1
Q
what is CF? demographics and epidemiology
A
- an inherited disease
- 1/2500 births
- inherited in an autosomal recessive fashion
- most common genetic disease in the UK caucasian population
- carrier frequency = 1/25
- P(both parents are carriers) = 1/625
- median lifespan = 49.1 years
2
Q
what is the chance of having a child with CF when one parent is a carrier and one parent has CF?
A
50%
3
Q
what is the chance of having a child with CF when both parents are carriers?
A
25%
4
Q
where is the mutation found in CF and what is the most common mutation?
A
= found on chromosome 7 in the CFTR gene
- F508del = a deletion of 3 adjacent base pairs at the 508th codon - loss of a phenylalanine
5
Q
role of CFTR protein
A
a chloride ion channel in the cell membrane of epithelial tissue cells
6
Q
normal use of CFTR protein
A
- upon activation, the CFTR protein channel opens
- causes intracellular chloride ions to leave the cell
- as a result, the extra cellular environment becomes negative
- therefore intracellular sodium ions leave the cell via sodium channels
- reduces the level of intracellular NaCl
- creates a water potential, so water moves out via osmosis, improving the quality of cellular mucous secretions
- under normal conditions, the secretions aren’t thick as they have sufficient water content
7
Q
how is CFTR activated and what kind of protein is it?
A
- ABC protein — have a modular structure with 4 conserved domains
- has an additional 5th domain called the Regulatory domain which regulates the activity of the protein
- when it is phosphorylated the channel can open
- also requires the binding of ATP to the protein (binds to ATP-binding domains)
8
Q
mutated CFTR protein in sweat glands
A
defective channel = inability to reabsorb chloride —> reduced reabsorptions of sodium ions and water —> excess loss of salt and elevated levels of NaCl in sweat
9
Q
mutated CFTR proteins in all other exocrine glands (+ how does it lead to organ damage)
A
defective channel = inability to transport chloride out of cells —> reduced secretion of chloride and water —> accumulation of intracellular chloride —> more sodium ions are reabsorbed —> more water reabsorbed —> formation of hyperviscous mucous —> accumulation of secretions and blockage of small passages of affected organs —> chronic inflammation and remodelling —> organ damage
10
Q
how does a defective CFTR protein lead to inflammation?
A
the thick mucus provides a prime environment for the growth of bacteria
11
Q
give examples of CF treatment
A
- diet supplemented with high-calorie/proteins foods — high energy to compensate for the increased energy demand
- given pancreatic extract/enzymes
- large doses of vitamins A,D and K and liberal use of salt
- daily chest physiotherapy
- maintain breast-feeding for as long as possible — boosts infant’s immunity, provides them with the much needed calorific nutrition
- maintenance of good hygiene
12
Q
what does daily chest physiotherapy entail?
A
- postural drainage = trachea is inclined downwards and below the affected chest area — drains fluid from lungs
- percussion and vibrations = clapping of chest with percussion cups — helps loosen and mobilise secretions
13
Q
name some symptoms of CF
A
- excess coughing
- hypoproteinemia
- anaemia
- fatty, foul-smelling stools
- ravenous appetite but poor weight gain — thin
- salty-tasting skin due to higher levels of chloride in sweat
- frequent coughing and wheezing
- difficulty in breathing as mucus clogs airways
14
Q
what is hypoproteinemia and why do patients with CF develop it? what does it result in?
A
= decrease of protein in the blood (less is digested as the thick music blocks the pancreas from secreting digestive enzymes)
- results in swelling (odema) due to the accumulation of fluid in tissues —> INFECTION
- a result of malabsorption
15
Q
how does CF affect the pancreas and what is the consequence?
A
build up of thick mucus prevents the pancreas from secreting digestive enzymes —> malnutrition and poor growth
16
Q
how does CF cause pancreatic disease?
