Greg Smith - What we are learning from a virus that reprograms our nervous system

[Music] this lecture is being brought to you in part by the generous gifts of these sponsors well welcome everyone and thank you for coming I think I should start off by saying that we're living in an amazing times art kubbs just won the World Series and we have a brand new president-elect and you won't come to listen to me talk about herpes I'll let you choose which is the most amazing of the three but with that let me thank particularly ken Ford for this wonderful invitation and for Nancy and Ken for graciously welcomed me to pencil beautiful Pensacola last night we had a wonderful dinner it's my first time here and I've been thoroughly loving it and if you don't if you will allow me I imagine most of you are not virologist and don't think about virology on a daily basis as do I so I'm going to give you my very short version of virology and 101 in preparation for learn I'm going to tell you about our research and let me start you off with a filter for me this is where it all begins some of you may have heard of the name pastor ER but you may not have heard of Chamberland he is the inventor of a porcelain filter it was unglazed and porcelain is porous and you can actually produce it so the pores have different sizes and by doing this they revolutionized sanitation because and this is what they look like these are small versions here these are some of the original ones these have a channel right down the middle of the cylinder and you can put them into this canister and when it's fitted you can put water through that at high pressure the water will then start seeping in from the outside of that filter that porcelain enter the central channel and come out the bottom of the canister you now have bacteria free water if you have access to this in 1884 you don't have to worry about cholera or many other diseases it was high tech at the time and you can see it won the gold medal at the health exhibition at 1884 now it also inadvertently led to a discovery that revolutionized biology and that started in Russia by this individual named Dmitri Ivanov ski and what he had found he was studying a plants called tobacco tobacco crops were suffering because they would get this model or mosaic disease on their leaves it's not good for production and what he had discovered is you could isolate the SAP from an infected plant and then put it on to a healthy plant and that plant would then become diseased and this was the first step to demonstrating an infectious pathogen for that plant but the revolution was that he took those chain went on pasture filters from eight years ago and he took the SAP and he filtered it through there and took the filtrate that exited the bottom and put it on to the plants all pathogens should now be gone and yet the plant became diseased and mosaic and so he as he described it he discovered what he called a filterable agent something that passed through this filter the first phrase that was used to describe what we ultimately call viruses Friedrich Lafleur did the same experiment several years later he didn't actually put a cow into the filter he just took the serum the little misleading I apologize for that but he took serum from an infected cow cows that was suffering from hoof-and-mouth disease passage it through the filter and sure enough the receiving cow got sick and there's a quote that I just like to read to you because I think it really puts a lot of context into what's going on at the time and how phenomenal this was if for other for further experiments carried out by the Commission confirm that the filtrate effects are as it indeed appears to be the case in fact caused by minutely living things then one could very well assume that they are causative agents of numerous other infectious disease of and in animals emphasis on man and yes the unspeakable experiment was then done these are soldiers from the spanish-american war 1901 this is a reason why you do not want to be part of the enlisted especially back then they were suffering from a terrible disease called yellow fever and so the experiment was done and they were offered a hundred gold piece for perk taking saying that if you don't partake you'll probably get the infection anyway you miles will take it and yes it works who was a success people died and back then that kind of a success was rewarded by having a very prominent hospital named after you Walter Reed Hospital now please don't make don't think this that I'm disparaging Walter Reed except to say I mean certainly accomplished much more than just this one instance but it does give you some perspective on how our society works once upon a time compared to now and yet although they knew that there were these filterable agents that could come out of those out of those filters no one could actually see what they were and in fact way back in 1683 anthony van luna hook had image microbes for the first time in water using his home a microscope a single lens that he ground himself by hand and he made some of you may have noticed that just last month google had a doodle on their homepage celebrating its 382 fourth birthday but no one no one was able to see these agents now let me shift for you and we'll come back to that and give you one last piece of perspective i'm sure many of you have heard of this Franklin Delano Roosevelt founded this does anyone know why you all know why very good you know my young students have no idea but that's absolutely right he had polio and he also was concerned about the spread of polio through the nation and so he started this saying that if everybody can contribute a dime to the cause we can find a cure to this terrible disease and notice the poster highlighting that both boys and girls are equally affected and however it strikes it random the twin girls only one of them has the crutches that led to an incredible influx that open the doors to modern virology in 1949 Enders and his colleagues were able to produce cultures of mammalian cells and a plastic glass dish in a petri dish in the laboratory and keep them alive these are particularly b-cells you're seeing here or in my lab these are from a monkey but you can use human cells I'm sure some of you have heard the stories of Henrietta Lacks and those are human cells they would look just like this in the microscope but the important thing here was he was not only able to culture these cells and then culture polio in these cells and by doing so this ultimately led to the development of the Salk vaccine that's a Nobel winning prize right there nowadays you just take it for granted and I just want to point out as long as we're here that for those of you have not looked down a microscope before that we're actually seeing it there's a number of cells and the cytoplasm I have indicated as is the outside of the cells the major volume of that cells we're all new proteins are made by the cell and the nucleus has the genetic information and if you want to see viruses not only without all you to produce the bottom to look at but now you can take this wonderful new instrument called the electron microscope and you can produce incredible images and see those filter