 The last thing that Dr. Marek said to me as I was coming up here to introduce her as well, by moving down to the front row at least there's one thing less to trip over. Well, I've already proved that it's still possible to trip over things. However, it is a very great privilege and a great personal pleasure to introduce Dr. Marek. We have somewhat similar backgrounds. We both left a cold rainy England, she from Cambridge to go to San Diego, and I came to North Carolina when I first came over here. And our paths crossed again in Rochester, New York, and I have to of course add New York to that title here in Minnesota where she was on the faculty at the University of Rochester when I first joined the faculty at the University of Rochester. So it's a particular pleasure for me to be able to introduce Dr. Marek. From Rochester in 1979, if I remember the date correctly, she went to Colorado to the University of Colorado Health Science Center to take up a position there, and she has been an investigator of the Howard Hughes Medical Institute there since 1986. Those of you that have read her biography in the brochure will know that it would take up most of her time speaking if I go through that. So I will just mention a few of the highlights of her achievements, perhaps foremost of which is the fact that she was elected several years ago to the National Academy of Science at a time when both young people and young women were not being elected regularly to the National Academy of Science, and many of us are very proud of the fact that she was elected to the National Academy. Her talk this afternoon emphasizes much of the work that she has done over the years, and she has been, you heard Dr. Manasara this morning, while Dr. Marek has taken another part of the immune system and made it understandable at a level that 10 years ago or 15 years ago we just could not have dreamed about, and as a biochemist standing here it's a particular delight to know that she's going to talk about the T-cell receptor and some of its biochemistry and immunology. So with no further ado I will introduce Dr. Phillip Marek and the title of her talk, T-cells and Health and Disease. Dr. Marek. It's really very wonderful to get the opportunity to talk to you all today. I think for two reasons, well for one reason, and that is that as a basic scientist there are two things that, and I expect to those of you who are scientists too know this, that there are two things that really give you pleasure in life. One of them is to go in the lab and look at this big machine and it's spitting out the results, and all of a sudden you realize that you know something from the results that you never had any idea that it existed before, so you discovered something new, that's really very exciting, and then the next thing that's really, the next thing that you do under those circumstances and it's the next thing that gives pleasure is to rush out and tell everybody else what it was that you found so they can all share in this wonderful discovery that you've made. So I hope that some of you will follow along with what I'm talking about today and get some pleasure out of the beauty of the science that I'm going to talk about. Not because we discovered it, but because it was there already to be discovered and already very beautifully arranged and set up so that it functioned very well. And what I'd like to do today is to follow along some of the points that Dr. Banasra have already introduced you to this morning. In particular, this idea that one of the things the immune system has to do in each of us is detect that foreign things have come into the body so the immune system has to be able to recognize just about all the foreign things that could enter your body. It has to have foreknowledge of what those things might be or has to anticipate them. On the other hand, simultaneously, the immune system must not be able to recognize any of the things that are in the body already, any of self. So the immune system must tell the difference between everything else and you. And if it doesn't, the crisis will either be that it doesn't recognize anything at all and you won't be immune to anything, or the crisis will be that the body, the immune system starts to recognize self. And then we get one of this collection of autoimmune diseases that Dr. Banasra mentioned earlier on, juvenile diabetes or multiple sclerosis or myosthenia gravis or lupus or one of many, many diseases that some of you probably thought about it yourselves. So what I'd like to explain to you today is how the immune system does this, how it tells the difference between self and anything else. And I'd like to review some of the basic properties of the immune system so you can understand what I'm talking about and some of these will be points that I make will be reminiscent of what Dr. Banasra has already mentioned, but I'd just like to remind you about these points. So the first thing I need to do in order to explain how we tell the difference between ourselves and everything else is to explain to you how the immune system works when it's functioning normally and they're going to have a series of cartoons here to help you understand this. And what you need to know is that immunity is driven by lymphocytes. These are the small little boring cells that float around in your bloodstream that everybody thought were doing absolutely nothing for years and years and years. And the critical thing about lymphocytes is that they have receptors on their surface, not drawn to scale here. This is my husband's picture. He took this picture. Actually there doesn't seem to be a little red dot going out. These