Cell Death and Disease / Department of Pathological Physiology1
Cell Death and Disease
Jan Balvan Ph.D.
Apoptosis / Department of Pathological Physiology2
Regulated Cell Death in Disease
Programmed cell death pathways can become pathological and promote chronic inflammation and many
other diseases.
Damage-associated molecular patterns, which are associated with various diseases, are host-derived
molecules that might be released from dying cells and then function as danger signals, enhancing local
inflammation.
Lytic forms of cell death, such as necroptosis, pyroptosis and NETosis, are highly pro-inflammatory
owing to the release of cell contents, which can contribute to inflammation.
Even apoptotic cell death, which is typically non-inflammatory, can cause pathology when upregulated or
when cell debris is improperly cleared, resulting in the release of inflammatory mediators.
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Apoptosis / Department of Pathological Physiology3
Apoptosis
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Ligation of the TNF receptor 1 (TNFR1) receptor
can result in either pro-survival signals or proapoptotic
signals.
These events lead to phosphorylation of RIPK1
(via the IKKs and TAK1), preventing the formation
of RIPK-dependent death complexes, and activation
of canonical NF-κB signalling, which promotes the
transcription of pro-inflammatory cytokine genes as
well as genes encoding anti-apoptotic factors such
as cellular FLICE-like inhibitory protein (cFLIP).
Typical apoptotic cell death during development or
homeostasis is non-inflammatory, and the cell debris
is rapidly cleared. Intrinsic apoptosis pathways that
lead to the activation of caspases instigate
mechanisms that actively suppress potentially
inflammatory responses. For example, the apoptotic
caspase cascade functions to prevent type I
interferon production during BAX/BAK-dependent
apoptosis; furthermore, during apoptosis, genomic
DNA is cleaved into small fragments, and HMGB1
remains bound to chromatin, limiting its proinflammatory
potential.
Apoptosis / Department of Pathological Physiology4
Apoptosis in Chronic Inflammation
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Apoptosis of neutrophils is another critical step in the resolution of acute
inflammation. Defects in apoptosis and a failure to clear inflammatory neutrophils can
prolong inflammatory responses (e.g. Rheumatoid arthritis).
Apoptosis is key to developing immunological tolerance, as negative selection in the
thymus and bone marrow removes self-reactive T cells and B cells by apoptosis.
Too much apoptosis causes inflammatory disease.
If a large number of apoptotic cells cannot be cleared rapidly by efferocytosis, cells undergoing apoptosis can
progress to secondary necrosis, with concomitant membrane permeabilization and release of proinflammatory
cell contents. Excessive cell death can also result in tissue atrophy or a loss of barrier function,
which might contribute to inflammation
The skin and gut are exposed continuously to commensals and pathogens, making them more vulnerable to
inflammatory consequences. Loss of barrier function in epithelial surfaces can result in prolonged exposure to
PAMPs and a more intense inflammatory milieu. Increased cell death, both apoptotic and necrotic, has been
associated with chronic inflammation in inflammatory bowel disease (IBD) and is particularly prominent in
paediatric patients.
Apoptotic bodies, suggestive of excessive apoptosis, have also been found in colonic biopsy samples from
patients with ulcerative colitis. Furthermore, skin diseases such as lichen planus, eczema, acute and chronic
graft versus host disease, bullous pemphigoid and chronic diabetic skin ulcers feature excessive apoptosis.
Rheumatoid arthritis
Graft vs host disease
Apoptosis / Department of Pathological Physiology5
Apoptosis in Chronic Inflammation
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Too much or too little apoptosis causes inflammatory disease.
In SLE (systemic lupus erythematosus), FAS polymorphisms that lead to defective apoptosis are thought to
contribute to the persistence of autoreactive T cells and B cells that generate autoantibodies. Interestingly,
an alternative SLE associated FAS mutation is associated with the increased apoptosis of activated
lymphocytes.
The fact that both decreased and increased apoptosis can cause essentially the same disease highlights
how context-specific dysregulated apoptosis can be.
Apoptosis and necroptosis are intimately
linked. Necroptosis is triggered by many of
the same stimuli as apoptosis and seems
to function as a backup, or ‘emergency’
type of cell death when apoptosis is
inhibited by pathogens.
