36 Rawan Almujaibeldoctor2016.jumedicine.com/wp-content/uploads/sites/6/2018/01/Genetics... ·...
Transcript of 36 Rawan Almujaibeldoctor2016.jumedicine.com/wp-content/uploads/sites/6/2018/01/Genetics... ·...
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- 36
- Rawan Almujaibel
محمود الحربي-
- Nafith
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The Extracellular Matrix
Extracellular Matrix is a fluid that contains proteins secreted by cells and
contains high carbohydrate content and it fills the spaces between cells
and binds tissues together.
There are types of proteins in the ECM called linker proteins not
secreted by the cells. Their job is to link the proteins in the solution to
the cells to form a network of proteins. This network connects cells
together, it connects proteins together and it connects cells to proteins.
This network is what makes the extracellular matrix a jell-like structure.
The extracellular matrix has two main components: the basal lamina and
connective tissue
Basal laminae: it is found beneath the endothelium and the epithelium.
These two types of cells rest on the basal lamina FROM SLIDE: It is a
Thin, sheet-like, structure upon which layers of epithelial cells rest. Also,
it is surrounds muscle cells, adipose cells, and peripheral nerves.
Connective tissue: It is loose network of proteins and carbohydrates
underneath epithelial cell layers where fibroblasts are distributed.
The connective tissue has proteins which help in connecting the cells
that were already in it to the basal laminea or they connect cells to each
other or even with other proteins.
If you have a close look on the connective tissue in bones, cartilage or
tendons, will find they are closely related to each other. However, the
type of secretions and type of deposited materials inside each of them
are different, so that makes them have their own unique properties.
For example: if you compare bones and tendons, you will find that
bones are harder than tendons due to deposition of calcium in bones.
Also, tendons are made mainly out of fibrous proteins which make them
softer than bones. Cartilage is made of high concentration of
carbohydrates (which makes it more viscous and allows it to return to its
normal shape if it is changed).
So the property of the tissue in general depends on the secretion and
the deposition of substances in the extracellular matrix.
FROM SLIDE:
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Matrix Structural proteins
-Tough fibrous proteins embedded in a gel-like polysaccharide ground
substance.
- Basal laminae is formed of matrix components that differ from those
found in connective tissues.
• Tendons: mainly fibrous proteins
• Cartilage: high concentration of polysaccharides (gel)
• Bone: hardened by deposition of calcium phosphate crystals.
There are 3 components of the ECM:
1) The matrix proteins
2) Polysaccharides
3) Adhesion proteins (link the matrix proteins to each other and link the
cells to the matrix as well); different tissues have different types of these
proteins
Now we are going to talk about the 2 main proteins that are secreted in
the extracellular matrix . . .
1- Collagens (is a glycosylated protein- its function to increase the
viscosity in the extracellular matrix) :
• The most abundant proteins in mammals (25% of the total
protein mass).
• Long, stiff, triple-stranded helical structure made of three α
chains.
(Their shape is triple-stranded; has 3 polypeptide chains wrapped
around each other. The amino acid structure is triplicate and the
glycine is repeated every 3rd amino acid, so the content of the
glycine is 1/3 of all amino acid in collagen).
• A basic unit of mature collagen is called tropo-collagen.
(Collagen is produced as a pro-collagen by fibroblast cells that are
in the extracellular fluid, so at the end it becomes mature to be
tropo-collagen. When we have the mature form of collagen, it will
attach and link with other collagen to form collagen fibrils then it
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forms fibres to be more condense and tough which can be seen
under light microscope).
Pro-collagen (immature) Tropo-collagen (mature) Fibrils
(more complex) Fibres (more condense and tough).
• Contents are rich in glycine (33%), proline (13%), and hydroxyl-
proline (9%).
(proline can be hydroxilated by enzyme called prolil-hydroxylase
and their co-factor is Vitamin C. If we don't have Vitamin C the
helical content of the collagen will be damaged. Also proline can
be modified by endoplasmic reticulum and in Golgi).
• Contains hydroxylysine (attachment of polysaccharides).
(Lysine can be hydroxylated in the collagen to become
hydroxylysine. The hydroxylysine is attachment for
polysaccharides).
• Cross-linking of chains via lysine and aldolysine via the action of
lysyl oxidase.
(The hydroxylysine can be oxidised by an enzyme called lysyl
oxidase which transfers it into aldehyde form "aldolysine", that
permits cross-linking in between collagen fibres and therefore we
will get very strong collagen).
Types of collagens:
Until now it is reported 28 types of collagens produced by very
different genes. Collagens are classified according to their types
(type I, type II… type 28)or according to their adopted shape.