lymphocytes each have receptors on their surface and we've only put one receptor on the surface but actually there's probably about 20,000 of these things on the surface of each lymphocyte. And what this diagram here illustrates is that for each lymphocyte the receptor is slightly different. It's slightly different in amino acid sequence. It's a protein. So each lymphocyte has about 20,000 of these receptors on its surface and they differ from one lymphocyte to another. And if you can imagine the magnitude of this you and I have about somewhere around a million million, 10 to the 12th or more of these lymphocytes in our bodies and each one of them more or less has a different receptor on its surface. So we're talking about a lot of different kinds of receptors here. Individually different from one cell to another. And as I expect probably many of you know when you get an infectious entering the body, for example this virus particle we've drawn here, by chance it happens to be able to bind to the receptors on some of these lymphocytes. In this case this lymphocyte right here, it fits into the amino acid sequence. It's able to bind to the amino acid sequence of the receptor on this lymphocyte. And the consequence of this is that this binding reaction right here between the virus and the receptor on the lymphocyte makes the lymphocyte divide and you end up with this clonal expansion of a lymphocyte which was able to recognize this foreign invading particle, the virus particle. And what happens is that at the end of the response we have many many more lymphocytes bearing receptors which can deal with this virus than you had when you started with. This can be an expansion of say a thousand fold in number. And that's in fact how you make an immune response to an antigen, a virus or whatever. And these cells at the other end here they're what's called effectal lymphocytes. They help you get rid of this virus particle in a number of ways that I don't have time to go into. Now this scheme which was for the immune response which was devised in the 50s by a number of people, MacFarlane Burnett, Neil Scioni and especially David Talmage who's in Denver right now developed this theory which is called the clonal selection theory is the basis for how we understand immune responses to occur. We have lots of lymphocytes. Each one has a different receptor on its surface and the antigen pulls out the ones that combine to and help deal with that particular antigen. Selects out the clonal cells. Now you can see that there's a problem with this system as far as self is concerned and that is that we believe that these lymphocytes and we know in fact generate receptors on their surface of random specificity. We don't know ahead of time what these receptors are going to be able to recognize and that means that you could generate lymphocytes bearing receptors that can recognize you and how do we deal with that problem given that the possible outcome of recognizing you is a whole collection of potentially damaging lymphocytes that can kill your cells or create inflammatory lymphocytes inflammatory hormone like molecules that could damage you. So why don't we generate lymphocytes that bear receptors that can recognize us and originally there were two hypotheses basically. One possibility is that for some reason or another we simply don't have the genes to code for the proteins to code for the receptors that could recognize bits of us. So we're just intrinsically unable our lymphocytes are just intrinsically unable to recognize us because we don't have the DNA to code for anti-self. That's one possibility. Another possibility is that somehow or another the immune system learns what self looks like and deals with the cells that could recognize self. So we know that we don't lack the genes we don't lack the DNA to code for proteins that can recognize us and the reason we know that is that my lymphocytes are very well able to reject tissue from my children actually they'd like to reject my children more often than not or my parents and of course my children's lymphocytes and my children are only too eager to reject me conversely. So and we know that we all share genes with each other so it would be almost impossible since I reject my lymphocytes would reject my close relatives tissues so effectively. It's almost impossible to imagine that genetically I lack the ability to recognize myself where I can certainly recognize my mother and my kids and so on very well. So this is sort of ruled out by that kind of reasoning the possibility that we lack that but there's actually another of course some very beautiful sets of experiments which tell us that the way we distinguish ourselves from everything else is because we learn to do that and the first experiment that was done to prove this was done by Ray Owen in 1945 which was the year I was born actually and what he showed was that if you have brothers and sisters calves who now we're talking about cows calves who are brothers and sisters normally they can reject skin grafts between each other very well but if they happen to have come out of the same pregnancy and they shared a placenta which that happens sometimes for calves they won't reject skin grafts one from the other the two twins don't reject skin grafts so somehow in utero the two calves learned that the other or at least they thought they learned that the other one was self too and their immune system won't reject the other calf anymore so we know that tolerance to self it's called tolerance the lack of ability to recognize self is something that your immune system learns now what I'd like to tell you about today is how the immune