Cell Death 1

From CellBiology

Cell Death 1



We have discussed the cell lifespan and how cell's are born. These next two lectures will discuss how cells die in response to stress, pathological conditions and by programmed cell death. This first lecture will also introduce the concept of cellular recycling and the response to cellular stress. The next lecture Cell Death 2 will focus on programmed cell death by apoptosis.


Lecture Slides: 2017 Lecture PDF

MH - note that content listed below will not match exactly current lecture structure but has been selected as having similar content

Links: 2017 PDF | 2017 | 2016 | 2015 | 2012 | 2010 | 2009 | 2008 1 slide/page (view)

Recent papers  
2016 - Recommendations from the International Harmonization of Nomenclature and Diagnostic Criteria (INHAND) Apoptosis/Necrosis Working Group. "Historically, there has been confusion relating to the diagnostic nomenclature for individual cell death. Toxicologic pathologists have generally used the terms "single cell necrosis" and "apoptosis" interchangeably. Increased research on the mechanisms of cell death in recent years has led to the understanding that apoptosis and necrosis involve different cellular pathways and that these differences can have important implications when considering overall mechanisms of toxicity, and, for these reasons, the separate terms of apoptosis and necrosis should be used whenever differentiation is possible. However, it is also recognized that differentiation of the precise pathway of cell death may not be important, necessary, or possible in routine toxicity studies and so a more general term to indicate cell death is warranted in these situations. Morphological distinction between these two forms of cell death can sometimes be straightforward but can also be challenging. This article provides a brief discussion of the cellular mechanisms and morphological features of apoptosis and necrosis as well as guidance on when the pathologist should use these terms. It provides recommended nomenclature along with diagnostic criteria (in hematoxylin and eosin [H&E]-stained sections) for the most common forms of cell death (apoptosis and necrosis). This document is intended to serve as current guidance for the nomenclature of cell death for the International Harmonization of Nomenclature and Diagnostic Criteria Organ Working Groups and the toxicologic pathology community at large. The specific recommendations are:Use necrosis and apoptosis as separate diagnostic terms.Use modifiers to denote the distribution of necrosis (e.g., necrosis, single cell; necrosis, focal; necrosis, diffuse; etc.).Use the combined term apoptosis/single cell necrosis whenThere is no requirement or need to split the processes, orWhen the nature of cell death cannot be determined with certainty, orWhen both processes are present together. The diagnosis should be based primarily on the morphological features in H&E-stained sections. When needed, additional, special techniques to identify and characterize apoptosis can also be used." PMID 26879688

more papers

Cell Lifespan


  • Neutrophil - 6-7 hours circulating, 4 days in tissue
  • Red blood cell - 120 days
  • Brain neuron - 50 - 100 years


  • embryonic/fetal cells may have different lifespans
  • produce cells that are selectively lost during development of tissues.
    • nervous system produces many more neurons than in adult.
  • programmed cell death or withdrawal of growth factors.
  • other cells "lost" through fusion events.
    • skeletal muscle - myoblasts fuse to myotubes.

Cell Recycling

There are a number of different cellular mechanisms and processes for reusing cellular components or removing abnormal products. These pathways can also be utilized during periods when the cell is placed under specific or limited growth conditions. Autophagy is an important part of this process, but will be covered under the Cell Death section of this current lecture.


Figure 6-88. Two general ways of inducing the degradation of a specific protein Figure 6-86. The proteasome

Ubiquitin-mediated protein degradation
  • Protein complex that degrades cellular proteins by proteolysis
  • located in nucleus and cytoplasm


  • regulate protein levels and degrade misfolded proteins


  • 26S proteasome Mr 2000 kDa
    • two 19S regulatory caps -ATPase active sites and ubiquitin binding sites
    • one 20S core hollow structure - catalytic core




Calpain superfamily members (image from Calpain for Modulatory Proteolysis Database)


  • proteolytically processes, rather than digests, substrates to transform and modulate their structures and activities
  • proteolysis in embryogenesis, regulation of key enzymes, structural proteins and in proinflammatory responses
  • mediates MMP2 expression and cell migration (fibroblasts and leukemic cells)
  • contribute to cell death in neurons by cleaving essential cytoskeletal proteins
  • tissue damage in response to pathological events (myocardial infarcts, stroke, atherosclerosis, and brain trauma)
  • Deregulated calpain activity following loss of Ca2+ homeostasis
  • intracellular free calcium concentration (Ca2+i) regulator in some late apoptotic signaling

MH - calcium will be covered again in signaling.


