T Cell Role in the Pathophysiology of Heart Failure

December 21, 2021

Information

December 16, 2021

Yale Pathology Grand Rounds

MariaPilar Alcaide, PhD

ID7311

To CiteDCA Citation Guide

00:00So. Good afternoon everyone.
00:03Thank you for attending a
00:06year pathology gram one.
00:07Sending a series.
00:09It's our great pleasure to invite Doctor
00:12Killer or Katie to speak at our grandma.
00:16I met at scientific conferences and I'm
00:19impressed by her outstanding mechanistic
00:21research in immunology and cardiology.
00:25Cloud received her pH D from the Autonomous
00:28University of Madrid, Spain, she.
00:31Performed her postal research
00:33at Brigham Women Hospital,
00:36have a medical school.
00:37She studied her faculty position
00:39as an assistant professor at the
00:41Department of Medicine, Tufts University.
00:43She has been attending
00:45resources professor since 2019.
00:47She's the program director of Immunology
00:51graduate program is an endowed chemist,
00:54and Joann,
00:55where professor and the interim
00:58vice chair of department
01:00immunology at Tufts Class Group,
01:03has made important discoveries in the
01:05area of mechanism T cell trafficking.
01:08Their research efforts are focused
01:10on understanding why and how T cell
01:14subsets and in gaseous interaction.
01:16They also study intrinsic properties of
01:19the vascular in the cilium that modulated
01:22key cell and leukocyte recruitment
01:24in the T cell trafficking and survival.
01:27Another exciting line of risk in
01:29class group is about inflammation,
01:31heart failure to recent research has
01:34contributed significantly to your paradigm
01:37shift in understanding of heart failure.
01:39Putting T cell inflammation as a measure
01:42of player in this heart failure disease.
01:46So exciting research program have
01:49led to multiple impactful papers
01:52such as circulation journal,
01:53experimental medicine,
01:54JCR inside a TVP and etc.
01:58Pillars research has been has
02:00been funded by NIH one brand.
02:03She has also showed great leadership in
02:06science by serving our editorial board
02:08such as Junior Clinic investigation,
02:10Fast, GMC and etc.
02:14Additionally,
02:15she has been serving the American Heart
02:18Association during past many years.
02:20She has been the HCA basic cardiovascular
02:22Science program chair since last year.
02:26Without further ado,
02:27let's welcome Killer.
02:28To give us her seminar entitled T
02:31Cell Role in the passive Physiology
02:33of heart failure, can I thank you?
02:37Thank you very much.
02:38You ring for the invitation.
02:40It's really nice to be here and also
02:44thank you to all those of you who
02:48I've met with this morning because
02:50I've returned your science because I
02:52I've been really enjoying, you know,
02:54all the things that you're doing here.
02:56So as giving said,
02:58I'm going to focus today's talk on the
03:01aspect in the lab where we study the
03:04role of T cells in the pathophysiology.
03:07Of heart failure I have no disclosures
03:11and basically this is a cartoon that
03:14summarizes the general theme of our lab,
03:17which is how the moon system impacts
03:19cardiac and vascular health.
03:21So as you being said,
03:22I train in immunology and then I
03:25further train in vascular biology,
03:27but we know that immune cells
03:31they really need to traffic into
03:33tissues to do their functions.
03:36But then once.
03:37In addition,
03:38they need to interact or crosstalk with
03:42all the different resident cells in order
03:46to modulate homeostasis or pathology.
03:49In the case of injury.
03:51So for today's talk,
03:52I will focus on what we've been
03:54learning recently in the lab from
03:56our work and also from the work
03:58of others of how this interaction
04:01between adapted and innate immunity
04:03contributes to cardiac remodeling.
04:06In hard and I I place a circle
04:10here because this is mainly where
04:12these interactions between adapted
04:14and it made immune cells happen.
04:17This is what T cell antigen
04:19recognition starts in the lymph nodes.
04:22But towards the end of the talk I
04:24will show some new data is still
04:26unpublished where we really find that
04:28they're very similar interactions
04:29that are also happening in the
04:31heart and that they might modulate
04:34correct Physiology this way.
04:36So as many of you probably know,
04:38heart failure is very complex
04:40and it's multifactorial.
04:41So to tackle mechanisms,
04:42we need to start in a simplistic way.
04:46But we also need to understand
04:48the full complexity.
04:49So what do we know is that
04:51regardless of the etiology,
04:53whether it was triggered
04:54by any ischemic event,
04:56such as a myocardial infarct
04:59or non ischemic event.
05:01The heart remodels and the characteristics
05:04of the failing heart are increased.
05:07High level curricular pressures and
05:11then a hypertrophic cardiomyocytes
05:14fibrosis and these results in
05:17systolic and diastolic dysfunction.
05:19And we've known since the 50s that
05:22systemic chronic inflammation
05:23is associated with pretty much
05:25all of the causes of all of the
05:28etiologies of heart failure.
05:30I'm just going to set up my timer
05:32here to make sure that we're.
05:33Runtime here.
05:35But unfortunately this by this knowledge,
05:37for many,
05:38many years today none of the
05:40anti-inflammatory therapies for clinical
05:42trials that were initially launched
05:44to tackle a pro inflammatory cytokines,
05:47such as TNF.
05:48And more recently,
05:49with the counters trial
05:51island bed and none of them,
05:53this is the anti TNF therapies
05:55that are very efficient in treating
05:57out immune diseases and chronic
05:59inflammatory diseases did not
06:01work in heart failure and there
06:04are more recent
06:05promising data with the Cantor's trial,
06:08although it's still early to tell
06:10whether it has really benefited in in
06:13some of the outcomes of heart failure.
06:16So what we know is that there are no
06:19anti-inflammatory antifibrotic therapies
06:21that have been successful today,
06:23and we know from many organ systems
06:26that inflammation or immune cell
06:29activation and fibrosis go together or
06:32have some overlapping functions as well.
06:35So the first question that we
06:37asked several years ago was is,
06:39is there cardiac information?
06:41Besides systemic chronic inflammation
06:43again going with the concept that if the
06:46immune cells traffic to an inflamed issue,
06:48do they exert their functions by
06:51communicating with the tissue
06:53or stroma cells?
06:55And then if that was the case,
06:57do the cardiac infiltrated muscles
06:59contribute to the hallmarks
07:00that we see of heart failure,
07:02such as correct fibrosis?
07:03And does that have any impact
07:05on cardiac dysfunction?
07:06And obviously we're very interested as
07:10basic scientists in understanding how.
07:13So the first experiment that we did
07:15this this is this was published.
