immuno . lec 9
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Wednesday, 13/7/2011
Ziad Al-Nasser
Saad Aldeen Mohammed
Lymphocyte activation
and hematopoiesis
9
30
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The Doctor said that we gonna change seat numbers after the first exam
which is on Saturday 23/7/2011. And it will be at 8:15 AM.
Yesterday we were talking about antigen presentation , the role of MHC
antigens 1 and 2 in the presentation process ,the antigen presenting cells
"APCs" (macrophages, B-cells ,dendritic cells) ,MHC restriction phenomena ,
and we said that : what really determines whether class 1 or 2 MHC will be
involved ,whether T-helper or T- cytotoxic cells will be involved, is by the
way or mode of trafficking of the antigen through the APC .
And we said that we have 2 pathways:
1-Intracellular.
2- Extracellular:which could be: endocytosis(which involves toxins and
vaccines) or phagocytosis (which involves bacteria) . And we said that the
outcome of that is the production of antibodies in the case of endocytosis
or activating macrophages to get rid of intracellular bacteria such as
"mycobacterium tuberculosis".
And we said how antigen when taken in, it will be processed and cut by
proteases into small peptides ,and how it will bind to class 2 which is
produced in the endoplasmic reticulum (ER).
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And also we said how the class 2 will be covered by the invariant chain and
then it will pass though the Golgi apparatus ,and how the antigen will bind
to the MHC 2 in the acidic vesicle and then it will be expressed on the
plasma membrane so the T-helper cells that has T-cell receptor and CD4
which is really very important can come and bind with its ligand (the
antigen +MHC 2).
And then we talked about the other pathway which is the cytosolic where it
involves the intracellular viruses, tumors as well where we require
cytotoxic T cells to kill the virally infected cells or tumor cell.
So the virus will be processed and cut to small pieces and these pieces
antigen have to be passed through the ER where MHC 1 antigens are
located by transporter enzymes (ex:- TAP1 and TAP 2 ).
Then it will bind to MHC 1 antigens and transported to be expressed on the
plasma membrane to be recognized by CD8 cells .
We have talked also about evasive organisms like herpes viruses, adeno
viruses which can manipulate these MHC 1 molecules so they are successful
pathogens.
We talked about DNA vaccines and so when we need to develop acquired
immune response and memory cells related to T- cytotoxic cells we
introduce the DNA of the organism and it will be processed by the
intracellular pathway (as if it has developed within the cell) and T-cytotoxic
cells will be activated .
And we said that this is the major arm for the activation of T-helper and T-
cytotoxic cells by MHC restriction but we have other important factors we
will talk about which are the cytokines which will be produced by the APCs.
Then we started talking about lymphocyte activation, and we said that we
have to understand the biological and biochemical events that will take
place in the activation process to understand why immune suppression
could take place, how can we induce an immune suppression, mechanisms
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of action of immune suppressive drugs that we need for successful
transplantation.
Transplantation will not be successful until we suppress the immune
system.
So we said that the activation of T or B cell requires a stimulus, which is the
binding of antigenic determinant with the receptor on the cell. It's not just
binding, its a cross linking that will take place which is very important.
So in this antigen, as long as binding and cross linking are taking place then
response will take place and if you want to stop that action then antigen
has to be cleared or that cell has to be going into apoptosis and so on.
After binding and cross linking you need a signal to be developed ,and this
signal will be initiated by the clustering process ,and seconds after that,
signal has to pass through ,and we call that (transduction) ,and the signal
passes through the activation of certain proteins that will be present on
these molecules (ITAMS).
so these proteins will get phosphorylated by other proteins and become
activated to attract other proteins like a cascade like the complement and
the blood clotting.
Remember that a cascade is a pathway that is every single step leads to the
next step till we reach the final result.
Finally the activated proteins that will develop will start biochemical
pathways within the cell to amplify the signal so the outcome will be a
cellular response ,and the biochemical reactions will end up in the
activation oftranscription factors.