  • Calpain (EC, Clan CA, family C02) is an intracellular Ca2+-dependent cysteine protease
  • ubiquitously distributed
  • shows limited proteolytic activity at neutral pH.
  • 13 genes in mammals
  • heterodimer large 80 kDa catalytic and a small 30 kDa regulatory subunit

Links: CaMPDB Calpain for Modulatory Proteolysis Database | PMID: 17608959 | Physiol. Rev - The Calpain System | Schematic diagram of a model for turnover/degradation of proteins assembled in myofibrillar structures in striated muscle

Cell Stress

The term "cell stress" can cover many different issues. It is used here to describe two specific circumstances: when normal cellular processes may function abnormally (protein misfolding), or under specific limiting growth conditions (starvation). Under these conditions the cellular response is graded, initially to correct the problem (Unfolded Protein Response) or reuse existing resources (Autophagy).

Endoplasmic Reticulum Stress

  • Endoplasmic Reticulum functions
    • protein synthesis
    • lipid metabolism
    • calcium regulation (Ca2+) storage, release, signaling

Abnormal protein folding can lead to the Unfolded Protein Response (UPR)

  • accumulation of misfolded proteins
  • aggregate in the ER lumen
  • causes ER stress

Unfolded Protein Response (UPR)

Unfolded protein response to bacterial pore-forming toxins

MBoC - Figure 12-56. The unfolded protein response in yeast

  • decrease in the arrival of new proteins into the ER
    • preventing additional protein misfolding and overloading of the organelle
  • increase in the amount of ER chaperones
    • chaperone BiP disassociates from and subsequently activate three signal transducers (Ire1, PERK and Atf6)
    • increasing the folding capacity of the ER to deal with misfolded proteins
  • increase in the extrusion of irreversibly misfolded proteins from the ER
    • subsequently degradation of these proteins in the proteasome
Unfolded protein response cartoon.jpg
  • Ire1 - Inositol Requiring 1 (Ire1α and Ire1β) is a transmembrane kinase/ribonuclease
  • PERK - PRKR-like Endoplasmic Reticulum Kinase phosphorylates the α subunit of eIF2, causes global translation attenuation
  • Atf6 - Activating Transcription Factor 6-like proteins (Atf6α, Atf6β, CREB3L2) transcriptionally initiate a gene expression program that includes cell cycle arrest

If all the above UPR steps fail, cell death by apoptosis is triggered.

Classification of Cell Death

Essential versus accessory aspects of cell death: recommendations of the Nomenclature Committee on Cell Death (NCCD) 2015 (note this is a new update of the 2009 recommendations below) PMID 25236395

  • 'Accidental cell death' (ACD) cells exposed to extreme physicochemical or mechanical stimuli die in an uncontrollable manner, as a result of their immediate structural breakdown. Such an unavoidable variant of cellular demise.
  • 'Regulated cell death' (RCD) can occur as part of physiologic programs or can be activated once adaptive responses to perturbations of the extracellular or intracellular microenvironment fail. Initiated by a genetically encoded apparatus, correlating with the fact that its course can be altered by pharmacologic or genetic interventions.
  • ‘Programmed cell death' (PCD) is used to indicate RCD instances that occur as part of a developmental program or to preserve physiologic adult tissue homeostasis.