allegiant's for the first time and no one ever was able to see them because they are amazingly small this is from a colleague of mine in Germany dr. Thomas mittleider and this is the virus we're going to talk about this is a herpes virus seen by transmission electron microscopy and what that simply means is this fear the spherical viral particle has been sliced right down the middle and you're seeing what it looks like it's like a layered onion so the center of that that dense core is DNA it's the call that the viral genome and the circle that goes around that is actually a polyhedron it's a 20-sided object so we call that in a constitution well mathematicians call it an icosahedron and if anyone had children or again the grandchildren who played role-playing games they would call that a 20-sided die okay and so right around that shell that holds the DNA is a layer of protein I should say the shell itself is called a capsid it's very rigid but the protein that surrounds it is much more floppy and then round that defining the whole thing is a lipid bilayer so an oily lipid around it and that has a lot of proteins that span through that layer and ultimately we'll make contact with cells and so just to summarize we have four different layers starting with the genome in the center of the particle now I like to refer to this as a nanomachine and there's a number of reasons for that but let me just give you one illustration so when a virus like a herpes virus first comes in contact with a cell whether it be in a dish in the laboratory or in the human body it will first make contact and then it goes into a search mode looking for a very specific protein that will lead to a triggering event now these particles don't have our not metabolic but they are filled with potential energy you can imagine them as a Rube Goldberg machine they're put into a configuration that's ready to trigger if the right event occurs and when the trigger occurs something happens the first thing that happens here when it binds to his receptor triggers its membrane to fuse with the membrane of a cell and that leads to the deposition of the internal components of the viral particle into the cytoplasm of the cell including the capsid shelled the polyhedron and the surrounding softer tengaman protein that then is able to very effectively travel inside the cell to the nucleus where docks at openings called nuclear pores and injects its genome into that cell upon this event the cell is infected productively and if it's in your body you now have herpes for life now any of these steps can go wrong right it's something may trigger incorrectly at the wrong moment the particle may be defective and if that but if everything goes right delivery of genetic information will cause more viral particles to be made in the nucleus of that cell and that amplification will lead to a lot of bursts of viruses that can now go out and infect more cells and spread the infection if it doesn't go right the virus doesn't really care too much because they've got the numbers on their side thousands of viruses per droplet in this mess right now I have to apologize because I know something what time before this night is over somebody will sneeze don't feel bad I promise you we will all survive ok so that's the basic idea of how the viruses were working at the cellular level but now let's talk about us and disease so when you first come exposed to a virus like this these neuro invasive viruses and I'll define that term in a moment the first thing that will happen is you'll get with the virus replicating at the body surface this often will lead to some kind of sore or lesion typically for the for herpes simplex virus type one around the mouth this is a typical presentation I know it looks terrible but it's not at all actually this is a childhood presentation called gingiva stomatitis and most it's least as a number of fever blisters all around the mouth and this will take care of itself you don't need any treatment really it won't it will go away of course the infection never goes away that the person will now seem perfectly healthy but later in their lives perhaps weeks later or decades later they'll get a recurrence infection and often presents a little bit more subtly this is the typical cold sore or herpes lady Alice I'm sure many of you have seen this kind of presentation that somebody before it's fairly common it's not terrible people don't go to the hospital for it but it is a social stigma of course and this is a presentation of what's actually going on so imagine the curve blue lines represent how much infectious materials on your lip and it's going up the orange down below is when that individual actually is presenting disease that you could look at them and say oh they've got something I can see sores around their mouth there are contagious before that happens this is the success story for the virus later will recur you'll get the cold sore presentation but notice the third one the third one I've plotted out here is actually the most common that people start shedding viruses on their lips and nobody even themselves well know eighty percent of you on average in this room are infected by herpes simplex virus most of you don't even know it and yet even those are you can say well I've never had a cold sore that a cementec shedding is very likely you and that's why the virus is in all of you for that because it's so successful at doing that and that's why I'm standing up here okay so to understand how that recurrent disease is working now we got to find that term neural invasion when the virus first comes up to a person's replicating in peripheral cells it amplifies like we've discussed and then it gets exposed to nerve endings in that tissue and it's eloquently able to enter those neural nerve endings and transport long distance within the shaft of the nerve to the neuronal cell of the peripheral nervous system in this case this would be a sensory neuron when I tap my lip and I can feel my lip it's those neurons that are communicating that information to my brain and that's where the herpes virus is going and once it gets entered appositives is DNA it goes into a hallmark type of infection called latency only herpes viruses truly do this but they completely shut down and go dormant and so they're basically hibernating in our nervous system for the rest of our lives but at some moments often during an emotional stress think midterm exams it will reactivate and progeny viral particles will be made in that neuron and then the amazing thing I think here is that they are sent down the axon in the opposite direction that that parental virus came in perhaps years or decades before very efficiently they'll infect the skin from the underside and you'll have that cold sore presentation sometimes they can go to the eye herpes simplex keratitis is one of the leading causes of infectious blindness in the developed world and the worst case scenario but fortunately quite rare is the virus can spread it to the central nervous system and when it gets into the brain and amplifies you now have herpes simplex encephalitis which is not it has it does not have good prognosis chances are the individual will have brain damage