T regulatory cells are critical to active peripheral tolerance. They actively suppress the other lymphoid cells rendering them energic. Recall from the cytokine lecture, the TGF-beta, up-regulates the adaptive immune system. However, TGF-beta with IL–21 and IL–6 directs the immune system to an adaptive attack. And TGF-beta plus retinoic acid supports the expansion of T regulatory cells and essentially calms aspects of the immune system. T reg cells are t cells that recognize an antigen and then block and attack on cells with that antigen. They may render B cells and other T cells energic. If you remove T reg cells or you suffer a mutation that prevents you from making them then you will suffer a massive autoimmunity. For a long time immunologists had postulated the existence of cells specifically tasked with blocking an immune response. But evidence for such cells proved elusive and the search for them languished until Dr. Shimon Sakaguchi identified and characterized a population of cells capable of down-regulating the adaptive immune system with antigenic specificity. Dr Sakaguchi currently works and teaches at the University of Osaka. He's part of a team of highly respected immunologist who present the edX course, The Immune System, New Developments in Research. In this course, you can take an in-depth look at a series of topics that I just get to introduce. You can also look at how these researchers obtained the complex information that helps us understand the workings of the immune system. So I hope what follows gives you some background and whets your appetite for a closer look at this clinically important area of research. Now, what is it that makes a T regulatory cell a T regulatory cell? All right, because like Th cells they have an alpha beta T-cell receptor. They develop in the thymus and they display the co-receptor CD4. Like Th cells they respond to IL-2 by clonal expansion and activation. Here is the IL-2 cytokine with four alpha helices. It binds to the IL-2 receptor. And for the IL-2 receptor to bind this with a strong affinity it needs all three of its subpeptides. That is it has an alpha, a beta and a gamma and the complete receptor then will bind IL-2 and set off a chain of events that leads to clonal expansion and activation of the t cells. The complete IL-2 receptor is a heterotrimer. The alpha unit is sometimes called CD25, the beta is CD122 and the gamma, CD132. Okay, in addition to having this in the plasma membrane, a T regulatory cell also has a form of selectin. L-selectin, again it's named is CD62L, and this is a molecule that specifically sticks to parts of the epithelium to help the T regulatory cell extravasate to where it's needed. And in particular, it binds very efficiently to the HEV that is the high endothelial venules that precede secondary lymphoid organs. Another thing is that this guy starts out with high levels of CTLA-4, something that's usually induced later in most Th cells. And the other interesting thing is, is that the CTLA-4 doesn't seem to suppress them but rather allows them to tie up a lot of the B7 that might stimulate some other T-cell. Unlike other Th cells, they don't need to up-regulate the alpha or CD25 sub-unit upon activation. That is most CD4 cells come equipped with a constituent of expression of these beta and gamma sub-units but must synthesize the third or alpha peptide before they can produce that high affinity IL-2 receptor. T regulatory cells on the other hand are ready from the get go and this has two important consequences. T regulatory cells get a head start over other Th cells. If a new anagen shows up, this means the first response would be one that represses an attack and an assortment of danger signals from the antigen presenting cells can overcome this. But basically this gives you a chance to kind of sniff it out and see if that antigen really represents its red or not before you up-regulate a whole bunch of immune cells. T regulatory cells will sop up most of these circulating IL-2 at least initially and that produces a more deliberate response. The IL-2 receptors have high affinity that means they will bind to most of the IL-2 initially present. And unless they are down-regulated by danger signals, that will slow down or stop the response of the rest of the adaptive immune system. Now, it turns out that there there are actually two categories of T regulatory cells, nTregs and iTregs. And the n stands for natural, because these are produced in the thymus and are released in to the circulatory system as functioning T regulatory cells. The iTreg cells are induced so they are really flipped over, probably from Th cells in the secondary lymphoid organs. If you start out with a naive Th cell, that naive Th cell, depending on the cytokine stimulation that it gets, can go along and develop into several pathways. If one of these cells winds up in the lymph nodes and gets the right kind of signal, it will then also become a T regulatory cell. Both types of T regulatory cells have a number of properties in common. They have a transcription factor Foxp3 which is both necessary and sufficient to induce the development to T reg. The Fox P3 is part of a basic category of helix-loop-helix proteins and here is an example of a classic version of that. You can see, you've got alpha helix, and then here you have this loop. And then you have more alpha helix that's attached, and it's going to be actually binding into the major groove of the DNA molecule. Now the winged helix is a rather interesting version of this because in a sense you make it by truncating the alpha helices that extend out right here and, so let's look at this again. So here I have the truncated alpha helices, but these alpha helices, instead of extending out, will kind of bend back. And also bind into the major group of the DNA molecule. And that means you're going to have something where you have a lot of alpha helix binding into the major groove, and