CAN EVERYBODY HEAR ME EMPLOY
OKAY, WE WILL GET STARTED.
SO IT'S A TRUE PLEASURE FOR ME
TODAY TO INTRODUCE 1 OF OUR OWN
AND ACTUALLY A COLLEAGUE OF MINE
FROM THE NIDCR, THIERRY
GAUTHIER, HE RECEIVED HIS DEGREE
IN MOLECULAR BIOLOGY DURING
TUMOR DEVELOP AM.
HE LATER PURSUED A Ph.D. AT
THE UNIVERSITY OF
[INDISCERNIBLE] IN
[INDISCERNIBLE] FRANCE STUDYING
OUR PHAGOCYTOSIS COULD REGULATE
INNATE IMMUNITY AND HE JOINED
THE LAB AT NIH IN NIDECKER CR AT
TWEBT 18 -
-IN 2018, HE HAS FOCUSED ON
THE ROLE OF MODULATING FUNCTIONS
AND THE CONCEPT OF SEPSIS.
HE ALSO STUDIED THE ROLE OF
ANOTHER TGF SUPER FAMILY, ACT 95
A AND HOW IT HAS MACROPHAGES
DURING INFECTION TO IMMUNITY AND
HE'S TRYING TO UNDERSTAND THE
LINK BETWEEN TGF BETA AND,
METABOLISM AND VARIOUS CONTEXT
AND HE AIMS TO TARGET THESE
PATHWAYS TO DEVELOP NEW
THERAPEUTICS FOR AUTOINFECTION,
IMMUNITY AND CANCER.
IT'S I PLEASURE TO HAVE YOU
TODAY.
NTHANK YOU SO MUCH FOR THE
INTRODUCTION.
I WOULD LIKE ALSO TO THANK THE
ORGANIZERS FOR GIVING ME THE
OPPORTUNITY TO PRESENT THIS WORK
EMPLOY SO I'M GOING TO JUMP
RIGHT IN AND TODAY I WILL TALK
ABOUT MACROPHAGES AND AS YOU
KNOW, THEY ARE INNATE CELL
TYPES.
THEY ARE IMPORTANT TO HUMAN
IMMUNE RESPONSE, SO AS AN
EXAMPLE, THE MACROPHAGES IN THE
LUNG WILL BE EXTREMELY IMPORTANT
TO CLEAR THE PATHOGENS,
MACROPHAGES IN THE LIMP FOADS
WILL BE IMPORTANT TO HELP
MOUNTING A PROPER ADAPTIVE
IMMUNE RESPONSE BUT WHAT'S
REALLY FASCINATING ABOUT THOSE
SETS IS THAT THEY'RE ALSO VERY
IMPORTANT TO MAINTAIN
HOMEOSTASIS AND SO A BREAK IN
THE FUNCTION COULD BE
DELETERIOUS, AND SO IT'S VERY
IMPORTANT TO CLEAR THE SEN
ILLEGALSEN ENSEL THROUGH RED
BLOOD CELLS EMPLOY SO IN OUR LAB
WE ARE STUDYING THE ROLE OF
TGFBETTA.
BUT NOT THAT MUCH INNATE HUMAN
RESPONSE SO I WILL JUST
INTRODUCE A LITTLE BIT OF TGF
BETA.
BELONGS TO THE TGF BETA SUPER
FAMILY OF PROTEIN AND IT'S
COMPRISES SEVERAL MEMBERS LIKE
THE BMPs, HERE, THE ACTINS AND
THE TGF BETA IT'S THE MOST
DRIEBED MEMBER OF THE TGF-B, AND
IT'S IT NEEDS TO BE ACTIVATED
AND THEN CAN BIND TO THE
RECEPTORS TGF-B AND 2 IT WILL
ACTIVATE DIFFERENT SIGNALING
PATHWAYS SO THE MAIN SIGNATURES
THAT WILLING PATHWAY THAT
INVOLVES THE SMADS, THE
ACTIVATION OF SMAT 2 AND 3, BIND
TO SMAT 4, TRANSLOCATE TO THE
NUCLEUS WRRKS IT CAN ACTIVATE
REFRESESSED GENE SUPPRESS AND
CAN BE SHARED BY ACTINS, TGF-B
CAN ACTIVATE SIGNALING PATHWAYS.
