COVID-19 and the Impact on the Central Nervous System

hello everyone nice to see all of you for our first uci biocide virtual dean's distinguished lecture uh before i introduce today's uh lecture and our guest speaker i just want to uh thank everyone in the biocide dean's office for helping put this uh virtual lecture together and for making the event uh success as uh many of you know this lecture was born out of a desire for us to better inform our local community here in orange county and southern california about all the cutting edge research being done in the school biological sciences at uc irv at uc irvine obviously because of the virtual nature of this we're able to even expand beyond our traditional borders as many of you know in the past these distinguished lectures have been hosted in a very large venue on campus and while i regret that we can't be on the campus right now we wanted to uh continue the series and after all what better time to shine a light on life science research during an unprecedented global pandemic as many of you know as biologists we have been fervently studying and many aspects of this novel coronavirus disease and in fact our school's organizing theme of mind body world continues to position us to make progress against covet 19 and future pandemics the uh importance i think of supporting life science research and education has never been more compelling uh than it is uh right now and we appreciate all of the all of you joining us on zoom today to learn more about the effects of coronavirus on the central nervous system as a matter of fact i can't think of a more timely topic and just as i was about to come on i got a little notification from cnn that just said that uh recently california became the second state in america to surpass over a million cases of covet 19. and so with that i think we're very lucky to have our own uh speaker professor thomas lane uh who joined the uci school biological sciences faculty back in 1998 and is now a chancellor's professor of neurobiology uh and behavior he has uh a true expert on coronavirus infection and has a very prolific and a very detailed research career studying the effect of coronavirus on the male on the brain for over two decades uh he is a well regarded uh scientist he is eagerly sought after to be a collaborator by many of us on campus and throughout the nation and he has played many roles at uci including serving as director of the multiple sclerosis research center and uh associate director of the institute of uh for immunology he's been awarded a national uh ms society collaborative center award and a cirm early translation award and today tom will share with us his research about coronavirus about learning how sars cov2 the virus that causes cobit 19 gets into the brain and what his team has discovered for its impact on conditions like diabetes and cardiovascular disease so let's all please welcome professor tom lee okay uh let's see i think everybody can hear me frank uh thank you so much for that extremely kind introduction um as well as the opportunity to talk about something that's clearly very important to us as well as a number of people and this is uh examining the effects of covenanting on the central nervous system i would also like to extend my sincere thanks to everyone that's been able to attend the lecture today i know everybody's very busy so thanks for taking the time and uh coming out to hear what we and others at the on the uci campus are doing all right so what i'd like to do is uh provide an outline of the talk this afternoon and i'm going to begin by providing an introduction to coronaviruses then i'll move into sars cov2 which is the ideological or causative agent of covid19 discuss some of the aspects related to the virology and immunology of this disease then move into cns conditions or symptoms that occur in some individuals following sars cov2 infection and then uh finally i'd like to conclude as frank pointed out on some of the pre-clinical mouse studies that we've been doing using sars cov2 infection of susceptible mice and specifically looking at the cns disease that occurs in these animals and then finally i'd like to uh conclude with some perspectives and then i'd be happy to answer some questions from the audience okay so um coronaviruses as this title implies this is a virus that we've been living with for decades if not longer and here are uh four transmission electron micrographs depicting uh sars kodi one sars cov2 mers cov and then uh mouse hepatitis virus which is a mouse coronavirus and the point of me putting these images up here is that although these are different types of coronaviruses you can really see from the images depicted that how these viruses acquired their name they had these solar crowns or coronas and this uh is the result of the spike glyco protein shown here in this cartoon which extends furthest from the surface of the virion and i would imagine most of you have probably heard of the spike glycoprotein because when social media is discussing the efficacy of vaccine or vaccine development most are talking about directing immune responses that target the spike glycoprotein because this mediates attachment to the host cell receptor and i'll talk about that in a little more detail later in addition to the spike glycoprotein there are additional structural proteins that are very important you have the membrane protein which is shown here as well as the envelope protein and these proteins all work together to encircle or protect the rna genome which is shown here by this large black line and the coronaviruses are single-stranded rna viruses um in the virology world you have dna viruses and rna viruses and then retroviruses but caprona viruses are rna viruses and they are among the largest known not just in physical size but really what this relates to is the size of the rna genome so these are roughly 31 kilobases or 31 000 base pair or base pairs in length and the four corona i'm showing here are a member of the beta coronavirus family there are actually four families of coronaviruses alpha and beta coronaviruses are um primarily able to infect and replicate in mammals and then the gamma and delta coronaviruses are primarily restricted to either fish or birds and so for the beta coronaviruses because they have a wide range of animal hosts which include humans obviously but also uh pigs cattle rodents such as mice and rats bats and camels they have a variety of different diseases that can be initiated in these animals upon infection and um the symptoms can be varied and they can be either benign that is really no evidence of infection at all or they can be much more severe which can include pneumonia diarrhea peritonitis which is inflammation of the perineal cavity or the gut as well as neurologic disease this is primarily characterized by encephalitis which is an inflammation of the brain and this can be accompanied by neurodegeneration so just very quickly um there are a number of neurotropic animal coronaviruses um that means coronaviruses that are able to infect and replicate within the central nervous system and one of these is marine coronavirus and as frank alluded to for the past two decades our laboratory has really devoted a lot of attention in understanding um the immune mechanisms that are elicited in response to infection of the central nervous system of susceptible mice by these marine coronaviruses that either help control the virus or contain it as well as potentially contribute to neurodegenerative disease in addition we've explored for quite some time molecular mechanisms that govern neuroinflammation in response to infection of the cns if you own a cat you've probably heard of a feline infectious peritonitis virus this is a very serious disease in cats it does induce this uh inflammation of the gut and it is under many circumstances lethal but the virus itself is cns penetrant it can get into the central nervous system of these cats and also cause problems and then finally another example i've included is the porcine encephalomyelitis virus and so this is a real problem of the swine industry where these viruses can get into the cns and cause or just wreak havoc uh for uh farmers that build pig herds all right but the coronaviruses that you're probably most