if they survive and survival rates are very low so that's a very unfortunate thing and now let me say for those of you do have cold sores don't go running out of here screening that you're going to die it's extremely rare but because 80% of the population has the virus the numbers game is pretty high and so this is a very significant problem for healthcare now most of the forms of disease are dependent on the virus to transport in those nerves and their fibers and so this is one of the things that my research program has been focusing on for a number of years oh yeah I haven't even mentioned what all the viruses are besides herpes simplex virus type one which is a causative agent of cold sores there's type 2 which is basically the same virus but it favors the genitals and the game and the ganglia that sensory neurons that in fact that projects to the to the genitals so that's herpes type 2 a lot of people colloquy say there's the above the Balaton that below-the-belt versions of the virus when I was in Australia for us at herpes meeting the kids the students there made shirt saying herpes virus is down under varicella-zoster virus a lot of people don't appreciate is herpes but in fact it is it's causes chickenpox and shingles of course and it's quite related to these other viruses I won't be talking about that further but I'm happy to talk about it afterwards I want to mention one other that most you probably have not heard of most herpes viruses actually have co-evolved with their hosts so the herpes viruses of humans uniquely infect humans in nature of the herpes virus of pigs in fact pigs and horses infect horses all of us all of these species have their own collection of herpes viruses that have evolved with them and are tailored to infect them very effectively but once in a while some of them can jump and particularly if the hosts are closely related to one another herpes bee also known as monkey be in fact certain types of monkeys including rhesus macaques and when the virus jumps from a rhesus macaque to a human it almost always causes that fetal and stuff a bit of disease is so rare normally but it now becomes the default outcome and if herpes simplex virus type one our cold sore virus gets into the rhesus macaque it will do the same thing to the monkey this brings up a very important concept something if nothing else I would love for you to take this away and that is that if there was a selective advantage for these viruses to actually kill us they would the fact that monkey bee can get into us and drop you means that there's nothing about your immune system that is so amazing that it can defend us against everything but we don't have to worry about that because these are under evolution selective pressure and if they do kill you they're basically burning down their own house and they go with it so those about any viruses I gain those properties throughout evolution have gone extinct and what we're left with is something much more benign but it leaves an interesting I think thought process at least a hypothesis and that is when we think about what's going to happen when I'm going to tell you about these infections you don't have to think about how we're fighting it off all the time so successfully but rather how the virus is making sure we don't die no disturbing I was hoping I would leave an impression with you okay so one of the things that I love to do in my lab is tinker and to tinker with any kind of biology particularly a virus it taught it means that you're going to start playing with DNA and so when I was fairly early my career one of the first things that we did is come up with a way to genetically manipulate these viruses and I'll just take you through a very short version of how this is done you can isolate these particles and extract the DNA out of them and so this would be a circular plasmid of DNA and you can put this with you a couple of tricks we were able to get it to stable maintained and bacteria now you might be thinking oh my gosh what is he doing and the whole point of this is that for decades the sign that the life sciences a scientific community have been using certain bacteria like e-coli to propagate DNA and manipulate it and once we have this in E coli we could use tools to start changing it and we could develop some very specific tools so that we could take any kind of change we want program it that way isolate the DNA and then put it in set million cell culture the DNA itself if delivered into the nucleus of cells is considered is called infectious you don't need anything else from the virus just that you need to kickstart the process those initial cells that take up the DNA will start producing viruses that will spread and spread and so you can harvest all that infectious material and study it one thing that we really like to do is put in what's called a fluorescent protein so a lot of marine organisms you may some of you may know fluoresce at nights like some jellyfish and so you can take those proteins and put them into other organisms including the virus and make them fluoresce so if you shine the right kind of light on them they will produce a different color light that you can then look at through a microscope each one of these dots is an individual fluorescent defiers particle that we made this case fluorescein red now I want to emphasize that each one of these dots you can actually see the pixels from the camera that captured the image is smaller than any given pixel there these particles are so small there's no way using lights light microscopy you could ever resolve them so they're actually a sub microscopic but that's where it would be and the light is just refracting out kind of like if you look through a telescope at a star far away you see the diffraction of the star but you're not actually seeing the surface at the star you're just seeing a point source diffracting light same thing here okay so intracellular transport in the axon nerve fibers then is the topic I want to get at now and of course the real experiment to do is if we could is look at how those viruses are transporting down those nerve fibers I have yet to get all my students to volunteer for this experiment but you can do something fairly easy and routine and then what we like to do is get eggs from a local farm that are fertilized and you can actually get neurons out of those little tiny chicken embryos and grow them in the dish in the lab and so that central ball of brightness are all these neuronal cell bodies are sensory neurons and they regenerate out there are nerve fibers called axons radially away from the cell body cluster and you can then infect that in the dish an image just a little tiny region of those fibers traditionally by electron microscopy microscopy this is something you might see so this is a zoomed in version of just one of those axonal nerve fibers and you might notice towards the bottom it looks like a little hexagon with a dark Center there's a herpes virus particle it's on its way it thinks it's going to your trigeminal ganglion from your left but it's full because it's in a dish the lines that you see in the background are called microtubules and they're basically stretched out ropes imagine if