system learns what self is because it's actually a very simple and straightforward message as you'll see and quite clever the way biology has set this up but I have to tell you a little bit about the receptors on cells before I do that and the first thing I'd like to introduce is to remind you of something that Dr. Banathara told you earlier on today and that is that the immune system in fact actually has three different kinds of receptors three different kinds of lymphocytes and each lymphocyte bears an entirely different type of receptor for antigen on its surface so we have in us as human beings about a million million B cells and each B cell has on its surface a slightly different version of this molecule which is an immunoglobulin molecule that's an antibody molecule very similar to the kind of antibodies that you have floating around in your serum so B cells bear immunoglobulins and then we have these two different kinds of T cells this kind over here the gamma delta cell which I don't understand so I'm not going to talk about that and then this T cell here the alpha beta T cell which was the kind of T cell that Dr. Banathara was talking about and it's the cell that I'm going to discuss today as well included in this group of T cells are the CD4 bearing cells the ones that I think Dr. Gallo might discuss a little bit later on when he talks about AIDS infections because the target of the HIV of the immunodeficiency virus is a kind of this type of T cell an alpha beta bearing cell but what I'm going to say about alpha beta T cells applies to some extent or another also to B cells and I haven't any idea whether it applies to gamma delta cells so we won't worry about them now I need to tell you a little bit about the T cell receptor the alpha beta receptor and how it works the alpha beta receptor on T cells the question is how do you get all these millions and millions of different slightly different specificities out of these receptors how do you construct a receptor which on one cell has a particular amino acid sequence a particular structure and on another T cell is similar but not identical how's that done and it's the way it's done it's done genetically in rather a complicated way that I don't want to go into but basically it's done in a in a similar manner to a kind of a cheap Chinese restaurant and what that means is you know when you go to I don't know if this happens in the state so much but when you go to a cheap Chinese restaurant they say you should take choose one possibility out of this collection of A things like spare ribs or whatever and then one you can choose one thing out of column B etc etc and put together your dinner by combinations from different lists and that's basically what the T cell does it can choose for the alpha chain it can choose a V it's called V alpha out of a pool of about 50 or 60 different V alphas it can choose just one and then it can choose a J alpha and there are about 50 different J alphas and it can choose just one of the 50 and then there's this thing called N which I don't want to talk about too much but there's about 400 different choices for N so a T cell is choosing is making its alpha chain out of one of 50 and one of 50 and one of 400 and if you multiply that together the number of possible ways of putting an alpha chain together is very large so it has just like you could put your Chinese dinner together in a lot of different combinations the T cell does the same thing for its alpha chain and likewise for its beta chain it puts it together in a lot of different combinations and in fact this is a sort of ballpark figure it's plus or minus two orders of magnitude right they're probably around 10 to the 14th different ways that we could assemble a T cell receptor so that's why I say out of the million million T cells we have probably none of them have exactly the same receptor they've all made slightly different choices so I hope I made that clear the diversity of the receptor is put together by choosing one of many combination one of many possibilities at each of these positions so that's the first point I want to make about the T cell receptor that it's put together with these little segments each of which is different from one cell to another and then the other thing I need to tell you about the receptor and this is something that Dr. Banasra pointed out before is that the job here's the T cell up here see it's something huge right up here and the way the T cell receptor works is it sees little fragments of antigen little peptide fragments of antigen bound in the groove that Dr. Banasra shows you of the major histocompatibility complex molecule these are proteins that are on every surface the surface of many cells in the body and the way we've drawn this you can see here's the surface of the T cell receptor made up of all these segments that I said the T cell had a choice about and all of these segments contribute to the ability of the T cell up here to bind to this little antigenic fragment a little chunk of flu virus or polio virus or whatever it is bound in the groove right here so the point is that for most antigens flu or polio or chicken pox or tuberculosis or whatever ragweed um the T cell has to have exactly the right combination it has to have chosen the right V alpha and the right V beta and the right J beta and so on in order to be able to recognize that peptide so if you remember I told you there were lots and lots of different specificities what this means is that in an unprimed in animal in a normal individual who hasn't had this particular chicken pox virus before for example the frequency of responding T cells is very very low it's