familiar with either directly or indirectly are these four human coronaviruses and these are really associated with mild seasonal seasonal respiratory illnesses um and so there are four of these as i indicated 229 e and oc 43 these were discovered in the 1960s at the university of chicago as well as the nih and then after the first sars cov1 if you will infection that occurred in 2002 this resurrected a lot of interest in coronavirus research and then two additional coronavirus were discovered in hong kong and then the netherlands and so these collectively are referred to as the common cold coronaviruses or ccc and roughly about 15 of upper respiratory tract infections are associated with um these four coronaviruses they um from a clinical standpoint they're almost indistinguishable from rhinoviruses which are another quote unquote common cold and what's interesting though about 90 percent of people tested to have antibodies to at least one of these particular viruses so we've all probably been exposed but what i want to transition to and use as a springboard and for the rest of the talk are the coronaviruses you've definitely heard about and so uh i'm gonna again i'll begin with this severe acute respiratory syndrome virus or sars which is now known as sars kovi one or sars one and so this emerged in 2002 and late 2002 the first known cases of an atypical pneumonia were described in fushon china by february of 2003 china had reported over 300 cases of these acute respiratory syndromes occurring in the guangdong province by april of that year it was identified as a coronavirus as the causative agent and then it spread to different countries around the world but ultimately it was contained in terms of how this got into humans it's clear now that bats were the original host of the virus but then at some point it was able to spread into civet cats as well as raccoon dogs and so these are animals that are often found in these exotic wet markets at various locations throughout china and it was during these wet markets potentially that it jumped into humans all right and that that in itself is not that big of a deal because a lot of viruses from jump from animals into human but this is a respiratory virus and the main thing that's very important to keep in mind once it jumped into humans it was then able to spread human to human through aerosolization of respiratory droplets containing the virus importantly no antiviral therapies are effective against this sars1 and there are no vaccines to date and i found this cover of time magazine and note that it's may 5th uh 2003 but so many things are relevant in this cover to what we're dealing with right now with sars2 uh we're all wearing masks we're curious about mechanisms by which the virus spread and then how scared should you be all right and so we've all um asked these questions and certainly we're all wearing masks every day um here is just a map that i compiled from the who that represents the communal cases from 2002 to 2003 for sars and um you can see that it was heavily concentrated in china but it did spread usually by air travel to other countries around the world and continents um including north america uh with canada being uh having on slightly more cases than the us um but by the near the end of 2003 who uh indicated this this this virus was contained all right and so one reason why it was even though there weren't any antivirals or vaccines is because the viru the virus is very hard to transmit you had to be very sick in order to spread the virus and the primary people that were affected by the virus were either family members that lived in close proximity with an infected individual or health care workers all right and then so once that was discovered you could quarantine or isolate infected patients and that really dampened the spread of the virus to where now we really have no problems with this so now let's jump to 2012 and this is when the middle eastern respiratory syndrome or mers coronavirus emerged first reported cases appeared in saudi arabia in 2012 or september of 2012 and this virus was primarily confined to countries either in or near the arabian peninsula and then in november of 2012 the virus uh that was isolated from these people was identified as a novel coronavirus this was a paper that appeared in new england journal of medicine and similar to sars one bats uh were are the original hosts however in this scenario it spread to camels and then camels are the vector to humans and again so once humans acquired it they were able to efficiently spread it from human to human similar to sars one there are no antiviruses or antiviral therapies available or vaccines i acquired this image from uh nih website um i like this because this depicts mers coronavirus particles colored in purple they're not really purple that have been infecting a cell monolayer in the laboratory these are called vero e6 cells but it really kind of shows how the virus can spread from cell to cell and as i indicated this virus um there weren't that many cases about 2500 cases yet it had uh close to 900 deaths and so this is a case fatality rate of 37 and i forgot to mention this for sars won the mortality rate was around uh 10 um but here again as you can see the um virus is confined uh almost exclusively to saudi arabia but also in areas surrounding this other countries did get involved but it was very minor hit again why because the virus doesn't spread very efficiently you have to be very sick with symptoms in order to spread the virus family members again and healthcare workers were the primary patients or people that were affected by the virus and i just want to put one thing in perspective if you consider the entire number of cases that we just discussed with sars1 and which is i think there were 8 000 cases total and 2500 cases here that's almost a daily um infection rate or a fraction of the daily infection rate uh in the united states alone for covet 19. which now allows me to bring up covet 19. so as i indicated the causative agent is a another coronavirus called sars cov2 it causes the disease coronavirus disease 2019 or covid19 and i put this article up because i think it really captures what's very important and the title is another decade another coronavirus and i think that's very important because we had stars one in the early 2000s mursco v 2012 and we still are dealing with this virus right now it's not been eliminated and then sars uh cov2 this article was written by stanley pearlman who is a outstanding clinician scientist at the university of iowa who is a internationally recognized expert in coronavirus pathogenesis um so just a quick background on this i i captured this image because i actually remember reading this article in science which is a very high profile scientific journal and note the time stamp january 3rd 2020 and what caught my eye was a novel human virus pneumonia cases linked to seafood market in china caused concern and so i immediately thought well this sounds a lot like sars one and indeed if you look at the time frame frame here which i'm sure everyone is familiar with because we all live through it we had december 31 2019 the who is informed of pneumonia cases of unknown cause in china by january 7th of 2020 this identified cause of pneumonia was a novel coronavirus two days later on the january 9th first death was reported in muham china by february 11th of 2020 the virus was named sars kobe 2 and the who names the disease coven 19 or coronavirus disease 2019 by march 11 2020 uh it's declared a pandemic status by the w.h.o um this is just an image or images i captured from johns hopkins uh public coronavirus or coven 19 website i'm showing the distribution of cases around the world this is the united states showing the distribution of cases and then finally i thought this was a great image captured from the san francisco chronicle at the beginning of the pandemic stay at home which is of course uh something we all did now the one thing that um sars one mars covi and sars2 have in common is they are primary again a respiratory disease that in which the lung is involved and in severe cases it's uh people will develop an interstitial pneumonia and die from that all right so that's what they have in common um i'll go over some other aspects