you're an astronaut's and you didn't have a propulsive jet pack but there was a rope and you could grab it hand over hand and pull yourself along while you're floating that's precisely what happens inside of his cell there are proteins that act as motors and they go hand over hand on these ropes these microtubule ropes to move the viruses somehow were out using those motors that this already has to move now when you're seeing there is a virus that has entered the sound fuse like I described for you before so it's only the capsid shell attached to some of those surrounding soft tagamet proteins that's what's in that image but here's where the fun begins at least for us is that I told you we can put fluorescent proteins onto this with a fluorescent protein you don't have to fix these cells and do these bombardment of electrons that are course not compatible with life you can do a nice imaging with a light microscope at those diffraction spots and watch the process happen each one of these spots is an individual herpes virus capsid on the run after has entered the nerve ending as heading towards the new the neuronal soma soma cell body where the nucleus is ultimately deposits genetic code and takeover notice that the motion is very recall precise --iv meaning it's continuous sometimes all its little pause it almost regains immediately motion to the right switch is inward bound sometimes on those they actually take a couple steps backwards and so the motion is handli bi-directional but the virus favors the inward routes the route that's going to be productive for it it's a very interesting type of motility as it turns out it's not only using cellular proteins to move but it's cracking a whip is as it turns out now you can image the fate of those particles by moving the microscope objective to the center we like to call into the lab downtown here's what downtown looks like these are neural soma they're all crowded together and if you image that all those dots lining in circles are around the nucleus of those neurons here's the overlay you can imagine that those cells like in the bottom-right illustration are half caps has now docked at these pores of the nucleus and they are having the viral DNA injected in and being taken over now one of the neat thing you can do with fluorescence live cell microscopy is you don't just have to image fluorescence of the capsid or one part one component of the virus at a time you can do two so in this case we've made a virus that encodes a protein that gives off green light or green fluorescence and the capsid shell itself is red and so what we're looking at here is a time-lapse staggering image where we image the red capsid and then we image the green and then the red back and forth as fast as we can but it while this is happening the virus is on the run so it looks like as they're moving that the colors are displacing but that's a temporal artifact just because it's moving so quickly the point that I really want to make to you is that this is an example of a protein that stays associated with that Cassatt as it goes down that axon and it's a so you can see in the illustration that I've noted that some proteins in that tagamet layers state capsid associated and some come off and are lost and this is the kind of experimentation that we did to be able to define that why is that important while the proteins that are bound to the capsid during this whole process could very well be the proteins that make it happen think of the capsid itself as a big oil tanker and there's little proteins bound to this the tug boats are really doing all the work and moving and moving it around and so here's something I think is particularly fun you can now take that protein and remove it from the virus genetically and now you have just that protein alone with no other viral material that you can express inside of a cell and what does that protein do by itself well if you tweak it just right and you can reveal its hidden secrets and hopefully this will play there we go this is not an infected cell but what you hopefully can see here is this single viral protein which has been made fluorescence is moving around inside the cell not just randomly but almost as if it's on tracks it's moving on those ropes hand over hand and there's so much of it in the cell you can actually see the ropes themselves being labeled indirectly by all that moving protein of the virus this protein is an effector of that long-distance transport that the virus uses to get into our nervous systems here's a prettier example of that using a process called confocal imaging and now you can actually see the individual spots all moving on these rope-like tracks now remember this is just a single protein that's normally bound to the CAF's that the capsid itself isn't even here I'm going to come back to this at the end of my lecture so just remember we now have that one protein that has this function identified okay but now I want to get back to really that I think the most interesting thing and that is the disease and how we deal how we can deal with it so this is this is a question that I've been thinking about all my career and I certainly don't like credit for it I was a year old when I think when Robert and sorry Richard Johnson and centric men said it must be explained for example why a virus causing no more than a cold sore and one person can produce a fatal encephalitis in another and I do think that's an absolutely fascinating question and something we need to figure out for to protect people and recently a discovery was made in New York and there's from a laboratory by an individual named John LaRon Casanova who is originally from France and he had discovered that we have a protein in our nervous system called toll-like receptor 3 now the name doesn't really matter suffice it to say that what he had discovered is he would he's a pediatrician and a molecular geneticist he traveled the world to find cases of adolescents herpes simplex encephalitis and then got skin samples from these people and then did brute force sequencing of all their DNA and try to say is there something about these poor kids that's different than all of us who don't only get a cold sore and don't get this severe form of disease and he found this they are deficient in the genes that encode this one protein but what does it do so then a collaboration began that I was thrilled to join the cats in live allowed I just mentioned was collecting patient tissue skin cells they are passed them on to Harvard who produced what they call induced pluripotent stem cells some of you may have heard the stem cell the controversy behind it these are not those kinds of cells these are actually cells not from an embryo but from skin of an adolescent child and they're turned into something that's more like a stem cell so it's pretty nice technology and the Sloan Kettering can then actually differential in to a neuron like cell but remember they have all the genetic code of that patient even though they aren't now these kind of artificial neurons they even ship them to us at Northwestern and we finished the process of differentiation and infect them and say what's going on and here's the cool thing this is what