AND SO, THIS TGF BETA THAT I
MENTIONED WE HAVE BEEN STUDYING
IN THE LAB HAS BEEN DESCRIBED TO
MODULATE IMMUNITY, OUR LAB BUT
NOT ONLY HAS DESCRIBED THAT
TGF BETA CAN INDUCE REGULATORY
T-CELL GENERATION BUT ALSO CAN
INDUCE TH17 AND TH9 CELLS AS
WELL AS REPRESS TH1 AND 2 AND
CDA CYTOTOXIC T-CELLS SO THIS IN
THE CONTEXT OF IMMUNITY AND IN
THE TERM OF INNATE IMMUNITY,
MUCH LESS IS KNOWN, SPECIALLY IN
THE CONTEXT OF MACROPHAGES,
NEVERTHELESS, THEY ARE STILL
PAPERS OUT THERE, THAT TGF BETA
COULD BE A MODULATOR OF THE
FUNCTION, THE TGF BETA PROMOTE
ANTIINFLAMMATORY MACROPHAGES, IT
CAN ALSO REGULATE MIRROR IMAGE
IMRAIGZ EMPLOY IN THE CONTEXT OF
HOMEOSTASIS, TGF BETA CAN
PROPROTEIN COMPLEX TISSUE REPAIR
AND THE RESOLUTION OF
INFLAMMATION, SO I'VE BEEN
STUDYING THAT QUITE A LOT DURING
MY Ph.D., ESPECIALLY IN THE
CONTEXT OF [INDISCERNIBLE], AND
TGF BETA CAN ALSO CONTROL TISSUE
REPAIR AND REGULATING MICROGLIA
DEVELOPMENT AND FUNCTION AND
FINALLY, TGF BETA SIGNALING IN
MACROPHAGE SYSTEM DIFFERENT IN
TUMOR DEVELOPMENT ESPECIALLY IN
TUMOR METASTATIC DEVELOPMENT.
SO THERE ARE A LOT OF QUESTIONS
UNANSWERED TILL, SO IN A BETTER
CONTEXT, TGF BETA IS NOT KNOWN
DURING BACTERIA WILL INFECS.
THE MOLECULAR MECHANISMS BY
WHICH TGF BETA PLAYS IN INNATE
CELLS IS NOT UNDERSTOOD.
AND THE ROLE OF TGF BETA IN
REGULATING MACROPHAGES IN
METABOLISM IS BASKLY UNKNOWN.
SO THAT WILL LEAD ME TO TALK
ABOUT 2 PROGECS WE HAD IN THE
LAB, THE FIRST 1 IS ABOUT HOW
TGF BETA CAN CONTROL THIS IN
METABOLISM IS THE SECOND 1 IS
ABOUT THE ROLE AND ACTIVATION OF
THE INFLAMMATION, SO I'M GOING
TO START TALKING ABOUT IMMUNO
METABOLISM AND SO, IMMUNO
METABOLISM IS QUITE COMPLEX,
IT'S INTRICATE PATHWAY THAT
INTERACTS WITH 1 ANOTHER AND
TODAY WE WILL TALK MORE
SPECIFICALLY ABOUT THE
GLYCOLYSIS, AND SO THE DPLI
COLSIS START WITH THE BREATHER
OF THE TBLUICOSE TO WHAT IS
TRANSPORTERS, AND IT WILL BE
DEGRADED BY DIFFERENT ENZYMES
THAT WILL LEAD TO A GENERATION
OF PYRUVATE AND IT HAS 2 CHOICES
IT CAN ENTER THE MITOCHONDRIA
AND FEED THE TISSUE CYCLE AS
WELL AS THE [INDISCERNIBLE] AND
IT CAN ALSO BE DEGRADED INTO THE
GENERATION OF ENZYMES AND THIS
IS CALLED AEROBIC FLI COLSIS
COLSIS AND IT HAS BEEN
DEMONSTRATED TO FOR THE
IMPORTANT PATH WAIL IN THE SET
WHEN IS THEY'RE ACTIVATED TO
GENERATE ENERGY ESPECIALLY TO
PROVIDE FAST GENERATION BUT ALSO
BECAUSE ALL THOSE METABOLITES
THEY CAN HAVE OTHER SIGNALING
PATHWAYS AND THEY CAN ALSO FEED
OTHER METABOLIC PATHWAYS.