that set stars cody two apart from the other two coronaviruses but first i just wanted to go over very quickly the immune response to sars cov2 infection because a lot of us have been hearing about different aspects of the immune response antibody responses they come up they go away and this is also very important with regards to the development of effective and safe vaccines and while i'm concentrating on stars kobe 2 and the immune response that ensues this actually is very similar to many different types of viruses okay so here we have um infection by sars kobe 2. it goes into the lungs and what happens very quickly within primarily days you have the virus is taken up by a variety of different cell types called antigen presenting cells and here is a dendritic cell or dc and the job of these cells is to engulf or capture the virus and then what it will do is two things it will migrate back to lymphatic tissue or lymph nodes and so if you're sick you know you have swollen lymph nodes and they get they get inflamed that's because your immune system is developing a response to that infectious aging so these antigen presenting cells like dc's go back to the lymph nodes and what they do is they present antigens or viral proteins or peptides to uh components of the adaptive immune response okay so that these apcs are antigen presenting cells are components of the innate immune response and their job is to communicate with cells of the adaptive immune response and specifically t cells and then subsequently b cells so very quickly they're just different types of t cells some that elaborate or secrete a lot of cytokines that allow for cell to cell communication other t cells help regulate or dampen inflammation other t cells that are elicited are cytotoxic t cells or ctls what the job of these cells uh the exclusive gel almost is to kill virally infected cells and then you have another t cell subset called follicular helper t cells and what these cells do is they provide information to b cells that have this antigen receptor on its surface these cells expand and can secrete antibodies that are specific for viral proteins and in this cartoon we're showing that antibodies can be specific for the spike glycoprotein shown here and why that is important is because if you have antibodies that can bind to the spike glycoprotein it can inhibit the ability of that virus to infect a cell and subsequently spread okay now this is part of the acute immune response and upon resolution of infection many of these lymphocytes will go away but you will have two remaining populations of t cells and antibodies secreting cells or b cells and these are called memory t cells or b cells and why are these important because if you become re-exposed to a virus that you've generated immune response to these memory t cells are rapidly expand and migrate to the site of infection to very rapidly eliminate the invading viral pathogen probably before you become symptomatic and so the question that really is important with regards to sars code v2 is a are we generating sufficient populations of memory lymphocytes so that if you have survived or gotten through covet 19 if you are re-exposed to sars kobe 2 are you protected are these memory lymphocytes sufficient to protect you from re-infection and this is extremely important all right in addition this whole formation of memory cells is the basis of vaccination and so what people all around the world companies around the world are trying to elicit or build vaccines that generate effective memory populations of cells to protect us from subsequent re-infection or just infection primarily all right now clinical features of covet 19 i'm sure we're all familiar with this but age is a major risk factor in general if you are relatively young as defined as less than 50 years of age which technically i'm not young anymore um you are either asymptomatic or will have very mild disease mild disease meaning either fever fatigue or fatigue or dry cough you might get a mild pneumonia but itself it'll resolve now as you get older and or have co-morbidities which means like a pre-existing heart condition or metabolic diseases you have an increased likelihood of developing more severe conditions including shortness of breath breath and in severe cases you might have to be admitted to the hospital into an intensive care unit close to 70 years old or greater with multiple comorbidities so again obesity diabetes heart conditions then you have a much greater chance of having severe disease uh being hospitalized developing what's known as ards or acute respiratory disease distress syndrome which is a massive inflammatory response in the lungs that's a result of overt cytokine expression you've probably heard the term cytokine storm well that mediates this inflammatory response in the lung and this can be fatal in certain people in addition cardiac injury can occur in elderly individuals as well as multi-organ failure all right now what is the receptor for sars cov2 because this is important in understanding the biology of the virus so all viruses need to get into a host cell in order to replicate they need to get into the cell release their uh genome and then pirate or employ host cell machinery in order to replicate and make progeny virus no different for sars kobe 2. it utilizes a receptor that's known as angiotensin converting enzyme 2 or ace2 and so how you have to think about this is basically if you needed to get into a locked house you have a key and you insert it into a lock open it up and you go in it's no different for a virus getting into a cell the spike glycoprotein is the key the lock is ace2 this engagement of these two allows then subsequent entry of the sars cov2 virus into the host cell and then the viral gema genome is deposited there is a hierarchy in terms of expression of ace2 protein and mrna in different tissues throughout the body and this to some extent will dictate the ability of the virus to spread oh one other thing ace2 under normal circumstances regulates normal vascular integrity and function and in fact there are drugs available right now that target h2 for people that have problems with hypertension all right so um what does sarosco v2 or covet infection of the lungs look like this these images i'm showing here are the result of a wonderful collaboration i've had for years with a truly outstanding clinician scientist dr rob edwards who is a professor in the department of pathology here at uci he has been able to perform autopsies on individuals that passed away from coven 19. this down here is lung tissue acquired from a 60 year old male that had extensive comorbidities and what i want to first focus your attention to are normal lungs so this is a just normal lung tissue that was acquired from an individual years ago that we cut to look uh what an uninfected lung looks like and really what i want to want you to look at under low power as well as high power is that overall there's a reduced cellularity and there's a lot of air right and that's what you want your lung to look like right because lung is for gas exchange in contrast if you look under low magnification of this individual that passed away from covid19 you see not a lot of air but an enormous amount of inflammatory cells that have migrated into the lung and then the brown staining here is uh viral rna and all of this work was done by the staining and everything was done by an outstanding uh second year graduate student hema oliveria that's in my laboratory that um she's just really a workhorse and she did all the staining here and what you can appreciate is this this massive involvement of a number of cells that are infected by virus um and that ultimately led to the passing of this individual okay so what are some of the unusual clinical manifestations of coven 19 that sets us apart from sars 1 and mars kovi one is there is a heart disease that's been reported in individuals as well as kidney damage there is also inflammation of blood vessels uh you've probably heard quite a bit about stroke and this is the result of thrombosis or coagulation that can occur there also are hyperinflammatory syndromes that can occur in both adults as well as young children and then a very important aspect that's related to sars kobe 2 or coven 19 is that