIF), released by mitochondria, is capable of inducing apoptosis independent of caspases (Candé et al., ). These steps can be inhibited by the antiapoptotic members of the Bcl‐2 family of apoptosis regulators. Finally, spontaneously for massive and perhaps some other cells, and otherwise if for any reason apoptosis is not evoked, cells destroy the bulk of their contents through autophagy. This may lead to the death of the cell with or without any sign of apoptosis.
Figure 2. Cell death during the morphogenesis of the mouse limb. (a) Embryonic day 13.5 mouse forelimb stained with Nile blue sulphate. Regions of cell death, indicated by the dye picked up by phagocytes, are obvious in the interdigital regions, along the apical ectodermal ridge, and along the anterior (inner) side of the limb. (b) Micrograph of the interdigital region of a similar limb, showing apoptotic cells identified by a positive TUNEL reaction (magenta stain). (c) A semi‐thin section of a similar section, in which apoptotic cells, many within phagocytes, are identified by the condensation of chromatin that gives nuclear material a very dense appearance when stained, as here, by toluidine blue or any other acidophilic dye.
Figure 3. Localised cell deaths during morphogenesis in zebrafish. (a) Dying cells revealed by acridine orange staining in the lens of a 25‐h zebrafish embryo. Focus is at the interface between developing lens and lens epithelium. (b) Lens stalk retained between lens and epithelium if apoptosis is blocked by inhibitor of caspase 3. (c) Caspase‐3 activity, revealed by presence of fluorescent inhibitor zVAD‐FMK (carbobenzoxy‐valyl‐alanyl‐aspartyl‐[O‐methyl]‐fluoromethylketone or Carbobenzoxy‐Val‐Ala‐Asp‐Fluoromethylketone), in what will become the vent (anus) of a 48‐h zebrafish embryo. (d) Phase‐contrast image of the same embryo, illustrating the developing vent. If caspase activity is inhibited, the vent will not open. Notes: L, lens and R, retina. Figure courtesy of Nathaniel Abraham.
Figure 4. Cell death along the anterior–posterior axis in the zebrafish larva. (a) Dying cells scattered across an approximately 20‐h zebrafish embryo, revealed by staining with acridine orange. (b) Death of Mauthner cells (large cells regularly spaced along the spine) and a few cells in the tail of an approximately 96‐h zebrafish larva. Photographed by Richard A. Lockshin © R. Lockshin 2010.
Figure 5. What makes a cell die: Not all deaths are apoptosis. Often a massive accumulation of autophagic vacuoles (here observed as green GFP‐LC3 positive structures), which can be seen in close spatiotemporal relationship to the mitochondria (red) precedes the onset of cell death. Scale bar: 20 µm.
Figure 6. A point of no return (PONR) for apoptosis has been previously described in context of mitochondrial depolarisation and is often preceded by changes in the mitochondrial network. These changes can now be morphometrically assessed in great detail, revealing mitochondrial loop formations or shifts in the fission and fusion equilibrium. Mouse hypothalamic GT1‐7 neuronal stained with JC‐1. This ratiometric fluorescent dye indicates polarised (red) and depolarised (green) regions of the mitochondria. SR‐SIM acquisition and three‐dimensional rendering reveals a distinct mitochondrial network that may dynamically shift between a fission (a) or fusion state (b). Depth colour coding (c) reveals the spatial organisation in z, with orange indicating the most superficial mitochondria. Often also circular mitochondria are observed, that can, due to super‐resolution, be quantitatively assessed (d).


Alles AJ and Sulik KK (1989) Retinoic‐acid‐induced limb‐reduction defects: perturbation of zones of programmed cell death as a pathogenetic mechanism. Teratology 40: 163–171.

Berry DL and Baehrecke EH (2007) Growth arrest and autophagy are required for salivary gland cell degradation in Drosophila. Cell 13: 1137–1148.

Blackstone C and Chang CR (2011) Mitochondria unite to survive. Nature Cell Biology 13: 521–522.

Candé C, Cohen I, Daugas E, et al. (2002) Apoptosis‐inducing factor (AIF): a novel caspase‐independent death effector released from mitochondria. Biochimie 84: 215–222.

Cole LK and Ross LS (2001) Apoptosis in the developing zebrafish embryo. Developmental Biology 240: 123–142.

Dekkers MP, Nikoletopoulou V and Barde YA (2013) Cell biology in neuroscience: Death of developing neurons: new insights and implications for connectivity. Journal of Cell Biology 203: 385–393.

Ernsberger U (2009) Role of neurotrophin signalling in the differentiation of neurons from dorsal root ganglia and sympathetic ganglia. Cell and Tissue Research 336: 349–384.

Fuchs Y and Steller H (2011) Programmed Cell Death in Animal Development and Disease. Cell 147: 742–758.