So transcription and translation protein are going to develop ,and these
proteins will activate the cells, so they will multiply ,differentiate and
produce cytokines.
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(Remember that the most important cytokine is IL-2 which is
differentiation factor produced by T-helper cells, and any suppression of
this factor will have immune suppressive outcome.
If and only if you have missing enzymes in the biochemical pathways thenthe patient is going to have immune suppression_ theSCID syndrome
where you have deficiency in enzymes that could be important in gene
rearrangement like rag1 and rag2 .
Also if there is deficiency in one of the enzymes required for activation
process the same thing will happen).
Sooooo the process starts here by an antigen recognition starting the
response ,then the signal has to be transduced through the activated
(phosphorylated) proteins, and then we will have activation ofbiochemical
pathways in a cascade pattern ,and the outcome of the biochemical
pathways is the activation of gene transcription so differentiation
,proliferation and production of
cytokines will take place.
B-cell Activation:
Lets now look in more details at B
cell (figure 11.1) :
* The antigen has to bind to a
monomer ofIgM ,and of course
there must be a cross-linking ofIgMs
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,one will not be enough.(BCRs are distributed all over the cell surface)
* we have the invariant chains , we have 2: Ig (alfa),Ig (beta).They are
accessory molecules to the BCR that are required for the signal to pass
through , so if they are not present the signal cannot pass through. Theseinvariant chains have specialized proteins (ITAMS) by which the signal is
transduced. By the way (ITAMS) are present in many other cells like T cells.
* the final outcome of the activation and the biochemical pathways in B-
cells and T-cells are the same, the difference is just in the nature of the
proteins that act as messenger molecules for signal transduction events.
T-cell Activation:
If you look at the T-cells (figure 11.2):
* We have the TCR that we have talked
about.
*we have CD3 which is required for the
passage of the signal. Here we have 4
peptide chains on the surface
(gama,delta,epsilon,epsilon) .
* On the other side we have another 2
chains ( zeta ,zeta) also required for the
signal to pass through by the means of
ITAMS , and here _in zeta chains _we
have multiple ITAMS which act as
proteins where the tyrosine is
phosphorylated.
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* phosphorylation of these ITAMS is seem to be very important ,,,but how
they are phosphorylated????
The initiation of the B and the T cell activation starts by thecross linking of
the receptor molecules by multimeric antigen ,and as we said that just one
binding is not enough ,the antigen must react with more than one receptor
at the same time.
B - Cell receptor:
We have membrane immunoglobulin which is a monomer of IgM act as a
receptor. Also we require an invariant protein chain Igand Ig and why we
require those??
>for mIg expression and signal transduction.
>for the ITAM immunoreceptor tyrosine based activation motifs the
protein that are needed to signal to go through .
T - cell receptor:
Its composed of two polypeptide chain or .CD3 invariant protein chains ,
four polypeptide chains two , , and two on the other side we called
them . All have the ITAMs.
the accessory molecule around the receptor are needed because of the ITAMs
that get phosphorylated .
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initiation of B-cell activation:
cross linkinng of receptor molecules by multimeric antigens in B-cells . we have
many B-cell receptor distributed all over the B-cell , and when antigen binding it
will be cross-linked between many receptor.
the second signal is needed we call it co-stimulatory signal .
intitiation of T-cell activation:
T-cell receptor occupation by an antigen on APC presented by class II MHC .Second co-stimulatory signal provided by by APC which could be a cytokine .
Accessory molecule : CD 11a , lymphocyte function antigen (LFA-1) binds to
CD54 or ICAM-1 as ligand and integrin .
So B-cell crosslinking and T-cell binding ag with MHC .
Co-receptor molecules in B-cell activation:
the antigen has to bind more than one receptor (cross-linked ).
Start with invariant chain protein Ig and Ig.
We have accessory molecules on the side :
CD81. CD19 : which only present in B-cell so if I want to target B-cell I can target
the CD19.
CD3 in B and T cellsbind to antigen cross linking in B cell.
