Geen diatitel

1
Human Molecular Genetics IV. Genetics of common
diseases/ Multifactorial genetics
2
genetics of common diseases

• coronary heart disease (CHD)
• atherosclerosis, hypertension
• cancer
• obesitas
• diabetes
• asthma
• schizophrenia
• dementia

3
genetics of common diseases

• rarely monogenic
• most often (i) polygenic and (ii) multifactorial

a phenotypic trait determined by (i)
interaction between several genes/loci,
each with a small additive effect (ii) influence
of environmental factors
4
Genetics of common diseases
multifactorial inheritance

• continuous
• no specific phenotype eg. length
• discontinuous
• specific phenotype eg diabetes,CL/CP
• critical balans, treshhold when crossed, the
• phenotype appears, severity phenotype

y
threshold
affected dividuals
5
Genetics of common diseases
multifactorial inheritance (discontinuous)
general population liability curve
y
threshold
affected dividuals
6
Genetics of common diseases
Evidence for multifactorial inheritence

• family studies

• increased incidence of a disease in particular
  families
• common environment check not related
  individuals (spouses)

• twin concordance studies

• dizygous (DZ) vs monozygous (MZ) twins
• common environment twins raised in different
  environment
• concordant both affected or neither affected
• genetically determined MZ similarly affected, DZ
  not
• environmental MZDZ

• adoption studies, population and immigration
  studies

7
Genetics of common diseases
Evidence for multifactorial inheritence
Sufficient evidence is obtained for
genetic susceptibility for a given common
disorder Which strategies for disease gene
identification can be followed? Example of CHD
what causes do we know for the disease?
8
Genetics of common diseases
Identification of genes involved in common
diseases

• methods
• linkage analysis using whole genome scans
• association studies using SNPs
• candidate gene analysis
• biochemical analysis
• combined approach
• study material
• families/affected sibs family members
• numbers/selection/clinical diagnosis/pheno-
• copies/
• animal models
• numbers/more homogeneous genetic background

9
x2
x1
x1
x2
x1
x2
10

• linkage
• genes are on chromosomes and thus assumed to be
• linked during transmission from one generation
  to
• another
• in reality linkage only holds for relative small
  
• distances due to meiotic crossing over
• linkage analysis follow the pattern of
  inheritance of
• polymorphic markers in pedigrees in which a
  disease
• phenotype segregates
• 1 recombination 1cM
• RFLP, minisatellites (VNTRs),
• microsatellites or (CA)n repeats

11
linkage odds of linkage likelyhood for
linkage/no linkage LOD logarithm of the odds
ratio for linkage LOD score gt3 significant
linkage lt-2 no significant
linkage haplotypes sets of alleles on a small
chromosome segment
a
A
B
b
c
c
12
association studies
13
association studies and LD

• linkage disequilibrium combination of closely
  linked
• alleles, referred to as haplotypes, originating
  from a
• single ancestral chromosome
• apparently contradictory with the expected
  random
• association assuming the occurrence of random
  CO
• over many generations
• cause founder mutations, recent mutations
• studied by polymorphic markers (RFLPs, CA
  repeats
• , more recently SNPs)
• study of inbred strains of mice or rats

14
(No Transcript)
15
Association studies for detection of disease
genes using linkage disequilibrium

• Can we use SNPs for association studies in man
• SNPs common and rare
• SNPs coding and non coding
• analysis of haplotypes and LD using SNPs
• computer simulation and experimental data
• suggest that LD extends only a few kb away
• from SNPs
• other data suggest gt 100 kb
• reasons for discrepany
• small studies
• different populations

16
Association studies for detection of disease
genes using linkage disequilibrium
Reich et al. Nature Genetics May 2001 rather
large blocks of LD interspersed with
recombination hot spots
17
Association studies for detection of disease
genes using linkage disequilibrium

• Study design (Reich et al. Nature Genetics May
  2001)
• 19 different chromosomal regions anchored
• around a coding SNP
• finished sequence for at least 160 kb (North
  European)
• frequent minor (less common) allele
• allows cross population comparison
• possible with modest sample size
• useful in search for common diseases
• resequencing of 2 kb region at
  0-5-10-20-40-80-160 kb
• 272 high frequency polymorphisms
• calculation of allele frequency and LD