you can have asymptomatic spreaders all right and what that means is that you can acquire the virus you do not have any symptoms that would make you aware that you have been infected so you don't have a fever a runny nose etc etc yet you can shed or spread the virus through respiratory droplets all right which is the primary means of spread okay and that's in stark contrast to sars one and mers co-v where you had to be very symptomatic in order to spread the virus in addition there's an increased frequency of neurologic disease or conditions in individuals with covid19 that set it apart so that is very different from sars one and mers co-v there was some mild cns involvement but not very extensive but there weren't that many cases right so it wasn't really studied that much but neurologic symptoms are common in coven 19. and so these could include uh in quote unquote uh benign symptoms which are loss of sense of taste and smell although that must be extraordinarily disorienting if you have this in addition you individuals complain of dizziness and headaches again stroke is a very uh serious problem seizures have been reported and then again inflammation of the brain so this is encephalitis or meningitis and what meningitis is is simply immune cells have migrated to the meningeal sheath that encircles and protects the brain in addition loss of myelin uh which are is a lipid structure that protects axons this has also been reported to occur in some individuals all right but a very i think disturbing and very important facet that's associated with um covet 19 are these individuals that have acquired the name long haulers and so i captured the cover or the title of an article from stat that appeared in august and it really sums it all up long after the fire of a covid 19 infection mental and neurological effects can still smolder all right and i apologize for the text in the slide but i think this is a very important component of this disease that we really need to understand better so these novel coronavirus symptoms in some individuals can last weeks or even months and again these are referred to long haulers and the one thing they have in common they've recovered from covid19 and test negative for virus primarily through nasal swabs right doesn't mean the virus might not be persisting within the central nervous system which is something i'm very intrigued about now about 10 of cobin 19 pitch patients can become these long haulers all right and interestingly this can affect anyone all right either young old healthy no comorbidities co-morbidities and it's been detected in patients who had severe forms of the disease that were hospitalized or those with very mild symptoms and the the symptoms are not simply restricted to neurologic conditions there are a spectrum of symptoms that are distinct from cns type disorders but because this lecture is focusing on the neurologic aspects of uh covet i'm just listing here some of the symptoms of these long haulers within the context of neurologic disease these can be headaches which can be mild or but can be debilitating in some people the recurrence or the emergence of loss of taste and smell even if these people did not originally have these symptoms the most common comp complaint is a brain fog all right where you just feel like you're in a fog memory loss and then excuse me difficulty concentrating now i just added this because there's a really nice paper that came out recently in bio archives from tony weiss corey who's a really sophisticated neurobiologist at stanford university and actually tony and i were postdocs together at scripps um and what he reports in this study that in looking at gene transcription profiles in very specific neuron subsets in the brains of cova-19 patients he reports the expression of a constellation of gene subsets that have been associated with problems in cognition and then very interestingly schizophrenia as well as depression and i think the one thing i want to point out is that this number about 10 percent of the coven 19 patients well you might say okay 10 that's not that many but if you look worldwide i think we're over 50 million people that are infected or have been infected with the virus and you know most of them will survive but what are the long-term consequences of this particularly if the virus is able to persist within the cns and i think this is something that's really important to study all right so what are the questions that we as well as others around uci and as well as the world are asking with regards to covet 19 and neurologic disease so um what are the mechanisms by which this occur and so how i've broken this down is are there direct mechanisms so in other words can the virus infect and replicate within the cns or cells specific to the cns which can either directly kill the cells or alter their function now we know from the literature that's emerging and that i've studied neurological consequences of cns infection we can detect viral rna in the cerebral spinal fluid as well as in the cns vasculature in addition there are some reports of virus in the parenchyma or deep cns tissue is there an acute if you have an acute cns infection by the virus does this result in enhanced or increased neural inflammation and this results then in tissue damage of the brain alternatively or collectively can virus persist within the central nervous system and does this result in chronic neuroinflammatory conditions and here's where we can use infection with mouse coronavirus uh to inform us on some of these processes or some of these events because mouse coronaviruses love to persist in the cns and as a result there is chronic inflammatory responses and ensuing neurodegeneration um alternatively you can have indirect mechanisms by which infection with sars kobe do could or two could induce neurologic disease and what i mean by that is simply in some patients you have this acute inflammatory response in the lungs all right and this is mediated primarily by soluble or pro-inflammatory proteins called cytokines and chemokines and these can be secreted from the site of infection but they can enter the central nervous system and do quite a bit of damage and or alter neurologic function and so these two mechanisms are not um exclusive all right they could be combined or um it could be separate and distinct but um i think these are things we need to figure out okay so uh very quickly before i get into some of the data i want to talk about um there are a number of neurotropic viruses or human neurotropic viruses that exist that are capable of infecting and replicating within the cns these include members of the rna virus family dna viruses as well as retroviruses no doubt i'm sure all of you have heard of some of these viruses like west nile polio hiv herpes zika and again we have here the three human coronaviruses or zoonotic coronaviruses that we've talked about mild involvement with mers and stars one but potentially a lot more involvement with stars kobe 2. and here in this cartoon i just want to talk about four main cell types that we feel might be very important one is the astrocyte the other is the neuron this is the neuron cell body and these are long axons that project from the body of the cell that are utilized by the cell to transmit either electrical or chemical symptoms are signals that allow the cells to communicate with one another you have oligodendrocytes oh by the way these axons are protected by a myelin all right and so the loss of this myelin sheath can be devastating and you probably this occurs primarily in patients with multiple sclerosis right but it seems to also be happening in people with uh some people that are infected with codeine and then we have oligodendrocytes these cells make the myelin sheath within the central nervous system and then we have microglia and i'm going to come back to microglia in just a minute um so what happens in the cns following infection the idea is very simple um you inhale sars cov2 and presumably undoubtedly it goes to the lung and that's where it establishes a beachhead or a primary site of infection but keep in mind if you're in inhaling the virus particularly through the nose it can go up and infect the olfactory bulb uh within the brain and um the olfactory bulb does a number of things