fluorescent imaging of cells look like at a lower magnification the first panel the blue on the left are the nuclei of the cells in the culture dish so that's where the genetic code is the next one are those rope-like microtubules they don't look like individual ropes because we're at low magnification so they're all bundled together at this low mag and then the third channel in red is the key one this is a fluorescent fires that we made in the laboratory and we infected these cells in the dish and notice at the top there's no evidence of an infection there's no fluorescence being made but at the bottom there is and the difference is you can clearly see on the label on the left is at the top one it was dried from skin cells of a healthy individual like you or me and the bottom one came from one of these poor kids who got herpes simplex encephalitis and lack the protein tlr3 and in culture you can mimic the susceptibility these neurons have been made to look like brain neurons the very neurons where the encephalitis would have occurred the fact that you can mimic that in a dish is a really phenomenal and I was thrilled to be part of this team I don't get the credit for that except to say that we just confirmed that results using our methods but it it gets a little get more interesting in a moment so now down at the bottom they've recently been able to make sensory peripheral neurons these are the neurons that normally get infected in all of us and where the virus will establish that lifelong infection and sure enough the virus loves to infect these cells and doesn't matter if it's coming from a healthy individual or the patient the virus infects these cells beautifully so what we're seeing is that we can fully replicate the normal neurotropic properties of this virus in a human normally we'll go into that try ganglia and it does but it won't go to the brain in a healthy individual and it doesn't up there either but if you take it if you look at the patient's then all bets are off the virus can infect everything consistent with the development of encephalitis in a person so what is this tr3 sensor protein I was telling you with well I threw a little illustration on there right next to that viral particle on a cell let's bring it up and so it is a surface protein it spans through the cell membrane and reaches the outside of the cell and it can do a couple of interesting things but for some reason that we don't understand it seems to be able to sense there's a herpes virus nearby and we're still trying to figure out exactly how it senses that but what it normally would be able to do is send off a cascading signal to this inside the cell basically saying get ready we have a problem get the battlements up and in a Cell that cascade that goes to the nucleus leads to the production of a protein called interferon which literally means interfere with viral replication that's where the name comes from and so this puts your cells into a nice protective State the defenses are armed it's ready to do battle so how is it that tr3 is stopping infection of brain neurons and healthy individuals like most of us well let's look north this so this is where we had a little fun so this was work done by a student in my laboratory and he said well let's work this backwards we know that we're getting a productive infection when we take the cells from a patient who had encephalitis and that's because as I just showed you that fluorescence imaging where you have lots of new capsules made in the nucleus of cells and they're all fluorescing and so the cells turn out that light up and we can see that so we know the cells get productively infected and so now we do some more genetic shenanigans I don't know yeah this is a reminder of that and so we do art of genetic shenanigans and we can start getting at these very specific questions and let me show you how this gives you a sense of a couple months in the lab so the next question is well if there you can't in fact a cell from a healthy individual can they even get their genes delivered into the nucleus of that cell and get and get to that stage and so to ask this question we made a virus that turns the cell's nucleus red if the genetic code gets delivered to that cell so if the genetic code is gone through the nuclear pore it will make a new red fluorescent protein the nucleus will turn red and you can image that and sure enough if you look at the patient's cells they're turning bright red bright red nuclei but the healthy ones are staying dim so that means that in a healthy person it can't even get its genes to the nucleus can it actually go down those nerve fibers to get to the nucleus in the first place for this we turn back to that kind of imaging I showed you before up top is the tr3 deficient patient they're moving retrograde ly down those axons you can see that nice process of motion there's lots that you can't see the axons themselves but there's lots going through that field of view that they're moving in the bottom is actually playing the helping individual has nerve fibers in that field of view or the cells have nerve fibers in that field of view but there know is no virus motion so the virus can't actually even go down the nerve fiber in a healthy individual and the end result like I showed you before getting to the nucleus and docking at the nucleus you see there's nothing there the healthy cells that are cells from the healthy individual are blocking all of this so what about that very first step well this was this required a little bit of extra work on our part to figure out how to do this but let me show you I think you might enjoy this we made a virus that carries an an enzyme from a bacterium called beta lactamase beta lactamase is the thing that bacteria used to cleave penicillin so that they don't get killed by the antibiotic it's their defense against that tactic an artificial molecule has been made shown down in the bottom left called CCF - the pertinent thing is that it has what's called a beta lactam ring so the chemistry under - let me just point this out is right here in red this very constrained bond is high energy and if beta lactamase is there it will cleave it and that molecule will split in two and the nice thing is that molecules actually fluorescence so when you shine light on that has CCF - inside of it normally you'll get you'll excite with a certain wavelength of lights and then you'll get a certain wavelength of light back out but if it Cleaves you get a different wavelength of light because the light energy is not transferring between these two parts of the molecule and so by doing that you can ask is the virus actually getting in because the virus excuse me the virus is in carrying that protein as part of its egg event that gets deposited in the cell upon fusion and so here's the basic results if you take a virus encodes beta lactamase and put them into an healthy the h or patient cell you can see that the fluorescence ratio is changing and that is indicative that the virus is in fact fusing into the patient cell but not into the healthy cell and just as controls if you don't infect the cells you don't see any background and if you infect the cells of the virus that doesn't actually encode