Thursday, Hussein Ziad Al25 10 Hematopoies 14/7/2011 Qasim Nasser is

About the exam :

Just to keep reminding you of the first midterm exam on the twenty third of July, and the material that decided to be included will be determined few days before the exam and then we will see how we are going to synchronize the dental as well usually we do have a separate exam but we will see doctor Ammar , we could have a unitized exam and we will talk about that when time comes so no problem .

Review for chapter # 11

Yesterday we talked about the T and B cells activation , all the stages that the B and T cells undergo through and how a stimulus has to bind to the B and T cell receptor, and how a cross-linking is

going to take place ,and how we have accessory molecules that are needed for the passage of signal. for the B cell, the Ig and Ig that have ITAMs which are very important ,and we talked about some of the molecules that are indicated on the surface of the B-cell that you can target and define as a B cell like the:- CD19, if you remember and CD21 which can bind to one of the complement components, and how those can help in the activation process. and also we talked about the T cell accessory molecules ,CD 3 ,Ig, IgDelta- and Ig ,and how those they have the ITAMs that are required here for the cellular signal production, and then how the receptors when occupied, how the signal passes and then how proteins like Lyn for example in the B cells or Lck in the T cells that they have to come with the CD4 or CD8 in the T cells to bind to the ITAMSs and get activated ,and then they will activate the Syk here is the ultimate result band in the T cell ZAP-70 .????? and how those intiate the amplification in certain biochemical pathways that we have talked about and the PLC-GAMMA is going to be activated , the diacylglycerol pathway and the influx of calcium and then the calcineurin that is going to be activated .there is a previous step we call it the immunophillin and this is the stage where it binds to the cyclosporine and preventing the formation of the calcineurin and also the GTP-pathway . A final outcome, you will see that the activation and production of transcription factors ,and these transcription factors initiate the proliferation and differentiation and the cytokines production. just simply how these the B cells and T cells can be activated and if any of those enzymes is missing like the x-linked sever combine immune deficiency when one of the enzymes for PTKs of the proteins is missing then the activation will be stopped and the antibodies can't be produced so you need to provide that patient with immunoglobulins or you can do the gene therapy and insert that gene that is going to code for that protein that is needed for the T or the B cell activation .

Chapter 12 (Hematopoiesis) :

And then we start talking about the hematopoiesis : the process,how blood cells in general are formed, and we said that we have the main factory which is the bone marrow, and if the bone marrow is shut down for one reason or another ,then red blood cells precursors , platelets precursors , white blood cells precursors myeloid and lymphoid are not going to develop, so the bone marrow is the most important factory that will provide all the blood components .and when it's shut down, we have a condition we call it pancytopenia and pancytopenia involves all the cells that are going to be shut down . and the first cell that will develop there and give you a mother cell we call it stem cell .the stem cell is self-generating and it has markers that are not present on other cells. we said CD34 and these cells get modified into more advanced cells we call them progenitor cells and in mature cells some of these markers will be added and deleted as you will see. So the stem cell is a very important cell that we can use in order to get the progenitor cells and mature cells , and we can take them as I've said from the bone marrow and cord blood and if the patient's

bone marrow is shut down for one reason or another then we can expose this patient to radiotherapy , cytotoxic drugs and then we can substitute with bone marrow by transplantation. Or we can add stem cells and that they come from the bone marrow and cord blood differ from the embryonic stem cells that they can develop also other organs and so on. And the challenge here is what is the triggering factor that is needed to make these stem cells differentiate into the cell line that we want it to be. this is the look into the future but now for the blood and blood component we are concerned with the bone marrow stem cells and the cord blood stem cells as well .