07:16This is back in 2015,
07:19but we wanted to say whether we
07:22could see cardiac inflammation
07:24in patients with heart failure,
07:25but we wanted to look at patients
07:28with non ischemic heart failure.
07:29Antonio Barish group at VCU in Virginia.
07:33He had elegantly demonstrated
07:34years before this that in response
07:36to myocardial infarction,
07:38there was decent infiltration in
07:40the human heart and interestingly.
07:42The diesels were infiltrated in the
07:46in the scar zone in the infarct zone,
07:49but also,
07:50so that goes along with a roll of
07:53the immune system during evolution
07:55to help healing.
07:57But interestingly,
07:58what they had found in inference
08:00was that there were also a lot of T
08:02cells infiltrated and remote zones.
08:04So we thought if we chose a patients
08:06that did not have any impact that have
08:08sort of like low chronic inflammation,
08:11where we see these as infiltrated
08:12in the heart.
08:13And this is exactly what we found
08:16here in Brown that they end stage
08:18nonischemic heart failure and
08:20heart samples which were taken as
08:22stated here from from there had
08:26significant diesel infiltration.
08:28Compared to non heart failure controls.
08:31And later on we did some experiments
08:33where we also wanted to look at what
08:36kind of pistols were infiltrated there,
08:38and we found that many of those
08:41teachers expressed the chemo keen
08:43receptor CXCR 3 and this is shown here
08:46by Red Arrows and quantified here.
08:49So this this made the basis to
08:52wanted to understand mechanistically
08:54what is thesis that expressed these
08:57receptors are doing within the heart.
09:01So our very broad hypothesis.
09:03110 and Evers joined my lab as
09:06postdoc was asking the question,
09:08is this purely an association or
09:10are they actually contributing or
09:12doing something in the heart and her
09:15hypothesis was that they would be
09:16doing something there in the failing heart?
09:20And to start doing this we have to
09:23choose a preclinical model and knowing
09:25that heart failure is very complex,
09:27there's no optimal or perfect.
09:31The clinical model that mimics
09:33all the symptoms of heart failure
09:36or how the disease develops,
09:38but we found that for nonischemic
09:40heart failure tack or transverse or
09:43reconstruction was a one where we could time,
09:47and for certain reasons,
09:48that we wanted to do.
09:49It was really a very good model to do so.
09:52Why? Because it induces pressure
09:54that mimics the pressure that heart
09:56failure patients have in the heart.
09:58Although the downside is that here.
10:01It induces a sudden pressure that then
10:04is restrained versus impatience as
10:06we know they developed progressively,
10:08but importantly,
10:09in this model we can basically track very
10:13nicely how cardiac hypertrophy develops,
10:15how cardiac fibrosis develops,
10:17and whether we can check in per cardiac
10:20function and at those time points.
10:22We could also look for diesel immune
10:25responses and correct diesel infiltration.
10:27So using this model I'm showing data
10:30from 4 weeks. Play time points.
10:32That is all summarizing schematics.
10:35Because we published this already.
10:38But what we found was that it
10:40specifically one type of thesis
10:42that are CD 4 positive T cells
10:45were infiltrated in the heart as
10:46early as two weeks post stack.
10:48And this is four weeks after Tak.
10:51And then,
10:51before they infiltrated in the heart,
10:54we saw a significant expansion of the
10:56medicinale draining lymph nodes that are
10:58the lymph nodes that drain the heart.
11:00And most of those T cells express
11:03interferon gamma so they were type
11:06one TT cells T H1 cells which also
11:09expressed the chemokine receptor.
11:116 year 3.
11:13And then we found that these
11:16infiltration was associated with at the
11:18time where mice developed fibrosis.
11:20As you can see here in pink,
11:21the collagen deposition and enlargement
11:24of the cardiac myocytes by H&E.
11:27And what we found using this mouse
11:30model was that if my eyes were
11:33genetically deficient in D zones
11:34and we use different models,
11:36diesel receptor alpha Nokia or MHC 2
11:39knockout or right to knock out what
11:42we found was that all the mice that
11:44did not have decent genetically they
11:47did not develop a cardiac fibrosis.
11:49We cannot see any College in
11:51the position here.
11:52And then when these mice were reconstituted,
11:54we see Excel 3 positive there
11:56from gamma positive.
11:57Keystones we could partially
12:00reconstitute the fibrosis.
12:01Certainly the provascular fibrosis.
12:03As you can see here,
12:05and we could reconstitute and or
12:07a lot of the cardiac dysfunction.
12:09Although this data also suggested
12:12that there had to be some cardiac
12:15antigen specificity involved to
12:17to induce the full induction of
12:19cardiac fibrosis and dysfunction.
12:21In these experiments, as I said,
12:23we reconstituted fibrosis,
12:25some parameters of systolic
12:27and diastolic dysfunction.
12:29But these cells that we put
12:31back into these mice,
12:33they were highly painful.
12:35Amatory but not antigen specific and
12:37this will link to the second part of my
12:41talk and why that might be important.
12:43How we can use that in in in to
12:47understand this complex syndrome.
12:50So the other thing that I needed
12:52was well now
12:52that we know that these T cells that express
12:55in there from gamma are increasing the
12:58lymph nodes under infiltrated in the heart.
13:02How can we see if they actually cross
13:05communicate with a cardiac residents?
13:07Because we saw that massive
13:09effect on cardiac fibrosis.
13:11We see it was a simple
13:13experiment to start with,
13:14which was isolating primary,
13:16correct fiberglass from Adele mites
13:18and then see isolated T cells.
13:21From the mediastinal lymph nodes of
13:23mice that were subjected to either sham,
13:25so control surgery,
13:27I should say that some might have the
13:30the open chest surgery and everything
13:33except for the construction to
13:36account for possible inflammation
13:38that happens during surgery.
13:40I know what you did is the Chico
13:43culture this and then she she
13:45coculture this indirect cultures or
13:47entrance once and the idea was that
13:49first he was going to see whether
13:51these teasers could adhere to the
13:54fibroblast and whether these fiberglass
13:56transform to myofibroblast and to
13:58do the readout for myofibroblast.
14:01We looked at alpha small muscle
14:04acting which is expressed upon
14:07fiber restaurants formation.
14:09So in those experiments,
14:11this is a representative image.
14:13You can see that a direct contact was
14:15required for these transformation,
14:17so these are two examples of
14:20transform my fiberglass that have
14:21a lot of T cells bound to them.
14:24The T cells were labeled in in green here
14:27and this is the quantification of the
14:31of of the red of alpha small muscle acting.
14:35So the conclusion from these
14:37experiments was that there was
14:38these communications between.