• 30 collagen genes that form more than 25 different types of
collagen that resist tissue stretching.
• Types:
- Fibrillar collagens (I, II, III, V, XI) – they look like fibres.
- Fibril-associated collagens (IX, XII, XIV, XIX, XXI) – they are
fibers in their shape but they have very small piece of collagen
that help to associated with other collagen fibres, or with
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network of the collagen, or with basal laminea, or with other
proteins.
- Network-forming collagens – they are forming network form,
not fibre form.
- Anchoring fibrils.
- Transmembrane collagens.
All these types are named according to the adopted shape of collagen.
1-Types of fibrillar collagens:
NOTE: one of the most important properties of fibrillar is very
rigid in their structure- we can find it in bones.
• Type I (90% of the total collagen component in the body): most
connective tissues (Long, aligned in parallel to each other, and
rigid (fit to be in bone structure)).
• Type II: cartilage and vitreous humor. It also makes fibres but
they are aligned randomly unlike type 1, which permits it from
adopting the rod-like structure (important in cartilage). However
that does not mean it is not rigid.
FROM SLIDE:
Smaller in diameter than type I and oriented randomly in the
viscous proteoglycan matrix
Rigid macromolecules, but compressible (to resist large
deformations in shape and absorb shocks)
• Type III: extensible tissues (skin and lung)
• Type XI: cartilage
• Type XXIV: bone and cornea
• Type XXVII: eye, ear, and lung.
NOTE: Each type has tissue localization; therefore these specific
areas apply and help in their functions.
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Another note: the doctor said that he put these as examples for
you to know that each type of collagen has its own tissue
localization, so I don’t know if they are for memorization or not.
FROM SLIDE:
Assembly of fibrillar collagens
• After secretion, they assemble into collagen fibrils (thin
structures visible in electron micrographs).
• Collagen fibrils aggregate into collagen fibres (larger, cable-like
bundles seen in the light microscope).
• Fibril-associated collagens: Collagen IX and XII: link fibrils to one
another and to other ECM components
• Network-forming collagens: Types IV: basal laminae constituent
• Anchoring fibrils: Associate network-forming collagens to fibrillar
collagens
How collagen is synthesised?
- Firstly it is produced and modified in the endoplasmic
reticulum then modification is completed in the Golgi.
NOTE: N-terminus and C-terminus stay on collagen even after
the modification has happened. They will be secreted outside
of the cells while they are on immature collagen (C- and N-
terminuses makes the collagen called pro-collagen and after
they are cut outside the cell, they are called tropo-collagen).
N- & C- terminus will be on the collagen until forming the 3
polypeptide chains which are not wrapped around each other
well. When they reach this stage they will get out of the cell in
this form (3 polypeptide chains not wrapped around each
other well). Afterward, there are N- & C- peptidase enzyme
outside the cells called procollagen peptidases, their function is
to cut the terminuses. The presence of terminuses in the
collagen (at the ends on the rod like structures) prevents the
collagen from aggregating and attaching to each other inside
the cells. That is the reason we have collagen free within the
cells.
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- After cutting the terminuses, they will be able to crosslink each
other with aid of lysyloxidase enzyme (lysyloxidase enzyme is
secreted out from the cell "their localization is out of the cell, it
won't work inside the cell).
FROM SLIDE:
Synthesis of collagen
• Individual collagen polypeptide chains are synthesized into
the endoplasmic reticulum (ER) as pro-alpha chains
• In the ER, selected prolines and lysines are hydroxylated and
some of the hydroxylysines are glycosylated
• One alpha chain combines with two others forming
procollagen
• During or following exocytosis, extracellular enzymes, the
procollagen peptidases, remove the N-terminal and C-terminal
propeptides forming tropocollagen (or simply collagen)
• Excision of both propeptides allows the collagen molecules to
polymerize into normal fibrils in the extracellular space.
What makes other type of collagens adapt different forms and shapes?
We all know that the structure of the collagen is composed of 3
polypeptide chains wrapped around each other in a helical manner. This
structure is not applied at the whole length of the collagen molecule.
For example, some types are helical in their shape which adapts a rod-
like structure which gives the proteins its strength, and then they are
spanned by amino acids residues which separate the helical region and
followed by another helical region is separated by another amino acid
and so on. This makes the protein not to be rigid in the whole structure,
you will find some areas are less rigid and some flexible as well. Because
of this, there are different types and shapes of collagen.