So the hematopoietic stem cells set in self-replicate and we can have many of those and then they can develop into myelo-lymphoid progenitor , common lymphoid progenitor or common myeloid

progenitor , the lymphoid will give you the B and T cells and the myeloid will give you the erythrocyte , the platelets , the granulocytes and the monocytes and what really make those progenitor cells differentiate into that, line you need a cytokine that is going to initiate that part of process ,for example the red blood cells here need erythropoietin that comes from the tissue as you know and so on here IL-7 and colony stimulating factor CSF , granulocyte-monocyte CSF , granulocyte CSF alone to produce granulocyte or monocyte CSF you will get monocytes , macrophages and so on . so we talked about that if you remember the last time about the blood cell lineages and the hematopoietic stem cells develop the CD34 they don't have a T cell receptor , the CD3 as well , they don't have the CD 19 on the B cells so you can see how those markers they will be added up later at certain stage. if you see CD19 or CD3 on stem cell that stem cell must be abnormal and you can sometimes consider that as a marker of abnormality or malignancy and so on . we talked about those markers that are present on early cells and you could see them later in the circulation where they are not supposed to be in the circulation and these are markers of malignancy and this is really very important for stem cells, we look only for the CD34 and we use that as a target to separate the stem cells and to count their number with the flow cytometry and the "FACS" machine that we talked about . remember that the hematopoietic stem cells in the bone marrow originally come from the liver , in the yolk sac start and then the liver during embryogenesis and then they shift into the bone marrow where they start to give you the others under the influence of growth factors as we said like colony stimulating factors, and these colony stimulating factors can induce the production of these progenitor cells as I said GM , G or M will give you granulocytes and monocytes , granulocytes or monocytes alone and so on . so they induce the differentiation and these cytokines sometimes are produced from the T helper cell , the T helper cell when gets

stimulated like for example IL-3 can act on the bone marrow and stimulates the bone marrow to start the process of hematopoiesis of the neutrophils line for example , macrophages line and so on . remember the CSF and we could use those as medicine and we genetically engineer those and you can use them as inducers when the patient is having neutropenia for example and you will increase the number of neutrophils , you can give that patient G-CSF for example so the bone marrow starts to make lineage that leads to production of neutrophils and so on .

so this is the follow up : * we start with hematopoietic stem cells in the bone marrow and proginator cells still in the bone marrow .

Here for B cells and some of those can go to the thymus gland and give you T cells , they start to mature and to be trained to perform specific function and then they come out as mature stem cells so mature cells as B cells for example ,they come out of the bone marrow but you don't see mature B cells in the bone marrow ,we see those in the circulation and the same thing here progenitor cells when they enter the thymus gland they are immature and they develop in the thymus gland but when they come out of the thymus gland they come out as mature T cells .so what really stamps the cell as B or T is the bone marrow for B and thymus for T .

Mature ready cells we call them virgin lymphocytes, they go to the secondary lymphid organs to perform the function that they were designed to do and the rest the neutrophils , the eosinophils for example , the basophils , the monocytes , the macrophages the same thing they come out of the bone marrow and when they are in the circulation they are mature or in the tissure for example as mast cells and so on .

So remember this which is really very important , the differentiation for example from one stage into the other requires an activating or growth factor and here you can see the IL-7 rule that usually secretes from the stromal variants in the bone marrow and how this is important if IL-7 is missing or receptors for IL-7 is missing that the patient is going to have sever combined immunodeficiency as simple as that .

so this is the common lymphoid proginator where here it will differentiate to either T cell or B cell , in the bone marrow B cell , in the thymus T cell which requires IL-7 . and what really makes that proginator to go and differentiate into natural killer cell is non specific and the receptor CD14 and CD16 as an immunoglobulin receptor is not known here but remember that the specific cells that we are talking about here going to the thymus gland and give the T cells whether T helper or T cytotoxic , whether T helper 1 or T helper 2 , when I say T helper 1 and T helper 2 it depends on the cytokines that are going to be produced according to the need whether these cytokines are needed to the production of antibodies or the cytokines are needed for the production of cellmediated immunity , this is the determination here to be noticed between T helper 1 and T helper 2 and you know here the T cytotoxic cells how they are going to operate in killing the virally infected cells and the tumor cells as will like those of natural killer cells but the natural killer cells they differ in that they don't have T cell receptor , don't have CD3 and non-specific but they look like a lymphocyte , we call them large granular lymphocyte which is the old terminology for them.