The biochemical phenomena that accompany RCD may be harnessed to classify it into a few subtypes, which often (but not always) exhibit stereotyped morphologic features. Nonetheless, efficiently inhibiting the processes that are commonly thought to cause RCD, such as the activation of executioner caspases in the course of apoptosis, does not exert true cytoprotective effects in the mammalian system, but simply alters the kinetics of cellular demise as it shifts its morphologic and biochemical correlates. Conversely, bona fide cytoprotection can be achieved by inhibiting the transduction of lethal signals in the early phases of the process, when adaptive responses are still operational. Thus, the mechanisms that truly execute RCD may be less understood, less inhibitable and perhaps more homogeneous than previously thought. Here, the Nomenclature Committee on Cell Death formulates a set of recommendations to help scientists and researchers to discriminate between essential and accessory aspects of cell death.

Classification of cell death: recommendations of the Nomenclature Committee on Cell Death 2009


Classified by cell death mode and morphological features (modified from Table 2)


  • Elimination of cytosolic organelles
  • Modifications of plasma membrane
  • Accumulation of lipids in keratohyalin granules in stratum granulosum
  • Extrusion of lipids in the extracellular space
  • Desquamation (loss of corneocytes) by protease activation

Cornified envelope - formation or ‘keratinization’ is specific of the skin to create a barrier function. Although apoptosis can be induced by injury in the basal epidermal layer (e.g., UV irradiation), cornification is exclusive of the upper layers (granular layer and stratum corneum).


  • Lack of chromatin condensation
  • Massive vacuolization of the cytoplasm
  • Accumulation of (double-membraned) autophagic vacuoles
  • Little or no uptake by phagocytic cells, in vivo

Autophagic cell death - defines cell death occurring with autophagy, though it may misleadingly suggest a form of death occurring by autophagy as this process often promotes cell survival.


  • Cytoplasmic swelling (oncosis)
  • Rupture of plasma membrane
  • Swelling of cytoplasmic organelles
  • Moderate chromatin condensation

Necrosis - identifies, in a negative fashion, cell death lacking the features of apoptosis or autophagy. Note that necrosis can occur in a regulated fashion, involving a precise sequence of signals.


  • Rounding-up of the cell
  • Retraction of pseudopodes
  • Reduction of cellular and nuclear volume (pyknosis)
  • Nuclear fragmentation (karyorrhexis)
  • Minor modification of cytoplasmic organelles
  • Plasma membrane blebbing
  • Engulfment by resident phagocytes, in vivo

Apoptosis - is the original term introduced by Kerr et al. to define a type of cell death with specific morphological features. Apoptosis is NOT a synonym of programmed cell death or caspase activation.


Epidermis cartoonCornified Epithelium

Caspase-14 and epithelial cornification

Occurs in the skin epithelium and occurs in the upper layers (granular layer and stratum corneum)

  • Elimination of cytosolic organelles
  • Modifications of plasma membrane
  • Accumulation of lipids in keratohyalin granules in stratum granulosum
    • F-granules (histidine-rich) are large, irregularly shaped granules
    • L-granules (sulphur-rich)
  • Extrusion of lipids in the extracellular space
  • Desquamation (loss of corneocytes) by protease activation
  • Apoptosis can also be induced by injury to the basal epidermal layer
    • UVB irradiation, chemicals, cytotoxic cytokines

Caspase 14

  • enzyme, cysteinyl aspartate–specific proteinase
  • recently identified as having a role in this process
  • activation correlates with cornification
  • not clear yet exact role in filaggrin processing, there are 2 theories
    • cleave filaggrin fragments - leading to further degradation into free amino acids by another endo- and/or exopeptidase
    • directly or indirectly activate an endo- and/or exopeptidase - further processes smaller filaggrin fragments


Autophagy types
Membrane events involved in autophagy
Possible membrane sources of isolation membrane

The Cell - Lysosomes in phagocytosis and autophagy

Autophagosome formation Movies PMID: 18725538

See also Lecture - Endocytosis

Autophagy Processes

There are several different classifications of autophagy this is probably the simplest.