FROM SLIDE:
Fibrillar vs. network-forming collagens
• The structure of network-forming collagens is interrupted by one or
two short non-helical domains, which makes the molecules more flexible
than fibrillar collagen molecules.
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• Network-forming collagens are not cleaved after secretion and
therefore retain their pro-peptides.
• Network-forming collagens do not aggregate with one another to form
fibrils in the extracellular space.
NOTE the Dr. skipped the information (below), I am not really sure if
we are going to be asked about them:
Premature assembly is inhibited
• The potentially catastrophic assembly of fibrils within the cell does not
occur because:
• The pro-peptides inhibit fibril formation
• Lysyl oxidase, which catalyzes formation of reactive aldehydes, is an
extracellular enzyme.
Collagen-related diseases:
Collagen is highly cross-linked in tissues where tensile strength is
required such as Achilles tendon. If cross-linking is inhibited, the tensile
strength of fibres is greatly reduced, collagenous tissues become fragile,
and structures tend to tear (skin, tendon, and blood vessels).
1-Scurvy:
It is an acquired disease (person can acquired it through the life)
It happens due to deficiency of Vitamin C it affects lysyloxidase &
prolilhydroxylase enzymes ( Vit C is important in hydroxylation process).
As a result of this:
-The helical formation of the collagen will be affected; therefore with
the temperature inside the body the helical formation is changed.
-lysyloxidase won't work properly; therefore the cross-linking won't be
proper, so the collagen will be very weak and arteries wall will be weak
as well, which makes the person to bleed very easily. Also, type 1 will be
affected as well leading to fractures.
From SLIDE:
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Deficiency of vitamin C results in insufficient formation of hydroxyl-
proline and, hence, poor synthesis of collagen, formation of unstable
triple helices.
Non-hydroxylated pro-collagen chains are then degraded within the cell.
Symptoms: skin and gum lesions & weak blood vessels.
2-Osteogenesis imperfect (OI) (Brittle-bone disease):
It is a defect in the formation of collagen caused by genetic
mutations.
It is rare disease caused by imperfect bone formation. A genetic disorder
of several forms that cause fragile, soft, brittle, and easily broken bones
There are four types of Osteogenesis Imperfecta designated as type I
through type IV
• Type I: the mildest form of the condition.
• Type II: the most severe form that results in death in uterus or shortly
after birth.
• Milder forms generate a severe crippling disease.
3 types out of 4 (type 1, 2, and 4) are autosomal dominant which you
need only one copy of the altered gene is sufficient to cause the
condition.
People who get this disease are more likely to be exposed to fractures,
due to weak collagen. These patients rely on the minerals deposited in
the bone to endure forces, and minerals alone are not enough to protect
the bone. So these minerals need support, which is given to them by
collagen. That is why defective collagen causes increases bone fractures.
Osteogenesis Imperfecta patients get blue sclera because the collagen
inside the eyes is very weak and is separated from each other, so the
venous blood behind the sclera appears and gives the patient the blue
appearance in the eyes.
FROM SLIDE:
Mutations of OI
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• Mutations in COL1A1 (it is mainly happened in collagen TYPE 1) and
COL1A2 genes that interfere with the assembly of type I collagen.
• Defective collagen weakens connective tissues, particularly bone,
resulting in the characteristic features of OI.
3-Chondrodysplasias:
It is a disease caused by mutations that affect chondrocytes function of
making type II collagen, characterized by abnormal cartilage (because
cartilage has a lot of type2 collagen), which leads to bone & joint
deformities.
The whole cartilage in the body will be affected, and any problem that is
associated with cartilage problems are possible to arise. Cartilage won't
be produced properly, type 1 or 2 collagen won't be functioning properly
and the associated carbohydrates in the cartilage won't move well.
Some patients can't walk well because their cartilage will be shortened
little bit, so they are not compressed well. Then over period of time we
will find degeneration (problems in movement of the whole body due to
the affect in the joints).
4-Ehlers-Danlos Syndrome:
It is a disease affecting the collagen formation. It is a heterogeneous
group of disorders that affect the skin, bones, blood vessels, and other
organs. Mutations happen in many types such as type I, III, or V collagen
and affect the synthesis of collagen. Also, it happens in processing
enzymes like pro-collagen N-peptidase, or lysyl-hydroxylase.
NOTE: it is a multifactorial disease. Strong collagen gives rigidity and
weaker collagen gives more flexibility.
Due to weakness of the collagen it will affect the joints (which are
covered by tendons) and make them very flexible. Also, the skin will be
hyper-extensible.
FROM SLIDE:
The signs and symptoms vary from mild to life-threatening.