something like one in a million or one in a hundred thousand T cells is able to recognize that virus likewise for your own peptides for example for a peptide from your big toe or whatever it is we imagine that the frequency of T cells if you hadn't already been immunized with that big toe antigen would be very very rare one in ten to the fifth or so what that means is it's tremendously difficult to figure out what's happened to that T cell if you made a T cell that could recognize an antigen in your big toe how could we figure out what it was doing was it there was it present or not um is it in the body or is it just sort of sitting around ignoring your big toe because it's so rare in the population we can't tell which T cell to look at so the question is how do we devise a tool so that we can follow what happens to T cells that can recognize ourselves and uh a few years ago John Kappler and I happened on a tool it was a complete accident that turned out to be very useful and this was the special kind of antigen we discovered it's called a super antigen and you can see that regular bits of flu or TB or whatever antigen it is that's entered the body the foreign substance usually lie in this special little groove here but these super antigens that we discovered have a totally different way of interacting with the T cell receptor they bind onto these peculiar molecules these MHC molecules right here and they clamp the whole thing up the side by binding to a piece of the T cell receptor called V beta and they don't care about all these other variable bits they're only interested in this one little bit called V beta now in human beings we have about 50 different V beta genes 50 or 60 different V beta genes all together what that means is that about 2 percent of our T cells or more in fact can be recognized can recognize a super antigen because they have the right V beta on their surface the super antigen will grab them and so in man we can follow very easily the ability of a super the interaction of a super antigen with a T cell because it's easy to see what 2 percent of T cells are doing or at least it's tremendously easier to see what 2 percent of the cells are doing than one in a million cells is doing if you see what I mean and fortunately for us over the years people have made antibodies against these V's these different V betas and so we can actually follow T cells bearing particular V betas by seeing whether they react with antibodies that may seem a little complex but actually I have to tell you that technically it's much much easier to do than it was before and I didn't tell you this but in mice which is the animal we work with most of the time because we can't usually get human beings to volunteer to get immunized with these and you'll see why in a minute in mice there are only about 20 V betas so that means that 5 percent of the T cells in a mouse bear any given V beta or thereabouts and that's a piece of cake to see by comparison we're one in a million cells so you'll see the kind of experiment that we can do with this very useful reagents in a minute I thought I'd tell you just a few things about super antigens before we go any further the first thing is is that they have to bind to these MHC molecules which I don't want to go into in any depth right now and the second is just to remind you once again that the way these special antigens react with T cells is just through that special part of the receptor called V beta and now I need to tell you what super antigens are and some of them in fact everybody in this audience has met a super antigen I'll guarantee you because they are made by bacteria a whole collection of made by streptococcus and staphococci and they are the major cause of food poisoning in this country a staphococcal enterotoxin B for example was first worked on in Madison and sequenced in Madison it's a fairly medium-sized protein it's the kind of protein that when you eat an egg salad that's been sitting out too long makes you feel really bad for a short space of time for about a day it's that kind of food poisoning I'm talking about and we think in fact that the food poisoning has something to do with the fact that these toxins engage T cells and maybe I'll come back to that later on anyway so staphococci make about 11 or 12 different toxins which are super antigens one of them is the toxic shock toxin which gained some notoriety in the early 80s when relied tampons were brought in it was the cause of a considerable amount of toxic shock an epidemic of toxic shock in this country and the other point I want to make about these particular kind of toxins is that they have specificities for different V betas and in order to explain that I have to say that if you remember I told you that the T cell has a choice of one out of 20 V betas or whatever it is each V beta is different in sequence and the scientists who study them have numbered them so in mice we have V betas 1 through 19 and each number designates a slightly different amino acid sequence slightly different structure for that V beta so staphococcal enterotoxin B for example reacts with T cells in mice bearing V beta 7 and the members of the 8 family and in man if you were to eat staphentrotoxin B the T cells in you that would get all upset about this would be those bearing V beta 3 12 etc etc whereas in mice for example another staphococcal enterotoxin called SEA engages T cells bearing V beta mouse V beta 3 now it's not only bacteria that make super antigens there's also a collection of viruses that make super antigens and the most well-known of those are some viruses in mice which called which cause breast cancer they're