CD21 we call it complement receptor 2 , which the same receptor forEBV viruses H type viruseswhich cause infectious mononucleosis . but
here CD21 can bind to CD3 when complement activated .
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All of them they have to be phosphrylated on cytoplasmic tails where the
ITAMs located .
Co- receptor molecules in T cell
activation
CD4 (co-receptor chain) orCD8(co-receptor disulfide
linked dimer) .
CD4 and CD8 areimmunoglobulin supergene
family and they have tails .
CD4 binds to class II MHC. CD8 binds to class I MHC . A protein ofSRCfamily we call it
LCK has to bind to the CD4
,when this is taking place the
CD4 can come closer to class II
MHC.
The same thing here when theLCK binds to CD8 it will come closer toclass I MHC for the activation to take place .
The LCK is associated with TCR if and only ifCD4 or CD8 bind to MHC .Soif genetically you are missing SRC or LCK protein then the process will
stop and you will be immune suppressed .
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Signaling events:PKTs protein tyrosine kinases
and PTPs protein tyrosine
phosphatases as well LYN (in B-cell) and LCK (in T-cell)
phosphorylate the ITAMs .
In B-cell >>When theITAMs in Ig and Ig get
phosphorylated the become
active and bind another protein
called SYk .
Then the SYK will getactivated by LYN .
The SYK will initiate thebiochemichal pathway .
In T-cell >>when the ITAMs inCD3 and chain phosphorylated by
LCK which carried in the tail of CD4 ,
they become active and bind protein
called ZAP 70 .
Then ZAP 70 will getphosphorylated by LCK .
The ZAP 70 will initiate thebiochemichal pathway.
So its all connected as a cascade type
of mechanism of action .
So now we have two protein as an ultimate outcome are going to form the
SYK for B-cell and the ZAP 70 for T-cell .
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Amplification through signaling pathways:
Same pathway inB and T cells
Phosphorylationand activation of
phospholipase C
(PLC), this is triggered
by syk or ZAP70, and
the activation of
phospholipase c will
start the biochemichal
pathways 3 pathways
formation of what we call diacylglycerol and protein kinase C andphosphorylation of several other protein and activating of transcription
factor include NK-B.
formation of inositol 1,4,5 - triphosphate IP3 which lead to release ofstored Ca and level of Ca will increase and this is called influx of Ca in to the
cell and this pathway will activate a protein called calcineurin and the
calcineurin will go and activate transcription factors with in the T helper
cell.
In this step we can use immunosuppressive drugs , like cyclosporin orthe tacrolimus ( these two are the most common used in transplantation
immunology ), they interfere with the production of calcineurin, when you
give these drugs they form the protein or the immunophilins >>> those
interfere with the function of calcineurin >>> so calcineurin cannot be
formed.
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The activation of ZAP 70 or SYK will activate the third pathway whichcalled GTP binding protein or RAS ,Ras activates MAP kinases that
activates transcription factors like AP-1 .
If as I said, any of those enzymes are missing or any of those proteins that
we have like Lyn, Lck, then the patient will be immune-suppressed; and
for example if a patient dont have a protein like in a gamma globulinemia
so patients B-cells will not have the ability to produce immunoglobulin, so
this patient will have immune deficiency, so we will not have humeral
immune response and the only way to help this patient is to give him
immunoglobulins or to treat him with the gene therapy and replace the
infected gene for that particular protein.
Response:
>This is what I was talking about the production of the nuclear factor (NF-
AT, NF-kB, and AP-1) acts on many lymphocytes genes to activate their
transcription.
>This prepare B-cells for proliferation and differentiation and T-cells get
prepared for enhance expression of cytokines as IL-2 IL-2 the most
important.
>Immunosuppressive drugs can modulate this response like cyclosporine &
tacrolimus. You should know these two drugs and there function on
Calcineurin
So this is simple the activation of T or B lymphocytes.