A
- the pancreatic ducts become blocked with mucus (pancreatic secretions are dehydrated and thick)
- pancreas continues to produce digestive enzymes
- these abundant enzymes damage the pancreatic tissue —> fibrosis of pancreas until it is no longer able to produce enough enzymes to properly digest food
17
Q
what is the effect of the pancreas no longer being able to digest food properly?
A
- malabsorption of nutrients — malnutrition
- lowered absorption of fats = diarrhoea, weight loss, malnutrition
18
Q
what is the 2nd most common mutation leading to CF?
A
= missense mutation changing Glycine at position 551 to an aspartate reside
= G5510
= reduces chances of the CFTR channel opening
19
Q
name 5 methods of diagnosis for CF
A
- chest x-ray — looks for lung infections and dilated airways filled with mucus
- sweat test — measures amount of NaCl in sweat
- blood test — blood cells screened for CF gene
- screening test
- analysis of faeces — tests for pancreatic enzymes
20
Q
what is an abnormal sweat test result, indicating CF?
A
> 60 mmol/l
21
Q
describe pseudomonas aeruginosa
A
- gram negative
- bacillus (rod) shaped
- ‘opportunistic pathogen’
- produces toxic proteins that enter and kill host cells
- resistant to many antibiotics
22
Q
symptoms of pseudomonas aerugiosa infection
A
- inflammation
- sepsis
- fever
- elevated respiratory and heart rates
23
Q
what is meconium ileus? what are the symptoms? what can it lead to?
A
= obstruction of the bowel caused by thick abnormal meconium
- distension of the abdomen or vomiting
- can lead to bowel perforation, a twisting of the vowel, or inflammation and infection of the abdominal cavity
(98% of babies with MI have CF)
24
Q
how are mutations expressed?
A
mutations are expressed as proteins — happens through the protein synthesis of the mutated gene
25
describe the stages of transcription
- DNA helix unwinds using RNA helicase
- one of the strands acts as a template strand
- RNA polymerase binds to the DNA of the gene at the promoter
- RNA polymerase always builds in the 5’ to 3’ direction (binds at the 3’ end)
- complementary base pairing occurs
- RNA strands have U instead of T
- RNA polymerase stops transcribing when it reaches the terminator
- splicing occurs — introns removed from pre-mRNA by small nuclear ribonucleoproteins (snRNPs)
- mRNA formed, leaves nucleus through nuclear pore
26
describe the stages of translation
- mRNA molecule attaches to a ribosome
- there are free molecules or tRNA in the cytoplasm, carrying anti codons and a specific amino acid binding site
- tRNA molecules bind with their specific amino acids and bring them to the mRNA on the ribosome
- anticodon on the tRNA molecule pairs with its complementary codon on the mRNA
- ribosome moves to next codon
- peptide bond forms between amino acids
- translation stops when a stop codon is reached
27
describe the process of PCR gene amplification
1. heat to 95 degrees — breaks H bonds, forming single-stranded DNA
2. add free DNA nucleotides and DNA polymerase
3. add primers
4. cool mixture to 55 degrees — allows primers to bond and form short lengths of double stranded DNA at either end of the sample
5. DNA polymerase extends the primer DNA to synthesise the complementary DNA sequence
6. temp raised to 72 degrees = optimum temp for DNA polymerase
7. DNA polymerase adds free nucleotides and amplification ends when it reaches the other end
8. subsequent heat denaturation of the double-stranded DNA, followed by annealing of the same primer sequences to the resulting single-stranded DNA, will result in the synthesis of further copies of the target DNA.
28
what can DNA probes be labelled with?
- a radioactive marker — location can be seen on exposure to photographic film
- a fluorescent marker that emits a colour on exposure to UV light (FISH)
29
what can DNA probes be used for?
- locating a specific gene for genetic engineering
- identifying the same gene on different genomes
- identifying the presence of absence of an allele for a particular genetic disease
30
what is genetic counselling?
a communication process that deals with the human problems associated with the occurrence of risk of occurrence, of a genetic disorder in a family. The process aims to help the individual or family to understand, choose, and adjust
31
what are the 2 methods of chromosomal analysis?