called mouse mouse mammary tumor viruses MTV and one of the viruses that I'm going to mention later on is a virus that's passed through the mother's milk from the lactating mother to her baby in the milk and this virus carries with it a super antigen which engages T cells in mice bearing V beta 14 but these mouse mammary tumor viruses are especially interesting because over the course of the years some of these viruses have picked themselves up and put themselves into the DNA of the mouse themselves they've what's called integrated themselves so there are mice running around in your backyards which have integrated into their own DNA some of these mammary tumor viruses and they carry with them super antigens so as far as these mice in your backyards are concerned the virus in their DNA is them they can't tell the difference between the virus and themselves because they grow up with it it's in their DNA just like anything else in their DNA so we can use these viruses that are making super antigens that are actually in the mouse DNA itself to study what happens when a mouse T cell develops and sees a super antigen which is specific for that T cell now I'll try and illustrate that on the next slide first of all I want to tell you the people who did the work I'm going to talk about today as John Kapler and myself we run a lab together and our postdocs who were involved in this work were Yong-Won Choi and Terry Finkel and Lesha Gignardovitz and Jim McCormick and some of the work I'm going to talk about was in collaboration with Brian Kotzen if I have enough time and another postdoc who mysteriously didn't show up on this slide whose name is Xavier Palliano so how do we tell the difference between ourselves and everything else given that we could make lymphocytes that could recognize us so basically over the years there are three theories that have come forward to account for tolerance to self and they are one possibility is that T cells while they're developing go through a stage where if their receptors get engaged by something that cell will die and since self is always present when T cells are developing self is always present all T cells while during development will go through this screen of whether or not they can react with self and if the clonal elimination theory the clonal deletion occurs they will die during that developmental stage I don't know if I said that clearly enough but I'll say it again later on another possibility is that contact with self during this sensitive developmental stage will cause the cell not to die but simply to become inactivated so it'll still sit around but it won't actually die but it won't be able to do anything threatening to the host anymore because it's inactivated by this contact during an early developmental stage and another possibility that's been suggested is that somehow or another potentially auto reactive T cells are kept at bay by other cells that keep them under control called suppressor cells so what I'd first like to show you is some evidence that at least for many self antigens this first mechanism clonal deletion is a very important one in maintaining self tolerance in order to do this I first have to explain to you how T cells develop T cells are formed in the thymus this gland right here in your chest and they go through the cells enter the thymus from the bone marrow or the yolk sac or fetal liver and at that point they don't have any receptors on their surface at all and when they get to the thymus the environment of the thymus induces the appearance of receptors each different on one T cell from another on the surface of these cells and then they go through three or four days of stage here which doesn't have very much receptor on its surface it's called an immature thymocyte and then the cells go through a process that Dr. Bonaster I've alluded to called positive selection that I won't discuss and they turn into mature lymphocytes first in the thymus medulla right here and later on in the periphery in the spleem and lymph nodes and peripheral blood of the animal so the T cell develops in the thymus and it goes through an immature phase in the thymus so now this is a complicated slide to explain what we have on this axis is the percent what we can do is look in individual mice and ask how many cells do they have how many T cells do they have which have on their surfaces for example this V beta V beta 6 and in a normal animal it's around about 12 percent of all the T cells bear V beta 6 on their surface now some mice are infected not are infected have in their DNA integrated into their DNA one of these mammary tumor viruses I told you about that makes a super antigen which can react with T cells bearing V beta 6 so that the T cells in that animal are growing up in the constant environment of this mammary tumor virus making a super antigen specific for V beta 6 and we can ask what percentage of the T cells in that animal bear V beta 6 and the answer is it's much less than the expected percentage instead of 12 percent which is what we've called 100 percent right here it's very confusing picture instead of 12 percent is less than 1 percent and this is we're looking at the periphery of the animal now in the lymph nodes and spleen so that the expression of this super antigen that as far as the mouse is concerned is self in the animal has caused the disappearance of all the T cells or most of the T cells bearing V beta 6 in that animal now when did that happen did it happen when the T cells got to the periphery or did it happen in the thymus and this is a slide that shows you that it happened in the thymus if you could just follow along here this is