Now we gonna continue the process we have talk about almost all the
adaptive Immune response T-cells, B-cells, antigen recognition molecules
and interactions
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Now we will start talking about the factory; how those lymphoid cells in
general in the second line of defense i.e. the phagocytic cells, the innate,
the adaptive how they develop and we have a comprehensive look into
how those cells co-operate together as one team; when the innate immune
system finish the job it could stop here but if it couldnt then how the
adaptive immune response is going to take place.
Hematopoiesis: CH#12
So these cells we are going to talk about mainly they develop in the bone
marrow and the originating cells we call them the stem cells, and those
stem cells then can differentiate into the main types of cells that we keep
talking about; either the lymphoid cells B, T cells , or the myeloid themacrophages, neutrophils, granulocytes, eosinophils, basophiles, platelets
all of those come from the bone marrow. And these cells develop in the
yolk sack and then they go to the bone marrow after that. So the origin of
the stem cells is the bone marrow
We can get stem cells from the fetal blood i.e. cord blood. And these stem
cells they can reshape and they can provide us with all the different cells
that we need.
How to get these stem cells and how to separate them from others?
How can we separate stem cells, take them, identify, use them to give us
the things that we need?
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By flow cytometry; by the FACS machine you remember florescent
activating cell sorter, those stem cells have special markers that they do not
share with others. One of them is called the CD34; so you can target the
CD34 then u can separate them and use them in therapy.
So the stem cells in order to differentiate and go in many stages, they
require growth factors and cytokines as well, and these growth factors,
colony stimulating factor CSF and cytokines the come from the Stroma
interconnecting tissue of the bone marrow.
And these CSFs you can use them in the lab as well; you can harvest them
and use them as therapy E.g. if a patient bone marrow is suppressed and
cant produce stem cells then I can use CSF to activate the bone marrow to
produce those cells and develop into progenitor cells and then I keep doing
those.
So in each stage we have those growth factors and cytokines that will be
used and finally both cells will develop please look back to the last figure
which is monocytes macrophages, mature cells, lymphocytes,
granulocytes, erythrocytes and platelets and each line of these will require
some of the growth factors and activating factors and so on.
So the process of Hematopoiesis, what do we mean be that?
The process whereby all blood cells are formed i.e. white blood cells,red blood cells.
How those will develop?
Remember the origin is the human bone marrow which is the sourcemeans we activate the bone marrow and all of these cells will come out of
the bone marrow.
The bone marrow is the main factory _remember that_, any suppression
for the bone marrow by tumors for example: leukemia, lymphoma or
use of drugs like chloramphenicol can suppress the bone marrow then
you will be severely immune compromised and you are going to have
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pancytopenia; all the blood elements the platelets, red blood cells,
white blood cells they are going to be suppressed.
Leukocytes Lymphoid + Myeloid.Then Mr. Ali interrupted the lecture and the Doctor said changing places
makes him feel that there is something illegal going on.
And then he added that the most important thing for medical student is to
be honest until improve otherwise role of thumbLOL.
Any try to occupy the seat of others is a crime based on students laws and
regulations. BTW I was one of the criminal in this lecture in order to do this
tafree3 :P .
Back to the lecture:
So we have lymphoid line that will lead to T cells, B cells and another sub of
that is the natural killer cells line. And the myeloid line that gives u
macrophages and the granulocytes and so on.
Then well be talking about lymphoid tissues and organs and cells of the
immune system and you will see how we can classify those into:
>primary organs which is the main factory where the cells will originate
they will be trained and they will get the specific function of a B cell or T
cell; and that is why we call it B cells from the Bone marrow or the bursa of
FabriciusInbirds, the bursa of Fabricius is the site ofhematopoiesis, a specialized
organthat is necessary forB cell development in birds.Mammalsgenerally do not
have an equivalent organ; thebone marrowis often both the site of hematopoiesis
and B cell development.wiki and T cells for the thymus gland. When the cells
go from the bone marrow to the thymus gland they will be trained to
recognize self from non-self and then they will go to:
>secondary lymphoid organs as what we call virgin T lymphocytes ready
for exposure to foreign antigen and then we have the immune response
where the macrophages taking place.