AND SO, THIS IS NOW VERY WELL
STUDIED THAT CHANGES IN
METABOLISM CAN AFFECT THE
CHANGES IN HUMAN CELLS,
ESPECIALLY MACROPHAGES AND WHAT
IS THE CURRENT DOGMA, THAT WHEN
MACROPHAGES BECOME
PROINFLAMMATORY, ESPECIAL
LEECIAL IN RESPONSE TO LPS AND
INTERFEREON GAMMA SO THEY WILL
START TO ENHANCE THE GLYCOLYSIS,
THEY WILL HAVE A BROKEN TISSUE
CYCLE AND A DECREASE
[INDISCERNIBLE] ACTIVITY, SO
MITOCHONDRIA RESPIRAIGZ AND THAT
WILL HELP THEM SUSTAIN AN
INFLAMMATORY PHENOTYPE FOR
CYTOKINE, CHEMOKINES, MIGRATE,
[INDISCERNIBLE] AND SO ON AND SO
FORTH.
SO THAT WILL HELP THEM SUSTAIN
THEIR ENZYME INFLAMMATORY
FUNCTION EMPLOY SO BASICALLY THE
GLYCOLYSIS SUSTAINS INFLAMMATION
IN MACROPHAGES.
AND SO WE WANTED TO ASK A VERY
SIMPLE QUESTION, WHICH IS BEST
TGF-B TO REGULATE IMMUNE
FUNCTION BECAUSE IT'S A VERY
IMPORTANT QUESTION.
SO WE STARTED WITH THE
MACROPHAGES AND WE GIVE THEM
TGF BETA AND WE RUN SPRMS.
SO INTERESTINGLY WHEN WE GIVE
TGF BETA TO MACROPHAGES NOT MUCH
HAPPENED SO WE THOUGHT MAYBE WE
MEDE TO ACTIVATE THOSE A LITTLE
BIT LIKE T-CELLS ARE STIMULATED
WITH ACTIVATION AND WE CHOOSE
LPS WHICH IS A LIGAND THAT HAS
BEEN VERY WELL KNOWN TO ACTIVATE
TBLI COLSIS, SO I SHOULD GIVE
LPS ENHANCE GLYCOLYSIS, BUT THEN
EXTREMELY SURPRISINGLY TO US
WHEN YOU GIVE THEM A COMBINATION
OF TGF BETA THEY CAN FURTHER
ENHANCE THE DPLI COLSIS, SO WE
COME FROM THE ENDS WE OBSERVE
THE UPTAKE OF GLUCOSE BY USING
FLUORESCENT ANALOGUE, INCREASED
AFTER TGF BETA TREATMENT IN
THOSE MACROPHAGES AND THERE IS A
REFLECT ANT IN THE SUPERINATENT,
AND IT WAS UPREGULATED AFTER
TGF BETA TREATMENT AND OVER ALL
WE SHOW THAT TGF BETA CAN
PROMOTE FLI COLSIS COLSIS AND
MEDIATE REACTION.
SO WE WANT TO ASK IS IT IN
GENERAL WHEN MACROPHAGES ARE
STIMULATED?
SO WE STIMULATE WITH DIFFERENT
TLR LIGANDS AND PREINFLAMMATORY
CYTOKINES AND IT SHOULD GIVE
MACROPHAGES LIKE
[INDISCERNIBLE], YOU CAN ENHANCE
TBLI COLSIS IN RESPONSE TO
TGF BETA IT'S ALSO TRUE IN
RESPONSE TO 848 WHICH IS A CLR7
LIGAND AND IT'S ALSO TRUE WITH
TNF ALPHA WITH IS DEMON TRAITING
THAT TGF BETA AND THAT'S A
GENERAL PHENOMENON.
