EEL Webinar: The relation between microbiote and endometriosis
Speaker: Paola Vigano
Moderator: Attila Bokor
All right good evening everybody and welcome to tonight’s eel webinar um tonight we’ll be having an extremely interesting and controversial topic and um we’ll be discussing about the relationship between endometriosis and microbi um as you got used to it we’ like to thank our sponsor G and richer for their
Support for El webinar and for that please allow me to um share short video to make a medicine you need to shift your perspective to see success comes from a series of Innovations without the dedicated work of our highly trained researchers no such progress would be possible it is today that
Experts need to think about the diseases of tomorrow and premium quality requires the latest technology all these aspects come together to create new treatments that will improve the health of millions this is what we work for every day Gideon RoR health is our mission okay now it’s my pleasure to um
Present our tonight’s um moderator um actually he doesn’t need any presentation he’s an um e Executive Board member profor AA Boer from the zal vice University in Budapest um who’s involved in so many projects um and he’ll be moderating tonight’s interesting topic AA thank you for being here thank you alen thank you
For your kind words and and our guest today is Professor paa veano I think she’s really an expert an authority of the field everybody knows Professor veano he uh he has an ex very extensive work on endometriosis and she’s also presently the IVF the head of IVF Center at the
University Hospital in Milano and she’s also the the co-editor in Chief of the Journal of endometriosis and uterine disorders the official Journal of SE she has published over 250 articles 250 I think this is very impressive and she has more than 5,000 citations already so she used to be also
Editor Associated editor of human reproduction update and human reproduction as well the top journals in Reproductive Medicine um Professor veano very recently was recognized among the top four World experts in endometriosis by expertscape ranking in paloalto so I think she really doesn’t need any more explanation so I’m very
Happy that she accepted our invitation thank you professor vgo and could you please uh share uh your knowledge with us whether is there any link between g m microbiota and and endometriosis I think this is really a very interesting question thank you very much fantastic presentation thank you thank you for the
Kind invitation I’m going to share my Screen do you see my screen yeah okay okay can go can I start yes please okay so thank you no I can’t right now I cannot see it oh sorry let me See okay now now it’s perfect it’s working okay thank you very much again for this kind invitation so today we will my ah sorry this is these are my conflicts of interest in the last three years and today uh the plan of this presentation uh is the following first
Of all we will talk a little little bit about the general aspects of the microbiome then I will try to show you some breakthrough of the microbiome resarch in other field just to understand what is happening in other uh Fields then we will talk about the limitation of microbiome studies because
I would like to communicate to you to inform you about the limitation and to have a critical U way to consider this study then we will talk about microbiome and endometriosis and the data and finally we will discuss about this very uh interesting study on fusobacterium and endometriosis and we we’ll try to
Draw some conclusion so uh about the general aspects of the microbiome um the interesting the microbiome has is has increased substantially in the last year and this was due to the human microbiome project which was a US NIH initiative which was a set up to understand the microbiota involvement in human health
And disease and the project was launched in 2017 with two phases the first phase was to characterize the microbiomes of human uh of healthy human sub at five major body siid using 16s and metagenomic sequencing and then in 2014 the NIH launched the second phase of the project project which was called
The integrative human microbiome microbiome project uh whose goal was to characterize microbiome and OST from free Cordes of microbiome Associated condition using omix technology the free condition that the NIH want to study were pregancy and preter birth U with the aim to understand the microbiome changes during the gestational period
And how microbiome could influence the neonatal microbiome the mother could influence the Neal the second condition with the inflammatory bowel disease with the goal to understand how the gut microbiome uh can change in women in adults and children suffering from inflammatory boow disease and the third condition was type two diabetes to
Accept the shift in microbiome of patient compared to not affected uh individuals here you have a this slide describe the biog geography of human microbiome and if You observe the various composition of the bacteria in the various uh body uh sides bacteria are these in the legend you can see that
They are in different colors and you can realize that for instant V bacteria which fuzo bacterium which is here in Orange um is substantially present in the oral cavity and not in the other District of the body and uh all um everybody knows that the microbiotic composition from the same site of