so this slide I want you to see of differentiation and the rule of IL's in that and its -IL- receptor and how important is to further go and produce the mature virgin lymphocyte so the development of the lymphoid cells of the T cell line occurs in the thymus gland , so if the thymus gland is missing you're not going to have the T cells as simple as that . the T cell-mediated immunity is going to be affected there will be no more T helper cells and so on . and we will talk about that when we talk about the significance of the thymus gland and you will see the thymus gland is mature when we are born and has done all the work and it keeps working till we become adults and then it will get necrosis and disappear , so if we remove the thymus gland nothing is going to happen because it has almost fulfill all its job while, if you remove it while the baby still inutero ,we will have immune deficiency and we have congenital anomaly that the thymus gland isn't developed here like the syndrome we call it DiGeorge syndrome where the pharyngeal pouch #3 is going to be missing then the patient is going to have severe cell-mediated immune deficiency , we will talk about that later . lymphoid progenitor and the rule of IL-7 in the differentiation and the lymphocytes is going to the thymus gland mature cells will come out of the thymus gland we call them virgin cells because they haven't been exposed to the antigens yet . lymphoid progenitors are in the bone marrow and mature B cells go to the circulation , you don't see mature B cells in the bone marrow , the same thing for natural killer cells they come out of the bone marrow and what really makes this differentiation into NK cells is unknown so we could get SCID here if we have IL-2 that is missing or IL-2 receptors that is missing . IL-7 or its receptors and as I said if they are missing . if you remember we talked about the IL-2 as the main growth factor and differentiating factor which is very very important, so if that

10

for one reason or another IL-2 is missing- in T cell development nothing will be happen then we could get SCID , and we have to remember the stem cell transplantation because of all the lineages that we have talked about is going to develop and what we need is to support the growth factor for the line that you want here , if neutrophils are missing then you have to supply the CSF the G-CSF for example and so on . and any of those genes that code for IL-2 or IL-7 or their receptors are missing we could replace them by gene therapy , we can identify the that gene and insert it into that patient and the patient starts to develop the receptors for the cytokine s that are missing . What about the lymphoid cell lines ? the B and T cells morphologically they look the same and you have to know that and if you look at them under the microscope you can't differentiate whether this B or T cell , but you will say this is a lymphocyte . how are we going to differentiate the T cell from the B cell ? from the markers on their receptors , the IgM monomers of the B cell, the BCR , the Ig , Ig , the CD19 and so on ,these are going to be B cell that is responsible for antibody production and you know that by now that they differentiate into plasma cells , B cell has on the surface MHC class II but why ? ** because B cells they act as an antigen-presenting cell APC at the same time remember the CD19 that is specific marker , the CD21 that we have talked about the complement receptor as well and the plasma cell . The T cell morphology and how to identify the T cell , the same thing is here from the markers on its surface , if it has CD4 then it's T helper , if it has CD8 then it's T cytotoxic , if it has CD3 it is simply T cell and so on . CD8,TCR and its function against viruses and intracellular infection we are talking here about T cytotoxic and in T helper the cytokines will be produced . Natural killer cells NK ,they look like lymphocyte but they are not specific but they have the same function as the T cytotoxic and

11

they are the main surveillant cells in our bodies , they have no antigen receptor , no TCR and this is very important so considered as innate , and no memory cells for NK cells against virally infected cells and tumor cells , and the virally infected cells they get activated by IF (interferon) they (the virally infected cells) produce IF's and this is very interesting, and if you take the NK cells and expose them to IF's they get activated so we call them activated killer cells so you can activate them and give them back to the patient that has tumor cells or virally infected cells and so on .

And what about the other cells lineages ? As we said common myeloid proginator will give you the the granulocytes , the erythrocytes and what really makes them differentiate to these lines are the IL's or the GF's that we have talked about , for example what really differentiate common myeloid proginator into eosinophils is the IL-5 (this is literally what the Dr said) so when we have eosinophilia I know that I have IL5 that has been produced by T helper cells to induce the bone marrow to produce eosinophils that we see in response to the parasitic infections and allergies as well.