the mice these are mice which have that MTV that encodes a V beta 6 reactive super antigen you can see they have a very low percentage of the mature thymocytes bearing this antigen bearing V beta 6 and they have a reduced less than expected number of the immature thymocytes bearing this V beta so the summary of this kind of experiment is that thymocytes do indeed go through a stage right here while they're immature about halfway through their immature phase the point of this red line right here in which they become sensitive to deletion and if their receptors at that time become engaged with something that can react with those receptors that is a signal for death of those thymocytes and therefore those particular cells because they're dead can no longer proceed through to the mature thymocytes stage and to the periphery and we see this is a deletion of that particular subset of cells so in this particular case self has caused clonal deletion of potentially reactive from T cells in the thymus so just to summarize what I've told you so far and to make sure to really ran this point through what this experiments have shown is that during their development T cells go through a stage when if their receptors bind antigen bind something those cells will die because our self is always there self is always present all developing cells as they go through this kind of adolescent rebellious stage will get checked out for their ability to be killed by antigen by self and if they their receptors react with self those cells will die and the critical point is that infectious things from the outside like for example polio virus are not always present in the body the child is born without not infected with polio virus and therefore during the early stages of that child's life its thymus will produce T cells that can react with polio virus antigens later on when the child becomes confronted with polio vaccine I hope or polio virus perhaps those T cells that are now already mature can help respond to and get rid of the virus as I showed you on a much earlier slide at the same time of course the T cells which are developing right at that very moment now I think the polio virus is self because it's in the body at the very time that those particular cells are developing and they'll die but it doesn't matter because by that time the child has already accumulated enough T cells mature T cells to help get rid of the virus so that's the theory to account for the data that I've just shown for you that I've just shown you about clonal deletion and I have sort of some sort of hysterical slides right now it is possible that not all antigens get into the thymus for example you could imagine that things that were in your big toe how would they ever get to the thymus well some of them do get carried back to the thymus on the surface of macrophages or whatever or B cells or something and and maybe able to tolerize T cells there but perhaps there are some kinds of antigens that don't get carried back very well for the thymus and how do we keep our lymphocytes tolerant to those and especially critical in this regard are various antigens that are in your brain or perhaps in your joints and I should tell you at this point that it's not clear that we are in fact tolerant to the antigens that are in our brain but I'd like to deal with a few more antigens that are elsewhere in the body first and so because we now have these tools the super antigens to help us in our experiments I'd like to illustrate this question of peripheral tolerance how we become tolerant to antigens outside the thymus with a couple of experiments here now in this case we've taken a mouse and injected an adult mouse and injected it with one of those bacterial super antigens this is SeA which if you remember in mice stimulates T cells bearing Vbeta3 so we injected the mouse with SeA right here on day zero and then we asked what percentage of T cells in the lymph node and spleen have on their surface this Vbeta3 with which SeA can react and what you see is there's a big spike as the lymphocytes proliferate in response to this antigen in number and then they fall off in fact they a fair number of them die and disappear and we're left with some cells left over at the end here which is somewhat less than we started with so an acute confrontation with antigen goes almost pretty much as you would have predicted you see a response as the lymphocytes divide and then they die away but if we put the antigen in chronically that is sort of every other day pretending as though it was an antigen in your big toe and you kept on challenging lymphocytes with it what we see is that if we put in a lot of it there's a spike of response but if we put in just a little antigen the lymphocytes just disappear the Vbeta3 bearing cells just disappear and eventually dwindle away to nothing so there's some kind of mechanism whereby a chronic antigen which is what self is causes the disappearance of target cells in the periphery and I have to tell you now we have no idea what this mechanism is and we certainly would like to know what it is because you could imagine the therapeutic value of understanding this mechanism in autoimmune diseases if we knew what the antigen was and we knew how to get this chronic disappearance business to work we perhaps could deal with autoimmunity much more effectively than we do right now so the immune system once again is dealing with potentially autoimmune C cells on the basis that the antigen is being continuously administered to them and as I said we don't understand what this phenomenon is but it looks like something real if and we wish that we understood it better