  1. macroautophagy - (also called autophagy) non-selective sequestration of cytoplasmic substrates
  2. microautophagy - lysosomal invagination and ingestion of minor portions of the cytosol
  3. chaperone-mediated autophagy - involves recognition of specific substrates by the heat shock cognate 70 (Hsc70) machinery
  • to remove abnormal cytoplasmic organelles and components, it is also a stress response
    • cellular self-catabolic process "eating oneself”
  • initial sequestered in a structure called a phagophore
  • which then closes into a double membrane vesicle the autophagosome
    • some autophagosomes formed in a PI3P-enriched (phosphatidylinositol 3-phosphate) membrane compartment dynamically connected to the endoplasmic reticulum
  • an autophagosome fuses with a lysosome
  • Regulated process of the degradation and recycling of organelles and cellular components
  • Resulting in organelle turnover and in the bioenergetics of starvation
  • Could result in cell death
    • through excessive self-digestion and degradation of essential cellular constituents

Is autophagy a cell survival or a cell death pathway?

Suggested that autophagic cell death is "cell death with autophagy rather than cell death by autophagy" see Figure 1

Autophagy Summaries

Some Recent Findings

This sub-section not examinable. - This material below are new research findings on Autophagy and the origins of the membrane for autophagosome biogenesis.

Mitochondria Supply Membranes for Autophagosome Biogenesis during Starvation

  1. Autophagosomes are derived from mitochondrial outer membrane during starvation
  2. Lipids, but not most proteins, are transferred from mitochondria to autophagosomes
  3. Mitochondria-ER connections are required to form autophagosomes during starvation
  4. Mitochondrial contribution to autophagosome assembly is unique to starvation

Movies: JCB - Apg5 helps form autophagosomes Autophagosomes form as cup-shaped organelles that engulf large parts of the cytoplasm. As shown by Mizushima et al., Apg5, part of a ubiquitin-like conjugation system, localizes to the forming autophagosomes, and is essential for their formation.

Autophagy model in satellite cells.jpg

Model of autophagy role in muscle stem cells (satellite cells) during ageing.


Neuron necrosis apoptosis.jpg

Necrosis ethidium homodimer marker
  • Greek, nekros = corpse
  • pathological cell death from extrinsic injury
    • tissue damage
  • autoimmune insulin-dependent diabetes?
  • irreversible


  • tumor necrosis factor, double-stranded RNA, viral infection or bacterial toxins
  • does not shut down of protein synthesis
    • occurs in apoptosis due to caspase-dependent breakdown of eukaryotic translation initiation factor (eIF) 4G, activation of the double-stranded RNA-activated protein kinase PKR, and phosphorylation of its substrate eIF2-

Early stages

  • cell and organelles (mitochondria) swell (oncosis)
    • (Greek, onkos = 'swelling') previously described as a separate form of cell death
  • due to disruption of plasma membrane
  • cell contents leak out leading to inflammation and necrosis

Late stage

  • loss of cell membrane integrity
  • finally cell disintegration
  • cell lysis can also trigger an inflammatory response
    • leading to further inflammation and damage
    • triggering a cycle of death

Bacteria - S. aureus

  • pore-forming secreted toxins can induce necrosis

Prothymosin-alpha 1 (ProT)

  • inhibits necrosis
  • switches cells from necrosis to apoptosis

Programmed Cell Death?

Until very recently this has been exclusively about apoptosis, covered in Cell Death 2.

There is a theory though that there is no such thing as "unregulated" cell death and that even necrosis may involve a regulated program. In addition, a novel cell death pathway has been identified in neutrophils in the fight against pathogens.

Neutrophil Extracellular Traps


Neutrophil extracellular traps

Neutrophil extracellular trap EM01.jpg

Bacteria caught in Neutrophil Extracellular Traps (NETs) PMID 22945932

Neutrophils in circulation are targeted by cytokines to migrate into infected tissues, where they activate, and engulf pathogens into a phagosome.

A second defense mechanism has recently been described Neutrophil Extracellular Traps (NETs)

  • composed of chromatin decorated with cytoplasmic proteins in the extracellular space
    • bind Gram-positive and -negative bacteria, as well as fungi
Links: Image of Process | Image - C. albicans induced NETs in pulmonary infection | Neutrophil Extracellular Traps Contain Calprotectin


See Time-lapse recording of neutrophils undergoing NETosis

Neutrophils were stimulated with PMA, and a z stack was generated for 2 h and 25 min on a confocal microscope. Directly labeled antibody fragments against NE (green) and chromatin (red) in the supernatant depict formation of NETs in the final phase.

