musculoskeletal disease (OA)

Question 1

what is the structure of TYPE1 collagen

Answer 1

made up of 3 alpha chains with a tri amino acidrepeat og Gly- X-Y
exisits as a triple helix rope like sturcutre
(tropocollagen)

type 1 collagen is made up of 2 chains of alpha 1 encoded by COL1a1
and 1 chain of alpha 2 encoded by COL1a2

Question 2

what is a dominant negative muation

Answer 2

mutation where the gene loses its function but also prevents other genes from perfroming thier function as well

in osteogenesis imperfecta, muations in collagen have a dominant negative efffect

Question 3

during the synthesis of collagen which stage is commonly associated with OI (transcribed, translated, mrna, prtoein folding…)

Answer 3

so the post transational modisification includning proline and lusine hydroxylation and glycosylation of hydroxylysine can get slowed down or inhibited in OI

Question 4

what is OI? can you name some symptoms of the disease?

Answer 4

autosomal dominant disease
genetic mutation in either in COL1a1 or COL1A2 gene which encdoes type 1 collagen
there are 4 main types with mild to lethal phenotypes and brittle bones is the main symptom

other symptoms include defromed bones, reduced mineralisation and beaded ribs which can lead to respiratory issues

Question 5

how can OI be diagnosed?

Answer 5

clinically or radiographically but now genetic testing is more common
> fractures or bowing of long bones
> reduced height and macrocephaly
> flat midface

hearing lose, blue schlera
> barrel chest
> scoliosis
> ostepaenia (low mineral density)
> bone biopsy reveals low bone density

Question 6

what is dentiogenesis imperfecta and how can we diagnose?

Answer 6

abnormality in dentin gene which can lead to abnormal tooth development
> thin enamel
> tooth discoloration
>narrow pulp chambers
> teeth ground to gum level

Question 7

there are 4 different types of OI. How are they distinguished

Answer 7

they are separated depending on severity and if they are excluded or included type
> this means if the abnormal gene product is incorporated into the matrix or not

Question 8

briefly overview : type 1 OI

Answer 8

excluded type so its mild in severity so symptoms include hearing loss, blue sclera
the matrix contains 50% of normal amount of collagen but its all normal so there are very little bone deformities

mutation in gene results in a null allelle and results in COL1A1 haploinsufficieny

Question 9

briefly overview : type 2 and type 3 OI

Answer 9

included type and lethal so symptoms include dentinogenesis-i, beaded ribs, short stature, bone deformities, low mineralisation of skull

type 3 is severe and progressive ans shares the same

this is because the matrix contains both normal and abnormal type 1 collagen

Question 10

briefly overview: type 4 OI

Answer 10

also the included type but milder in severity so symptoms include some hearing loss, normal sclera, mild/moderate bone deformity and dentinogeneis-i is quite rare

as is type 2+3, there is reduced secretion of collagen so the matrix is deficient and defective but its different to 2+3 as its caused by point mutations in COL1a2 only!

Question 11

why is the included types of OI more severe than the the excluded type/type 1 OI

Answer 11

this is due to the dominant negative effect.
Mutant gene product not only loses its own function but also prevents normal gene products from functioning correctly.

often affects multimeric proteins such as collagen which is made from more than one alpha chain
> type 3 and 2 collagen made of 1 alpha chain/1gene
> type 9 collagen made up of 3 alpha chain/3 genes
and then collagen fibers aggregate too so a mutation in one chain will affect the overall structure too

Question 12

alongside if OI subtype is included or excluded type, what else can influence the severity of OI?

Answer 12

• Which gene involved - COL1A1 more severe than COL1A2
• What amino acid replaces the glycine - glutamate or aspartate more severe than alanine and serene
• Where in the gene is the mutations - C-terminus more severe disease

Question 13

how can we treat OI?

Answer 13

antireabsorbative therapy - reduced the osteoclast activty but efficacy only mild in children

anabolic therapies - uses recombinant human growth hormone to help promote bone formation

drugs and antibodies

Question 14

which mutatios cause the most severe froms of OI
A missense in c terminus of COL1A1
B nonsense in n terminus of COL1A1
C missense in c terminus of COL1A2
D nonsense in  n terminus of COL1A2

Answer 14

OPTION A
`
missense mutations in col1a1 in the c terminus region as that is where the collagen chains twist to form tropolcolalgen
missense mutations would mean the abnormal collegen would be incorporated into the matrix

Question 15

which chondral dysplasias show loci heterogeneity?
A sticklers syndrome
B achondroplasia
C muliple epiphyseal syndrome

Answer 15

option c and also option a too!

mutations encoding type 9 collgen, sulphate transporters
mutations in different gene loci

achondroplasia shows mutational heterogeneity as the mutations is with the FGFR3 which is in the same loic

Question 16

how can you calculate the probability of abnormal molecules follwiing a COL1A1/2 mutation? what are they?

Answer 16

probability of normal gene+ mutant A1 + mutant a2
> three alpha chains make up collagen
now there are 2 x COL1-A1 genes so either one or both of these chains can be mutated = 75% abnormal
now there are 1 x COL1-A2 genes so its either mutated or not = 50% abnormal

Question 17

what is the main cause of Type 2 OI? what does this result in?