12

They are professional in killing parasites and usually they form around 2 - 4% of the granulocytes but when you are infected with a parasite you will see may be 20% , 30% or more the same is when you have allergies because they have some mediators that can neutralize the histamine and so on . they have almost the same mediators as those of the mast cells we will be talking about this later just remember the IL-5 is responsible for their production in the bone marrow .

What about the mast cells ? mast cells can be present on tissue in general and the skin as well and those they have receptors for which antibody ? IgE antibody but which part of the antibody is going to bind to the mast cells ? the FC receptor so you can imagine the IgE antibody will bind with the mast cell receptors and when that happens we say that the patient is sensitized he FC receptor so you can imagine the IgE antibody will bind with the mast cell receptors and when that happens we say that the patient is sensitized , so if the patient is exposed to another allergen, it will bind to the IgE on mast cell and cross-linking will take place and mast cells will be activated and then degranulation is going to take place , histamine, heparin , leukotrienes and platelet activating factor PAF and so on, all of them will be excreted. And cause smooth muscle contraction and edema and inflammation that the patient could have or suffocation and anaphylaxis , we will talk about that later. So you can see mast cells, the GM-CSF and IL-3 as well will be responsible her for the production of mast cells make, most of the neutrophils as well can here induce its production . What about the myelomonocytic progenitor cells and the macrophages lines here , the interdigitating cells and so on for the

13

APC , You will require the GM-CSF (G and M togrther) and IL-3 or GCSF or M-CSF alone. And that will lead to production of macrophage and neutrophils or interdigitating cell. What about platelets and erythrocytes ? Erythropoietin very important to induce the bone marrow to produce the RBC's lines, the erythroid cells which is nucleated but when develops into RBC it will not be nucleated , this really very important because if you want to see a viral infection the viruses infect the cell and utilize all the protein synthetic apparatus of the cell to produce the virus , so RBC's can't be infected with viruses. So viruses we have a virus called parvovirus B19 , it infects the erythroid progenitor that has nucleus but doesn't infect RBC's, but RBC's can be infected with other infectious agents the malaria , parasites , plasmodium malaria we will talk about the infections to the RBC's and so on . Erythropoietin will come from kidney to stimulate bone marrow to produce RBC's. What about platelets?? It will come from megakaryocytic by stimulation of thrombopoeitin. So the myelomoncytic progenitor will lead to neutrophils under GCSF . or monocyte under M-CSF. Or dendritic cell which has a lot of arms and can work as antigen presenting cell under GM-CSF . So neutrophils they are granular. The main enzyme in it is peroxidase and it responsible for what we call respiratory burst and also phagocytosis. And we have 10 to the power 9 produced daily by the bone marrow. And the half life of them is only 6 hours, so they don't survive much. They will come to the site of inflammation by the chemotactic factor and make the phagocytosis . we have two types of granules in it primary and secondary. Primary will be there and secondary will be

14

produced later. primary contain myloperoxidase and elastase. Secondary contain lysozyme and collagenase.

Basophiles act as a mast cell in the same way. Monocyte they can live longer and do phagocytosis and also antigen presenting cell. They are like cross bridge between innate and specific immune response. And don't forget to remember the interleukin molecules and the other molecules and the source of them and the biological activity of them. The doctor started to read exactly from the table 12.5 and say to concentrate on them.

For the CSF the source are very similar. From macrophage fibroblast and endothelial cells. The biological effect of them is to stimulate cell proliferation.

15

M-CSF for monocyte and macrophage . GM-CSF for myeloid progenitor. G-CSF for neutrophils proliferation. For interleukins # 3 and # 5 are produced by T cell , #4 produced by T cell and mast cell , #7 by stromal cell of bone marrow. Function of them :_ ** *IL-3 : activates multiple hematopoietic cells *Il-4 : stimulates B cell to produce IgE . T helper 2 cells differentiation and also mast cell *IL-5 : stimulates eosinophils . and activates B cells for Ig production "IgG" *Il-7 stimulates T-cell progenitor proliferation and differentiation and it is an essential growth factor for mature t cell. So without IL7 t-cell cannot be differentiated . So far we talked about the cells of the immune system so what about the organs of the immune system. We said we have primary lymphoid organs that they are responsible for production and differentiation between self and non-self antigen of cells. So the process of eliminating the self reacting lymphocytes occurs in the bone marrow or in the thymus gland. The lymphocyte that are produced after the deletion of self reacting lymphocyte we call them nave or virgin lymphocyte. They are looking for exposure with antigen and the do this in the secondary lymphoid organs. Like spleen, lymph node and so on. So the main function for secondary is to be the battle ground with the antigens. That's why when we get infected the lymph nodes will be enlarged and pain full !!