18
Association studies for detection of disease
genes using linkage disequilibrium

• Results (Reich et al. Nature Genetics May 2001)
• relatively large blocks of LD
• why?
• Study of Yorubans, Nigerian population
• common ancestry with NE around 100.000 yrs
  ago
• similar allelic combinations at short
  distance
• half length LD is less than 5 kb
• Consequences
• genome wide LD mapping probably possible but
• limited resolution, choose other populations for
  
• refined mapping

19
Mouse models

• history 1900 inbred strains of mice
• until 1970s difficulties in finding the
  responsible defects
• later linkage analysis, positional cloning,
• genetic maps of mouse and man al lead to
• identification of single gene mutants
• development of powerful statistical programs
  lead
• to quantitative trait locus (QTL) analysis
• advantages through inbreeding and controlled
• environmental factors (eg nutrition)
• knock out and transgenic mice

20
Genetics of common diseases

• situation 1 (CHD)
• rare monogenic disorders
• known (metabolic, biochemical) pathways
• situation 2 (obsesitas)
• mouse models with monogenic traits
• previously unknown metabolic pathways

21
Genetics of common diseases coronary heart
disease

• frequent - high incidence, important impact on
  public health
• environmental and behavioural changes
• increasing age

• single gene disorders (rare) vs polygenic
  disorders (common)
• multifactorial inheritance
• multiple and complex genetic factors interacting
  with environment
• Duchenne muscle dystrophy vs infectious disease

22
Genetics of common diseases cardiovascular
disease - atherosclerosis

• hart attacks (infarct), stroke (thrombosis) and
  peripheral
• vascular disease
• occlusions in large and middle sized arteries
• late onset as a result of chronic damage of
  vascular
• endothelial cells
• involved elements LDL, thrombocytes,
  macrophages, lymphocytes,
• smooth muscle cells of intima, invasion of fat
  particles in blood
• vessels and formation of fibrocellular
  atheromatuos plaques

23
(No Transcript)
24
(No Transcript)
25
Genetics of common diseases CHD -
atherosclerosis

• CHD coronary heart disease
• multifactorial, no Mendelian segregation
• genes involved in lipoprotein level, blood
  pressure,
• diabetes, obesitas, blood coagulation, immune
  system,
• blood vessel reactivity
• environmental factors smoking, nutrition,
  exercise
• interindividual variation in disease
  susceptibility
• predictors of risk

- blood lipids (cholesterol) - blood pressure -
blood coagulation factors
26
Genetics of common diseases cardiovascular
disease
lipoprotein metabolisme
http//www.mmip.mcgill.ca/unit2/cianflone/lect48nu
trition.htm
27
Exogenous pathway
Esterifaction of fatty acids and cholesterol
Uptake of remnant particles in liver by LDL
receptor/LRP and chylomicron receptor
Triglycerids - cholesterol Fat soluble
vitamins phospholipids apoB48/apoCI-III/apoAIV
Remnant particle enriched with cholesterol
esters apoB48 apoE/C
Release of fatty acids (FFA) in peripheral
capillaries Mediated by Lipoprotein lipase
(LPL) and co-factor apoCII
28
VLDL particle central triglycerids and
cholesterol packaged
with phospholipiden and one apoB100 molecule
29
Exogenous pathway
Esterifaction of fatty acids and cholesterol
Uptake of remnant particles in liver by LDL
receptor/LRP and chylomicron receptor
Triglycerids - cholesterol Fat soluble
vitamins phospholipids apoB48/apoCI-III/apoAIV
Remnant particle enriched with cholesterol
esters apoB48 apoE/C
Release of fatty acids (FFA) in peripheral
capillaries Mediated by Lipoprotein lipase
(LPL) and co-factor apoCII
30
Endogenous pathway
Synthesis of triglycerides and cholesterol in the
liver
Partial conversion of IDL to LDL by hydrolysis
of triglycerides to cholesteryl-ester, removal
of apos except apoB100 Partial clearance of IDL
in liver by LDLR and apoE
Assembly of triglycerides and cholesterol with
phospho- lipids/one apoB100 molecule and many
apoC/E molecules into VLDL particles
IDL
Formation of VLDL remants (IDL) after removal of
triglycerids
31
Endogenous pathway
excess LDL molecules oxidise Attract macrophages,
transform into foam cells upon LDL uptake,
oxidation
Membrane and steroid hormone synthesis
32
Exogenous pathway dietary lipid absorption and
transport