Apoptosis / Department of Pathological Physiology6
Acute versus Chronic Inflammation
Acute inflammation is a healthy response to a danger
signal, such as trauma, stress or infection.
Individual cells respond to such insults by initiating
acute inflammation, but amplification loops derived
from the surrounding cells are probably required to
generate tissue and immune responses.
Once an insult is dealt with, the resolution of
inflammation is vital to restore cellular and tissue
homeostasis.
Failure to deal with an insult or resolve acute
inflammation results in amplification loops that
generate damage-associated molecular patterns
(DAMPs).
Persistent amplification loops can generate a
vicious cycle of inflammation, leading to
disease.
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Apoptosis / Department of Pathological Physiology7
Regulated Cell Death and Inflammation
Acute inflammation is a defensive response, usually initiated through the detection of an extracellular stimulus by local
immunoreactive cells, and mediated by cytokines, chemokines and other second messengers. The initiating signals can include
damage-associated molecular patterns (DAMPs) or pathogen-associated molecular patterns (PAMPs), which activate cellular
receptors on, or within, individual cells within exposed tissues (e.g. macrophages). Such stimuli include endogenously derived crystals,
such as cholesterol or urate, and exogenous particulates, such as silica crystals. In response, signalling molecules induce and regulate
varying degrees of local and systemic inflammation, chemotaxis, cell proliferation and tissue repair.
Cytokines often form amplification loops that increase the production of inflammatory mediators, including themselves, orchestrating
local tissue and systemic immune responses. Cytokines, such as TNF or other so-called ‘death ligands’ of the TNF superfamily, can
also induce cell death programmes via death receptors that in turn generate more DAMPs. Certain DAMPs, such as high mobility
group protein B1 (HMGB1) and ATP, serve important functions in tissue regeneration following acute inflammation; these DAMPs can
recruit stem cells and immune cells to the site of inflammation, promote proliferation and differentiation of tissue-resident stem cells
and stimulate angiogenesis.
Once danger or damage has been dealt with, resolution of acute inflammation is essential for restoring homeostasis. Failure to resolve
the inflammatory response can lead to chronic inflammation. The amplification loops that are necessary to ensure a sufficient acute
inflammatory response can instead promote excessive inflammation and persistent immune responses, resulting in ongoing tissue
damage and eventually, chronic inflammatory diseases.
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Chronic inflammation is an important feature of a wide range of diseases that affect barrier surfaces, including inflammatory diseases
of the gut (such as Crohn’s disease and colitis) and of the skin (such as eczema and psoriasis). Common rheumatic conditions such
as, rheumatoid arthritis (RA), psoriatic arthritis (PsA), osteoarthritis (OA), ankylosing spondylitis (AS) and chronic tophaceous gout are
also associated with varying degrees of chronic inflammation. However, compared with inflammatory diseases of the gut and skin, the
aetiology of other chronic inflammatory diseases associated with sterile inflammation is less clear.
Apoptosis / Department of Pathological Physiology8
Cell death is both a cause and a consequence of inflammation.
A cell exposed to an inflammatory stimulus has several potential
fates:
survival, production of inflammatory cytokines and possibly
proliferation;
non-inflammatory or even anti-inflammatory apoptotic cell
death; or
pro-inflammatory programmed cell death such as necroptosis
and pyroptosis.
Phagocytes clear non-inflammatory apoptotic cells by
efferocytosis, preventing progression to late apoptosis and postapoptotic
necrosis.
Functional apoptotic machinery can inhibit necroptosis.
Post-apoptotic necrosis, necroptosis and pyroptosis lead to the
uncontrolled release of damage-associated molecular patterns
(DAMPs) and inflammatory cytokines into the extracellular
environment. DAMPs and cytokines can then function as proinflammatory
stimuli, perpetuating a cycle of inflammation and
cell death.
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Cell Death and Inflammation
Apoptosis / Department of Pathological Physiology9
Cell Death and Inflammation
Lytic forms of programmed cell death, including necroptosis,
pyroptosis and NETosis, are thought to promote inflammation
by releasing damage-associated molecular patterns(DAMPs),
and other inflammatory mediators, into the extracellular
environment.
Apoptosis counters the effects of DAMPs by ensuring the rapid
removal of a dying cell and its contents before the contents are
released.