Neutrophil extracellular trap cartoon.jpg

  • requires generation of reactive oxygen species (ROS) by NADPH oxidase
  • lobulated nuclear morphology lost
  • euchromatin and heterochromatin distinction lost
  • all the internal membranes disappear
    • allowing NET components to mix
  • NETs emerge from the cell as the cytoplasmic membrane is ruptured
    • process distinct from necrosis or apoptosis

Chronic Granulomatous Disease (CGD)

  • mutations in NADPH oxidase
  • cannot activate this cell-death pathway or make NETs
  • these individuals are susceptible to infections

Links: Medline Plus - CGD | OMIM 233700


  • Necrosis traditionally considered an accidental cell death that is not subject to cellular regulation.
  • at least a part of necrotic cell death may be executed through a regulated mechanism (involves RIP1, a death domain-containing kinase)
  • activated upon stimulation by tumor necrosis factor α (TNFα), FasL, and TRAIL (same ligands that can activate apoptosis in the death receptor pathway).

Neural Cell Death Decisions

Neural programmed cell death.jpg

Three mechanisms of Death[1]

Apoptosis Autophagic cell death (ACD) Necrosis
Pro-apoptotic signals induce Bax translocation to the outer membrane of mitochondria. Bax-mediated pore formation leads to the release of apoptogenic cytochrome c (Cyt c) into the cytosol and apoptosome formation. The apoptosome activates procaspase-9 and catalytically active caspase-9 induces activation of downstream effector caspases, caspase-3 and -7 (Caspase 3/7). Phosphatidylserine (PS) exposure at the cell surface is required for the clearance of apoptotic cells. In the adult neural stem cells (NSCs), p53, Bim and PUMA have been implicated in activating apoptosis. In addition, the pro-apoptotic proteins, Bax and Bak are the key regulators. Mcl-1 antagonizes the pro-apoptotic proteins, and therefore, considered as a critical anti-apoptotic protein for the survival of the NSCs. In extrinsic apoptosis, a death ligand (TNFα, Fas, or TRAIL) binding activates death receptor and induces DISC complex formation near the receptor. Upon DISC complex-mediated activation of caspase-8, intrinsic and extrinsic apoptosis converge at the level of the executioner caspase cascade. In adult NSCs, PED/PEA-15 represses the activation of caspase-8. Release of Ca2+ from the ER and subsequent transfer to the mitochondria promtes the commitment of NSCs to cell death. Autophagy is induced when cells are starved of nutrients or survival factors. Atg7 regulates the maturation of autophagosome and initiates the lipidation of LC3 (also called LC3 II). Cargos subjected to degradation are degraded in the autophagolysosome. AMBRA1 and Beclin-1-induced autophagy is inversely correlated with apoptosis in adult NSCs. Under insulin-deprived condition, the adult hippocampal neural stem (HCN) cells succumb to ACD wherein the cell fate is under the control of GSK-3β activation. Inhibition of GSK-3β phosphorylation (p-GSK-3β) induces ACD. The negative regulator of ACD is calpain, which also mediates the crosstalk between apoptosis and ACD. Extracellular ATP or death receptor activation rapidly induces RIP1/RIP3 necrosome formation. Necrotic cell death results from the depletion of cytoplasmic ATP due to mitochondrial dysfunction. A purinergic P2X7 receptor-mediated necrosis induction has been reported in adult NSCs. However, regulator of necrosis in adult NSCs has not been identified to date

Atypical Cell Death

This section is not as relevant as "typical" cell death sections and is based upon the 2009 recommendations. A variety of historic and specialised terminologies that have arisen within the scientific literature.