Different individuals are more similar than the microbioma composition of different side of the same individual another example for instance here in Violet you have the fuus filum which includes the lactobac which is dominant in the vagina now we know that the microorganism and the OST communicate in numerous way so uh including the
Production of metabolism and the modulation of the immune system which is very important but also through a direct Exchange of biochemical and genetical material between the microorganism and the OST and in turn the OST can influence the microbiome in in a variety of ways for
Instance in the gut it can be the diet but also the pum modulation while in the vaginal for instance it can be the sexual abits but how uh can we uh we um um measure the microbiome how can be the microbiome be evaluated so usually the genetic material is extracted from
Samples for instant FAL samples and they are prepared for sequencing uh and then the genetic sequencing are filtered and compare to a nonn database to identify to which microorganism that sequence belong to and then from here different multivariate analysis can statistical analysis can be performed so you can do
An analysis of abundance that tells you how many percentage of the microbiome are made up of a specific organism you can do an analysis of biodiversity that tells you what amount of individual bacteria from each of the bacterial species is present in your microbiome but you can do also an
Analysis of function that can identify difference in metabolic function and you can also do a principal component analysis that characterize a sample and compare it to other samples so uh an extraction and different way to analyze the data it is also important to underline that of course the microbiome and the
Microbiota is are important but also their metabolizes are important for instance the Gat metabolis the gut microbioma seems to have an important role throughout the body and is very important to some aspects of women’s health because the microbiota of the gut consist of um B microorganism that can
Produce an enzyme which is called glucuronidase and this enzyme can deconjugate estrogens when they were about to be excreted causing them to re-enter into the body and remain active for all the function negated by estrogen and for instance this is very important for the composition of the vaginal microbiome because the vaginal
Microbiome is affected by the estrogen because the estrogen can stimulate the production of glycogen and G glycogen in the vagina can promote the dominance of the lactobac so um I wanted to spend few uh slide also to um to to understand what is happening in another field which is
Very important and I choose as an example the um study of the microbiome in the connection between the gut and the Brain probably those of you that has seen the that has U participated to the West Congress the world Andis concer in Edinburg may have uh seen how this
System works because there was a presentation there but uh uh in summary uh neuroscientist have a specific interest in this in understanding the role of the G microbiome for for the brain and there are two big challenges in the study of this interaction one is to actually understand and underpinning
How the the Gap microbion can send a signal to the brain and second since they have uh very interesting results on mice they are trying to understand how much these the interesting data in mice can be translated into humans it’s it’s very important to underline that to gain evidence that the
Microbiome is involved in a system uh a good strategy is to take out the microbiome from the system and see what happens and this is the the basis for the use of The Germ free animals to answer many question of the microbiome these germfree mice have been a study a
Lot for the connection between the got microbiome in the brain but I want to focus the the attention on this kind of mice because they have been also used in endometriosis and just as a a an interesting point the these germ free animals have been shown to live longer so what the
Neuroscientist have found brilliantly found is that this germ animals mice have a variety of change in the brain a lot of change they have an hyper permeation in the prefrontal cortex of the brain and particularly this feature is m m is a characteristic only of the
Male mice then these G free mice have a dis regulation of the neurogenesis of the hippocampus neurogenesis these mice have changes in a variety of NE neurotransmitters including gab and serotonin they have changes in the immune cells in the brain they don’t function correctly in the absence of the microbiome and finally
These germfree animals um have a disrupture in the blood brain barrier so a lot of changes in this animal without the microbion that influence every aspect of this life of the mice alive uh but also um um all the all the function that are based on the brain which is an incredible uh
Demonstration so let’s go on a little bit on uh uh what are the limitation of this microbiome study I want to underline that there are some limit limitation not only in endometriosis but in every field that should be known uh to critically evaluate the studies on uh
Microbiome and I want to share with you this um fantastic paper uh U published in nature biology that have um listed all the myth and misconception on microbiome that everybody should know first of all one of the myth about the microbiome is that the microbiota out