than we do I'd like to point out one other thing about this mechanism and this is a little difficult to grasp from this slide so I'll just tell you and that is that we can interfere with that disappearance in response to peripheral antigens if we put in an inflammatory agent at the same time so we're chronically giving this mouse staphocontrococcal entrotoxin A for example and making the T cells bearing VB to 3 disappear and if at the same time we give the mouse some kind of really strong stimulus an inflammatory stimulus in this case an adjuvant called complete Freund's adjuvant they don't disappear after all so maybe that's another phenomenon that has something bearing on autoimmune diseases and how they stop so to summarize what I've told you so far acute exposure to super antigens of peripheral mature T cells causes them to expand and then disappear a little bit and if we expose mature T cells chronically to a super antigen and presumably a regular antigen too this causes them to die and in an inflammatory process such as complete Freund's adjuvant an inflammatory agent can interfere with this deletion and we are thinking along the lines that some phenomenon like this or related to this may have some bearing on the induction of autoimmunity in patients now I don't think I have time to show you this so I'll skip it and just come to the summary and that is that what I didn't have time to show you is that under some special circumstances instead of getting deleted T cells can get inactivated so basically what I've described so far is this mechanism of clonal deletion adenolgium in the thymus or in the periphery self can cause the disappearance of target cells and also under certain circumstances um self can instead of courtesing these target cells to disappear can just make them inactive and what I haven't discussed with you is this third mechanism of suppression and I'm not going to do that today what I wanted to finish up with very briefly is some description of how this is relevant to disease in man and what I'm going to show you is very hyper their real data but the explanation of the data is pretty hypothetical as you will see and a little complicated so I hope you'll follow along with the results now if you remember in man I told you there are about 60 different V betas and each T cell bears a different V beta in this assay that I'm showing here we only actually measured 20 of them or families of 20 so what's shown on this slide is the percentage of T cells in two individuals that bear different V betas on their surface so for example you can see that both of these individuals have a fair number of T cells bearing V beta 13.1 which is just a name for an amino acid sequence on their surface so this is a commonly used V beta by these two people and by most of the people in this audience um and I just wanted to point out a couple of things of this about this of general interest the V betas are not sort of equally often used some of them are used very frequently like this 13.1 by T cells and some of them are hardly ever used like we none of us have any T cells hardly at all bearing V beta 10 and I don't know why it should be such a poor despised relative but it just isn't often used the other point I wanted to make is that these two individuals are not identical for example the dark one the dark person right here has hardly any T cells bearing V beta 3 and the hatch person here who's me who's infinitely superior has many more T cells bearing V beta 3 um in her circulation and we don't know what that what caused that we don't know the significance of that it's just a fact right now uh and the scientific and medical consequences of that we don't understand now what I wanted to show you here is a patient this is some experiments that the Xavier Pagliata postdoc in our lab and Brian Kotzen published about a year ago to dealing with the T cell repertoire in people with rheumatoid arthritis which is an autoimmune disease of the joints as I expect some of you know and he Xavier was trying to figure out whether there was anything peculiar about T cell receptors about the T cells in patients with rheumatoid arthritis so what he did was he collected the T cells from the peripheral blood and the synovial fluids that is the rheumatic joints of patients uh of the same patient and compared what kind of T cell was in the peripheral blood to the kind of T cell that was in the synovial fluid and in the dark bars the dark bars show the percentage of T cells bearing different V betas um in the peripheral blood again he was looking at V betas 1 through 20 and the hatched bars show the same V betas measured in the synovial fluid the percentage of T cells bearing them and what you can see is that on the whole things are rather similar except V beta 14 um where there's definitely a lot more T cells in the synovial fluid than there are in the peripheral blood bearing this V beta and this wasn't true simply for this one patient it was true for every patient we looked at this is a collection of room patients with rheumatoid arthritis right here the dark bars in every case of their peripheral blood and the hatched bars is what's happening um in their synovial fluid and you can see for all of these there are more V beta 14s in the synovial fluid than in the peripheral blood here are some control patients who had a different kind of arthritis in their joints not rheumatoid and here are a collection of individuals who didn't have arthritis and we can't look at the T cells in their synovial fluid because first of all it's not ethical to go sucking things out of people's knees if they don't have something wrong with them and