  1. Mitotic catastrophe - death mode occurring either during or shortly after a dysregulated/failed mitosis. (can lead either to an apoptotic morphology or to necrosis)
  2. Anoikis - induced by the loss of the attachment to the substrate or to other cells. (historical term)
  3. Excitotoxicity - death occurring in neurons challenged with excitatory amino acids, such as glutamate, that leads to the opening of the N-methyl-d-aspartate Ca2+-permeable channel, followed by cytosolic Ca2+ overload and activation of lethal signalling pathways. (overlaps with other types of death such as apoptosis and necrosis)
  4. Wallerian degeneration - take place in the nervous system in which part of a neuron or axon degenerates without affecting the main cell body. (neurons affected by Wallerian degeneration remain alive)
  5. Paraptosis - originally introduced to describe a form of programmed cell death morphologically and biochemically distinct from apoptosis. (still unclear whether paraptosis represents a route of cell death that is truly distinct from all others)
  6. Pyroptosis - macrophages infected with Salmonella typhimurium. (apical activation of caspase-1, but not of caspase-3)
  7. Pyronecrosis - necrotic cell death of macrophages infected by S. flexneri. (whether pyroptosis and pyronecrosis play any role outside of the innate immune system)
  8. Entosis - form of ‘cellular cannibalism’ in lymphoblasts from patients with Huntington's disease. (default pathway that is unmasked exclusively when other catabolic reactions are suppressed)

Paper Links: PMID 18846107 | PMC2744427 | Cell Death Differentiation - 2009 (2005 was previous recommendations)

Alternative cell death mechanisms in development and beyond PMID 21123646



Molecular Biology of the Cell

Alberts, Bruce; Johnson, Alexander; Lewis, Julian; Raff, Martin; Roberts, Keith; Walter, Peter New York and London: Garland Science; c2002

Molecular Cell Biology

Lodish, Harvey; Berk, Arnold; Zipursky, S. Lawrence; Matsudaira, Paul; Baltimore, David; Darnell, James E. New York: W. H. Freeman & Co.; c1999

The Cell- A Molecular Approach

Cooper, Geoffrey M. Sunderland (MA): Sinauer Associates, Inc.; c2000

Madame Curie Bioscience Database

Chapters taken from the Madame Curie Bioscience Database (formerly, Eurekah Bioscience Database) Eurekah.com and Landes Bioscience and Springer Science+Business Media; c2009

Search Online Textbooks


"Apoptosis is an international peer-reviewed journal devoted to the rapid publication of innovative basic and clinically-oriented investigations into programmed cell death. It aims to stimulate research on the basis of mechanisms of apoptosis and on its role in various human disease processes including: cancer, autoimmune disease, viral infection, AIDS, cardiovascular disease, neurodegenerative disorders, osteoporosis and ageing. The editors intend to encourage the development of clinical therapies against apoptosis-related diseases."
"Autophagy, is a unique peer-reviewed journal with an international audience that covers the following topics: Macroautophagy, microautophagy, specific organelle degradation (e.g., pexophagy) and additional autophagic processes including chaperone-mediated autophagy; The molecular mechanism of autophagy including the characterization of structural proteins and structure/function relationships; Signaling and autophagic regulation; The role of autophagy in diseases including cancer, neurodegeneration and myopathies; Developmental roles of autophagy and its connection with aging; Autophagy in microbial invasion and the immune response; and Autophagy in cell death."



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2017 Course Content


Lectures: Cell Biology Introduction | Cells Eukaryotes and Prokaryotes | Cell Membranes and Compartments | Cell Nucleus | Cell Export - Exocytosis | Cell Import - Endocytosis | Cytoskeleton Introduction | Cytoskeleton - Microfilaments | Cytoskeleton - Microtubules | Cytoskeleton - Intermediate Filaments | Cell Mitochondria | Cell Junctions | Extracellular Matrix 1 | Extracellular Matrix 2 | Cell Cycle | Cell Division | Cell Death 1 | Cell Death 2 | Signal 1 | Signal 2 | Stem Cells 1 | Stem Cells 2 | Development | 2017 Revision

2017 Laboratories: Introduction to Lab | Fixation and Staining |

2017 Projects: Group 1 - Delta | Group 2 - Duct | Group 3 - Beta | Group 4 - Alpha

Dr Mark Hill 2015, UNSW Cell Biology - UNSW CRICOS Provider Code No. 00098G

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