that bacterial enzyme you don't see the change either so it only happens when the virus can get in and it has that foreign enzyme and carry along with it to give you that readouts the virus is getting into the patient drive cells and so what we conclude is the virus can interact somehow with the cell surface of a brain neuron and that T tlr3 protein senses it and immediate it's up a force field and it's got to be close to instantaneous because when the virus is coming in and wants it's going to fuse very quickly and this counter tactic to prevent the fusion is going to be very fast nothing like that's ever been described before and we don't know yet exactly how the cell is able to do that but it's unique to our central nervous system neurons our peripheral nervous system neurons don't do this so HSV is tuned to infect peripheral sensory neurons where it goes latent causes our lifelong infections but it's also tuned to avoid the neurons of the central nervous system and thereby prevent the formation of encephalitis and lethality it's happy we're happy but I'm not completely happy I still would like to stop it altogether and so I would like to bring you back to this experiment where we took that one protein and put it into cells and saw that it could transport along those ropes along those microtubule ropes all by itself and the reason that was an important experiment for us is now we don't have to be a virologist anymore we can be much simpler we can now start studying the mechanics of that protein it's all by itself which makes our lives a lot easier and so from that we've now been able to start expressing variants of the protein and try to understand basically by dinging on this part of it dinging on that part how is it actually achieving this function here's one more example of what I've been showing you all along this is a wild type virus with a red fluorescent polyhedral capsid shell it's moving down axons and in a plastic dish heading to the neural soma or it's going to inject it's the aims of a nucleus I'd like to call this presentation this this phenotype as a marathon runner these are viruses on a mission they're moving at about two microns per second that's probably an arbitrary number it means that by the time it's leaving the cells of your lip and going into your nerve by to the time that it gets into those neurons then your try general sensory ganglia is about 10 hours and it's running the whole time that's pretty good run as we started digging on those proteins that were associated with the capsid shell that when it enters we found we could debilitate this I like to call the motion you see down below the drunk marathon runner you'll notice that the particles are no longer predominantly heading to the right although some quite are the motions still there but now they're bouncing around they often go the wrong direction seem to get lost and the nice thing about this is that this is over a very short distance in a microscope when you think about the distance from here to about the center of your head where your sensory ganglia are that's an extraordinary long distance that's the ten-hour track that the viruses have to take and acting like that they're never going to make it so why is that interesting let me end with you with what we're recently starting to do I'm very excited by this I hope this is gonna work we're developing what we call the non-invasive herpes viruses vaccine and so basically using the discoveries that have been made in the laboratory or making these viruses that are Noman are able to invade any part of our nervous system not just the central but they can't even get into the peripheral branches of our nervous system anymore so these viruses then when they in fact so what so let me just restate what I've already said and I think you all know but make sure on the same page if a normal virus infects us or replicate it in that peripheral skin tissue and then some of them will go down our nerves and get to the cell bodies of the sensory neurons and inject the DNA and now you are infected for life what we have discovered is that the invasive machinery that visited viruses use is specifically not on the surface of this of the viral particle like many people have been thinking for decades but on the inside it's that inner surface that is revealed when the virus enters a cell that contains the specific proteins to achieve this feat so we like to call these new to recombinant viruses they no longer have the neuroinvasive component but when applied to the surface skin they replicate they're just as well as normal virus we would from nature they just can't get in the nervous system so they can't run and they can't hi and what's nice about that is that if they get stuck out there that gives our immune system time to kick in and the mutant mammalian immune system is a beautiful thing if given a chance and so the immune system will destroy it it will sterilize it and it will remember and what we're learning is that it will now prevent not just disease at the surface but it prevents subsequent viruses from ever getting into our nervous systems this is not a fun experiment but I think it's someone that is worth sharing nevertheless there are some viruses I told you like monkey B when it gets into human is extremely lethal this is a virus called pseudo rabies virus it's a virus normally found in pigs but it will jump to other farm animals it causes something called the Matich and cows it costs at something called pseudo rabies and dogs it jump it silly to anything and go straight to the brain of that other animal other than the pig and kill it by encephalitis with the exception of all higher primates which includes almost everybody in my lab so it's a fairly safe but deadly virus to study and when you give this to a mouse just gently inoculate it the mouse will unfortunately succumb to that infection typically just over two days that's that first plot on the left right over fifty hours but if you vaccinate with one of our neuter viruses and wait different amount of days before we challenge with a wild-type isolate of a nasty virus you can see the animals are starting to survive longer and longer and by two weeks not only are they surviving the infection but they're showing no symptoms it's like you give them water when we have isolated tissue from these animals to say is there any evidence that despite the lack of disease that the virus is still getting there at all so far our initial results are no we cannot detect any evidence of viral material in the nervous system of animals now this of course is just an initial experiment it's an artificial experiment done in a rodent but I think it leads to hopefully a useful tool in this battle I want to end by the same that there are a number of neural invasive viruses in nature many of which in fact humans that we deal with rabies polio which we mentioned from March of Dimes are two of the most common ones that people will recognize those viruses despite being nasty and potentially deadly or paralytic don't usually infect us easily during the height of the polio outbreak in the 50s only about one in 200 who got infected actually got pulling on myelitis for the most part it was an enteric