http://en.wikipedia.org/wiki/Birdhttp://en.wikipedia.org/wiki/Birdhttp://en.wikipedia.org/wiki/Birdhttp://en.wikipedia.org/wiki/Hematopoiesishttp://en.wikipedia.org/wiki/Hematopoiesishttp://en.wikipedia.org/wiki/Hematopoiesishttp://en.wikipedia.org/wiki/Organ_%28anatomy%29http://en.wikipedia.org/wiki/Organ_%28anatomy%29http://en.wikipedia.org/wiki/B_cellhttp://en.wikipedia.org/wiki/B_cellhttp://en.wikipedia.org/wiki/B_cellhttp://en.wikipedia.org/wiki/Mammalhttp://en.wikipedia.org/wiki/Mammalhttp://en.wikipedia.org/wiki/Mammalhttp://en.wikipedia.org/wiki/Bone_marrowhttp://en.wikipedia.org/wiki/Bone_marrowhttp://en.wikipedia.org/wiki/Bone_marrowhttp://en.wikipedia.org/wiki/Bone_marrowhttp://en.wikipedia.org/wiki/Mammalhttp://en.wikipedia.org/wiki/B_cellhttp://en.wikipedia.org/wiki/Organ_%28anatomy%29http://en.wikipedia.org/wiki/Hematopoiesishttp://en.wikipedia.org/wiki/Bird -
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Then one of the guys changed his place because someone was sitting on his
seat and the Doctor said it is complicated issue So we will see that how the mother cell originates in the bone marrow B
or T cells; the mother cell or the originator cell of the B cell originate in thebone marrow, lives in the bone marrow, trained in the bone marrow,
differentiate self from non-self also in the bone marrow and then it comes
out to the circulation going to the secondary lymphoid organs, and the
same thing for the T lymphocytes and those which go to the secondary
lymphoid organs where they will be exposed to the antigen.
And then you will see that the B and the T cells they keep circulating
among the secondary lymphoid organs.
And the secondary lymphoid organs are the spleen major secondary
lymphoid organ, lymph nodes all over our body, the adenoids and the
mucosa associated lymphoid tissue MALT.
These lymphoid cells have a target; there main target is to get exposed to
the antigen, if they managed to achieve that role then they will survive,
memory cells will develop and they will stay in our body for a long period of
time.
But if they dont do this function then they will commit suicidal. So all of the
lymphoid cells that come out of the bone marrow and the thymus gland
they will be circulating and trafficking among the secondary lymphoid
organs hopefully that they meat there counter antigen, if they do not then
they will disappear and they will die, and we will tell you more about
Lymphocytes recirculation or trafficking or what we call homing.
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In order to achieve that we will have Adhesion molecules that are present
on the cells that will go through the secondary lymphoid organ, and you
can monitor that through micro-cameras and you can see how these cells
adhere in the mucosal lining and then they can bass through the
endothelial cells into the assigned area of the secondary lymphoid organs
So her in the bone marrow look to the figure we have stem cells and
those stem cells they will keep multiplying and they require activation
molecules, growth factors. And those will go under the influence of the
colony stimulating factors and cytokines in what we call lymphoid and
myeloid progenitor.
progenitor means an intermediate stage cell from HayatStem cells progenitor cells differentiated cells
During that development certain markers are going to be added and some
will be blocks, and we will talk about these, and then they will be
differentiated those from the mother cells into the common progenitor into
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the B cell, the T cell and the myeloid progenitor that gives you the rest of
them development of the erythrocyte and the megakaryocyte and the
granulocytes
The Doctor stopped here and he will continue the next lecture.
Finally
THE END
I wanna Give very big thanks for:
EHAB DEEP: thanks a million brother, actually u did most of the work I
wont forget that
RAF@ TWALBEH: really no words can give you your position in my heart.
MOHAMMAD MEQDADI: I dont wanna thank you because you dont need
thanks :P
And also a very warm thanks for all my friends and to all 2009 batch.
Good luck all In your exams and in your entire life
DrSkodeh
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