SO FOR THE FLI COLSIS COLSIS IS
FINE, WHAT IS DIFFERENT, WHAT IS
THE TYPE OF FUNCTION EMPLOY SO
TO TRY TO UNDERSTAND THIS
QUESTION, WE PERFORM RNASEQ
ANALYSIS AND WE DID THAT IN
COLLABORATION WITH THE DOCTOR
AND, SO, AS CAN YOU OBSERVE
HERE, WHEN YOU GIVE TGF BETA TOO
MUCH IN RESPONSE TO LPS, IT
DRAMATICALLY REMODELED THOSE
MACROPHAGES AND IF YOU DEEPER IN
TERMS WHAT HAVE GENES CHANGE,
CAN YOU OBSERVE THERE ARE
SEVERAL JOONS BELONGING TO DOWN
REGULATION THAT ARE DOWN
REGULATED LIKE CHEMOKINES AND
FACTORS FOR EXAMPLE, WHILE IT
PROMOTES EXPRESSION OF
ANTIINFLAMMATORY FACERS LIKE
TGF BETA I ALGENERATED ACE 1.
THEY PROMOTE EXPRESSION OF
RGENERATED ACE 1 AND
DISCIPLINARY CREASE THE
EXPRESSION OF DNF ALPHA SO DEMON
TRAITING THE FACT THAT THE
RESPONSE TO THE BETA THOSE
ACTIVITIES MACROPHAGES HAVE ALSO
DECREASED INFLATION IN RESPONSE
TO THESE TGF BETA.
SO, THE NEXT QUESTION, WE WANTED
TO ADDRESS IS BY WHICH
MECHANISMS, BEST PROMOTE TBLI
COLSIS AND UNDERSTANDING A
LITTLE BIT IN THE SIGNALING
PATHWAY, AND SO IF I COME BACK
TO THE EARLY DATA I JUST SHOWED
YOU, WE NOTICE THAD 1 ENZYME
BELONGS TO GLYCOLYSIS WHICH IS
NAMED PFKL, AND THAT WAS THE
ONLY 1 THAT CAME UP AND PFKL IS
EXTREMELY IMPORTANT BECAUSE IT'S
CATALYZING THE RATE IMAGING
ENZYME THAT'S REVERSIBLE THAT
LEADS TO THE GENERATION OF
[INDISCERNIBLE] IN PHOSPHATE AND
SO ISSUES THE [INDISCERNIBLE]
PHOSPHATE WILL BE DIRECTED
TOWARDS THE LOWER STEPS OF DPLI
COLSIS, RATHER THAN BIOSYNTHESIS
AND THE FLI COLSIS COSALATION
PATHWAY.
SO WE COME FROM AT THE MRNA
LEVEL, THAT PFK WAS UPREGULATED
AND WE COME AT THAT AT THE
PROTEIN LEVEL AS WELL, AND THE
ACTIVITY WAS ALSO UPREGULATED IN
RESPONSE, DEMONSTRATING
BASICALLY THAT TGF BETA CAN
INDUCE THE EXPRESSION EMPLOY NOW
DOES IT REGULATE DPLI COLSIS,
SO, TO UNDERSTAND THAT POINT, WE
USE AN SiRNA DREKED AGAINST
PFKL EXPRESSION AND AS CAN YOU
APPRECIATE HERE, IN THE CONTROL,
TGF BETA IS ABLE TO PROMOTE MRI
COLSIS BUT IT'S NOT THE CASE
ANYMORE IN THE CASE OF SIRRNA,
AND PREDICTION WHICH IS INCREASE
UP TO TGF BETA BETA TREATMENT
AND THE IT IS COMPLETELY
ABROGATED.
SO IT CAN INCREASE IN A PFKL
MANNER.
SO HOW DOES IT GET EXPRESSION?
SO THE FIRST THING WE TRY TO
ADDRESS IS WHETHER THERE IS
INVOLVED BECAUSE AS I TOLD YOU
IN THE INTRODUCTION, IT'S THE
SIGNALING OF TGF BETA SO 3
RECEPTORS WHICH WOULD INDUCE
PHOSPHORYLATION AND ACTIVATION
THAT CAN USE GENE EXPRESSION.
BUT IF YOU--IF WE USE SMAD 3
KNOCK OUT, TO INDUCE EXPRESSION
IS ABSOLUTELY NOT CHANGED.
AND THEN, THE FLI COLSIS COLSIS
WITH THE CELLS, TGF BETA IS ABLE
TO PROMOTE FLI COLSIS COLSIS, BY
TGFBETTA IS INDEPENDENT OF SMAD
3 EMPLOY SO WE LOOK FOR A FEW OF
THEM BUT WE FIRST REALIZE THAT
mTOR IS A VERY SPECIAL PART OF
METABOLISM AND MACROPHAGES AND
THAT CAN REGULATE INFLAMMATION
AS WELL.