pathogen yeah swallow it it replicates in your intestine you pass it out good sanitation and washing hands does wonders but only one in 200 so it was an aberrancy when that virus went into the nervous system and in fact when it did go in it was under no it gained no selective benefit for the process it didn't spread to other people better in fact it for spread works because now it's stuck in the nervous system deep inside of you and it can't get out again very well rabies does in fact are nervous isn't very well and then it can spread from person to person or animal to animal but for it to spread your typical e-beam bit by a rabbit the vote you could take a bath in rabies viruses and it wouldn't hurt you it's only because the animal is dramatically damaging your skin and exposing muscle and nerve tissue directly to the virus that that virus can get in and do it's horrible damage to your nervous system these viruses just require a gentle kiss and I think that's another one of the phenomenal things about these viruses is that they have figured out this way to deliver genetic information to our nervous system and so despite all the detriment that they caused it's almost you can almost imagine a Star Trek future where one day we can actually just paint on something onto a person's lip and treat a genetic disorder from their family that's in the nervous system using technology that nature provides and with that I just want to quickly mention the people in my laboratory particularly Alexia Richards a postdoc and osa family while a fellow graduate students they did a lot of the work Alexia is working on the vaccine and ELISA Fahmy is doing the work on the central nervous system infections and I'd be amiss not to mention the people have supported financially this research on the top right and her wonderful collaborative science is a team effort I could never have done any of this alone Patricia solar's and Gary Picard are neuro scientists there's young on Casanova I mentioned is the pediatrician collecting tissues from around the world Lawrence Tudors turning them into neurons and Luigi neurons Lowe is turning them into these adduced pluripotent stem cells and make all this possible and my immediate left is Alexia into my mirror right is oh so foamy Lila fo and with that I'd be happy to take any questions and thank you for listening [Applause] okay will your vaccine work with somebody that's already infected and the second question is what's your timeline for developing an effective vaccine we're trying to go as fast as you can so the the first answer is will it be able to treat somebody who has already been infected and the answer is it's not going to cure that individual once herpes has its foothold in the nervous system it's going to take some pretty fancy technology that I can't even imagine you have to get it out of there the genetic code is now entwined with our genetic code in those neurons and that's a tough challenge but by boosting our immune response we can make potentially make reactivated events later in that person's life less severe and I think an important one to consider is a pregnant mother and so one in two thousand births in this country the mother inadvertently exposes the newborn to herpes simplex virus type 2 in the birth canal and the infections that happen in the neonates can be mild not that mile but recoverable to severe and I would just love the idea of being able at one point to say first trimester this person's never had HSV - and that's important because it's the mother to get it contract HSV - during pregnancy that are most at risk to passing it to the newborn and so if they don't have it give them the vaccine perhaps on the lip let them develop their response and by the time she's ready for birth there won't be a problem that's that's my dream I don't know if that's actually gonna work but that might be one way to go about it timeline for the vaccine so we have just some Northwestern is being very supportive about this they have actually put me into a mentorship program in Chicago where we're working with entrepreneurs and businessmen to try to figure out how to form a company to try to get this to market but it's in its infancy at the moment we're just getting started thank you in recent months the IHMC has been blessed with a number of lectures concerning epigenetics and new nutrition science that allows certain foods to engage the enzymes and cells enzymes and all the DNA within the cell will allow the cell to intentionally make its own any oxidants at exponential levels there's that whole potential for the immune system getting greater resistance to viruses yeah that's a great question and you know this is a real burgeoning field people been looking mostly at bacterias thus far and the idea that are not only we have a back to roam that is our flora that exists with us but a viral of all these different viruses that exist particularly in our test ins is I think going to be a very fascinating piece of research going forward but at this point we don't know yeah when I was a child I got these canker sores in my mouth I mean all the time it was just debilitating yeah and my mother took me to the family physician and he recommended a series of smallpox vaccinations I took the smallpox vaccinations and they went away and I mean to this day maybe I'll get a canker sore once every six months or so okay but my daughter had the same thing when she was born and we probably took her to six different physicians looking for someone that could recommend something no one could recommend anything and since there's no more smallpox vaccination is there anything to help combat canker sores in the mouth so canker sores can be caused by many different scents and so the problem is is that it's not always quite clear let me take a step back and to say I am a PhD researcher and not a clinician so take this with a grain of salt please but I I will say that canker sores can be caused by many different things herpes viruses like simplex is one and so probably I'm not even familiar with smallpox being a major cause of this but one thing is interesting about smallpox is that when you vaccinate without agents it can often cause cross protection to other agents and so it might have been an indirect effect that helped you as far as your daughter now I don't know what to say that if it is simplex then all you need is some a cycle of your a drug that is commonly prescribed to treat active lesions with simplex it doesn't cure the disease by any means but lessens the symptoms of the outbreak and it's a wonderful drug it's probably one of the best drugs that medicine has ever developed because it has very little to no side effects even if overdosed accidentally so you might want to talk to your doctor about a possible prescription there but unfortunately I can't say more than that yeah anyone else okay I think okay that's fine all right here thank you sure is a relationship between stress and the reoccurrence of herpes absolutely yeah I imagine most people have ever had a cold sore wall you're probably not Ian yes right now yeah and so yeah with with kids in college and school it's right around exams it doesn't have to be emotional stress the