very important things one of which is controls your sense of smell and we know from studies in my laboratory and others that you can take mouse coronavirus for example and administer it intranasally to animals and the virus loves to get into the brain and replicate within the olfactory bone so it's not entirely not possible now um i've had um a very uh new but i can tell this is going to be a great collaboration with dr bill young who is a professor here in the department of pathology he was kind enough to share some pathological features that occur in people that have passed away from covid some of these it doesn't look like much but collectively it can be very damaging one is swollen astrocytes you see these large uh cells that are kind of fusing together these are astrocytes that become enlarged or swollen alternatively you can have what's called red neurons and what this simply is is the result of acute ischemia all right and these types of neurons can be distributed throughout the brain and it can be very damaging uh in terms of neurological uh function now more severe examples are provided by a paper that came out in 2020 and this is from claudia luconetti's laboratory who is a fantastic neuroscientist at the mayo clinic and she had this beautiful paper showing that in an individual that passed away from coven 19 extensive myelin loss um in white matter tracts of the brain that was then associated with extensive axonal damage and this can have devastating consequences for neurologic function in individuals so what i'm trying to get across is the notion that um the neuropathology is extensive we're still learning quite a bit about it because we just need more and more samples in order to better understand this all right so what are we doing in my laboratory and i when i say my laboratory i want to re-emphasize the fact that this the subsequent studies i'm going to be shown is a complete team effort all right from individuals around the country as and more specifically here at uc irvine all right and so the first question is or the concept i want to get across is that we can use other viruses to model sorrows kobe 2 biology and pathogenesis and the advantage of this one of the key advantages of this is that experimental work can be performed in what's known as a bsl2 or biosafety 2 laboratory all right and what that simply means is you don't have to put on the quote unquote spacesuit and go into a negative pressure room in order to do experiments with the virus you can do experiments in a regular biosafety cabinet that's been certified in a laboratory here is uh one of my graduate students colin pachow who's working in the bi uh in our laboratory working with the virus i'm going to introduce here and this is for better ways to describe it it's a recombinant virus the backbone of the virus it's a vsv which stands for vesicular stomatitis virus but i don't want you to worry about that what i want you to know is that this virus has been engineered to express the spike glycoprotein of covid19 or soros cov2 on its surface all right and so this virus was developed in the laboratory of sean whelan who's chair of microbiology at washu and then um we have uh sean was kind enough to let us use this virus and then subsequently we've been working very carefully or closely with dr bert semler and dr alex blenn uh dr semler is the former chair of micro and molecular genetics here um in the school of medicine as well as he is the director of the center virus research and he is a very elegant uh viral virologist and bert and alex have been very helpful for our laboratory and assisting us in growing the virus and sequencing the virus and what have you but you can use this virus to do a number of important experiments um for studying aspects that can be related to sars kobe 2 infection including determining the number of cell types in the cns that are susceptible to infection and so the question that we first asked is can we infect different human cell types so neurons astrocytes or microglia with this virus that has on its surface spike the spike protein of coven 19. all right we can then look for virus in the cells and measure their growth so this work would not have been possible without a fantastic collaboration i have with dr leslie thompson and dr matt blurton-jones both of which are professors in neurobiology and behavior here at uc irvine and the wonderful support staff that they have in their laboratories they provided us human neurons so charlie here was able to give us human neurons for our work and these are just beautiful looking cells um andrea was able to provide us with human astrocytes andre and charlie work in dr thompson's laboratory and then christina ii was able to provide us with human microglia and christina works in dr blurton-jones laboratory and so to cut to the chase we infected these different cell types with our um our vsv stars2 spike virus and what we found after 72 hours was very interesting we could clearly see infected neurons and how we could see this is because this particular virus has been engineered not only to express the sars2 spike glycoprotein but also a reporter gene called uh gfp or green fluorescent fluorescent protein and so you can see here clearly a number of different neurons that have been infected by the virus and under high power you can see that these cells are well they're really beautiful in my opinion you can see the virus extending down these long processes shown here extending far out and so this work uh was done by colin in the laboratory as well as hema oliveria they really worked extraordinarily hard to get this system up and running and i'm truly indebted to their hard work again we could look at the growth of the virus in these neurons you can see at 24 48 and 72 hours there's a natural increase in the number of infected cells but what was interesting is that we could not detect infected cells in either microglia or astrocytes arguing that at least within this experimental context neurons were the primary cells infected in addition i'll just tell you and not show you the data that we could block infection of neurons by this virus by blocking ace2 which again is the the human receptor uh that binds the spike glycoprotein okay so those are experiments that we have ongoing with regards to um working in the bsl2 but we also have initiated experiments working in the bsl3 so in other words how do you do experiment with sars kobe 2. so one uh it's much more involved you have to work in a high containment biosafety lab 3. the bsl3 is used for research or diagnostic work that involves microbes that are primarily aerosolized including stars cov2 and so i think there are several very important things i need to point out before i show you some of the data that we've acquired one we have two state-of-the-art bsl3 laboratories on the uci campus and uci is the designated national training center for bsl3 work around the country so in other words people from all over the us come to uc irvine to learn how to work in a bsl3 and this has been made possible by um gary landucci who has um is truly outstanding and how he's raised the visibility of bsl3 work not only on this campus but nationally as well as internationally and he deserves an enormous amount of credit for providing us investigators the facilities that we need to do this work in addition eric guandique and ando subba eric is the director of the high containment bsl3 laboratories anju is the university safety officer we have worked extraordinarily closely with gary eric and anju because working in the bsl-3 requires extensive training anything we do requires a standard operating procedure that we work and edit and then rehearse before we go into the bsl3 i also would be remiss uh not to mention dr ilhelm musoudi who is a professor in molecular biology and biochemist she is extraordinarily sophisticated virologist and neuro or immunologist who's doing some cutting-edge work in covid 19 looking at different patient populations and then finally and i will move on from this but i just have to recognize these people because they're so important for our work i've introduced hema who's a second year student there's also susannah furman a second year graduate student in my lab yu ting chang who is a fourth year graduate student and