DEGENERATION AND SENESCENCE
! always check questions and conclusions at end of ppt
APOPTOSIS .........................................................................................................................................3
Brief history of cell death ..................................................................................................................3
Biochemistry of apoptosis: Apaf-1 and role of mitochondria ...............................................................6
Biochemistry of apoptosis: Bcl-2 family members ............................................................................ 10
Biochemistry of apoptosis: caspases .............................................................................................. 15
Regulation of apoptosis .................................................................................................................. 21
Therapeutic targeting of apoptotic pathways .................................................................................... 23
THE INFLAMMATORY RESPONSE ........................................................................................................ 29
Innate immune receptor signalling .................................................................................................. 35
Cell death checkpoints in the TNF pathway ...................................................................................... 43
Methods to study inflammatory signalling ........................................................................................ 51
UBS and DUB in innate immune signalling ....................................................................................... 56
Targeting TNF in inflammation and cell death ................................................................................... 60
PYROPTOSIS ..................................................................................................................................... 64
Systemic chronic inflammation (SCI) and its role in human diseases ................................................. 64
IL1B as key cytokine connecting innate immunity and inflammatory disease ..................................... 64
pattern recognition receptors in innate immune system ................................................................... 65
Inflammasomes: signalling pathways and role in diseases ................................................................ 65
GSDMD pore formation and role in IL1B secretion ............................................................................ 67
functional assay for differential diagnosis of FMF ........................... Fout! Bladwijzer niet gedefinieerd.
FERROPTOSIS ................................................................................................................................... 69
Process of ferroptosis ..................................................................................................................... 69
Role of ferroptosis .......................................................................................................................... 72
Treatment strategies....................................................................................................................... 73
AUTOPHAGY ..................................................................................................................................... 75
Mechanisms of protein degradation ................................................................................................ 75
Macro-autophagy ........................................................................................................................... 76
Physiological relevance of macro-autophagy ................................................................................... 80
Autophagy-associated pathways ..................................................................................................... 81
Phagocytosis: clearing of apoptotic cells............................................................................................. 83
A brief history of apoptosis and efferocytosis ................................................................................... 84
Seminal work on efferocytosis in C. elegans ..................................................................................... 84
Current state of efferocytosis research ............................................................................................ 85
Clinical implications of efferocytosis ............................................................................................... 86
1
,POSSIBLE THERAPEUTIC IMPLICATIONS ............................................................................................. 89
Many ways to die ............................................................................................................................ 89
Pore forming mechanisms .............................................................................................................. 89
2
,APOPTOSIS
− Homeostasis: cell survival cell death
BRIEF HISTORY OF CELL DEATH
? why is it so important to look back in science
• Roots of actual knowledge
• Relevant and successful research questions
• Simple – basic concepts = basis: still relevant
• Fundamental science takes time
• Appropriate technology and methods
• Many different model organisms
• Continuous process
19 T H CENTURY: SPORADIC DESCRIPTION OF THE OCCURRENCE OF CELL DEATH
− Rudolf Karl Virchow: cellular pathology