Billions of cells undergo apoptotic cell death daily without
provoking an inflammatory response. It might be that apoptosis
and lytic forms of cell death have evolved to provide opposing
effects on inflammation.
The availability of inhibitory drugs that target the pro-inflammatory cytokines involved in
inflammation amplification loops has transformed the treatment of many inflammatory
diseases. The success of TNF antagonists such as etanercept, infliximab and adalimumab
has confirmed the importance of TNF in the pathology of many chronic inflammatory
diseases, including RA, PsA, AS, psoriasis and inflammatory bowel disease. TNF can amplify
the initial inflammatory response by transcriptionally upregulating the production of
additional cytokines and chemokines. Thus, the presumption has been that TNF inhibitors
function by dampening these amplification loops.
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation:
breaking the cycle to treat rheumatic disease. Nat Rev
Rheumatol 16, 496–513 (2020).
Apoptosis / Department of Pathological Physiology10
Necroptosis
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Necroptosis is mediated by the necrosome, a complex consisting
of receptor interacting serine/threonine-protein kinase 1
(RIPK1), RIPK3 and mixed lineage kinase domain-like protein
(MLKL).
During extrinsic apoptotic signalling, following ligation of TNF
receptor 1 (TNFR1) or pattern recognition receptors (PRRs),
activated caspase 8 cleaves RIPK1 and possibly RIPK3,
preventing the formation of the necrosome.
Complete inhibition of caspase 8 (e.g. HNSCC), for example by
the long isoform of cellular FLICE-like inhibitory protein, enables
the formation of the necrosome and subsequent phosphorylation
and activation of MLKL.
Activated MLKL oligomerizes and translocates to the cell
membrane, generating pores that result in the release of cell
contents, cellular swelling and rupture, and necroptotic death.
Apoptosis / Department of Pathological Physiology11
Necroptosis and disease
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
homozygous loss-of-function mutations in RIPK1 have been described in patients with a range of inflammatory disease
phenotypes, including arthritis, lymphopenia and early-onset IBD. Freshly derived monocytes from these patients were
more sensitive to TNF-induced necroptosis, but, in contrast to RIPK1-deficient mice, intestinal inflammation was not
associated with increased apoptosis in those patients with early-onset.
These data suggest that genetic deficiency of RIPK1 can sensitize cells to necroptosis; however, given the pleiotropic
role that RIPK1 has in the immune system, whether cell death has a primary role in other human inflammatory diseases
is not yet clear.
Activation of the RIPK3–MLKL axis has been particularly observed in neutrophilic diseases, such as cutaneous
vasculitis, ulcerative colitis and psoriasis; although the extent to which necroptotic cell death is a cause, consequence or
an exacerbating factor in these diseases is unknown.
An emerging theme suggests that key mechanisms of neurodegeneration involve the interaction of cell-autonomous
mechanisms mediated by RIPK1 in CNS glial cells to promote the degeneration of axons and neurons in cell-nonautonomous
manners. This deleterious environment is established by activated microglia and astrocytes, as well as
infiltrating immune cells such as T cells and macrophages, through their ability to release disease-associated
inflammatory mediators and promote the vicious cycles of neuroinflammation and neurodegeneration. In this
environment, RIPK1 not only may act as a key effector of inflammatory cell death but also may mediate deleterious
inflammatory signalling to provide a feedforward mechanism to accelerate neurodegeneration.
Yuan, J., Amin, P. & Ofengeim, D. Necroptosis and RIPK1-mediated neuroinflammation in CNS diseases. Nat Rev Neurosci 20, 19–33 (2019). https://doi.org/10.1038/s41583-018-0093-1
Apoptosis / Department of Pathological Physiology12
Necroptosis and disease
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Yuan, J., Amin, P. & Ofengeim, D. Necroptosis and RIPK1-mediated neuroinflammation in CNS diseases. Nat Rev Neurosci 20, 19–33 (2019). https://doi.org/10.1038/s41583-018-0093-1
The necroptotic signalling pathway has been implicated in several pathologies based on disease models, which include retinal
degeneration, brain impact trauma, cerulein-induced pancreatitis, ethanol-induced liver injury, septic shock, multiple sclerosis,
Huntington disease and tumour progression.