these are the true heroes of the work i'm going to show you they are the people that are going in on a daily basis into the bsl3 working with animals collecting tissues infecting them etc etc and none of this work would have been possible without their dedication to doing this all right so what we primarily do in the bsl3 is infect mice with virus all right specifically we infect mice with um sorry well uh with stars kobe 2. but unfortunately it's not that easy because normal mice um do not they have ace2 but their ace2 is not compatible with um sars cov2 all right so they're resistant to infection for the most part so we've had to acquire transgenic mice called k-18 mice for reasons i'm not going to go into but these animals have been engineered to express human um ace too all right so we acquired these animals um from the jackson laboratories and then we use them for infection to look at different responses in the brain or lungs and you can use these transgenic mice uh for a number of important experiments one to evaluate antivirals as well as vaccine development anti-inflammatory responses as well as just simply to characterize responses that can occur with regards to defense and disease and here i would like to sincerely acknowledge the efforts of dr grant mcgregor grant is a professor of developmental uh developmental biology or developmental and cell biology here at uci um he's also the director of the transgenic mouse facility and then dr shamaka who is really works very closely with grant all of the work that we've done with the k-18 mice would not have been possible without their extraordinarily help in developing or allowing us to rapidly expand this colony of k-18 monks and so i'm truly indebted to them as well as the office of research for building the outstanding transgenic core facility that we have here okay so the experimental goals and i'm almost done is that we want to infect uh these k-18 mice uh with virus how we do this it's an intranasal root of administration so we simply anesthetize the animals and you put a little drop of liquid-containing virus on their nose and they inhale it and they're infected but we wanted to establish that this was a good mode of infection and then determine if this allowed for infection of lungs and brains others have certainly reported this but you really want to make sure you can do it yourself and then in addition we really wanted to evaluate the functional role of this cell i mentioned earlier microglia in both host defense and disease so microglia are considered the immune cell of the brain and we and others have shown that viral infection of the cns does activate microglia and this activation contributes to both host defense as well as disease by secreting these cytokines and chemokines that influence immune cell infiltration into the central nervous system and so just quickly highlighting the importance of this we published a paper uh earlier this year clearly showing that microglia impact host defense disease as well as repair in a model of mouse coronavirus infection of the cns these are the individuals that did the work that are shown here for charlie was a former student in my lab dominic just defended his phd colleen stone was a former laboratory manager of mine and then kim green is a great friend and collaborator i have here at uci who's an expert in microglia so again the question is do microglia influence host defense or disease in response to sars cov2 infection and so how we can address this is we can specifically ablate microglia in the brains of mice prior to infection with virus and then evaluate disease progression tissue damage as well as viral burden is in the brains and lungs and how we do this is it uh it's great and why is it great i'll tell you why it's great uh because we can use a drug called plex 5622 and this drug selectively depletes microglia upon administration to mice all right and the great thing about it is this drug is formulated in mouse child so all the mouse have to do all the mice have to do is eat the chow that contains the drug and it gets rid of microglia so this is actually what the plex 5622 chao looks like it actually is pink the control chao is shown here in white these are k-18 mice separate cohorts we feed either plex 56 2 22 chow or control child for seven days to the animals this will get rid of about 95 percent of the microglia in these animals we then infect them with virus intranasally and collect brains and lungs one and two weeks post-infection so this um shows hema susanna and uting prior to going into the bsl3 this is me i actually do all the infections i'm just an ardent believer if i'm sending these students into the laboratory or the bsl3 to do this work i need to be in there with them to do it and you can see it's quite the get up that we wear again kim green is a collaborator for these studies kim is smiling because he is not dressed in the bsl3 guard and because he's not going in all right so what happens when we do this well um again these are preliminary data we've acquired these over the past month and a half it's been an extraordinary uh long involved experiments but in the plex 56 22 treated animals that is they don't have microglia we start seeing an increase in clinical disease severity that's associated with weight loss all right and so that means these mice are sick and this argues the idea that plex 56 mediated ablation of microglia increases disease onset as well as severity in the sars cov2 infected animals and what's interesting here is now lung tissue that we acquired from these animals at day 8 and 9 post infection and this was work that we did in collaboration with rob edwards the the pathologist on campus and then the staining for the virus was performed by hema oliveria and what we're looking at is viral rna and what jumps out at us was animals on the control chow had much greater amounts of viral rna in the lungs compared to those animals that were infected with sars kobe 2 but treated with plex 5622 and you can actually quantitate this using quantitative pcr and this is what colin pachow has done in the laboratory um very rapidly i might say and you can see there's a quantitative difference in the full change compared to uninfected animals the amount of viral rna in the lungs of animals on controlled child versus plex 5622 now what's happening to the brains all right so here we're focusing on the arrow the area and circled in this sagittal section showing the mouse brain this is the olfactory bulb but this is the cerebral cortex and um mouse coronaviruses love the cerebral cortex all right um primarily because there's a lot of neurons and so this is kind of a low power image just showing that there's a lot more virus in the cerebral cortex of animals treated with plex 5622 compared to control chao arguing that uh the plex 5622 ablation of microglia increases viral burden in the brain and indeed you can look here and you can quantitate this again this is work done by colin and there's a dramatic increase in the amount of viral rna in the brains of plex treated mice versus control and so we just got this data monday um and again it highlights the incredible collaboration we have with rob edwards and then hema did all the staining for virus and this is odd to say but these are just beautiful images all right and so this is in the cerebral cortex of plex 5622 treated mice and what you can clearly see here are neurons we haven't technically immunophenotype these cells but we're in the process of doing this but these are neurons that have been infected and these are this is just a high power image of these cells here and you can see these uh from the cell body these long extensions uh extending that are infected which raises some really fascinating questions about how the virus spreads in an infected cell um in addition we see in the cerebral cortex these infected neurons but what i'm pointing out here is what's known as meningeal inflammation inflammation these are immune cells that are migrating into the meninges similar to what's been reported in patients with covet 19. in addition this is a perivascular cuffing or perivascular inflammation this is the vasculature within the central nervous system and these are the highways that the immune cells employ to traffic into the the brain in