- karyotyping: performed with metaphase spread and so requires a population of dividing cells (blood, amniocytes). It can only detect relatively large intrachromosome changes (deletions, insertions, inversions)
- FISH = ‘ Fluorescent in still Hybridisation ‘ — used to detect small changes in chromosome structure. Fluorescent probes target known sequences on chromosomes
32
what is a genetic disorder?
= an illness caused by an abnormality in the individual’s DNA. It is inherited or acquired
33
name the 3 types of point mutations
- missense
- nonsense
- frameshift
34
what is a missense mutation?
- amino acid substitution
| - effects on structure/ functionality
35
synonymous vs non-synonymous mutation
- synonymous = functionally silent and evolutionary neutral
| - non-synonymous = changes the protein sequences and is frequently subject to natural selection
36
what is a nonsense mutation?
- premature stop codon
- results in a truncated, shorter, unfinished protein
- the earlier in the sequence, the more likely it is to be detrimental
37
what is a frameshift mutation?
- a mutation that alters the reading frame or insertion of a stop codon
- radically alters protein sequence/structure/functionality
38
what are 3 molecular causes of genetic disorders? give examples of each
1. chromosomal abnormalities eg. Down Syndrome
2. DNA/single gene mutations eg. F508del — CF
3. multi factorial eg. neural tube defects (NTDs)
39
why is making a diagnosis of a genetic disease important?
- common cause of childhood morbidity and hospitalisation
- appropriate clinical management
- treatment and prognosis
- information for family
- risk estimation for future offspring and other family members
40
name 4 types of inheritance and give examples of each
1. autosomal dominant — Huntington’s Disease
2. autosomal recessive — CF, sickle cell anaemia
3. x-linked recessive — red-green colour blindness
4. x-linked dominant — Rett Syndrome
41
what does every diploid cell contain in respect to autosomes?
each diploid cell has 2 copies of all the autosomes
42
characteristics of an autosomal dominant inheritance
- disorder occurs where one of the 2 copies has a mutation and the protein produced by the normal form of the gene cannot compensate
- one mutant allele = disease
- males and females affected equally
- affected children have affected parents
- doesn’t skip generations
- two unaffected parents will not have affected children
43
in an autosomal dominant trait, what is the chance of inheriting the mutation (and so having the disease) from 2 heterozygotes? what is the chance of inheriting the disease as a heterozygote?
- offspring of heterozygotes have a 75% of inheriting the mutation and so having the disease - AA, Aa, Aa
- 50% chance of being a heterozygote - Aa, Aa
44
characteristics of an autosomal recessive inheritance
- disease only manifests when the mutant allele is present in a double dose eg. aa
- heterozygotes are carriers and show no features of the disorder
- clinical features = usually severe
- males and females affected equally
- skips generations
- 2 affected individuals always results in an affected child
- consanguinity frequent
45
what is pseudodominance?
when a homozygous individual (aa) marries a carrier (Aa) of the same disorder. children have a 50% chance of being affected (Aa, Aa, aa, aa)
46
what is co-dominance? give an example
when two allelic traits are both expressed in the heterozygous state. eg, blood group AB
47
characteristics of x-linked recessive inheritance
- diagonal inheritance
- daughters of affected males will be carriers
- fathers do NOT pass to their sons
- sons of female carriers have a 50% chance of being affected
- present in males and only in homozygous females (usually rare) (males more commonly affected)
- trait skips generations
48
characteristics of x-linked dominant inheritance
- transmitted by both sexes
- 2x more frequent in females
- transmission by male: every daughter affected, no son affected
- transmission by female: 50% of the children will have the disorder, 50% will be unaffected, equal chance of son or daughter being affected
- females less severely affected than males
49
what is population screening?
the process of identifying people who appear healthy but may be at an increased risk of a disease of condition
50
what are the 4 stages of Piaget’s theory of child development?
