INTRODUCTION
− gaining insights into the pathogenesis of the
disease phenotype → which gene is causing the
disease, disease mechanism
− identifying therapeutic targets and diagnostic
markers → identification of signalling molecules
− testing potential treatments
− when in vitro and/or computational models can’t
reproduce the biological complexity required to
solve the scientific question
− exosomes or extracellular vesicles as a model
− organoids = advanced models with respect to in
vitro and/or computational models for studying
human disease
− in vivo = organismal (patho)physiology, ex vivo =
cellular (patho)physiology
− criteria for choosing a model:
o human analogue appropriateness → anatomy similarities in the modelled disease context
o information transferability → similarities in physiology/organ function in the modelled
disease context
o genetic conservation of the target gene(s) to model the disease (key functional domains) and
of the mutation (inherited diseases) → screening for orthologues (genes in different species
evolved from a common ancestral gene by a speciation (lineage-splitting) event) and
paralogues (gene copies created by a duplication event within the same genome),
expression patterns in the model organisms compared to humans
o ease and adaptability to experimental manipulation → choosing the technique/method to
induce the disease model and for obtaining readouts from the model
o ethical and ecological implications
o budget and time
MOUSE MODELS
− advantages:
o best model for mammalian development processes
o closely related to humans
o 10-15 offsprings per litter and 1 litter every month
o sequenced genome
o 450 characterized inbred strains
o well-developed genetic manipulations
o short generation time, small, and an accelerated lifespan
− different embryo development
, − anatomy & physiology:
o comparable to humans, most processes are very conserved
o microbiome overlap and absorption processes in the tract
o same structures in the kidneys (glomerulus, tubules) → studying kidney exposure to
diuretics and drugs by looking at exosome transporters in the urine
o similar differences in male and female mice → studying sex and drug reaction differences
and hormonal pathways
o 90% murine brain is identical to human, very conserved regions → physiological processes,
Alzheimer’s disease
o behavioural studies → no self-reflection → most mental illnesses can’t be studied
DISEASE MODELLING METHODS
− non-genetic:
o easy injection, but stressful
o accurate oral gavage but stressful
o osmotic pumps provide constant flow
o food/water is the easiest but less accurate
− genetic models:
Allele type Description Most common used
Knockout Complete loss-of-function. Gene function by comparison to non-targeted controls.
Replacing the coding region of
Knock-in a known gene with an Used to study the phenotypic impact of this variation,
(nucleotide alternative sequence of the often pulled from human clinical observations.
substitution) same gene containing a Functional impact is allele specific.
specific mutation.
A targeting construct is used
to introduce a new variant into Used to study the role of the variant and/or express the
Knock-in a specific locus. May be an desired variant or new gene product, ex. humanized
(insertion) alternative variant of the mice or a recombinase under the control of an
targeted gene or a complete endogenous promoter.
new cDNA.
Co-expression of cre recombinase in the same cell will
Floxed LoxP recombinase recognition remodel the flanked region according to the
(inducible sequences are introduced directionality of the LoxP sites and controlled by a
knockout or flanking a targeted gene, exon tissue-specific promoter. In the absence of cre the gene
knock-in) or portion of a construct. is typically expected to function as wild type. Permits
tissue specific excision or expression of a gene. Allows
− gaining insights into the pathogenesis of the
disease phenotype → which gene is causing the
disease, disease mechanism
− identifying therapeutic targets and diagnostic
markers → identification of signalling molecules
− testing potential treatments
− when in vitro and/or computational models can’t
reproduce the biological complexity required to
solve the scientific question
− exosomes or extracellular vesicles as a model
− organoids = advanced models with respect to in
vitro and/or computational models for studying
human disease
− in vivo = organismal (patho)physiology, ex vivo =
cellular (patho)physiology
− criteria for choosing a model:
o human analogue appropriateness → anatomy similarities in the modelled disease context
o information transferability → similarities in physiology/organ function in the modelled
disease context
o genetic conservation of the target gene(s) to model the disease (key functional domains) and
of the mutation (inherited diseases) → screening for orthologues (genes in different species
evolved from a common ancestral gene by a speciation (lineage-splitting) event) and
paralogues (gene copies created by a duplication event within the same genome),
expression patterns in the model organisms compared to humans
o ease and adaptability to experimental manipulation → choosing the technique/method to
induce the disease model and for obtaining readouts from the model
o ethical and ecological implications
o budget and time
MOUSE MODELS
− advantages:
o best model for mammalian development processes
o closely related to humans
o 10-15 offsprings per litter and 1 litter every month
o sequenced genome
o 450 characterized inbred strains
o well-developed genetic manipulations
o short generation time, small, and an accelerated lifespan
− different embryo development
, − anatomy & physiology:
o comparable to humans, most processes are very conserved
o microbiome overlap and absorption processes in the tract
o same structures in the kidneys (glomerulus, tubules) → studying kidney exposure to
diuretics and drugs by looking at exosome transporters in the urine
o similar differences in male and female mice → studying sex and drug reaction differences
and hormonal pathways
o 90% murine brain is identical to human, very conserved regions → physiological processes,
Alzheimer’s disease
o behavioural studies → no self-reflection → most mental illnesses can’t be studied
DISEASE MODELLING METHODS
− non-genetic:
o easy injection, but stressful
o accurate oral gavage but stressful
o osmotic pumps provide constant flow
o food/water is the easiest but less accurate
− genetic models:
Allele type Description Most common used
Knockout Complete loss-of-function. Gene function by comparison to non-targeted controls.
Replacing the coding region of
Knock-in a known gene with an Used to study the phenotypic impact of this variation,
(nucleotide alternative sequence of the often pulled from human clinical observations.
substitution) same gene containing a Functional impact is allele specific.
specific mutation.
A targeting construct is used
to introduce a new variant into Used to study the role of the variant and/or express the
Knock-in a specific locus. May be an desired variant or new gene product, ex. humanized
(insertion) alternative variant of the mice or a recombinase under the control of an
targeted gene or a complete endogenous promoter.
new cDNA.
Co-expression of cre recombinase in the same cell will
Floxed LoxP recombinase recognition remodel the flanked region according to the
(inducible sequences are introduced directionality of the LoxP sites and controlled by a
knockout or flanking a targeted gene, exon tissue-specific promoter. In the absence of cre the gene
knock-in) or portion of a construct. is typically expected to function as wild type. Permits
tissue specific excision or expression of a gene. Allows