All result from defects in collagen synthesis and/or processing.
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Major manifestations are skin fragility & hyper-extensibility & joint
hypermobility.
• The most clinically important mutations are found in the gene of type
III collagen.
• Since type III collagen is a major component of arteries, mutations
affecting type III collagen result in fragile blood vessels.
Other symptoms include stretchy skin & hypermobile joints.
..................
Now we are going to talk about the second most important protein that
is secreted in the extracellular matrix...
2-Elastin (it is not a glycosylated protein):
- The main component of elastic fibres is elastin. It is highly
hydrophobic and rich in proline and glycine. It contains some
hydroxyproline, but no hydroxylysine.
Formation of the elastic network:
1- Secretion of tropo-elastin (same as tropo-collagen).
2- Assembly into elastic fibers.
3- The cross-linking via lysines (through lysyloxidase).
Elastin structure:
• The elastin protein is composed largely of two types of short segments
that alternate along the polypeptide chain:
• Hydrophobic segments, which are responsible for the
elastic properties of the molecule; and
• Alanine- and lysine-rich a-helical segments, which form
cross-links between adjacent molecules.
Function of elastic fibre:
• Elastin is the dominant Extracellular matrix protein in arteries.
• The normal elasticity of an artery restrains the proliferation of these
cells.
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• Abnormal or deficiency of the elastin results in excessive proliferation
of smooth muscle cells in arterial walls and narrowing of the aorta.
Elastin Tissue:
Elastin tissue is covered with a sheath of microfibrils (they are fibers that
have distinct glycoproteins, including the large glycoprotein fibrillin,
which binds to elastin and is essential for the integrity of elastic fibers).
When we have a problem in fibrillin such as mutation it will cause a
disease called Marfan's Syndrome.
Elastin-related disease:
1-Marfan's Syndrome:
This disease is happened due to mutated in fibrillin (so when the sheath
is not covering the elastin, the tissue will be very weak. For example, the
elasticity of the arteries will be decreased due to loss of the microfibrils
sheath so these arteries will not be able withstand the pressure that
comes from the heart, this leads to rupture in the main arteries)
Clinical features (FROM SLIDE):
A tall, thin build; Long arms, legs, fingers, and toes and flexible joints;
Scoliosis, or curvature of the spine; A chest that sinks in or sticks out;
Crowded teeth; Flat feet
(This piece of information was in the slide, but not mentioned by the
Dr.).
2-Emphysema (destructive lung disease):
Is a dysfunctional α1- antitrypsin deficiency that leads to increased
activity of elastase and it is characterized by destruction of alveolar
walls. α1-antitrypsin main function is to inhibit the elastase enzyme
which breaks down elastin, so if there is a deficiency in this enzyme, the
elastase will not be inhibited and it will destroy elastin in the alveolar
walls. A mutation happens by changing Lysine to Glutamic acid which
causes misfolding, forming aggregates & block of ER export.
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Cigarette smoking also inactivates α1-antitrypsin by oxidizing essential
Met (359) residue, decreasing the enzyme activity by a factor of 2000
folds.
..................................................
Now will move to talk about Extracellular Matrix Polysaccharides...
Glycosaminoglycans (GAGs) (it is the main component in the
extracellular matrix):
It is polysaccharides of repeated disaccharides. One is either N-
acetylglucosamine or N-acetylgalactosamine (amino sugar- which is
modifies by amino group). The other is usually acidic and it is either
glucuronic acid or iduronic acid. The latter two are highly negatively
charged and are sulphated most of the time (not always sulphated). One
of the types of glycosaminoglycans is proteoglycans which have
carbohydrates as their main content.
Most of the Glycosaminoglycans exist as proteoglycans (Cell surface
proteins with either transmembrane domains (syndecans) or GPI
anchors (glycipans) interacting with integrins).
1. Hyaluronan acid (not a proteoglycan): anionic, non-sulphated
GAG distributed widely throughout connective, epithelial, and
neural tissues.
Aggregan as a molecule (an example of them):
It is a large proteoglycan (core protein) consisting of more than
100 chondroitin sulfate chains joined to a core protein. Multiple
aggregan molecules bind to long chains of hyaluronan that
become trapped in the collagen network, forming large
complexes in the Extracellular matrix of cartilage mainly.
Why Glycosaminoglycans is the main thing in the extracellular matrix?
Because their negativity is very high due to the huge number of the
repeating disaccharides (one of them amino sugar and the other is
sulphated acidic in nature most of the time), plus the negativity that we
find in the carbohydrates adding the sugars. Also, the shapes that they
adopt specifically the branches that it has, gives you high connectivity in
extracellular matrix higher than any carbohydrates.