a bunch of extended loops. And the loops on these guys are larger and more random in their outward appearance. And so you actually get a structure that looks like this. Now if I just showed you that that would be just completely incomprehensible, but what's going on here is you're looking down the DNA molecule into this and you can see that the transcription factor which is a dimer, is held into the DNA molecule, in the major groove. And a lot of random coil loopy stuff extends out. So, we put together a metaphor for this. A visual metaphor because what this molecule kind of looks like is a butterfly attached to the DNA. That is it has the alpha helices are kind of like feet that bind to specific sequences of bases in the DNA. And it's kind of like a butterfly perched on those. And the loops extend outward and presumably they're going to attract other molecules that will change the regulation of the DNA of this cell. The net result being it's going to become a T regulatory cell. The next questions is, what induces the expression of Foxp3? Well, Foxp3 is induced by a STAT and in this case, this is the general way of up-regulating a STAT. You can see that something binds to your receptor which dimerizes, and then it auto-phosphorylates, and when it auto-phosphorylates, it attracts the STAT monomers which then pick up phosphates. When they picked up phosphates the STAT dimerizes and then enters the nucleus and up-regulates genes. Now here's the deal, though. We have a lot of different kinds of STAT. So depending on what receptors are activated, and what STATs are then activated, you get different sets of genes up-regulated. In particular the STAT that's responsible for up-regulating the production of Foxp3, is something called STAT5, would suggest that there are, [LAUGH] one, two, three and four that up-regulate other developmental pathways of a T cell other than T regulatory. So the next question is what activates this receptor up here at the top. An important signal that sends something down a T regulatory pathway is TGF-beta. And also one thing that would inhibit this pathway is inflammatory cytokines signals, IL-6 in particular. IFN-gamma which is the Th1 signal also shuts down the pathway as well. Now, this pathway is also promoted by IL-2. In the gut it's also promoted by retinoic acid and so there are ways of synergizing and modulating this particular response. And in particular a cell has to decide once it has the TGF-beta whether it wants to become a T regulatory cell or a Th17 cell, which will coordinate attack on extracellular pathogens. So, if I look at this thing, I see some interesting parallels. That is in some ways the nTreg and iTreg look like a version of central tolerance and peripheral tolerance. The differences in them are parallel in the differences in the way these tolerances are essentially set up. So the nTreg is formed in the thymus and that's a primary lymphoid organ. Ordinary would be having central tolerance in getting ready of anything that recognized a self–antigen. But in this case the nTregs are activated by self–antigen. That is, you're going to get the cells to down-regulate something that would attack your own tissues. Hence, then you set this up in a primary lymphoid organ. Induced T regulatory cells are a bit more like peripheral tolerance, in that they are determined after release, they're going to be up-regulated in the lymph nodes and spleen. And so basically these things also look at antigen. And if the antigen is presented in the absence of the danger signal that will activate the pathway to produce, an induced T regulatory cell. So that's a little bit like peripheral tolerance. The difference of course is with these two things you're activating particular cells instead of either apitosing or rendering them energic. But again, you're activating a repressor cell so this makes quite a bit of sense. Review the T regulatory cell. Okay, recall we had this wonderful cartoon showing our T regulatory cell with its stop sign, its dove of peace riding a Foxp3 and having the complete IL-2 receptor. The T regulatory cell will also be signaling using TGF-beta. In other words it responds to TGF-beta and produces more of it. It will also produce IL-10 and again, the IL-10 will tend to dampen any immune responses. This cell will also be removing the IL-2 by binding it to its receptor. It's going to be using it's CTLA4 to sop up the B7 and also will tie up the extravasation sites at the lead ins to the secondary lymphoid organs. So that's a summary of a little bit about how T regulatory cells work but there's one more thing that I'd like you to keep in the back of your mind. I'm want to briefly considered the role of the gamma-delta T cell, something that is both mysterious and complex. If you'll recall, it has this gamma-delta T cell receptor which is sort of bent and also tends to bind lipids. It also has as you can see a variety of other receptors and presenting peptides and we have it patrolling the epithelium down here where it is sitting, sort of of on a carpet. Now, this regulation of gamma-delta T cells leads to autoimmunity and typically this results from the pro inflammatory cytokines that we see being released here at the bottom. The IL17 and the IFN-gamma, both pro inflammatory and this tends to down-regulate the T regulatory cells. However, if I have a gamma-delta T cell and it receive retinoic acid it can actively promote tolerance and it will will release protective peptides into the gut. So, that mice that are mutant in gamma-delta T cells actually get colitis of they're not kind of prevented from getting colitis. Further studies of gamma-delta T cells show that they are not all the same and that there is a sub-population with inhibitory properties and selected depletion conserve in therapies for various disorder. I would like you to remember these guys as an insufficiently exploited resource for possible therapeutic procedures.