14:40Pieces of fiberglass that was required
14:43to induce transformation and then a
14:46mechanistically what we found was
14:48that this was TGF beta dependent,
14:50so it will block TGF beta.
14:52We inhibited this transformation that
14:54was not too surprising because we know
14:57that PDF better was it's a classic
15:00profit garlic cytokine that induces.
15:03This transformation and that is
15:05highly made by my fiberglass as well,
15:07but what was more interesting is
15:09that we found that T cells bound to
15:12the fiber rest through A4 integrin,
15:14and we come one in the fiberglass
15:16and then once bound,
15:18the diesels were induced and DJ better
15:21released by the myofibrils are not
15:23the other way around and this was also
15:26published so I wouldn't show a lot
15:28of the data there so I can focus on
15:30more recent data in in our lab as well.
15:33We're currently working on on
15:36further mechanistic insight
15:37into how we can prevent this.
15:40They released induced by the FIBERLESS
15:43upon the contact with the discus.
15:46So as a summary of background of
15:48why we became interested in this,
15:50we can see that in this transformation
15:53from the healthy heart to the failing
15:55heart using an experimental model,
15:57in this case of transverse article
16:00struction visit this activation,
16:02particularly of this T cell subset.
16:05And then we say that these are traffic
16:07to the heart and once in the heart.
16:09They crossed off with a fiberglass
16:11and induced cardiac fibrosis.
16:13We block these by either using
16:16agent mice that don't have T cells,
16:19or by using depletion diesel
16:21antibodies in wild I'm eyes.
16:23I didn't show the data by the way.
16:25We also did those studies.
16:27We prevent this transformation.
16:30We also found that the characters
16:35themselves they in response to
16:37pressure in response to duck even
16:39before the T cells get there,
16:42they can actually make chemokines that
16:45Kim attracts positive T cells and we
16:47did this using a reported mice for
16:50these schemes and and doing a time course.
16:53So we actually found that the
16:56fiberglass are actually functioning
16:58as a semi immune cell because
17:00they're releasing chemokines.
17:01Then they end up attracting first my
17:04load cells and then T cells to the heart,
17:07and then as I just showed,
17:09we found that they can regulate
17:12correct for groceries.
17:13So then when I question that we asked
17:16was well how and where this is being
17:19activated in the heart and this work
17:22was done by Jay Wanyama who was a
17:25graduate student in the lab and he
17:27was really interested in knowing
17:29this because he found he said well,
17:31if we found a specific antigens that might
17:34be relevant for the T cell immune response.
17:37Then one could think about in
17:39this in the future.
17:40Potentially immunizing for heart failure,
17:43right?
17:44There will be a long term goal or at
17:47least understanding whether this is
17:50what was this this activation happening
17:53and where over time it could be prevented?
17:57So to study that we use a reporter
18:00mice for T cell activation or
18:03T cell receptor engagement.
18:05So these are in order to be activated
18:07by antigen presenting cells,
18:09they need to recognize antigen,
18:11and in the case of CD 4 positive T
18:13cells they express the diesel receptor
18:16here and then dendritic cells.
18:18Are they the main antigen presenting
18:21cells express MHC two and they
18:24can capture antigen and induce T
18:27cell receptor signals and these
18:30reporter mice mimic.
18:32They're basically reporters
18:33of diesel receptor engagement.
18:35So the green are the cells are because
18:39they express N 77 which is downstream.
18:42The diesel receptor bound to GFP.
18:45So the greener the cells are.
18:47That's telling you that
18:48they're recognizing antigen.
18:49This expression is also transient,
18:53so if we see green cells,
18:56it means that at the time where
18:58we're harvesting those cells,
18:59they're recognizing antigen.
19:00But it might be that they recognize
19:03antigen and then they're not recognizing.
19:06Antigen at that point and then
19:08they lose expression.
19:09So in order to look at this,
19:11we basically did the time course
19:13of tag again early on,
19:15where is compensatory changes and then
19:18once is systolic dysfunction is established,
19:21and then we kept them for longer.
19:23That will mimic more chronic heart
19:26failure and what we did is we harvested
19:29the hearts and the medicinal influence.
19:32And the first thing that we did here.
19:35And we found that was very,
19:37very surprised,
19:38surprising,
19:39and the most interesting finding.
19:41I I I felt from from this story that we
19:44recently published was that we saw
19:46this T cell receptor engagement not
19:48only in the cardiac lymph nodes,
19:50but also within the heart.
19:53And as you can see here,
19:54you said the very bright GFP
19:57cells that increase overtime.
19:59So that's telling us that once
20:01the T cells infiltrate the heart,
20:04they must be vendrick cells.
20:06And potentially other cells that
20:08capture the antigen and induce
20:10decent expansion within the heart
20:12and that would be bypassing the the
20:14final trafficking that you need from
20:16the lymph node into into the heart.
20:19So we quantify this and as you can see,
20:22there's a significant increase of
20:25GFP positive active T cells in
20:28the heart that correlates with
20:30decline in systolic function.
20:32Measure here with fractions.
20:35So if we go back to how this teaser
20:38activation happened in the timers,
20:41we have a lot of high T cell
20:45receptor clonal diversity because our
20:48diesels are deciding what you know,
20:52depleting what's against self antigens
20:54and selecting for what we might
20:57need in the future if we get closer
20:59to the high in the Middle East and
21:01lymph node we will have the selected
21:03a pool of clones that will get expanded.
21:05If there is any immune response
21:07and then the question was what
21:09we were seeing in the heart,
21:10so our data using these reporter
21:13found that indicated that there was
21:15this expansion in the correct running
21:18lymph nodes as well as in the heart,
21:20and then we decided to do this
21:22receptor sequencing to get a closer
21:24look at whether these T cells or
21:25whether it might be recognizing.
21:27So this is the structure of the
21:29T cell receptor.
21:29Many of you probably know,
21:31but just as a brief reminder
21:32it has two Chainz,
21:34the alpha and the beta chain.
21:36And then they recombine in many
21:38different ways to form a specificity.
21:41Or these pocket to many different antigens.
21:43So by sequencing this Cdr three region,
21:47which is where these two chains
21:50get closer and form the pocket
21:53for antigen recognition,
21:54we could get a sense of whether we
21:56get in many different clones which
21:59will indicate high clonality and not
22:01not really an antigen specificity.
22:03There was anything in the heart,
22:05or whether we get.
22:08Enrichment so the results that
22:10we found was as expected.
22:11There was a lot of clones,
22:13many different diesel receptors
22:15sequences in the timelines.
22:18These are the inguinal lymph nodes and
22:20again we have a high clonal diversity
22:22and what we found was that the
22:24closer that we were getting to the heart.