response to infection and you can see this is loaded with inflammatory cells that are then migrating into the parenchyma to help control infection all right so to put everything in perspective and i'm concluding here uh um and again thank you for your attention uh cultured human neurons express ace ii and are susceptible to infection by virus expressing the cov2 spike glycoprotein importantly we can block infection by blocking ace ii cultured human microglia and astrocytes are resistant to infection at least under the experimental conditions that we've employed to date although we are continuing these studies intranasal inoculation of human ace2 transgenic mice that is the k-18 mice with sars cov2 does result infection of both the lungs and the brain this is consistent with other reports what we've determined is that neurons appear to be a primary target of infection by virus but we need to look at later times post-infection to determine if other immune cells or excuse me cns cell types are infected importantly i believe this is a unique observation that we've um that we're working on is that targeted ablation of microglia via plex 5622 reduces the viral burden in the lungs but increases the viral burden in the brain arguing that microglia may be important in controlling sars kobe 2 replication and spread within the brain so these are obviously areas that were looking all right this is my last slide because i think it's important to discuss where are we now all right and so what i've talked about today is just a very small portion of some of the amazing work that's going on on the campus of uci as uh frank alluded to at the beginning of the talk um there are a number of outstanding laboratories that are contributing to our knowledge base on the basic biology of stars cov2 with regards to both the virology and immunology clinical aspects associated with infection the development of novel therapeutics that can be employed that can limit viral replication and then finally vaccine development and i would say the majority if not all of these ongoing studies were initially facilitated by funding of coveted pilot grants that were provided through us to us through the uci office of research and why this is important is because this provides seed funding to investigators to then subsequently apply for a number of larger grants to the national institutes of health for example and so i gratefully acknowledge the craft covert awards from the office of research finally and this is my last slide i have to acknowledge several people cynthia manlepas is my laboratory manager and she technically is my boss she makes my job exponentially easier she runs the show beautifully she's in charge of the ordering she does everything and it just seems she's graceful and she uh works beautifully with everybody in the laboratory and none of the work that i would have been able to do would have been possible without her uh these four students that i've talked about extensively are truly amazing i cannot say enough good things about them they are amazing in what they've done with the work that we're doing and amber and dominic are also amazing but they're working on our mouse coronavirus projects rob edwards again and the experimental tissue research center that he directs none of this none of the images that i showed on human samples would have been possible again i also need to acknowledge bill and his contributions ilhelm and her contributions she's my partner in crime discussing experiments with covet 19 and her wonderful support staff leslie and matt and kim for being there to help provide tissues and reagents that we acquire i also have to acknowledge uh dean laferla um who's a tremendous dean in the support staff in the office of uh the biosci and for the sake of time i do have to again acknowledge gary eric and andrew for allowing us to get into the bsl3 charlie andrea and christina for their help in getting us all these samples um wonderful collaborators here that we'll be working with eric mike and craig stan pearlman who helped us provide by providing virus sean with also virus and other collaborators and i also would like to again acknowledge the funding for the laboratory and frank thank you again for this opportunity and i'd like to thank everybody for their attention thank you uh tom thank you so much i mean this was a tour de force uh not only in terms of the way you presented all of the material but just in terms of the amount of research progress that you have uh made i mean you get you have to remind everyone that essentially you started working on this march of 2020 and the uh you know progress is just impressive so congratulations to you and your team uh there as you can imagine a number of questions from many people and so we'll try to be respected of the time but see how many we can get through and i guess the most relevant question to ask right off the bat is what are your thoughts about the pfizer vaccine that was just recently announced yeah um i think it's great uh we need it um you know you the 90 effectiveness and simply what that means and as probably most people know it's effective in uh the um occurrence of symptoms right and what we don't know today and pfizer knows this right um uh is whether this is um really uh blocking infection uh reducing symptoms and there are a number of other important questions is the vit you know how effective is long-term memory that's evoked in response to the vaccine but it's a great start right and i think we all you know are beginning to breathe some sigh of relief and there are a number of other really compelling vaccines in the pipeline as well and so this is a great start and so i'm very encouraged by this can you maybe expand on what your thoughts are about immunity for uh sars kobe too i i mean if you go through this horrible disease do you have to look forward to getting it again uh you know that's that's the question right that we all are worried about you know i think frank the interesting point is believe it or not we can learn quite a bit from those common coronaviruses because um we know that over time your antibody levels wane all right so you know about six five to six years following an infection that the antibodies to these relatively benign upper respiratory tract infections can go away and you are susceptible to re-infection all right now whether this is going to happen with uh uh sarah's kobe 2 we don't know a lot of people social media has really grabbed onto this notion that antibody titers weighing over time but in the big picture antibodies titers do wane over time all right and that happens under most infections all right the main thing now is we just have to keep in mind that this disease although it seems like it's been an eternity it's been eight months right and we have a lot more to learn about the immunology the basic biology but we'll get there we're going to get there yeah i mean i think you you know that i would my phd was in virology and i worked on vesicular stomatitis virus myself and back in the 80s people would talk about how fascinating coronaviruses were that there were no really relevant human uh diseases that were impacted by them and that has certainly uh changed um so i i guess it raises a question someone let me raise a question someone brought up which was how was the original source contained in 2003 and are there any lessons that we can learn from that yeah um thank you for the question it was contained probably primarily because it was a really crummy spreader right because you had to be very sick and have symptoms in order to spread the virus and once that was determined that family members and healthcare workers were the primary people exposed you could easily quarantine infected individuals um and that's the same that's true with mers kobe so effective quarantining um masks were apparent obviously just from the photo of time but a lot of techniques that we're using now right but the problem is sars kobe 2 is much more insidious because you can be asymptomatic and very efficiently spread the virus right so a few of our professors have a very similar question so uh professor james hicks and robertson want to know if the commonly prescribed drugs that upregulate acetube receptors in the brain are possibly making people more susceptible