• absorption of fatty acids and cholesterol in
  intestinal aborptive cell
• esterification to triglycerids and
  cholesterol-esters, respectively
• transport to lymphatic system and into plasma in
  the form of
• chylomicrons (triglyceride rich lipoproteins)

33
Exogenous pathway dietary lipid absorption and
transport

• chylomicrons are large particles consisting of
• core of triglycerids and cholesterol-esters
• apolipoprotein apoB48 and small amounts of
• apo CI, CII, CIII en E and A-IV
• metabolised (hydrolyse) in peripheral
  capillaries to fatty acids as
• energy source for skeletal muscle tissue or for
  storage in fat cells,
• through the action of lipoproteine lipase (LPL)
  and apoCII as co-factor
• following release of triglycerids, apoA en apoC
  are transferred to HDL by LPL,
• chylomicron remnants (cholesterol-rich) are
  removed from circulation by
• LDL receptor en LRP (low density lipoprotein
  receptor related
• protein) mediated pathways in the liver

34
Endogenous pathway hepatic lipoproteins

• liver synthesises triglycerids and cholesterol,
  which together with residual
• dietary fat, fat-soluble vitamins and apoB100
• (1 molecule per VLDL partikel) are incorporated
  into VLDL particles and
• secreted into circulation
• aim transport of fatty acids from liver to
  other tissues
• functional form results through inclusion of
  apoE and apoCII en CIII from HDL
• hydrolyse and removal of core-triglyceride by
  LPL
• result VLDL remnants IDL (intermediate
  density lipoproteins)
• 1/2 absorbed by liver via apoB ( ligand for LDL
  receptor)
• 1/2 hydrolysed by hepatic lipase to LDL
  (cholesterol-ester rich)

35
hepatic lipoproteins (2)
LDL
- carries 60-70 of plasma
cholesterol, delivers cholesterol to peripheral
tissues and to the liver for further
metabolism and excretion in bile
(receptor mediated process) - 75
taken up by liver via apoB100 (ligand for LDL
receptor) - 24 to peripheral tissues for
membrane and steroid hormone biosynthesis
metabolic consequences of cholestrol uptake by
cells (1) decreased de novo cholesterol
synthesis, (2) increased conversion of
cholesterol into cholesterolester
(storage form of cholesterol) (3)
decreased expression of LDL receptors -
remaining 1 remains in circulation and can be
modified by oxidation, these oxidised LDL
particles can attract scavenger macrophages
which become foam cells as they ingest
these particles
36
DYSLIPIDEMIAS

• familial LPL and apoCII (LPL co-factor)
  deficiency
• - no hydrolysis of chilomicrons and VLDL
  resulting in
• hypertriglyceridemia, no increased risk for
  atherosclerose
• - 1/mio except in high risk populations (eg
  Quebec)
• - low fat intake

37
DYSLIPIDEMIAS

• FH, familial hypercholesterolaemia
• - defect in LDL receptor gene no LDL
  clearance from circulation
• (no r, precursor doenst reach the
  membrane, r doenst bind
• LDL, hundreds of different mutations)
• - HoZ (LDLx4-6) not older than 30 yr, HeZ
  (1/500)
• 1/2 heart attack before age of 60 yr
  (LDLx2)

• familial apoB100 defect
• - one single mutation
• - no binding of LDL to receptor
• - HeZ increased LDL 50-100
• - 1/1000

38
DYSLIPIDEMIAS
search for common variants in genes
influencing LDL content

• linkage studies for three genes involved in LDL
  metabolism
• in 150 families

CYP7 cholesterol 7?-hydroxylase, enzyme involved
in bile acid production

• other loci 1p34, 13q, 15q25
• Hyplip1
• - mutant mouse strain for
• familial combined hyperlipidemia (FCH)
  phenotype
• - triglycerides and/or cholesterol raised
  plasma levels
• - fine mapping of mouse locus
• - 13 candidate genes mRNA expression and
  sequencing
• - thioredoxin interactin protein

39
OBESITAS introduction (1)

• body mass index (BMI) gt30
• increased risk for NIDDM, hypertension, CHD,
• reproductive problems, etc...
• 1/3 Amerikan population, increasing problem
• in children
• interaction between genetic, environmental and
• psychosocial factors
• energy homeostasis