AND THAT mTOR HAS BEEN
REPORTED IN PAPERS TO BE ABLE TO
BE ACTIVATED BY THE TGF BETA
SIGNALING AND THE MACROPHAGES.
SO THE WAY WE CAN MEASURE
INTERACTION IS BY MEASURING
PHOSPHORYLATION OF THE DOWN
STREAM TARGETS OF S6/4 EBP1, SO
AS YOU CAN SEE HERE, WE CAN
PROMOTE PHOSPHORYLATION OF 6 AS
WELL AS FOR EBP1.
SO IF YOU LOOK AT THE ACTIVITY
BY USING RAP O MICEIN WHICH IS
AN INHIBITOR, CAN YOU ALSO GET
THE PFKL, THE EXPRESSION IS
DEPENDENT ON mTOR.
SO NOW IT'S A KINASE, IT CANNOT
DIRECTLY REGULATE GENE
EXPRESSION, IT NEEDS A
TRANSCRIPTION FACTOR BELOW, AND
SO WE AGAIN TRIED DIFFERENT
COMBINATIONS THAT CAME TO BE
NEGATIVE WUWE TARTED TO GET
INTERESTED IN [INDISCERNIBLE] SO
AGAIN, IS SUPPOSED TO BE AN
EXTREMELY REGULATOR OF THE
METABOLISM ESPECIALLY
MACROPHAGES AND SO WHAT WE COULD
NOTICE AND THAT WAS A LITTLE BIT
INTERESTING, THAT'S THE PROTEIN
LEVELS OF AFFECTED BY TGF BETA
BUT IT CAN BE PHOSPHORYLATED BY
TGF BETA IT'S SERIES POINTSINE
62 WHICH IS FORFORALATION AND
THIS IS THE mTOR AND ABROGATED
IN THE PRESENCE OF
[INDISCERNIBLE] IT CAN REPRESENT
THE TGF BETA IN THE SENSE AND IT
WE DO COMPUTATIONAL ANALYSIS,
THAT'S BINDING POTENTIAL BINDING
SITE INTOS THE PFK PROMOTER AND
IN RESPONSE TO TGF BETA SIERK
AMERICA I CAN BIND TO THE
PROMOTER AND THIS IS COMPLETELY
OBLIGATIONS BROIGATED BY RAPP A
MYOCIN, NOW IT'S RERESPONSIBLE
FOR THE CHANGES IN EXPRESSION
YES IT IS SO IF YOU USE A
INHIBITOR, YOU CAN INVERT THE
INDUCTION BY TGF BETA AND YOU
CAN ALSO BLOCK THE INDUCTION OF
GLYCOLYSIS IN THE PRESENCE OF
THIS INHIBITOR, A DEMONITRATION
THAT TGF BETA CAN REGULATE
EXPRESSION, AND mTOR DEPENDENT
MANNER.
SO I SHOWED YOU THE SIGNALING BY
WHICH IT CAN INDUCE PFK AND
GLYCOLYSIS BUT I SHOWED YOU IN
THE MEAN TIME, IT CAN DECREASE
INFLAMMATION IN THE CELLS, SO IF
THERE ARE ANY RELATIONSHIP
BETWEEN THESE CHANGES IN
GLYCOLYSIS AND INFLAMMATION,
ACTUALLY, IT SEEMS THAT THERE
ISN'T BECAUSE IF YOU USE THE
PFKL THAT I CANNOT USE BEFORE,
IF YOU REMEMBER, BUT THE ABILITY
OF TGF BETA TO DECREASE ALPHA
AND IL6 PRODUCTION, IS
ABSOLUTELY NOT AFFECTED BY THE
BLOCKADE OF PFK EXPRESSION,
DEMONSTRATE THAGOREAN YOU HAVE A
PROCESS, IN WHICH TGF BETA CAN
PROMOTE GLYCOLYSIS AND DECREASE
INFLAMMATION BUT THOSE PROCESSES
ARE COMPLETELY UNCOUPLED TO 1
ANOTHER EMPLOY SO HOW'S
INFLAMMATION REGULATED THEN, WE
CAME BACK TO SMAD 3 BECAUSE IT'S
STILL VERY IMPORTANT FOR TGF
BETA SIGNALING AND THIS TIME IF
YOU USE THE MACROPHAGES AND
INDUCE THOSE WHICH IS AN
ANTIINFLAMMATORY MOLECULE IS
COMPLETELY ABROGATED IN THE
MACROPHAGES.