Virus-induced necroptosis can be beneficial to host cells by limiting the lifespan of the cells in which the virus replicates or by
inducing necroptosis-mediated inflammatory responses.
Virus-mediated regulation of necroptosis can be detrimental to the host under some circumstances: for example, when viruses
induce necroptosis in immune cells that are required for infection control, such as CD4+ T cells during HIV infection. HIV-infected T
cells have increased necroptosis rates, which were correlated with a higher sensitivity to TNF-induced cell death and decreased
membrane-associated caspase 8 activity.
Bacterial infection.
Enteropathogenic Escherichia coli (EPEC) express the pathogenicity effector protein NleB1, which blocks apoptosis and necroptosis.
NleB1-deficient EPEC was unable to effectively colonize the host, which indicated that bacterium-induced cell death protected the
host organism. In agreement with bacterium-induced cell death being beneficial, RIPK3 deficiency, especially when combined with
FADD or caspase 8 deficiency, results in a much higher susceptibility to Yersinia infection.
By contrast, Staphylococcus aureus toxins induce RIPK3-mediated necroptosis in lung epithelium, which is detrimental to the host.
Other bacteria such as Salmonella enterica subsp. enterica serovar Typhimurium, Clostridium perfringens and Mycobacterium
tuberculosis have been shown to induce necroptosis in vitro (potentially, any organism that expresses TLR3 and TLR4 ligands can
induce necroptosis), but the actual role of necroptosis during in vivo infection by these bacteria is yet to be defined.
Apoptosis / Department of Pathological Physiology13
Pyroptosis
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Pyroptosis is inflammatory form of cell death, characterized by formation of pores in the cell membrane, the liberation of proinflammatory
cytokines such as IL-1β and IL-18, DNA fragmentation and chromatin condensation. Cellular swelling is followed by
loss of membrane integrity and the release of cellular contents (including any intracellular pathogens).
The physiological function of
pyroptosis is thought to be
prevention of intracellular
pathogen replication, control of
such pathogens through IL-1βmediated
and IL-18-mediated
inflammatory signalling and the
exposure of those pathogens to
extracellular killing mechanisms.
However, pyroptosis can also be
activated in response to DAMPs,
and sterile, DAMP-triggered
inflammation is a potential
contributor to inflammatory
diseases.
Apoptosis / Department of Pathological Physiology14
Pyroptosis and Inflammatory Disease
Anderton, H., Wicks, I.P. & Silke, J. Cell death in chronic inflammation: breaking the cycle to treat rheumatic disease. Nat Rev Rheumatol 16, 496–513 (2020).
Although pyroptosis is one physiological response to infection, as are apoptosis and necroptosis, inappropriate overactivation of
this pathway can lead to inflammatory disease. Furthermore, DAMPs can trigger formation of the inflammasome and
pyroptotic cell death in the absence of infection.
These characteristics are demonstrated in periodic fever syndromes, such as familial cold autoinflammatory syndrome, Muckle–
Wells syndrome and chronic infantile neurological cutaneous and articular syndrome, also known as neonatal-onset
multisystem inflammatory disease (NOMID).
These inherited autoinflammatory diseases present with diverse symptoms, including periodic fever, urticaria, joint pain and
swelling, headaches and conjunctivitis. They are collectively referred to as the cryopyrin-associated periodic syndromes (CAPS)
and are associated with gain-of-function mutations in NLRP3.
For some inflammatory diseases, inflammasome activation has been implicated, but pyroptosis has not been directly
demonstrated (Gout is caused by MSU crystal deposition in and around joints, leading to acute inflammation and intense
neutrophilic infiltration).
Other examples of diseases associated with inflammasome activation and the release of IL-1β and IL-18 include pyogenic
arthritis, pyoderma gangrenosum and acne (PAPA) syndrome and hidradenitis suppurativa.
GoutMuckle–Wells syndromePyogenic arthritisNOMIDHidradenitis suppurativa
Jemec, G.B.E., Hidradenitis Suppurativa. New England Journal of Medicine, 2012. 366(2): p. 158-164.
Apoptosis / Department of Pathological Physiology15
Pyroptosis and Disease
Cancer
Pyroptosis can not only inhibit tumor cell proliferation, but also form a microenvironment suitable for tumor cell
growth and promote tumor growth.