1. the sensorimotor stage = birth — 2y
2. the preoperational stage = 2–7y
3. the concrete operational stage = 7–11y
4. the formal operational stage = 12y+
51
what is the sensorimotor stage?
- birth — 2y
- the infant knows the world through their movements and sensations
- children learn about the world through basic actions such as sucking, grasping, looking and listening
- infants learn that things continue to exist even though they cannot be seen (object permanence)
- they are separate beings from the people and objects around them
- they realise that their actions can cause things to happen in the world around them
52
what is the preoperational stage?
- 2–7y
- children begin to think symbolically and learn to use words and pictures to represent objects
- children at this stage tend to be egocentric and struggle to see things from the perspective of others
- while they are getting better with language and thinking, they still tend to think about things in very concrete terms
53
what is the concrete operational stage?
- 7–11y
- children begin to understand the concept of conversation; that the amount of liquid in a short, wide cup is equal to that in a tall, skinny glass for example
- their thinking becomes more logical and organised, but still very concrete
- children begin using inductive logic, or reasoning from specific information to a general principle
54
what is the formal operational stage?
- 12y +
- the adolescent or young adult begins to think abstractly and reason about hypothetical problems
- abstract thought emerges
- teens begin to think about moral, philosophical, ethical, social, and political issues that require theoretical and abstract reasoning
- begin to use deductive logic, or reasoning from a general principle to specific information
55
what is an upper respiratory infection? give examples
an infection affecting your sinuses and throat
- common cold
- epiglottitis
- laryngitis
- pharyngitis (sore throat)
- sinusitis (sinus infection)
56
what is a lower respiratory infection? give examples
affects airways and lungs. in general are more serious and last longer than upper respiratory infections
- bronchitis (a lung infection that causes coughing and fever)
- bronchiolitis (a lung infection that affects mostly young children)
- chest infection
- pneumonia
57
what bacterium is most commonly associated with respiratory infections in individuals with CF beyond the first 5-10 years of life?
Pseudomonas aeruginosa
58
according to piaget, children learn through a process of assimilation and accommodation — what do these words mean?
- assimilation = the process in which the child uses current schemas to interpret its external world
- accommodation = the process of changing ones existing ideas/schemas as a result of new experiences/information
59
why is the gram negative pseudomonas aeruginosa often difficult to treat?
in large colonies of bacteria biofilms can form blocking the action of traditional antibiotics and protecting the colony from the environment
60
transition vs transversion mutation
transition = interchange of purines (A,G) or interchange of pyrimidines (C,T)
transversion =pyrimidines swaps with purine or vice versa
61
when does CF begin in a person?
at conception
62
what shape are coccus bacteria?
spherical
63
in the UK, population screening occurs in all births for what disorder?
phenylketonuria
64
what sites are there on a ribosome?
A,P,E sites — tRNA binding sites
65
CF mainly affects what organ?
the pancreas
66
why is gram negative pseudomonas aeruginosa often difficult to treat?
in large colonies of bacteria biofilms can form blocking the action of traditional antibiotics and protecting the colony from the environment
67
what conditions do gram negative bacteria such as pseudomonas aeruginosa thrive in?
human or wet conditions
68
where are ribosomes made?
in nucleoli within the nucleus
69
what contains the cell’s hereditary units?
the nucleus
70
in non-dividing cells, DNA is found where and in what form?
found in the nucleus in the from of chromatin
71
coccus bacteria are what shape?
spherical
72
how are mitochondrial disease inherited?
maternal
73
what is the appendix attached to?
caecum
74
what is the sweat test?
measures the concentration of chloride that is excreted in sweat. it is used to screen for CF
75
if a missense mutation codes for an amino acid that is chemically dissimilar, for example has a different charge, what is it termed as?
a non-conservative substitution
PBL SEM 1
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