SO HAVE YOU BASICALLY SHOWING
THAT TGF BETA REGULATES SMAD 3
AND YOU HAVE THIS UNCOUPLING,
TGF BETA CO PROMOTE GLYCOLYSIS,
BUT VERY NICE BUT THIS IS KNOW
VITRO.
WHAT'S HAPPENING IN VIVO.
SO TROO TOY TO UNDERSTAND WHAT
THE IS ROLE OF TGF BETA IN VIVO.
WE WENT TO THE CONTEXT OF SEPSIS
WHICH IS MIMICKED BY LPS
INVITIO, AND WE USED AN LPS
SEPSIS MODEL AND WE
INRECYCLINGED IP INTO THE MICE
AND THEN WE WORKED ON THE MODEL,
THIS IS A SEPSIS MODEL FOR REAL
INFECTION ONGOING TO THE PERO
TONIGHTIS AND SO IF YOU INJECT
LPS IN THE MICE, CAN YOU OBSERVE
THAT IN THE BLOOD OF THOSE MICE,
SO AT 6 HOURS NOT MUCH CHANGES
BUT AT 16 HOURS, WE SEE SOME
CHANGES.
AND SO, BECAUSE THERE IS A LOT
OF TGF BETA WE WANT TO BLOCK IT
SO WHAT IS HAPPENING IF YOU
BLOCK TGF BETA BY USING AN
ANTIBODY ABOUT 15 MINUTES BEFORE
INJECTION.
IF YOU BLOCK TGF BETA YOU CAN
PROTECT THE MICE OF THE MICE
WHICH IS SURPRISING BECAUSE
TGF BETA IS SUPPOSED TO BE
ANTIINFLAMMATORY, SO WE EXPECTED
THE OPPOSITE, SO JUST SHOWING
THAT PROMOTE BLOCKING IN DURING
SEPSIS COULD BE DELETERIOUS
MAYBE, SO WE INJECT THOSE INTO
THE MICE BEFORE LPS INJECTION
AND THAT TIME WE INYECT INTO THE
MICE YOU CAN ACTUALLY AND BUT IN
TERM OF INFLAMMATION, THERE IS
NO MAJOR CHANGES IF YOU LOOK AT
THE LEVELS OF TNP ALPHA, SO
DEMONSTRATING THAT IT IS DELETE
ERIOUS FOR SEPSIS DEVELOPMENT
BUT NOT BECAUSE IT CHANGES THE
INPOLICEMANNATION.
SO THEN IF IT'S NOT INFLAMMATION
COULD BE AND IF YOU REMEMBER
HERE, AND THEY ALSO HAVE
INCREASED AND WHICH IS AN
INHIBITOR TO KILL THE MICE WAS
COMPLETELY ABROGATED AND MICE
ARE PREDICTED FROM THERE.
DEMONSTRATING THAT IT IS
MEDIATED BY GLYCOLYSIS, SO HOW
DOES GLYCOLYSIS REGULATE THE
SURVIVAL OF THE MICE IN THIS
CONTEXT IN SO WE TESTED A FEW
PARAMETERS, LIKE TEMPERATURE,
WEIGHT, MOST OF THEM CAME BACK
NEGATIVE BUT WHAT WE NOTICED IS
THAT THE COAGULATION AFTER THE
TREATMENT AND THAT TGF BETA
TREATMENT THEY HAD ENHANCED
COAGULATION AND THIS IS VERY
INTERESTING BECAUSE COAGULATION
HAS BEEN DESCRIBED NOW VERY WELL
DURING SEPSIS AND COVID-19 TO
DRIVE THE DEC AND TO BE
EXTREMELY IMPORTANT FOR THE
DEATH OF THE PATIENTS.