22:27There more the decreased number
22:29of total clothes that we found,
22:32and the highest enrichment of the
22:35top 20 most represented groups.
22:37So today's my zaveri complicated
22:40analysis and we found that there was
22:43a restricted clonal pool in the heart
22:45and that the majority of the cells
22:47were represented by top 20 clones.
22:49So we started diving more into
22:51what this could be.
22:53So what are these enrich
22:55clones responded to in the
22:56hunt and this will be the working month.
22:59We know that this is happening and
23:01then we know that there's this close
23:04being expanded and what could those be?
23:06So we ended up focusing on Russ and then
23:10again because of the time limitation.
23:13I'm not going through
23:14everything that we went through,
23:16but it basically one hypothesis was
23:18that you could have a cardiomyocyte
23:20proteins right that the myocytes die,
23:23and then the fragments are picked
23:26up by adding percentage selves.
23:29But it turns out that in the attack
23:30model and like in response to my
23:32kardelen function where you see a lot
23:34of cell damage in the attack model.
23:37We don't see significant salvage early
23:39on and even later on at four weeks.
23:42But what we do see is high increases of
23:46intramyocardial reactive oxygen oxygen
23:49species which are labeled here in green.
23:53So we went back to literature and we
23:56hypothesized that maybe Rose could modify
23:58correct proteins that then form new
24:01antigens that are listed AT cell response.
24:04Why do we?
24:05Why would do we think that
24:07there was a hypothesis?
24:09It was because similar mechanisms
24:11had been described in the vasculature
24:13in the context of hypertension,
24:15where there was this formation of ice level.
24:19Glenda Lynn's,
24:20which are highly reactive intermediates by.
24:24Lipid peroxidation.
24:25That then can adapt to self proteins
24:27and create these new antigens.
24:29So we contacted the people who had
24:32done the scientists that had done this
24:36very interesting research hypertension
24:38David Harrison and Annette Kirabo,
24:40and this started a beautiful
24:42collaboration in which we were able
24:45to test this hypothesis in the heart.
24:48So basically we obtain a lot
24:50of reagents from their labs,
24:51and the first thing that we wanted
24:53to know is whether this matter
24:55in the human heart.
24:56So we went back or human heart failure
24:59sections and we use this one day 11 which
25:02is an antibody that recognizes proteins.
25:05Modified biologist was given to us
25:08by David and Annette and as you
25:11can see here we saw significant
25:13recognition in three different
25:16heart failure patient samples.
25:18And no signals in a non
25:21heart failure patient.
25:22We went back to the marsh model and in
25:24the marsh model we went back to these
25:27reporter my cells and T cell recognition.
25:31And we used the D1DD11 antibody
25:35that recognizes proteins
25:37modified by strategies in mouse.
25:39This was a different version of the antibody.
25:42And then they also send us some ISO
25:45so they can generate and I soils LG
25:50scavengers that could be used in in mice.
25:54So how does it work with experiments in tack?
25:57And then this is the structure
25:59of the eye surgeons calendar,
26:00and this is the control peptide.
26:02So the two Joba that I'll be
26:04showing is one with coverage.
26:06Those rose reactive proteins and then
26:09we tracked this activation and T
26:13cell receptor engagement over time.
26:15In some experiments we use tempo
26:18because it's an antioxidant,
26:20so it works upstream of the of the.
26:24Draws formation so just to make it
26:27simpler in for understanding the idea,
26:31this is a question that we were trying
26:33to ask and the idea is that in response
26:36to diagnose Ross and then this Ross
26:38induces the formation of this lipid
26:41peroxidation and this ISO, geez,
26:44that then they adapt to a cardiac protein.
26:47I'm from Disneyland pigeons that could
26:49be taken by the dreaded cells and
26:52being presented these cells to induce
26:55T cell activation and proliferation.
26:57So we could block these with temple perhaps.
27:00And if we block this, if this was true,
27:03maybe we would see less this
27:05activation or proliferation in heart.
27:07And if you block this with the
27:10ISO G specific scavenger,
27:12we could potentially block this and
27:13block this activation in the heart,
27:15and then of course the final question was,
27:17will this have any impact incorrect function?
27:22So the first thing that we did is we
27:24did those experiments in mice and then
27:27we isolated and Rick cells from mice
27:29treated with these AI soldiers scavengers.
27:32And as you can see here,
27:34this is the antibody that detects
27:36the isolated protein adducts.
27:38And as you can see with the four coma,
27:41which is the control and the
27:44control compound,
27:45we see that in drink cells
27:47express this and take a protein.
27:49But this is significantly
27:51inhibited when we scavenged.
27:55And then when we look directly in the House
27:57looking for teachers using the reporter mice,
27:59we found that only in those miles that
28:02were treated with the iceberg scavengers,
28:05we were able to significantly decrease
28:08this teaser activation within the heart.
28:11This is again GFP,
28:12because this underreported mines and
28:14I don't think I mentioned it earlier,
28:16but this is this receptor beta
28:18to make sure that we're focusing
28:20on the right distal population.
28:23So then how can OK?
28:25So now we know that under excels pick it up,
28:28but added functional inducing
28:30diesel proliferation.
28:32Here we found that there's
28:35less this activation,
28:36so the hypothesis is that when
28:38these are become activated,
28:39then they have to proliferate.
28:42So we tested this hypothesis ex vivo
28:45and we took the genetic cells from
28:48control mice and loaded them with either
28:51ice or GS or with a correct license
28:54that were taken from the tag mice.
28:58And then we coculture this and Rick
29:00cells that were incubated with these
29:02characteristics with teachers that
29:04came from medicinal lymph nodes
29:06from either sham or tag mouse.
29:09And again we were trying to
29:11mimic these response.
29:12And after four days we can
29:14look at T cell proliferation.
29:17So the way we look at T cell proliferation
29:19is because the teachers are labeled
29:21with a membrane guide that is CFC.
29:23And if they proliferate as you can
29:25see here to see if you see a die is
29:29diluted and this is just one example.
29:32If we combine some T cells with sham lysates,
29:37there's no proliferation,
29:38and if you put tactics so there's
29:41significant proliferation.
29:43But basically what we found here,
29:45this is only a representative experiment.
29:47Everything is quantified in the manuscript.
29:50But what we found was that only when
29:53the teachers came from mice with
29:56experimental heart failure and the
29:58proteins that cells were loaded with
30:01cardiac proteins that came from Tak
30:04only this combination is when we
30:06were able to see these new antigen
30:10presentation and decent proliferation.
30:12So did this have any impact in
30:15cardiac function?
30:15And these are echoes done
30:17by our collaborators.