Answer 17

point mutation/ SNP is the main cause. this results in the glycine being substituted
> reduced thermal stability
> slower rate of triple helix formation
> poor packaging into fibrils
> slower rate of collagen secretion from cell

Question 18

the amino acid that replaces glycine can affect the severity of OI. name some aa and how severely they impact OI

Answer 18

alanine and serine = mild symptoms
glutamate or aspartate substitution = severe symptoms s they disrupt the packing of the triple helix and reduce the thermal stability of collagen

Question 19

how does the position of the mutationa ffect the severity of OI?

Answer 19

if the mutation is at the 3’ end of the mRNA / c-terminus, there are more severe symptoms as its at the 3’ end where the triple helix form starts packing so there will be a LARGER disruption
so if this is disrupted then its highly likely there will be an increase in post translational modifications
> hydroxylation of Lysine and proline
> glycosylation of hydroxylysine

Question 20

what is the issue with overmodification with regards to oi

Answer 20

this overmodification/post transcriptional processing can reduces the thermal stability of the collagen and lead to increased intracellular degradation and slower secretion.
also over-modification molecules can lead to poor fibrillogenesis and these abnormal fibrils = poor crosslinking so act as poor templates for mineralisations

Answer 21

i

Question 22

briefly describe the process of endochrondral ossification// how your long bones form

Answer 22

cartilage forms as mesenchymal cells differentiate to chondroblasts which secrete cartilage and become embedded in the lacunae in cartilage matrix
osteoclasts allow vascular invasion and longitudinal growth of the woven bone in the midshaft and the growth plate forms and articular cartilage forms too

growth plate allows bone to continue to grow longitdinally

Question 23

what is the role of the growth plate

Answer 23

it regulates rate of bone length and allows the bone to continue to grow longitudinally.
seperates the diaphysis from the epiphysis

Question 24

describe the components of the growth plate

Answer 24

made up of a mix of resting, proliferative, hypertrophic, proliferating and reserve chrondrocytes but it has a distinct organisation

Question 25

how does the growth plate maintain this distinct organisation

Answer 25

with the help of cartilage matrix molecules
> type 2911 collagen
> aggrecan, type 10 and growth factors like VEGF and MMP13 in expressed in specific regions such as the hypertrophic and bone region
> there are also a range of different intrinsic chemical signals and hormones which allow cells to differentiate

> extrinsic signals like diet, GH, insulin, nutrition and skeletal load will also affect growth

Question 26

what happens if the differentiation of collagen is disrupted?

Answer 26

growth plate will not extend at its normal rate
> either be accelerated
> or be reduced (as seen in chondroplasias

Question 27

why are there so many different types of chondroplasias (150 types! :0)

Answer 27

as we know chondroplasias are due to defects in cartilage. now cartilage is made up of many components (collagen, proteoglcan, ECM proteins like COMP
so a mutation in any of these components will lead to similar clinical phenotype
> this is locus heterogeneity

Question 28

what do all chondroplasias types have in common?

Answer 28

All defects in cartilidge so cab lead to abnormal linear growth
they all lead to a skeletal deformity usually dwarfism due to a mutation with endochondrial ossification which alters and disrupts the normal function of the growth plate

Question 29

what is the role of cartilage in the ECM? How does it achieve these functions?

Answer 29

it forms a meshwork and gives the ECM mechanical strength and compression resistance
> can entrap large sulphated proteoglycans which can retain water
> articular cartilage allows for frictionaless movement between joints

Question 30

briefly describe achondroplasia. does it show locus heterogeneity?

Answer 30

Achondroplasis is caused by a GOF mutation in FGFR3 that disrupts normal diffeention form pre to hypertrophic chondrocytes.
this results in short limbed people

does NOT show locus heterogeneity as its just one gene that is affected

Question 31

What is DMD? What is the genetic basis

Answer 31

X linked recessive disorder
Due ax chrinose deletion at Xp21 deletion or trabsloaciton between X:21

Progressive muscular weakness and death in 3rd decade due to cardiac.and respiratory failure

Question 32

How voukd we detect the DMD gene

Answer 32

Use subtraction cloning where healthy DNA is cloned and compared to patient DNA to identify deleted region
This is a form of cytogenetics which is staining the chromosome

Or identity breakpoints in indiduals with X chromosome translocation

Question 33

If DMD is a xreccesive then why do females have it

Answer 33

Due to x inactivation which can favour DMD manjfedtstion

So if normal allele is inactivated then only the X:21 chromosome remains

Question 34

What is the longest human gene. Features of it, what does it emcode

Answer 34

DMD GENE

2.5mMbp Long, 86 exons and 99% introns
Mutation hot-spot between exon 43 and 55 where there are deletions

Encodes dystrophin which can be reduced/none in DMD or have low molecular weight in BMD

Question 35

Why is DMD more severe than BMd

Answer 35

Frame shift mutation so very low or no dydtophon expression so sever eclincal symptoms

BMD frame jietsl deletions so some dydtophon expression, maybe a lower molecular weight but less severe symptoms

Question 36

Where on DMD GENE is deletion xommon

Answer 36

43-55 most common mutations hot-spot and there the majority deletions occur