40
OBESITAS introduction(2)

• obesitas genes identified
• genetic predisposition
• availability of food, composition,
• excersise
• thrifty gene hypothesis (Neel, 1999)

41
OBESITAS introduciton(3)

• energy balance
• energy storage when energy intake is higher than
  total
• expenditure
• E-expenditure through physical activity,
• basal metabolism and adaptive thermogenesis

42
OBESITAS control of energy-intake and body
weight

• behaviour, autonomous nervous system and
• neuroendocrine
• short term start and stop eating due to hunger
  and
• saturation, controled by neural and endocrine
  factors
• long term eg by leptine hormone produced
• by fat cells
• CNS ligand-receptor signal-
• transduction pathways

43
OBESITAS GENEN
single gene disorders mouse models for
obesitas causative genes are identified
(agouti, fat, tubby, obese, diabetes)
Db/Db
Ob/Ob
obesitas hyperfagie hypogonadisme
44
orexigenic
agouti
MC4R
Ob
anorexigenic
leptine gereguleerde centrale melanocortine
circuit
45
neuropeptide Y/Argp (Agouti related
peptide) - endogenous regulator of energy
balance - feeding-inducing neuropeptides -
strong expression at nucleus arcuatus
(hypothalamus) - leads to suppression of MC4R
(melanocortin 4 receptor) - causes
increased food intake - decreases energy
expenditure - link with insuline is unclear, not
the dominant peripheral signal molecule -
first discovered orexigenic factor
46
neuropeptide Y/NPY
receptors - KO mouse normal
NPY/leptin dubble KO reduced effect of leptin
KO - KO for 6 known receptors obesity instead
of expected anorexigenic effect reveals
complexity of control mechanisms and
multifactorial control
47
agouti/Agrp
- gain-of-function Argp mutant mouse obesity
phenotype comparable with loss-of-function for
Pomc of Mc4r - Ay mutation ectopic expression
of agouti color, dominant obesity
syndrome, increased growth and
yellow hair color - related to Argp
48
leptin-leptin receptor - leptos thin -
hormone primarily produced by adipocyt - belongs
to cytokine family of proteins - is responsible
for complex neural respons incl hunger,
behavioural changes (search for food),
decreased metabolism, infertility...
49
leptin-leptin receptor
- communication concerning lon term energy
storage - other effects outside CNS decreased
triglyceride accumulation in tissues other
than fat tissue (eg muscle, liver),
contributes to insuline resistance - abscence
of leptin signal in the presence of food
causes obesitas - causes decreased expression of
NPY/Argp - induces starvation respons
50
leptin
- treatment by subcutaneous leptin injection - 2
families with leptin mutatie - AR - HoZ for
loss-of-function mutation
leptin-receptor
-1 family - HoZ mutation responsable for
truncation of the cytoplasmatic domain -
class I cytokine receptor
51
CART/POMC/?-MSH - CART cocaine and amphetamine
related transcript ?-MSH derived from
proopiomelanocortin (POMC) - CART and POMC are
induced by leptin (anorexigenic) - produced by
two neuronal populations within the
hypothalamus - POMC twchildren with HoZ or
compound HoZ for loss-of-function mutation
52
Mc4r - hypothalamic homologue of MC1R (receptor
in melanocytes) - KO melanocortin obesity
syndrome agouti but without yellow hair
color - mutation in humans are responsible for
4-5of obesity cases (haploinsufficiency,
not dominant negative)
53
Other genes

• neuropeptide processing enzyme
• - carboxypeptidase E exclusively in mouse
  (fat)
• - PC-1 discovered in man
• complex obesity syndromes in mouse and man
• probably as a results of POMC processing
• MC3R obesity in mouse
• UCP and BAT (brown adipose tissue)

54
QTL analysis in mice search for obesity genes

• effect of individual genes on energy
  expenditure,
• hyperphagia and fat storage
• study of the effect of dietary composition
• more than 70 loci identified in mice

blue whole genome scan human red mouse
QTLs green human monogenic mutations
55

• Breakthrough in genetic studies on common
  diseases
• ADAM33 gene in asthma (Nature 418426, 2002)
• G72 in schizophrenia (PNAS 9913675, 2002)