30:19That Medical Center Rob Landon,
30:21who is a cardiologist that whom I've
30:24I've been collaborating with for many,
30:27many years,
30:27and what we found is that
30:29these attack animals.
30:31You can see the flat line here and
30:34decrease historic function that is,
30:37is quantified here.
30:39Fractional shortening and
30:40the harsh from the two hobas.
30:42For them I struggle with this.
30:43Lavender it were very healthy
30:46compared to to the control.
30:51So to summarize this part,
30:53what we found was that in response
30:55to high level trickler pressure.
30:58There was a significant induction of
31:01intramyocardial rose in the heart
31:03and ended in Derrick cells that are
31:06here are picking up some of the
31:09of the proteins that are modified
31:11by brass induced eye surgeries.
31:14And then in the lymph node,
31:15we saw that these T cells respond
31:18to antigen and expand and then
31:20they can go back to the heart.
31:22And traffic to the heart.
31:23But perhaps what was more intriguing
31:26to us was that once in the heart.
31:29You sometimes bypass this later on
31:32within the heart because they can
31:34actually recognize antigens within the
31:37heart and be expanded there under.
31:40This has significant effects
31:42on cardiac fibrosis.
31:46And I I didn't show the kind of
31:49fibroglandular also had significantly
31:51decreased fibrosis and this is going
31:53to become more relevant for the next
31:56part of the talk where I will be
31:59talking about this critical antigen
32:01recognition that happens in heart.
32:04So for the last part of the talk,
32:08then we I will focus about these kids
32:12are correct fibroblast crosstalk.
32:15So as I showed in the first part of the talk,
32:17when the diesels Infiltrator hi,
32:19this is an image ex vivo.
32:21So these are fiberglass and culture
32:23with T cells and you can see the
32:26green cells are here and the blue
32:29little nuclei of the diesels and
32:31this is a large nuclei of fiberglass.
32:33So we had found that with the diesels,
32:37either DH one cells that were
32:39generated extra evil or T cells
32:42isolated directly from TAC mice.
32:44They bound to the fiberglass
32:46and once they bound they induce
32:48the transformation to alphasim
32:518 producing correct fiberglass.
32:53And then in in the second part of
32:55the talk I just recently showed you
32:58that I didn't present themselves
32:59and particularly in lyrics.
33:01Elves present antigen to T cells
33:04and there is this intramyocardial
33:07diesel receptor engagement.
33:10So then we we thought,
33:11well,
33:11they're not that many dendritic cells
33:13in the heart as compared to other cells,
33:16right?
33:16Is it possible that during this T cell,
33:20fiberglass crosstalk not only the T cells
33:23are telling the fiberglass to to induce TGF,
33:27beta and transform?
33:28But maybe the fiberglass because diesels
33:31are firmly adhered to the fiberglass.
33:34Maybe the fiberglass may be functioning
33:37as also an antigen presenting
33:39cell that is semi professional.
33:42So we went back to literature,
33:44and in this there's this growing
33:47field of struggle immunology,
33:48where the concept is that antigen
33:51presentation is no longer an
33:53exclusive domain for the lyrics.
33:55Also obviously then Derek cells
33:57are antigen professional antigen
34:00presenting cells,
34:01but they're also evidence that
34:03a stronger cells that support
34:05tissue architecture can serve as
34:08antigen presenting cells.
34:09Depending on the context.
34:11So this is an.
34:13In an example of fibroblastic
34:15particular cells in the lymph nodes.
34:18Timing is stomach cells can do that as well.
34:21There.
34:21There are many reports that
34:22show that two more infiltrated
34:24fiber blasts do that as well,
34:26and it's also a recent report in
34:28the Lang where Lang epithelial cells
34:30in the context of inflammation,
34:32can actually present antigen
34:35to certain diesels.
34:37So we hypothesize that cardiac
34:39fibroblasts may be functioning as antigen
34:42presenting cells, and that these.
34:45T cell receptor engagement that we
34:47were seeing in the heart was not
34:50exclusively due to the dirt excels,
34:52but also to cut fiber breads.
34:54And a wind.
34:55Emma let this work and then called it cower.
34:58In my lab was also contributed
35:01significantly to this project.
35:03So to remind you what an antigen
35:05presenting cell in order to be an
35:07antigen presenting cell as poor
35:09as an indirect cell.
35:10You need to efficiently internalize
35:13and process antigens.
35:15You need to display them
35:17bound to MHC 2 molecules.
35:20And then you have to present
35:22that at the cell surface and
35:24professional Apcs and Rick cells
35:26constitutively express all of these.
35:29MHC do is constantly expressed and then
35:33these costimulatory molecules CD80 or
35:36CD86 that are induced upon stimulation.
35:41So we started investigating whether correct,
35:44fabulous may fit into this category.
35:47So this is a way that we select correct
35:50fibrous in the heart with digest the hearts,
35:53and then then we acquire this
35:56and all the non fraction.
35:58We're having the filial cells.
36:01We'll have local sides and we have
36:04a correct fiberglass here here.
36:06Sorry so we have leukocytes here.
36:093045 positives City 31 positive and
36:12killer cells within the local sides
36:15you could look for any local side that
36:18you're interested in and within the
36:20double negatives not in the filling.
36:22No leukocytes.
36:23We use these marker to detect
36:26a cardiac fiberless mask 4.
36:29We also do this in Linux reporter mice,
36:33and that's where is indicated here.
36:35So these are Linux tracing mice
36:38where we could more more definitely
36:40get into the cardiac fiberless.
36:43So the first thing that we did is do
36:45they express MHC two and a baseline?
36:48They don't.
36:48But as soon as you activate them with
36:52interferon gamma you induce expression
36:55of MHC two and actually in the filling
36:57search for instance by people here
36:59at nearly the department of Pathology,
37:01German Barber,
37:02and others found that endothelial cells
37:05can present antigens to T cells as well,
37:09and they respond and express MHC two
37:11in response to interferon gamma.
37:12So this would be a.
37:14It's similar mechanism of expression.
37:17And then what we found it was
37:19that they do express customer
37:20little molecules that you need to
37:22trigger that diesel activation.
37:24They do express CD 80 and
37:27is not further inducible,
37:29induced by different comma but
37:32they don't express 86.
37:34We collaborated with Jenn Davies and
37:36their impact grad student in her
37:38lab at the University of Washington
37:40and exactly the same experiments
37:42using the Linux trace in mice.
37:44That is a reporter for correct fiberglass,
37:47as shown in here.
37:49And as you can see here,
37:50all the correct fibers that are shown here.
37:53The majority of them in response to it there,
37:55from gamma, they express MHC 2 here in red,
38:00so this is GFP and this is no inter
38:03from gamma. With their from them.