Numbers human cells by 10 to one which is not exactly uh correct uh this according to this paper um more detailed analysis indicate that the true figure albate still impressive is probably C to a ratio of one to one and it is it should be noted that the
Ratio is likely to vary from person to person and is dependent on factors such as the airort the size and the amount of FAL material that are present in the column so uh it is not as much as um numerous as we thought at the beginning another M misconception as about the
Microbiome study is that sequencing is uh unbiased um actually um although sequence be method has been transformed formative for microbiome research with this new uh um on approach they are not perfect biases can be introduced at every steps of sequence based study from sample collection to storage through laboratory
Base steps such as DNA structure to to choice of the pipelines for the bioinformatic and to choice of the database used for the analysis so the sequencing is not perfect always perfect and another myth about the microbiome underlined by this fantastic paper in nature microbiology is that most disease are
Characterized by a part of biome we always say that dead disease is characterized by a disbiosis a pathobiome and actually these ter these terms these biosis my Pome are simplistic um because microorganism and their metabolites are neither good or bad they merely exist their impact on on
Us as host are heavily dependent on contest microorganism or metabolized areitos in one context CA not not har in another and which is a very important uh point and uh um what I also would like to to underline is that uh we have a lot of
Condition in the brain the lungs in the liver in the intestin and so uh and so on what also systemic disease and one common team and in this in this organ we can have a multiple sclerosis parking for disease asthma liver cancer diabetes etc etc but what one common uh aspect
Among all these condition is that they results in increased levels of inflammation as you can see here and uh uh such inflammation can in turn theate the Gap microbiota reducing diversity and allow facultative anerobic to increase as you can see here in red and so in the study of the microbiota since
All the condition are associated with inflammation it is very difficult to distinguish cause from effect in in in this study and since we are talking about an inflammatory disease like endometriosis what is true for all these inflammatory disease can be uh relevant and important also for endometriosis and I want to present also
To share you to share with you uh the results of this study in which these author are a group of author that respond to a paper published in nature this paper this big paper publishing in nature uh found a strong correlation the the the paper in nature is this one and
These are all the bacteria and this this this paper publishing nature found a strong correlation between microbiome and 32 different cancer types and they created a machine learning predictor with a near perfect accuracy and distinguish among the different cancer some months later these author which are from his paper and their results are
Here check the sequencing of the nature paper and the strong error in the genome leading to millions of false positive findings largely because most of the sequencing identified as bacteria were indeed human so they have concluded that the microbiome based predictor for identify this 32 33 cancer in the nature study were completely
Wrong so sorry for this uh long presentation on the um General aspects of the microbiome and we can go now inside into the data on endom endometriosis and um I told you that ger free mice were also were used in St in the study of the uh the in the study of
The microbiota and actually uh these um authors did this very very nice uh paper very with very good experiment in which actually they have implemented a model using antibiotic IND microbiota deated mice and I apply them in the field of endometriosis so they surgically induced endometriosis in both controls and
Antibiotic uh uh and microbiota deated mice and uh they transplanted a a piece of uterus into the peronal and then 21 days later they evaluated the leion as and as you can see here lesion uh were in normal M were significantly larger and more vascularize than Vision present in the microbiota deated
Mice then they did another model of endometriosis induction because they injected endometrium into the peronal of both control and microbiota deting M and again uh they evaluated the vision 21 days later and again also in this model of endometriosis they found that that the control mice had lesion larger and
Greater in number then lesion of microbiota De plated mice supporting a role of microbiota in the development of the leion but then the authors would like to understand whether the gut microbiota was important for the endometriotic leion grow so to this aim they injected the uterine fragment into the peronal of
Um control and don’t and microbiota deated mice and then transplanted the fal samples from mice with and without endometriosis by Oral cavage in this mice and then uh 21 days later they evaluated the vision and what um I have to say that here when it’s written n e it’s FAL samples from non-endometriosis