Galluzzi L, Bravo‐San Pedro JM, Vitale I, et al. (2014) Essential versus accessory aspects of cell death: recommendations of the NCCD 2015. Cell Death and Differentiation 22: 58–73.

Glücksmann A (1951) Cell deaths in normal vertebrate ontogeny. Biological Reviews of the Cambridge Philosophical Society 26: 59.

Green DR and Levine B (2014) To be or not to be? How selective autophagy and cell death govern cell fate. Cell 157: 65–75.

Hengartner MO and Horvitz HR (1994) The ins and outs of programmed cell death during C. elegans development. Philosophical Transactions of the Royal Society of London, Series B: Biological Sciences 345: 243–246.

Hensey C and Gautier JA (1997) A developmental timer that regulates apoptosis at the onset of gastrulation. Mechanisms of Development 69: 183–195.

Hurlé JM and Colombatti A (1996) Extracellular matrix modifications in the interdigital spaces of the chick embryo leg bud during the formation of ectopic digits. Anatomy and Embryology 193: 355–364.

Hurlé JM, Ros MA, Climent V and Garcia‐Martinez V (1996) Morphology and significance of programmed cell death in the developing limb bud of the vertebrate embryo. Microscopy Research and Technique 34: 236–246.

Kerr JF, Wyllie AH and Currie AR (1972) Apoptosis: a basic biological phenomenon with wide‐ranging implications in tissue kinetics. British Journal of Cancer 26 (4): 239–257.

Kristiansen M and Ham J (2014) Programmed cell death during neuronal development: the sympathetic neuron model. Cell Death and Differentiation 21: 1025–1035.

Kroemer G, El‐Deiry WS, Golstein P, et al. (2005) Nomenclature committee on cell death. Classification of cell death: recommendations of the nomenclature committee on cell death. Cell Death and Differentiation 12 (suppl. 2): 1463–1467.

Leist M, Single B, Castoldi AF, et al. (1997) Intracellular adenosine triphosphase (ATP) concentration: a switch in the decision between apoptosis and necrosis. Journal of Experimental Medicine 185: 1481–1486.

Levi‐Montalcini R and Hamburger V (1951) Selective growth stimulating effects of mouse sarcoma on the sensory and sympathetic nervous system of the chick embryo. Journal of Experimental Zoology 116 (2): 321–361.

Liu Y, Shoji‐Kawata S, Sumpter RM Jr et al. (2013) Autosis is a Na+, K+ ATPase‐regulated form of cell death triggered by autophagy‐inducing peptides, starvation, and hypoxia‐ischemia. Proceedings of the National Academy of Sciences of the United States of America 110: 20364–20371.

Lockshin RA and Williams CM (1965) Programmed cell death: I. Cytology of the degeneration of the intersegmental muscles of the Pernyi silkmoth. Journal of Insect Physiology 11: 123–133.

Lockshin RA (1969) Programmed cell death. Activation of lysis by a mechanism involving the synthesis of protein. Journal of Insect Physiology 15: 1505–1516.

Lockshin RA and Zakeri Z (2001) Programmed cell death and apoptosis: origins of the theory. Nature Reviews. Molecular Cell. Biology 2: 545–550.

Loos B, Genade S, Ellis B, Lochner A and Engelbrecht AM (2011) At the core of survival: Autophagy delays the onset of both apoptotic and necrotic cell death in a model of ischemic cell injury. Experimental Cell Research 317 (10): 1437–1453.

Loos B, Engelbrecht A, Lockshin RA, Klionsky DJ and Zakeri Z (2013) The variability of autophagy and cell death susceptibility: Unanswered questions. Autophagy 9: 1270–1285.

Loos B, Du Toit A and Hofmeyr JH (2014) Defining and measuring autophagosome flux‐concept and reality. Autophagy 10. DOI: 10.4161/15548627.2014.973338.

Milligan CE and Schwartz LM (1997) Programmed cell death during normal development. British Medical Bulletin 52: 570–590.

Mirkes PE (1985) Cyclophosphamide teratogenesis. Teratogenesis, Carcinogenesis, and Mutagenesis 5: 75–88.

Muñoz‐Pinedo C, Guío‐Carrión A, Goldstein JC, et al. (2006) Different mitochondrial intermembrane space proteins are released during apoptosis in a manner that is coordinately initiated but can vary in duration. Proceedings of the National Academy of Sciences of the United States of America 103: 11573–11578.

Negron JF and Lockshin RA (2004) Activation of apoptosis and caspase‐3 in zebrafish early gastrulae. Developmental Dynamics 231: 161–170.

Oppenheim RW, Flavell RA, Vinsant S, et al. (2001) Programmed cell death of developing mammalian neurons after genetic deletion of caspases. Journal of Neurosciences 21: 4752–4760.

Osborne BA (1998) Apoptotic signaling pathways in lymphocytes. In: Lockshin RA, Zakeri Z and Tilley JL, (eds). When Cells Die: A Comprehensive Evaluation of Apoptosis and Programmed Cell Death, pp. 245–266. New York: Wiley‐Liss.