38:07Does this matter in vivo?
38:08So in vivo we did pack and we found
38:11that carrot fiberglass isolated from
38:14from this report device expressed
38:18MHC 2 and you can see it here.
38:21You can focus here and this is zoom vision.
38:24So this will be all in green.
38:27Are cardiac fibrosis and as you can
38:29see there are also other cells that
38:31could be in the filler cells in a
38:33small kappel Aries or the drink cells
38:35as we previously shown that Expressen EC2.
38:37I've been here.
38:39Definitely,
38:39the correct fiberglass are expressing MHC
38:43two in response to tech as well.
38:45And sometimes if you look you can find
38:48that T cells seem very close to this
38:52MHC 2 expressing correct fiberglass.
38:54We use all the models of cardiomyopathy
38:58and cardiac inflammation to see
39:00whether this was unique or not,
39:02and we used the tickers eye infection
39:05model of myopathy because we know
39:07that because I is a parasite,
39:10I didn't use his highly strong
39:12in there from gamma responses,
39:14and as you can see here in this model,
39:16the correct fabulous also expressed
39:18any todo and more more of them
39:21express MHC do and then at the MFA.
39:25The prizes and densities also higher.
39:29So then the next question was,
39:31well, let's see if they can.
39:34Take up the antigen processor and
39:36present that induce T cell proliferation
39:39and to do that we use a reagent that
39:42is do DQ of album and so this is a
39:45novel women protein that can be taken
39:48up by proteins and if it goes in the
39:51lysosomes with acidic lysosome pH,
39:53which is what you're required to
39:56process antigens it costs related
39:58degradation and becomes for S.
40:00And as you can see here,
40:02regardless of the interference.
40:04Treatment DQ over is processed
40:06by cardiac fibroblast.
40:08This is just one example of correct
40:11fiber rest treated with equal woman,
40:13but you can see here the comparison
40:15of a large correct fiberglass
40:17and obviously a smaller in size.
40:19Here the bone marrow derived
40:21cells that a process.
40:24So that was very exciting too,
40:26because then that means that
40:28if they're able to process it,
40:30they might be able to load it into MHC
40:33two and induce decent proliferation.
40:35So we did similar studies as
40:37what I showed before to to look
40:40for diesel proliferation,
40:41and we use in this case we use
40:44transgenic mice that are what they do,
40:46so these mice all the T cell
40:48receptors in the details express
40:50a receptor for available mean.
40:53And then we took this specific piece
40:56of argument and then on the other hand,
40:59we took it a wild diaper.
41:00Makes you do knockout.
41:01Correct fiber rest.
41:03Three of them within their front comma
41:05and treated them with normal woman.
41:07So in this Co cultures,
41:09if they carry fiberglass,
41:10are processing available mean?
41:11As I I recently showed with Valve woman.
41:15All these diesels with a receptor
41:18for Valve women should be able
41:20to proliferate and we did other
41:23experiments in which we use E.
41:24Coli,
41:25a bacteria that over expresses
41:27about women as well.
41:29And again,
41:29these are the readout of proliferation.
41:31If there is prolific,
41:33there's no proliferation.
41:34These teachers that I label with
41:36CFC will not dilute the die,
41:38so we wouldn't see any peaks any dilution,
41:42but if there is proliferation we will
41:44see this dilution of the membrane dye,
41:47but that's exactly what we saw here,
41:50so this is overwhelming protein.
41:52But here the fiberglass haven't
41:55been treated with interferon gamma,
41:57so they don't express MHC 2.
41:59Very little proliferation,
42:00but here what you can see is that
42:03when they you induce expression
42:04and you treat with over protein,
42:06there is a significant proliferation
42:09of diesel,
42:10suggesting that the fiberglass
42:12can induce diesel proliferation.
42:14And here's a demonstration of
42:16dendritic cells as a positive control,
42:18where we see a proliferation
42:20and as I said before,
42:22these are the professional
42:24antigen presenting cells,
42:25so they don't need to be pre
42:27treated within their from grammar.
42:28They express MHC 2. Constitutively.
42:31We also did these experiments,
42:33obviously with the MMC to knockout.
42:35Correct fiberglass to to show
42:37that this was a specific.
42:41So just to be.
42:44Over a convenience with this
42:47we do particularly proteins.
42:48So instead of putting a
42:50soluble of argument there,
42:52we collaborated with Carolyn Genco and
42:54Robert in our floor in order Department
42:57and they just happen to have these E.
43:00Coli that over expresses,
43:02so we try to correct fiberglass with E.
43:04Coli that had an empty vehicle
43:07or expressing involvement,
43:09and we saw that only when when E.
43:12Coli was expressing about woman we saw.
43:14Diesel is specific for ovum proliferate,
43:17and this is all quantified here,
43:20and this is the positive
43:22control with the sender excels.
43:25So going back to the cardiac pathophysiology,
43:29does this make?
43:30Does this have any effect on correct
43:33dysfunction or cardiac fibrosis?
43:36So in collaboration with Jenn Davies,
43:38the University of Washington,
43:40we obtain the TCF 21 Mercury
43:44Mirror mice decree driver.
43:46Please inducible and we
43:48crushed it with MHC to flux.
43:51And we generated the correct
43:54fiberglass specific deficient in MHC
43:57do and these mice are only deficient
43:59if you treat them with tamoxifen
44:01because it's an inducible line.
44:03So as you can see here,
44:05we corroborated that it when
44:07we treated with tamoxifen,
44:09we decrease the expression of MHC 2.
44:13Here, so this this is what we will be
44:15looking at and we decrease expression.
44:18Incorrect.
44:18Fabulous, but not in bone marrow Dr.
44:20Dendritic cells where MHC 2 levels
44:23remain compatible in the treated
44:26and not treated with oxygen.
44:28And then we looked at fibrosis and as
44:30you can see here there was significant
44:33fibrosis in the TACK control group
44:36than non democracies untreated,
44:38but when we treated with tamoxifen we
44:41reduce fibrosis significantly and this
44:44has an impact in fractional shortening.
44:46So here's that mouse with flattened.
44:50You know contraction here
44:52and this is the most mice,
44:55so again these are the ones that don't
44:58have image do in the fiberglass and
45:01they have preserved systolic function.
45:04So we looked in the lymph nodes
45:07to right because we wanted to
45:09see whether this was where.
45:11Remember that we're eliminating this in
45:14the in the cardiac fibroblast and they
45:17might be other cells that express TCF 21,
45:20although it is,
45:21it was described to be a
45:23driver for collect fibers.
45:24And what we find is that the T cell
45:27immune response in the lymph node
45:29is not altered by by this intact.
45:32And we also see.