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Now will move talking ABOUT Extracellular Adhesion proteins:
They link matrix proteins with one another and to the surfaces of cells.
The main protein is Fibronectin (secreted by fibroblasts):
It is the principal adhesion protein of connective tissues.
It is a dimeric glycoprotein that is cross-linked into fibrils by disulfide
bonds. It is secreted by the fibroblasts and binds to collagen and GAGs
that are already found in the extracellular matrix from one side. From
the other side they bind to transmembrane proteins (integrins) in the
plasma membranes of cells.
That’s how the network in the extracellular matrix is made. Firstly,
Collagen is secreted to give some strength to the extracellular matrix
which is not linked to anything. The GAGs are present and most of them
are attached to proteins (in a proteoglycan structures). Then helical
proteins attach to collagen in one side, GAGs in other side, and attach to
the cell as well through the integrin protein within the plasma
membrane (so the network is completed then).
Other main protein is laminin:
It found in basal laminae. It is T-shaped hetero-trimer with binding sites
for cell surface receptors, type IV collagen and perlecan. It has 3 chains:
A chain is in the centre and other 2 chains are B1 and B2 wrapped
around each other over the A chain.
Laminins are tightly associated with another adhesion protein, called
nidogen, which also binds to type IV collagen, all of which form cross-
linked networks in the basal lamina.
Cell-matrix interactions: Role of integrins
Any protein in the extracellular matrix will mediate its effect over the
cell through integrins. They are a dimer of 2 chains: α & β. The integrins
mediates the signals that are coming from outside the cell or going
outside from the cell (from nucleus to the cell membrane, extracellular
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matrix and so one). It binds to the cytoskeleton from inside, and binds to
the extracellular matrix from outside. Integrins work as dimers.
They are inactive at first then it will become active upon binding to
other molecules.
Integrins are a family of transmembrane heterodimers (α & β). They
anchor the cytoskeleton at focal adhesions and hemi-desmosomes.
What is focal adhesion?
When we talked about the cell and how it moves, we came across the
actin filaments they are polymerised in specific area in the cell. Why
does actin polymerize in that specific area? Because actin-binding
proteins are concentrated in that area. Why are actin-binding proteins
concentrated in that area? Because of signals that come from outside
the cell. These signals affect integrins by activating them, so integrins
localize at that area and actin-binding proteins bind to them. So this
complex recruits actin and ‘tells’ actin that it needs to polymerize in this
area. So actin polymerizes while it is bound to the actin-binding protein.
So focal adhesion is an area with collection of proteins.
FROM SLIDE: Development of focal adhesion:
1. Activation of integrin and binding to Extracellular matrix.
2. Recruitment of additional integrins forming focal complex.
3. Development of focal adhesion.
........................................................................
Cell to cell interaction
Cells interact with each other by junctions, for example by adherence
junction (like desmosomes), how do they interact by these junctions? ...
So, there are Actin filaments surround the cell from inside. They
communicate with the outer signals by attaching to cadheren ( a
transmembrane protein); and the cadheren is attached with another
cadheren from other cell and so on. So the interaction happens in this
way.
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Cell interaction devided into 2 types:
1- Homophilic interaction :(when the proteins within between the
cells are the same type we call this type as homophilic)
2- Heterophilic interaction : (when the proteins in between the cells
are different types we call this type of interaction is heterophilic)
We will talk about 2 of them ....
Selectins:
They are different types (all types called a group of selectins). It has a
role in leukocyte-endothelial cell interaction "attachment and squeeze
of leukocyte":
1-Rolling of leukocytes is mediated by selectin (L-selectin, P-selectin, and
E-selectin are attached to leukocyte, and then they facilitate the
attachment to the endothelium) .
2-Prior to invading a blood vessel, integrins are activated (the integren is
activated when the selectens and leukocyte are connected with each
other by sending signal to integren)
3-firm attachment allows invasion (when integren is activated it will
send signals to actine filaments to grow, then the leukocyte will squeeze
and attachment will occur).
Cadherins:
• Selective adhesion between embryonic cells and formation of stable
junctions between cells in tissues.
Types:
- Classical tissue cadherins
- E-cadherin: epithelial cells
- N-cadherin: neural cells
- P-cadherin: placental cells
- Desmosomal cadherins
- 7-Transmembrane cadherins
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As you can see there are lots information in this sheet I advise you to go
back to the slide and refer to the pictures and figures there... Sorry for
any mistake.
Best of luck...