Pampaloni F, Ansari N and Stelzer EHK (2013) High‐resolution deep imaging of live cellular spheroids with light‐sheet‐based fluorescence microscopy. Cell and Tissue Research 352: 161–177.

Penaloza C, Orlanski S, Ye Y et al. (2008) Cell death in mammalian development. Curr Pharm Des. 14: 184–196. Review.

Pérez‐Garijo A, Martín FA and Morata G (2004) Caspase inhibition during apoptosis causes abnormal signalling and developmental aberrations in Drosophila. Development 131: 5591–5598.

Sanders EJ and Wride MA (1995) Programmed cell death in development. International Review of Cytology 163: 105–173.

Sato M and Sato K (2013) Dynamic regulation of autophagy and endocytosis for cell remodeling during early development. Traffic 14: 479–486.

Sengelaub DR and Forger NG (2008) The spinal nucleus of the bulbocavernosus: firsts in androgen‐dependent neural sex differences. Hormones and Behavior 53: 596–612.

Tsukamoto S, Hara T, Yamamoto A, et al. (2014) Fluorescence‐based visualization of autophagic activity predicts mouse embryo viability. Scientific Reports 4: 4533.

Vega Thurber R and Epel D (2007) Apoptosis in early development of the sea urchin, Strongylocentrotus purpuratus. Developmental Biology 303: 336–346.

Wang J and Lenardo MJ (2000) Role of caspases in apoptosis, development, and cytokine maturation revealed by homozygous gene deficiencies. Journal of Cell Science 113: 753–757.

Wang MW, Wang F, Zheng YJ, et al. (2013) An in vivo molecular imaging probe (18)F‐Annexin B1 for apoptosis detection by PET/CT: preparation and preliminary evaluation. Apoptosis 18: 238–247.

Zaidi AU, D'sa‐Eipper C, Brenner J, et al. (2001) Bcl‐X(L)‐caspase‐9 interactions in the developing nervous system: evidence for multiple death pathways. Journal of Neuroscience 21: 169–175.

Zakeri ZF and Ahuja HS (1994) Apoptotic cell death in the limb and its relationship to pattern formation. Biochemistry and Cell Biology 72: 603–613.

Zakeri Z and Lockshin RA (2002) Cell death during development. Journal of Immunological Methods 265: 3–20.

Zakeri Z, Lockshin RA, Criado‐Rodriguez LM and Martinez AC (2005) A generalized caspase inhibitor disrupts early mammalian development. International Journal of Developmental Biology 49: 43–47.

Zakeri Z and Lockshin RA (2008) Cell death: history and future. Advances in Experimental Medicine and Biology 615: 1–11.

Zakeri Z and Lockshin RA (2009) Essentials of apoptosis: a guide for basic and clinical research. In: Yin X‐M and Dong Z, (eds). Cell Death: Defining and Misshaping Mammalian Embryos, pp. 409–422. New York: Humana Press.

Zheng TS, Hunot S, Kuida K and Flavell RA (1999) Caspase knockouts: matters of life and death. Cell Death and Differentiation 6: 1043–1053.

Further Reading

Abrams JM, White K, Fessler LI and Steller H (1993) Programmed cell death during Drosophila embryogenesis. Development 117: 29–43.

Domingos PM and Steller H (2007) Pathways regulating apoptosis during patterning and development. Current Opinion in Genetics and Development 7: 294–299.

Edinger AL and Thompson CB (2004) Death by design: apoptosis, necrosis and autophagy. Current Opinion in Cell Biology 16: 663–669.

Hardy K (1997) Cell death in the mammalian blastocyst. Molecular Human Reproduction 3: 919–925.

Hardy K (1999) Apoptosis in the human embryo. Reviews of Reproduction 4: 125–134.

Jurisicova A, Varmuza S and Casper RF (1995) Involvement of programmed cell death in preimplantation embryo demise. Human Reproduction Update 1: 558–566.

Khosravi‐Far R, Zakeri Z, Lockshin RA and Piacentini M (2008) Introduction. In: Khosravi‐Far R, Zakeri Z, Lockshin RA and Piacentini M, (eds). Methods in Enzymology. Vol. 446, pp. xxi–xxii. Maryland Heights, MO: Elsevier.

Lawen A (2003) Apoptosis – an introduction. BioEssays 25: 888–896.

Lockshin R and Zakeri Z (eds) (2003) When Cells Die II. A Comprehensive Evaluation of Apoptosis and Programmed Cell Death. New York: Wiley‐Liss.

Rojo C and González E (2008) Apoptosis in zebrafish embryos: removing cells from inappropriate locations. Zebrafish 5 (1): 25–37.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Zakeri, Zahra, Loos, Ben, and Lockshin, Richard A(May 2015) Cell Death during Developmental Processes. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0022094.pub2]