45:34Similar infiltration of character
45:36in these mice with tamoxifen.
45:39So working hypothesis now is that
45:42these are the conclusions right that
45:44in these crosstalk we have the T cells
45:48and the fiberglass communicating.
45:50And in this two week crosstalk
45:52we think there correct fibrils
45:53are Sentinel cells that can sense
45:56correct insults and directly boost
45:58the adaptive immune response.
46:00We think that there's a potential of
46:03moderating decent immune responses
46:04without impairing systemic diesel
46:06activation by the cells which could
46:09have undecided major suppressive effect.
46:11So the fact that we see.
46:14Similar activation in the in the
46:16lymph nodes that that tells us to
46:19think that these this is critical in
46:22the heart for the correct fibers.
46:25And then the overall summary,
46:26just to wrap up and leave some time
46:30for questions is that responses.
46:32I think we're pretty convinced with
46:34our work and a lot of the work,
46:36that of others that I've cited and
46:40that we we always site in in our papers,
46:42is the diesel immune responses
46:44contribute to the pathophysiology
46:46of nonischemic heart failure in
46:47many different ways, right?
46:49So we think that eleven took blood pressure,
46:52induces a significant levels of
46:56drugs and the formation of new
46:59antigens that trigger this activation
47:02in the heart. Within that limited repertoire
47:05of those T cells respond to ISO LGS.
47:08Modified cardiac new antigens and contributes
47:11to cardiac fibrosis and dysfunction.
47:13But we don't think these are the early
47:16antigens that diesels are recognizing,
47:18because if you recall from our data,
47:20even when we scavenge those icebergs
47:24modified proteins, we still see
47:26some decent activation in the heart.
47:29So we are doing a lot of more in
47:31depth analysis of single cell.
47:33He's service center sequencing and
47:34trying to get to what are those other
47:37antigens that might be induced response?
47:39And we also see that they're not the same.
47:43Backgrounds overtime,
47:44which might be very relevant to
47:46see how heart failure progresses,
47:48at least pretty nicley.
47:51And then the last conclusion from this
47:53is that these bidirectional actions
47:55between correct resident cells,
47:58in this case fiber rise and T cells
48:01contribute to correct this activation.
48:03My fibrous transformation and
48:05dysfunction under the correct
48:08fabulous expression of MHC 2 molecules
48:11is central for these response.
48:15So with that, I'd like to thank my lab.
48:19I think I've mentioned everyone who's
48:22done the work who's now moved on to new
48:26exciting research leading positions,
48:28and then this is the new members of
48:30the lab that are trying to pick up
48:32on all the good work that previous
48:35former members did in the lab.
48:37Our collaborators at the University
48:38of Washington,
48:39Vanderbilt or collaborators at absent as
48:43Medical Center and our funding sources from.
48:4718 and also from Dallas University.
48:51With that,
48:52I'll be happy to answer questions,
48:54but before that I'll make an announcement
48:56of a very exciting conference,
48:59which will hopefully happen in
49:01person is scheduled to be in person
49:04in Chicago and there will be a
49:06lot of interest in science,
49:07not only in information but a lot of
49:10cardiology and basic and traditional science.
49:14So I'll be happy to take any questions.
49:16Thank you for your time.
49:21Thank you Paula for the wonderful talk.
49:24So now we are open to questions.
49:28I can maybe stop sharing and.
49:33Either way, would you like me?
49:34Or maybe I can leave it open
49:35in case I need to go back to
49:37good? Yeah, good idea Harold, please?
49:40Yeah hi, I really enjoyed the talk.
49:44Wonderful stuff.
49:45A very simplistic question.
49:47So at autopsy when we see
49:49hearts from patients who have
49:52really horrible heart failure.
49:54I don't ever recall seeing a
49:56striking infiltrate of lymphocytes.
49:58Is it that we just get them
49:59at the end stage or or?
50:03I think so compared to other cells
50:05T cells they you don't see massive
50:08infiltration as you would see.
50:10For instance post MI in the
50:12in the in fact zone, right?
50:15And I think I think you don't
50:18need that many of them,
50:20so they're definitely sparse
50:22compared to other major cells.
50:24And then the other thing that I would
50:26say that we've seen when we take
50:28samples from Elbert issue is that
50:30the human heart is very large, right?
50:32So it it also depends where you take
50:36the the piece from, and we've seen.
50:40We've seen some samples that
50:43have more than others.
50:46I don't think it's the timing
50:48because we've done. I mean,
50:50we haven't looked at human hearts early on,
50:52other than those of those healthy,
50:54right, and those have noticed.
50:57But if there was a way to
50:59look inhuman in mice,
51:00you can track that very easily, right?
51:03And one thing that we don't see is that.
51:06And in the chronic phase.
51:08So if you take the mice way longer,
51:10we see that the diesels are more
51:12activated when we use the reporter mice.
51:15But we don't see more details
51:17of our own per say.
51:19So I think that's that's
51:21maybe why inhuman is tricky,
51:23but definitely is not a
51:24dominant cell in the heart.
51:26You have to.
51:27You'll have to find them,
51:29but not being dominant doesn't mean
51:31that they don't do a lot right,
51:34because if they're highly
51:35activated they can release a lot
51:37of factors and do do other things.
51:39But that's a great point, thank you.
51:41Thank you for your question.
51:44I have a question actually 2 questions.
51:49So wonderful talk again.
51:52So do you think this fibroblast
51:57CD4T cell interaction may also
51:59induce macrophage to invade or
52:03activate resident macrophage
52:06to contribute to the fibrosis?
52:09That's the first question.
52:10Second question is related
52:13with hardwood question,
52:14have you considered utilizing
52:17genetic heart failure model such
52:19as meising heavy chain mutation?
52:22Our foes reach you genetic HTM mice
52:25as well as dilated cardiomyopathy like
52:28muscle Lim protein knockout mouse.
52:32They have natural heart failure.
52:34Whether your hack information could
52:36be extended to genetic heart failure,
52:40which may mimic human heart failure
52:42more closely. What do you think?
52:44Yeah, those are two great questions,
52:46so I'll ask for the first one.
52:47The first one, you're totally right.
52:50We've seen that cardiac fibroblast.
52:52Really, ski machines that
52:54not only attract diesels,
52:55but they attract monocytes.
52:58And we we did find that actually
53:02before Sumanth Prabhu had now in
53:05Washington University of Saint Louis.
53:08He found that early on,
53:11and we've corroborated that the Maya
53:14size the CCR 2 positive Milo itself,
53:17so the haematopoietic Lee derived monocytes.