to infection yeah jim great question a lot of people interrogated that at the beginning but it kind of tailed off so if it had you would have heard about this to great extent so i don't think there's a really solid literature base in that right now now we can test that potentially in in these animal studies but i think i'm more interested in looking at anti-inflammatories and if that limits uh the spread of the virus or reduces the pathology so um that's an area that probably requires a little more investigation but it's not prominent in the literature right now okay um then if sars kobe 2 is able to enter the brain are there any antiviral drugs that can penetrate the cns and inhibit viral replication wonderful question and extremely relevant because um it's not clear if a vaccine will be able to evoke a memory or get virus in the brain so developing drugs that are seen as penetrant is not trivial um and so remvere supposedly has good penetrance into the central nervous system um but i don't know if that's been tested to great extent in the animal models and i think it's important to put this in the context of hiv because we have the cocktail of drugs that work great at reducing viral burden in the periphery but a lot of people that are hiv infected can have virus in the brain and it is a reservoir these drugs do not penetrate the cns and these individuals can some of them can develop what's known as hiv dementia all right so this is one of the areas that i'm very interested in investigating in my laboratory particularly within the context of long haulers right so what if people that have these neurological disease for years and years and years is it due to a persistent viral infection we don't know and this raises that really emphasizes the importance of characterizing drugs that get into the brain so another question relates to uh the findings of covet with individuals with epilepsy and what's known about that um you know it's it's probably a comorbidity what we don't know is if there's greater cnn's penetrance clearly neurons are involved with epileptic seizures and so are these seizures occurring or enhanced seizures occurring because of viral penetrance or infection of the cns and neurons to date that's pretty much the extent i know of this um and it's it's certainly an area that requires uh greater investigation okay uh excellent uh so maybe explain to us how uh lung um inflammation can exacerbate disease in the brain right so the easiest way to think about this is when in when all of us are infected by a virus or any type of microbial infection one of the first things that happens is we acquire a fever right and that fever is your body's first one of the first defense mechanisms because microbial pathogens don't like elevated temperatures how we raise our temperature in response to infection is via cytokine secretion in the periphery all right so certain cytokines like interleukin-6 and tnf alpha can get into the brain and go to an area of the brain called the hypothalamus which is the regulates your body's temperature right so it's almost like the rheostat so this is an example how peripherally derived cytokines can influence factors in the brain now in patients that have covid or severe covet infection they often have cytokine storm that's elaborated initially in the lungs these cytokines could then enter into the central nervous system and cause a number of different changes uh that can be either acute or potentially more permanent so that's that's the notion right there of how this can happen okay um rick robertson asks another question does the immune response depend on naive t cells and those of us who are getting older don't have many naive t cells left um well any type of a virgin infection you need naive t cells that are bearing you know cognitive antigen receptors that can expand to the infection right and so um certainly as you get older um the efficacy of your immune response wanes over time right we know that from vaccination trials and what have you but that's why i think the priority for these first round of vaccines will be for the elderly like you know this last week i was visiting my father who's 90 years old right and so um you know you want these people to be protected um so they the older you get the more effective your t cell and antibody responses are in addition your type secretion of interferon in response to infection also diminishes as you get older right and that's that's truly one of the first responses that are required to control any type of virus but that's also particularly true for soros covey too so uh let's make this the last question considering the hour and i guess it's the relationship between vaccine development and anti-viral medications uh and just uh what are your thoughts on that the relationship between those two obviously both are going to be critical in the response to covet um the big picture you need a vaccine right i mean if anything has proven over the past century is that vaccine vaccination is truly one of the greatest contributions of medicine has made to humanity so um we need a vaccine but in the short time there are uh you know antivirals can be very effective in limiting hospital stay or limiting even entry into the hospital okay and so a lot of investigators in the field right now are looking including our laboratory are looking at not only novel antivirals but repurposing fda approved drugs that are being used for other disease indications that you'd be surprised it could be very effective in limiting viral spread in covid patients so you know i can't say one is more in my opinion the vaccination is the vaccine is more important but it doesn't diminish the importance of developing effective antivirals and you know we're doing both primarily we're working on antivirals in my laboratory with some great collaborators in chemistry because chemists at the end rule the world so um that's the way it goes all right thank you tom that's a good way to and and unfortunately we can't get to the many many other questions um that are in the uh chat but i just want to remind everyone that um the 2020 dean's report was largely devoted to the covet 19 pandemic so if you're interested in learning more about not only what tom is doing but what other faculty in the school are doing please visit our website at bio.uci.edu and you'll find a lot more information you'll be able to download the dean's report and see what we're doing there and hopefully have some of your questions answered i also tom i want to just before we depart again thank you for this tremendous work uh that you're doing for being such an outstanding colleague and such a friend to uh so many of us and you pointed out the money that you got the pilot award that you got from the office of research and a significant amount of that also came from the school biological sciences and so i just want to make the point to the individuals in the audience that this is often times where your contributions to the school of biological sciences particularly to the dean's excellence fund allows us to support the kind this kind of novel innovative research and i think it's because of these funds that we were able to disperse to you and to other researchers in the school that we've been able to make so much great progress uh and not only on the research front but also to help our students so if there's anyone in the audience that's interested in helping or learning how you could become closer more closely affiliated with us uh and join our biosci ambassadors program these are not you know programs that require huge donations but just um contact either andy denuzzo or robin martinelli their information is up on the screen right now and you could see i think firsthand the direct impact that this kind of support has for all of society so with that uh again tom thank you very much and let me wish everyone a very safe and wonderful uh holiday season and we'll see you for our next dean's distinguished lecture in the winter quarter thank you all bye good night

Channel: UCIBioSci

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Keywords: education, university, UC Irvine, Biological Sciences, science, biology

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Length: 74min 21sec (4461 seconds)

Published: Wed Jan 20 2021