• Disease = physicochemical disturbance of cells ➔ alteration to function
• All cells arise from other cells
• Functional capacities reflect in structural changes
• Regulated and impaired homeostasis of the organism = basis for (patho)fysiology
Cell death = basis for pathology
o Necrobiosis: spontaneous wearing out of living parts – shrinkage
o Necrosis: accidental process – visible
− Walter Flemming
• Morphology of cellular structures in oogenesis: chromatin margination and fragmentation
• Chromatolysis in nucleus: dissolution of Nissl bodies in the cell body of neurons
− Ilya Mechnikov: phagocytosis
• In metamorphosis: induction and removal of dead cells
+ injury: DAMP (e.g. wooden splinter)
+ infection: PAMP (e.g. bacteria)
• Juvenile cells get removed and replaced by adult cells (e.g. tadpole tail)
3
, 1930 – 1970: PROGRAMMED CELL DEATH DURING DEVELOPMENT
− A. Glücksmann
• Programmed cell death: morphology of nuclei
• Feulgen staining: acid hydrolysis of DNA
− John W. Saunders
• Programmed cell death: limb development
o Limb bud and limb development
- PNZ = posterior necrotic zone in vitro culture: programmed to die
- PNZ = posterior necrotic zone transplanted: process suppressed
- PNZ = posterior necrotic zone: macrophages remove dead cells
! cell death occurs on the right time and the proper place
o Removal of interdigital webbing
− Christian de Duve: lysosomes = suicide bags – killing cells when ruptured
1970 – 1980: DESCRIPTION OF CELL DEATH AS A CELLULAR PROCESS – APOPTOSIS
− Sydney Brenner: C. elegans
• Multicellular model organism
o Short life cycle: days-weeks years (human)
o Complete genome known: 20.000 genes 22.000 (human)
o Mutagenize: forward genetics manipulation: reverse genetics
o Transparent – no need for fluorescent markers
o Fixed number of somatic cells = eutely
- Newly hatched larva: 558 somatic
- Hermaphrodite XX: 1090 somatic - 131 die (= 105 neuronal)
! 2000 germ cells
- Male XO: 1031 somatic
! 1000 germ cells
o Conserved pathways
o High throughput
• EMS = ethyl methane sulfonate: induces specific mutations
o Forward genetics approach: generates mutations ➔ discover genes
o Reverse genetics approach: known gene mutating
• Nobel prizes
o Genetic regulation of organ development and programmed cell death
o Development of the green fluorescent protein: GFP
o microRNA and the post-transcriptional gene regulation
− Apoptosis = falling of the leaves in autumn
• Functions
o Turnover in adult tissues
o Elimination of cells during embryogenesis
o Therapeutic tumour regression
o Physiological involution and atrophy of tissues and organs
o Triggered by noxious agents
• Morphology
o Nuclear and cytoplasmic condensation
+ Breaking up of cell to membrane-bound, ultrastructurally well- preserved fragments
o Apoptic bodies are taken up by phagocytosis
+ Involved in cell turnover in many healthy tissues
4
! always check questions and conclusions at end of ppt
APOPTOSIS .........................................................................................................................................3
Brief history of cell death ..................................................................................................................3
Biochemistry of apoptosis: Apaf-1 and role of mitochondria ...............................................................6
Biochemistry of apoptosis: Bcl-2 family members ............................................................................ 10
Biochemistry of apoptosis: caspases .............................................................................................. 15
Regulation of apoptosis .................................................................................................................. 21
Therapeutic targeting of apoptotic pathways .................................................................................... 23
THE INFLAMMATORY RESPONSE ........................................................................................................ 29
Innate immune receptor signalling .................................................................................................. 35
Cell death checkpoints in the TNF pathway ...................................................................................... 43
Methods to study inflammatory signalling ........................................................................................ 51
UBS and DUB in innate immune signalling ....................................................................................... 56
Targeting TNF in inflammation and cell death ................................................................................... 60
PYROPTOSIS ..................................................................................................................................... 64
Systemic chronic inflammation (SCI) and its role in human diseases ................................................. 64
IL1B as key cytokine connecting innate immunity and inflammatory disease ..................................... 64
pattern recognition receptors in innate immune system ................................................................... 65
Inflammasomes: signalling pathways and role in diseases ................................................................ 65
GSDMD pore formation and role in IL1B secretion ............................................................................ 67
functional assay for differential diagnosis of FMF ........................... Fout! Bladwijzer niet gedefinieerd.