M and here are samples from endometriosis m and you can see actually that FAL samples from mice without endometriosis that you can see here were significantly smaller and they develop less lesion a lower number of lesion compared to mice with FAL samples from mice with endometriosis and this was a way to
Indicate that the fal microbiota could U improve to favor the lesion development but then the authors um um ask themsel actually is uh if actually it’s only the the microb biota Deion depletion that in some way alter the uterine microbiome with the consequence that transplanted uterine fragments in
Microbiota the pl in mind are less able to form Vision then fragment in control mice so to test this possibility the author injected endometrial fragment from ble and microbiota deting mice in control mice and in peronal cavity control mice and what they observe is that mice receiving the endometrial
Fragments either from the basical or from microbioma the we might the velop lesion in similar number and volume and size and so they concluded they also they they the auo concluded that the uterine microbiota might be dispensible but not the Gap microbiota in for endometriotic lesion
Grow in mice and the results of these very well done study is that microbiota deting mice show reduce endometriotic leion grow the transplantation of Gat microbiotal Boral gavage of samples FAL samples from M endometriosis rescued the endometriotic Le should grow supporting the role of the G Mota while the uterine
Microbiota was shown to be dispensible for endometriotic leion growth okay fantastic data on M what are the evidence in human we will see rapid ly the paper in the vaginal of the vaginal microbiota in endometriosis women in women with without osis of the uterine microbiome and of the Gap
Microbiome so what about the vaginal microbiome in women wom without endometrosis I didn’t do a systematic review what I did is taken the paper publish on the vaginal microbiome in endometriosis and I have checked all the bacterial that are present in the the virus increase or decrease if there is a dominant of
Some bacteria in the vaginal in the vagina of women with invol osis and you can see that the results are quite different and apart from some concordance for KI and shella uh there are the results are quite um uh different among them and I don’t think we can conclude so far um something
About from this human data on the vagina what about endometrial microbiome before showing you the results of the uh of the endometriosis patient I want to underline the fact that the endometrial microb biome is receiving a lot of interest in the context of the Reproductive Medicine in
The IVF because there are some groups that um has studied a lot of the endometrial microbiome and they found a good correlation between the endometrial microbiome and the results of the and the IVF outcome this is only one only just one of these study in which the authors have investigated the
Reproductive impact of an altered endometrial microbiota in endometrial fluid on implantation miscarriage ongo pregnancy life rate and they have evaluated 35 women undergoing ex or oide donation treatment with at least one good quality embryos and then uh they uh classify the samples of the endometrial microbiome with um two machine learning
Models H and both model provided similar conclusion that the percentage of lactobac in the endometrial microbiome should be uh might be a significantly uh variable able to predict the life burth rate and these are the results in which you can see very clearly that when the endometrial microbiome has a dominance
Of laob Basil you have a pregnancy rate of 70% while the when the the endometrial microbiome uh lacto basil is is not do is not dominant the uh um pregnancy rate slow down to 33% what about endometriosis again I didn’t do a systematic review I just check the literature for for for the
Original papers and you can see that there are some bacterias ineter which is very interesting from a biologic point of view because endometrial should be a sterile environment but when we check all the paper together there are different abundance in different bacteria and again nothing is concordance and so it is very difficult
To do some results from this study and what about the gut microbiome again I uh uh I check the the the papers there are not a lot of paper on G microbiome and endometriosis it is interesting because on gut microbiome and endometriosis there are more reviews than original
Articles and but again you can see that of course there are a lot of bacteria but the different and the dominance of one over the others in patient with endometriosis compared to controls is absolutely not evident so again uh concluding something from this data is very difficult and we need more EV
And I want to to conclude with this fantastic paper on fusobacterium and endometriosis because it has been um paper that has gone to Media so it is a very very very good paper the experimental uh the experiment many many experiment many experiment have been done and uh now I I will try to
Summarize because it’s a little bit complex to read the paper so I would like to summarize the main point of this paper in order to clarify uh also the the enormous uh amount of um effort which is behind this paper uh first of all um um I would like
To share um the the main table of this of this paper which which is the characteristic of the patient included they including 76 patient without endometriosis and 79 women with endometriosis um they are all patient operated and they are all patient isct tomiz they have uh the uterus has been