stress can come in other forms so for instance a sunburn on the face can often reactivate it as well certain kinds of chemicals sometimes found in fruits when put onto the lips can also be irritating and be enough to cause a reactivated event the actual signaling that causes the viruses that are dormant to turn back on again and cause that recurrence cold sore is really still mysterious there are a lot of people researching it and we're getting some hints but there's a big black box there still all right next what is the relationship between and among the different kinds of herpes and particularly I'm interested in shingles and whether there is any correlation between whether you do or do not get cold sores and whether you will or will not get shingles uh-huh and should or should not have the shingles virus as immune as a vaccine or just have it internal it's a vaccine so I would again I'm not a clinician but I can't see why not to get the vaccine at this point it seems like it's very effective it was it's been used for a long time started off in Japan and so I have a long history there before it came to the West the shingles vaccine is the same thing as the chickenpox vaccine is just a higher dose one thing to know about those vaccines is like the one that I'm proposing here we refer to them as live attenuated so some vaccines are just dead pieces of pathogen and they can never do you any harm other than the fact that they're in irritants and and that's about it a live attenuated virus actually can do something while it's in you the polio vaccine was like that and it turns out people who get the oral polio vaccine two drops in the mouth this is how they're trying to eradicate it from the work from the world they actually will release infectious polio as a result of taking that because the virus will start to regain its virulence while it's in that individual and so lots of family members can sometimes be exposed to a nasty version of the virus chicken pox is much more stable but it does actually Rican tain its neuroinvasive property so what I'm talking what I told you about here today is new so the chicken pox vaccine and the shingles vaccine when you get it goes right down your nurse and it sets up a lifetime vaccine infection in your sensory nurse and those things can reactivate and so the only problem there is if at some point you have some kind of an unusual condition that would make that vaccine more virulent than it should be so for instance if you've become immunocompromised for some reason your immune system is dysfunctional that vaccine then could become problematic in fact they don't like to give the vaccine to HIV patients because their immune systems compromised and they can lead a complication so that's the only downside but right now based on all the data a lot of people in gaming's vaccines it's looking really impressive for the typical public and so I would just say go for it at this point answer the simplest question did I so the simplest question is as far as I know there's really no correlation there the viruses although they're related are quite different in how they infect us where they in fact us and I have never heard of any clinical study that said by having one you're more likely to get recurrences of the other okay yeah I have had what's now been dalek diagnosed his mother raised meningitis since about 1973 which causes the whole neural affects you basically get the stiff neck a horrible headache your brain swells I had it recurrently just constantly and then they was finally oh just about four years ago we figured out that it was probably mullah race cuz I wasn't dead so probably wasn't encephalitis nice it it appeared the first time right after I had actually had encephalitis mosquito-borne encephalitis and Middaugh ways occur with were actually herpes one lesion I still get it I take BellSouth clavier when I do which lessens the intensity of the attacks I mean I used to have to do things like take things to prevent having all kinds of very bad things happen but so it's a is helping control the neural affects as well it does control the neural affects but just wonder does that mean that the it's the tlr3 expression that's screwed up with me well probably not because if it was you probably would have had the encephalitic infection that's so that's what that was associated with now here's the thing that's a little tricky is that there are a lot of rare presentations of disease associated with any type of infection and you might be one of those unusual in the cases of something that most people will never present and the difficulty there is you can have someone like genre on cuts no traveling in the world finding these poor encephalitic kids but finding people like you enough of them so we can really say I mean everybody's got random mutations in their DNA that's just that's just part of be alive and so be now say well which one of these mutations are actually associated with this type of disease that this person has is difficult unless you have a huge cohort of people that all have the disease you can say this is what they all share in common and so if there's not enough people like you it's very hard to figure out netic Lee what's predisposing you to that kind of disease if it even is genetic for that matter alrights meningitis do you know you know I don't know off the top of my head I'm sorry yeah I have one last question sir here numerous viruses infect the the insect Kingdom in rather bizarre ways of the zombie ant for example yes are you acquainted with any similar virus infections in humans don't kill us but change the way we behave absolutely rabies rabies is the classic it's the prototype in humans it's it's a virus emits got to be from another planet it is just so crazy so it's normally in bats as it turns out as many of you probably know but if you get into many mammals from the bats including dogs and Coons and so when the virus gets into animals it starts off as this piercing bites either by the bat or rabid dog the virus then ascends up the motor neuron to normally caused your muscles to flex and as slowly works its way up into the brain and the crazy thing is it then goes first into a part of the brain called the amygdala and this is part of the brain that controls emotion and so when the virus gets there it makes you violence makes animals wants a bites you know for all anyone who happens to be a science fiction fan this is the zombie apocalypse right this is what they almost always say it is the science fiction stories that rabies is what guy got the got the world it's kids it's going into the brain is changing our behavior making us act this way or mostly not usually humans but mostly dogs but humans as well or other animals but then the craziest thing of all is that after the amygdala it travels into the salivary ducts and it's secreted in this live at just when it's made you aggressive and so the rabid animals biting and transmitting tons of viruses and injecting it into muscles of other animals it's is almost science fiction I'm not pleasant note let's thank our speaker [Applause]

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Length: 59min 49sec (3589 seconds)

Published: Thu Dec 08 2016