FERROPTOSIS ................................................................................................................................... 69
Process of ferroptosis ..................................................................................................................... 69
Role of ferroptosis .......................................................................................................................... 72
Treatment strategies....................................................................................................................... 73
AUTOPHAGY ..................................................................................................................................... 75
Mechanisms of protein degradation ................................................................................................ 75
Macro-autophagy ........................................................................................................................... 76
Physiological relevance of macro-autophagy ................................................................................... 80
Autophagy-associated pathways ..................................................................................................... 81
Phagocytosis: clearing of apoptotic cells............................................................................................. 83
A brief history of apoptosis and efferocytosis ................................................................................... 84
Seminal work on efferocytosis in C. elegans ..................................................................................... 84
Current state of efferocytosis research ............................................................................................ 85
Clinical implications of efferocytosis ............................................................................................... 86
1
,POSSIBLE THERAPEUTIC IMPLICATIONS ............................................................................................. 89
Many ways to die ............................................................................................................................ 89
Pore forming mechanisms .............................................................................................................. 89
2
,APOPTOSIS
− Homeostasis: cell survival cell death
BRIEF HISTORY OF CELL DEATH
? why is it so important to look back in science
• Roots of actual knowledge
• Relevant and successful research questions
• Simple – basic concepts = basis: still relevant
• Fundamental science takes time
• Appropriate technology and methods
• Many different model organisms
• Continuous process
19 T H CENTURY: SPORADIC DESCRIPTION OF THE OCCURRENCE OF CELL DEATH
− Rudolf Karl Virchow: cellular pathology
• Disease = physicochemical disturbance of cells ➔ alteration to function
• All cells arise from other cells
• Functional capacities reflect in structural changes
• Regulated and impaired homeostasis of the organism = basis for (patho)fysiology
Cell death = basis for pathology
o Necrobiosis: spontaneous wearing out of living parts – shrinkage
o Necrosis: accidental process – visible
− Walter Flemming
• Morphology of cellular structures in oogenesis: chromatin margination and fragmentation
• Chromatolysis in nucleus: dissolution of Nissl bodies in the cell body of neurons
− Ilya Mechnikov: phagocytosis
• In metamorphosis: induction and removal of dead cells
+ injury: DAMP (e.g. wooden splinter)
+ infection: PAMP (e.g. bacteria)
• Juvenile cells get removed and replaced by adult cells (e.g. tadpole tail)
3
, 1930 – 1970: PROGRAMMED CELL DEATH DURING DEVELOPMENT
− A. Glücksmann
• Programmed cell death: morphology of nuclei
• Feulgen staining: acid hydrolysis of DNA
− John W. Saunders
• Programmed cell death: limb development
o Limb bud and limb development
- PNZ = posterior necrotic zone in vitro culture: programmed to die
- PNZ = posterior necrotic zone transplanted: process suppressed
- PNZ = posterior necrotic zone: macrophages remove dead cells
! cell death occurs on the right time and the proper place
o Removal of interdigital webbing
− Christian de Duve: lysosomes = suicide bags – killing cells when ruptured
1970 – 1980: DESCRIPTION OF CELL DEATH AS A CELLULAR PROCESS – APOPTOSIS
− Sydney Brenner: C. elegans
• Multicellular model organism
o Short life cycle: days-weeks years (human)
o Complete genome known: 20.000 genes 22.000 (human)
o Mutagenize: forward genetics manipulation: reverse genetics
o Transparent – no need for fluorescent markers
o Fixed number of somatic cells = eutely
- Newly hatched larva: 558 somatic
- Hermaphrodite XX: 1090 somatic - 131 die (= 105 neuronal)
! 2000 germ cells
- Male XO: 1031 somatic
! 1000 germ cells
o Conserved pathways
o High throughput
• EMS = ethyl methane sulfonate: induces specific mutations
o Forward genetics approach: generates mutations ➔ discover genes
o Reverse genetics approach: known gene mutating
• Nobel prizes
o Genetic regulation of organ development and programmed cell death
o Development of the green fluorescent protein: GFP
o microRNA and the post-transcriptional gene regulation
− Apoptosis = falling of the leaves in autumn
• Functions
o Turnover in adult tissues
o Elimination of cells during embryogenesis
o Therapeutic tumour regression
o Physiological involution and atrophy of tissues and organs
o Triggered by noxious agents
• Morphology
o Nuclear and cytoplasmic condensation
+ Breaking up of cell to membrane-bound, ultrastructurally well- preserved fragments
o Apoptic bodies are taken up by phagocytosis
+ Involved in cell turnover in many healthy tissues
4