53:20They infiltrate the hard
53:22before the diesels do,
53:24and we found following following up on that,
53:27we found that the correct fiberglass
53:29they make CXCL 9 and 10 that are
53:33chemoattractants for diesels,
53:35but they also make a lot of C, CL two.
53:37So that makes a lot of sense
53:39that when they sense pressure.
53:41And they release the chemo kids.
53:43The second thing in that related
53:46to that question is that.
53:51Once they infiltrate,
53:52so we,
53:53we've found that the major source of the
53:56diesel chemoattractant proteins is not.
53:58The fiberglass is actually the Milo itself.
54:02So those I think it's all orchestrated.
54:05Basically they fibroblasts release
54:07chemo treatments for innate cells.
54:09Then they adapted cells come
54:12because there's also an interaction
54:13between a myeloid cells and the
54:16fibers that we cannot ignore.
54:17I didn't do your second question.
54:19I would love to look at these
54:22models of carry myopathy.
54:25We haven't looked because we we've never.
54:28We don't have the tools or or the mice,
54:31but I would love to to do it because
54:34I think it's it's very important
54:37and especially in those mutations
54:39that the myocytes are are working.
54:42And I did,
54:44at dysfunctional from very early
54:46on it spontaneously right.
54:47You could really track and I'm sure that
54:50there will be other antigens involved, right?
54:53So it might be that we will need to find
54:56out whether inflammation plays a role.
54:58It might be that it has
54:59nothing to do with information,
55:01but if it did, if it did,
55:03it would be easier to track whether the T
55:07cells might be recognizing proteins that are.
55:11You know that may be misfolded,
55:14or that their mutated due to
55:16the mutation in the myocyte,
55:17so that that would be a yeah, that would be.
55:20That's that would be an excellent Ave.
55:23Thank you any other questions?
55:33So yeah, I I,
55:35I'm very excited about the talk.
55:37I have another follow-up question.
55:40What do you think about
55:42stressed cardiomyocytes?
55:44They may release new entity to the
55:48to the identical cells or two so
55:50they they catch up the antigen and
55:53present to CD 4T cells because we
55:56have cardiac troponin T troponin I.
55:58Fragmented release in the injured heart
56:01and and also cardio my side when stressed.
56:05They may secrete teacher
56:07Beta 2 instead of beta one.
56:09We adapt be interesting avenues for you to
56:12yes so that so first hypothesis.
56:15When we started going after the antigen
56:18when J join my lab and he really wanted
56:21to look at this diesel drones right
56:24and our first hypothesis was I had
56:27just written a commentary on a paper.
56:30Looking at all these in altimmune
56:33myocarditis you know how character
56:35planning and myosin binding protein
56:38C and all these proteins that
56:42that are fighting people, right?
56:45So we thought that that that those
56:47who were going to be the ones but
56:50in the tag model because we didn't
56:53see death of Carrie myocytes,
56:55we didn't focus on that.
56:56But you're right, they might be.
56:58They might be that the stretch.
57:00Induces, as you could do that
57:02nicely with your model, right?
57:03Because you can stretch all these cells we.
57:06We don't have the ability to do that,
57:07but I think it's also possible because
57:10the myocytes see the fiberglass
57:12seed in between the myocytes, right?
57:14So there's all these literature that,
57:18and a huge field of research that
57:20people study kind of fiber as
57:23kind of myocyte communication.
57:25So it might be that those fragments
57:28are actually picked up by you?
57:30Know the Mayo side doesn't
57:31really need to die.
57:32It might be that it's a stretch
57:34and the fiber rest pick it up.
57:37And then the fiberglass percent,
57:38but that's purely an in speculation.
57:41We haven't.
57:41We haven't looked at that,
57:44but I think it's not only.
57:46I think this is very complex and
57:48it's not only limited to the
57:50T cell binding to fibroblast.
57:52I think there is a.
57:54Cross communication with like an
57:57orchestrated response there with my
58:00insides fiberglass and immune cells.
58:03Yeah, so we have resident go ahead.
58:07Yeah this is Jeff Squire.
58:10I just wonder you made a comment
58:12on your introduction that said
58:14that no immune intervention.
58:16No trial has produced any
58:18effect on cardiac failure.
58:22And I wonder whether there any
58:24observations in patients who receive.
58:27Chronic immunosuppressive therapy
58:29with any number of different drugs,
58:32whether there's any effect
58:34on cardiac failure.
58:36Yeah, so we did actually have to look at
58:40that because we made a long table of.
58:44Exactly looking at that right of,
58:46you know, these were the TNF blockers
58:49and these are other immunosuppressive
58:51agents and we didn't find any.
58:54I don't think there's been.
58:56There's been a small trials looking at that,
58:59and I think people have looked
59:02at method tracks and other drugs,
59:05but I don't think there's a detail
59:07investigation of what having the
59:09expectation would be that if you
59:11suppress inflammation it be good, right?
59:13But those drugs also have a lot of side
59:17effects that may be in patients with.
59:20Cardiac failure are no quick right,
59:24so I think we really need to dive into
59:27into not blocking inflammation generally
59:32and try to find a smaller pathways.
59:37I certainly agree, but I just
59:39wonder whether there's any
59:40evidence that you know what the
59:42effect of the immune system is
59:44on in clinically and inpatient.
59:46A lot of patients who get,
59:48you know steroids and get
59:50cyclosporine and other.
59:53I I don't recall all the details,
59:55but there is a very elegant review by dogmen.
59:59And Luigi Adamo that they published recently?
01:00:02Maybe? Maybe not.
01:00:03That recently, maybe a year ago in Nature,
01:00:06reviews, cardiology, and they have they.
01:00:09They did exactly that,
01:00:11and they reviewed all the literature
01:00:14in large trials, small trials,
01:00:16directly tackling immune mediators,
01:00:20or general immuno suppressors. And I,
01:00:23I recall that the conclusion is what I said,
01:00:26but maybe you know, maybe I mean.
01:00:29But I. I think yes, if you have time.
01:00:32That review was very detailed and it was.
01:00:35It was very nice to to read and they had the
01:00:38they reviewed the mechanistic part of it,
01:00:40but then they review all the patient trials.
01:00:43At the end.
01:00:45I believe it was in nature reviews,
01:00:47cardiology for sure and I don't
01:00:49know if it was 2020 or 2021.
01:00:52But
01:00:53thank you yeah. Yeah.
01:00:59So if if there are no additional
01:01:01questions so thank you so much
01:01:04Paula for this exciting talk.
01:01:05We learn a lot cardiology immunology,
01:01:08so thank you very much.
01:01:09Have a nice afternoon.
01:01:11Thank you for the invitation.
01:01:13I thank you all for attending bye.
01:01:15Bye bye thank you.