Chronic tumor necrosis caused by pyroptotic cell death of a small number of tumor cells in the central hypoxic
area of the tumor inhibits antitumor immunity and accelerates tumor development.
The acute inflammation induced by pyroptosis in the tumor microenvironment enhances the immune response
and suppresses the progression of the tumor.
Pyroptosis is also involved in the negative effects of radiotherapy. The AIM2 inflammasome played an
unexpected role in responding to radiation-induced DNA damage and induced caspase-1-mediated pyroptosis
in intestinal epithelial cells and bone marrow cells, which is one of the causes of radiation-induced
gastrointestinal and hematological toxicity.
Cardiovascular disease
NLRP3 and caspase-1 are the most widely involved in cardiovascular diseases. By affecting angiogenesis,
myocardial hypertrophy, destabilizing plaques on the arterial wall, myocardial fibrosis, endothelial injury and
other pathological processes, they play an important role in myocardial infarction/reperfusion, arrhythmia,
Kawasaki disease and other cardiac diseases.
Apoptosis / Department of Pathological Physiology16
Gasdermin-related therapy
Gasdermins are the effectors of
pyroptosis, and activation of
gasdermins might prevent tumor
formation. The researchers
developed a bioorthogonal chemical
system mediated by the probe
phenylalanine trifluoroborate (PheBF3,
a cancer-imaging probe that is
significantly and specifically
absorbed by tumors) at the cellular
and in vivo levels. The study
revealed that the pyroptosis of a
small percentage of tumor cells can
effectively regulate the tumor
immune microenvironment and
activate a strong T cell-mediated
antitumor immune response.
Apoptosis / Department of Pathological Physiology17
Regulated Cell Death in Heart Disease
Regulated and unregulated forms of cell death have been implicated in the pathogenesis of
multiple forms of heart disease: ranging from MI without reperfusion, MI with reperfusion
(I/R), heart failure of diverse etiologies, myocarditis, congenital heart disease, and others.
MI results from acute and prolonged deficits in the supply of oxygen, nutrients, and survival
factors to the myocardium relative to the demands of this tissue.
During acute MI, cardiomyocyte death is the central event in pathogenesis. Acute loss of these cells in the infarct zone drives subsequent adverse remodeling
in the remaining myocardium leading to heart failure.
much of this acute cell death occurs through regulated death programs and, therefore, may be amenable to therapeutic manipulation.
Recent data prompt reconsideration of the relative contributions of apoptosis and necrosis in I/R. Early studies
suggested that apoptosis was the primary form of cell death in this syndrome. In some cases, this conclusion was
based on genetic manipulations that were thought at the time to impact only apoptotic cell death but were later
recognized to also affect necrosis. Raising further questions concerning a role for apoptosis in I/R are data suggesting
that caspase-dependent cell death is less robust in adult as compared with fetal/neonatal cardiomyocytes
Apoptosis / Department of Pathological Physiology18
Regulated Cell Death in Heart Disease
In most studies, genetic and pharmacological inhibition of cell death signaling reduces cardiomyocyte death and
infarct size only in the context of reperfused MI (I/R).
Studies predating the regulated cell death era have traditionally considered unregulated necrosis to be the primary
mode of cardiomyocyte death in this syndrome based primarily on morphological criteria
Non-apoptotic cell death mediating cardiomyocyte loss during I/R.
MPT-mediated necrosis and necroptosis,
ferroptosis, although it remains unclear whether this program is functioning as an independent cell death modality or
an amplification loop for the necrotic cell death. Regardless, the molecules that mediate ferroptosis may provide new
therapeutic targets.
PARP-1-dependent processes also appear important in I/R. The significance of PARP-1 activation may be more related
to its depletion of NAD and disruption of cellular energetics.
The data regarding autosis in ischemic neuronal injury are intriguing, and encourage further study of this form of
autophagy-dependent cell death in myocardial I/R.
Pyroptosis may also contribute to I/R, but the major challenge is in understanding the cell types in which it is taking
place. Current data suggest that pyroptosis operates primarily in non-cardiomyocytes, bone marrow-derived cells
recruited to the heart during MI and perhaps cardiac fibroblasts, but further study is needed.
Dr. Martina Raudenská
Doc. Michal Masařík
Thanks for your
attention.