So granulocyte monocyte colony stimulating factor(GM-CSF) is for both the granulocyte and monocyte ,granulocyte colony stimulationg

16

factor(G-CSF) is only for granulocyte (neutrophils) and the monocyte colony stimulationg factor(M-CSF) is only for monocytes (macrophages) so those will make the progenitor cell to differentiate into one of these cells which we have talked about. For example the mast cells they have other triggering factors, (GM-CSF) and interleukin 3 and here the intermediate step is not really known, but mast cells will develop and you know mast cells and their role as inflammatory cells and they have the vasoactive amines responsible for getting rid of many parasites, they play a role in hypersensitivity reactions , based on the mediators that they have inside. The important thing to remember that the mast cells they have receptors here for IgE ,E receptor 1 and 2 ,and we have also receptor for , receptor 1 as well, while the macrophages they have receptors called CD14 and CD16 mainly for the Fc receptor that plays a role as I told you before for opsonisation, phagocytosis and so on.

Look at the figure above>>> And the same thing for eosinophils they have another triggering factor , interleukin 5 btw plays a role in the differentiation process i.e if you dont have interleukin 5>> eosinophils will not develop , so interleukin 5 makes eosinophils to take place . now we said that the mast cells activation or defense against parasitic infections . am sure you know all of that

17

information!! , the myelomonocytic progenitor cells that will give you the macrophages and dendritic cells and so on , the same thing here, interleukin 3 attached on the bone marrow remember that ! (GMCSF) that will lead to granulocyte colony stimulating factor neutrophils and so on for the dendritic cells and the macrophages under the influence of the same colony stimulating factor (GMCSF,M-CSF and interleukin 3) and then they will differentiate into either dendritic cells or macrophages. And we said that dendritic cells they look like a macrophage but they have more appendages as I said before and wherever they are located like macrophages we call them interstitial cells and tissues , interdigitating cells and lymph nodes, plasma cytoids , veil cells and so on we have different names for those , those they present antigens to T-helper cells mainly , dendritic cells and macrophages that they cross link also the innate and the specific immune response through the antigen presentation , through phagocytosis, opsonisation as well. And the difference as I told you between the macrophages and the neutrophils ,the neutrophils they have so many enzymes ,hydrolytic enzymes, myeloperoxidase, elastase, those they play a major role in the defense ,and later they develop collagenases as well. And these enzymes if they are deficient they then you are going to have neutrophils with normal number , but their killing ability is going to be compromised . and we will talk about these important points regarding immune deficiency when some of these innate components are deficient and what is the difference between those enzymes and the enzymes the eosinophils are having. Now eosinophils they look like neutrophils but the difference between them is based on the granules that they are having, they are more toxic , they are killer cells, those they are phagocytic cells , the eosinophils have heparin inside and they can kill parasitic cells as well as we see them in allergies, like those of mast cells they have IgE receptor on their surface as well, and we will tell you more about the role of neutrophils and eosinophils in the inflammatory process , but we have a very important difference between neutrophils and macrophages its that neutrophils are expired by the act , y3ne our body develops around 10^11 neutrophils per day !!!!!

18

imagine this number ! and half life of those is just 6 hours , they live for 6 hrs only! Continuous factory! So they are more important in phagocytosis compared to the macrophages .
-

The END -

And finally Mr. Ali Al.zu'bi ( the coordinator of science hall 2 ) Thanks a bunch for your great effort

Done By: Hussein Qasim

19