secondly they don't have any if you're a healthy individual you don't have any T cells in your knees anyway so that was a good reason not to do it apart from the ethical considerations right there but actually the point I really want you to notice from this slide is not the difference between the synovial fluid and the peripheral blood but the fact that some of these patients with arthritis had no detectable T cells bearing V beta 14 in their peripheral blood at all it was like they were all gone and that wasn't true for any of the normal controls that we looked at so there was something about this disease that was removing the T cells bearing V beta 14 um from the blood um and one explanation for this is that um there's a super antigen specific for V beta 14 sitting in the joints of these patients that's sort of sucking all the VB to 14 into the joints all the VB to 14 pairings T cells into the joints so there's nothing in the peripheral blood they're all in the joints well let me tell you there isn't enough room in your knees for all the VB to 14 bearing cells you have in your body simply wouldn't fit so that's probably not the explanation and in fact I don't have time to go into it we we know that that's not true it's not that there's a super antigen in the joints attracting all the T cells in there what the disease looks much more like what the phenomenon looks much more like is the expression of some sort of chronic super antigen in these patients bodies which has caused deletion remember I told you that chronic expression of antigen causes deletion of peripheral T cells so it's called all the VB to 14 cells in the periphery of these patients to disappear and the only ones that are left behind are a few that could be rescued because they're sort of hidden in the joints and responding to antigen in the joints so I've written the theory down on this slide I think here we go this is such an exciting way to show this slide what this slide is going to show you is the hypothesis we developed to account for the fact that some arthritis patients don't have any T cells bearing this particular VB in their joints and theory goes the theory goes like this that these people are wandering around fairly normal they have a few T cells in them already that have snuck through all these processes of tolerance that I told you about so they just didn't see joint antigen in their thymus and the joints are all nicely encapsulated so they never see the antigens the collagen or whatever it is that's in the joints in their periphery so a few T cells go through the thymus they mature and they come out and potentially these T cells could react to that individual's collagen antigens for example joint antigens if they ever saw them but because they're never exposed to the joint they never do anything much and they're not activated they're not excited about anything they're not doing anything then one day this individual catches something that chronically expresses a super antigen specific for VB to 14 and this causes the activation of all the VB to 14 cells in this individual and one of the things that happens when T cells become activated is they become much more mobile and they can travel around better so the theory goes that these T cells can sort of some of the mobile ones go through the joints and the VB to 14 cells that could recognize joint antigens stay there and initiate the process of disease the process of rheumatoid arthritis in the joints meanwhile the chronic expression of this VB to 14 specific super antigen deletes all the T cells everywhere else in the body because it's there for such a long time and they the T cells begin to think that the super antigen is self if we could skip that slide and just do I get extra minutes for this so here's the theory I don't want to go through it again you you heard it the these people who are rheumatoid arthritis is an MHC linked disease in in this country it's linked to DR expression of an MHC antigen MHC protein called DR4 all those individuals I showed you in fact were positive for the DR4 MHC type so what we're saying is in some people who are DR4 positive these T cells sneak through tolerance somehow or another and they sit around and don't do any damage all of a sudden the individual becomes invaded by an exogenous antigen a super antigen that's specific for VB to 14 it activates the cells the joint specific VB to 14 bearing cells whiz off to the joints and start to cause the disease rheumatoid arthritis meanwhile in the periphery the rest of the cells become deleted and then other T cells join the VB to 14 bearing cells in the joint and you eventually develop the symptoms of rheumatoid arthritis this is very hypothetical and almost certainly not right but it's a sort of idea on which I don't mean to belittle it but what I mean is that you're never right the first time but you sort of approach rightness gradually so this is a sort of a a hint about how these diseases might start it's not the absolute answer but it's the kind of thing that makes you feel as though you have your foot in the door and you can start prizing the door open a bit more and maybe see more glimmerings of the truth as you go further inside but what I wanted to finish off with is the last slide which describes how the immune system tells the difference between self and everything else because this is the absolute critical feature that I wanted you to take home with yourself with you and that is we tell ourselves from everything else because we are always there and the infectious agents are only there some of the time and that is the critical element which defines self from non-self thank you very much