Extracted and these um these support the the the age which is not very which is over 40 you can see that the both the age and the proliferative cycle of the women are at the borderline level of the significant between the two group but the last is the uh the the results from
The fusobacterium presence in the endometrium of the these women and they have 77% in women without endometriosis and 64% % in women with endometriosis so of course High statistically significant so it seems that fuz bacterium infection is characteristic of women with endo and uh then what did the author uh
Do the author did a lot of experiment as I mentioned and uh I would like to summarize them for for you so first of all that they they did expression profiling comparing between um endometrial fiberblast from patient with endometriosis fibroblast from ovarian endometriosis and also from fiberblast of women with
Endometriosis utopic endometrial from women with endometriosis and they found that the only genes that were confirm at Artic PCR level were was trans scaling you can see here very clearly that that trans scaling was highly expressed in fiberblast from ovarian endometriomas and lower in the utopic endometrial patient with
Endometriosis and even lower in utopic endometrial patient without endometriosis uh then they did an a Ser of experiment in vitro in which they demonstrated that the up regulation of trans scaling could promote fias proliferation but they also demonstrate the transforming grator beta one that you all know that it’s the most
Important molecule that allow the transformation from fiberblast to myofiber blast to start the fibrosis can be Beed by the expression of transc then they uh went to the animal model and so they infected the cenic doriz with f iium in a model of endometriosis and they injected the
Endometrium in the peronal cavity and actually they found that when the Fus bacteria when when the was infected with the Fus bacteria the lesion that was form in the recipient of the infected uter was uh greater bigger and uh multiple compared to the uh when
The indeter was not infected by the F of bacteria moreover they could demonstrate a abundant M2 mcroof infiltration a higher expression of transforming grator beta one and transr positive myo fi blast could be observed in endometrial trauma of uter that have been infected with the F bacteria and also in the
Endometriotic legion which was established by this endom infected endometrium into the mice and even more interestingly they treated the mice with antibiotics for a week and fusobacterial was no longer present in this m the M2 microf infiltration was reduced transforming of beta one expression what is used
Transing expression what R Us so were already reducing the endometrial or the M endometrial when they treated the um the with the antibiotic moreover the recipient might that receive the endometrium from the uh who of bacterium infected samples that were treated with antibiotics develop significantly fewer endometriotic Leisure than the control
Group without antibiotics here you can see it’s very summary what I’m telling you and here you can see that these are the FI blast here fi blast here in Violet and in Orange is the F of bacterium when the F bacterium is present you can have a n regulation
Transforming Factor beta one that start the fosis process but also increase in trans scaling that faos proliferation attack in Mobility while antibiotics can eliminate all these pathway and I want to show you finally only one this is a very clear and very uh simple experiment in which you can
See that they uh have this donor M and they infected the their endometrium with f bacterium and uh 56% of these bacteria actually have an infection in the endometrial while 44 no but they they use these M they are uterine uterine samples to have been inoculated into the recipient
Mind and when recipient mind receive the uh infected or not infected uter you can see that the number of the lesion and weight of the lesion are significantly increase when the uh the uter is infected by the F bacteria so uh the conclusion uh of my talk are
The following we have very strong uh evidence for a role of endometrial and D Gat microbiome in endometriosis but in animal model in human studies are few uh usually they have a small sample size and we are absolutely unware unaware on whether the microbiota function can be the cause of the
Consequence of the disease we have absolutely uh we need more evidence to to understand that the mechanism underline the possibility to to have a role in endometriosis in women with endometriosis on the microbiome and uh I really uh would like to you to understand that in evaluating this study
We uh based on what we have seen in another another field we have to be very critical on the collection and the sequencing bias that the study may have and I would like to thank you for your attention thank you very much it’s just very interesting so this actually this is
What exactly what I expected since I also read this paper on Fus of bacterium actually I I didn’t really understand everything so it was I think it’s very useful that you just summarized for us and indeed it’s I think it’s a good methodology and it’s a very very solidly
Written paper however I’m I’m not very sure that it will be very easy to reproduce it so yeah yes but I don’t want to say that it’s it’s so I I’m really looking forward for a for another other group to to reevaluate this data yes the the point is that you may
Reproduce the animal models the animal study and the reproduce you may reproduce the woman study so there are two different point of view uh maybe so we need data we need data absolutely in women because okay 76 women but uh we need more data on on that absolutely
Because also because uh that women was a is I women and we actually the the paper didn’t mention on how they collected that endometrium so we don’t know if they open the uterus and took the the samples or rather they passed through the vagina and the cervix which is not
Very good in the study of endometrial samples from the microbiome yes definitely um is there any question from the audience because we have um an quite a fairly High number of participants let me see so there is one question what is the upto-date Practical point of all that you have just said
Nothing maybe no no I I I would for me I would be very prudent on on all because you have seen that the human studies are very heterogeneous um so um I would stay there and see and not it’s not so easy to tell to a patient to take
Antibiotics you we can have resistance to antibiotics so I would stay there waiting for other evidence if they will they will for sure uh come other evidence yes so what do you think AA in my opinion I would be very surprised that if our patient just just would take some
Antibiotics and all disease will disappear so for me and if it’s true I’m very more than happy because we don’t do any more major surgeries on these patients so I think this is a very bright future but at the moment as I understand the results are quite
Conflicting and there is there is no solid evidence supporting the role of microbiota on the formation of this disase and also the progression of this disease as well absolutely and what you said I think it’s so so true and and I’m I’m still wondering that you have we
Have more review articles on microbiota than original ones so this is a new era of of meta analysis and um for me you know if you have a a very nice metaanalysis published in a in a top journal and it’s it it is based on very poor quality data so your meta analysis
It makes just no sense I agree totally totally I agree totally no there also a lot of narrative reviews but uh I mean it’s an interesting topic microbiome for sure but uh we we need also solid data otherwise we are talking about nothing yes and are you planning any study on microbiota paa
In Milano um we were we we were we were uh we are in the phase that to to see if we can replicate the human study because it’s not so difficult because also the didn’t the F excuse me the human study on fusac sorry fusobacterium fusobacterium the F
Because the fusobacterium study has not been done with a sequencing it was been done with ancr so it’s very easy to replicate the but you know the problem is how to take this endometrium because if you pass through the vagina really the contamination is so
Possible it is yeah last day I was in another Congress and there was a colleague that was talking about about an expert in microbiome that has evaluated the the um our papers in in OB gynecology and this microbioma expert thought that it was crazy to think that
Passing the the vagina and the service was nothing of course it is are important yes so there are a lot of lot of things to clarify but um yes but anyways I think this is still an interesting issue because I I also have the feeling that maybe diet has an
Impact on microbiome and we see that some patients of us of ours they if they follow a DI diet they just get somehow better however we have no evidence on on the use of of of diet so maybe diet in microbiome is or gut microbiome some it
Has some link so maybe in the future we will have some more interesting results and maybe we can give um a good answer to this question from the audience so if if there are no more questions um would you like to say some something for us what is this beautiful
Picture where is this taken this place this place is on the lake of ko on the lake because I live between KO and Milan so these are this is the place where I go in on Sundays on yes this is gorgeous yes beautiful yes thank you very much for
This kind invitation very much really thank you very much for your for your talk it was very interesting and we look forward to the new data and hopefully we can have some more ex exciting discussion on the role of the microbiome and and endometriosis so thank you very
Much thank you to you thank you thank you bye bye thank you before before closing the the webinar please allow me to share with you the upcoming um eel events so next year in 2024 from the 6th until the 8th of June we’ll be having the European andent showes Congress in
Buest and until then we’ll be having two more uh robotic and Metro surgery master classes the first one will be on the 2 and third of November in duburg in Germany and next year in March in uh in Bordeaux in France and and again in December at you are going
To uh host the next um e master class in uh Budapest um and I think we can now call this an evening and uh thank you all for being here and discussing this uh interesting and also controversial topic thank you very much have a nice evening and see you too
By byebye by right bye righte thank You