AND DISCOAGULATION WHICH IS
ENHANCED BY TGF BETA SO
ENHANCEMENT THAT THEY NEED LESS
TIME TO CO AGULATE THE BLOOD,
THAT'S WHY IT'S INCREASED
BECAUSE IF IT'S TO THE
[INDISCERNIBLE], THE INHIBITOR
YOU CAN REVERT BACK TO THE
TGF BETA EFFECT NORMAL LEVELS,
AND WE ALSO USE HEP A RIN, TO
BLOCK THE CO AGZULATION IN THOSE
MICE IN TERMS OF SURVIVAL, WHY
TGF BETA CAN INDUCE THE DEATH OF
THE MICE, IT'S ABROGATED IF YOU
TREAT THE MICE WITH HEPARIN.
THIS IS AXIS THAT IS RESPONSIBLE
FOR THE DEATH OF MICE IN THE
SEPSIS MODEL.
SO I WANTED TO TRY TO UNDERSTAND
A LITTLE BIT MORE BY THESE
MECHANISMS TO PROMOTE
COAGULATION AND SO WE CAME BACK
TO OUR RNA SEQ DATA AND 1 GENE
BELONGS TO COAGULATION AND IT'S
F13 A, AND SO, IT'S A VERY
SIMPLELIFIED SCHEMATIC OF
COAGULATION, BUT THE INTRINSIC
AND EXTRINSIC PATHWAY WILL LEAD
TO FIEB RIN AND IT WILL HAVE TO
BE STABILIZED AND CROSS LINKED
TO FORM A PROPER CLOTS TO
LOCALIZE COAGULATION AND THIS IS
THE ROLE OF F13 TO DISCOVER THIS
CROSS LINK.
AND IT COULD COME FROM IN
MACROPHAGES IN VIVO IN THE
SEPSIS MODEL, THOSE MACROPHAGES
THEY HAVE INCREASED LEVEL OF F13
EXPRESSION, AND THIS IS ALSO
TRUE INVITIO AND THIS IS
DEPENDENT ON TBLI COLSIS BECAUSE
IF IT'S G, YOU COMPLETELY
REVERSE THE EXPRESSION, THE
INDUCTION OF 13 F A IS
COMPLETELY ABROGATED.
DEMONSTRATE THAGOREAN TGF BETA
CAN PROMOTE EXPRESSION OF F13 A
1 IN A DEPENDENT MANNER AND IT
CAN DRIVEWAY THE COAGULATION
INTO THOSE MICE.
SO FINALLY, WE WANTED TO TRY TO
UNDERSTAND WHETHER THIS PATHWAY
COULD BE ACTIVE DURING HUMAN
SEPSIS, AND SO, WE WERE ABLE TO
INTERROGATE SIPPINGLE SET RNASEQ
DATABASES FROM THE BROAD
EN--STRATEGIESITUTE THAT HAD
BEEN PUBLISHED AND SO HERE THIS
IS A SEPSIS COHORT, SO THIS IS
MACROPHAGES, MYELOID CELLS LET'S
SAY TO BE MORE PRECISE AND SO
THEY WERE ABLE TO GATHER
DIFFERENT TYPE OF PEASHTS,
HEALTHY VOLUNTEERS WITH LIFE
INFECTION, NOT LIFE THREATENING
SUCH AS UTI OR PATES THAT END UP
IN THE E. R. AND HAVE SEPSIS,
FOR THOSE PATES THAT HAVE SEPSIS
THEY ARE EXPRESSION OF FTA 1 AND
TGF BETA RECEPTOR 1, AND WE DID
A SIMILAR ATHAT WILL SIS IN A
COVID-19 COHORT BECAUSE COVID-19
PATIENTS THEY BASIC LYE DIE OF
SEPSIS AND WE COULD OBSERVE THAT
IN MYELOID CELLS MAINLY, AFTER
COVID-19, THEY HAVE EXPRESSION
OF F13A, AND DEMON TRAITING THAT
THIS DISCOAGULATION CAN BE USED
BY TGF BETA AND BE ACTIVE DURING
SEPSIS IN HUMAN.
SO TO SUMMARIZE THIS FIRST PART
OF MY TALK, WHAT I SHOWED YOU, I
HOPE I CONVINCED YOU, TGF BETA
IN MACROPHAGES IN CONTEXT OF
INFLAMMATION, CAN INDUCE
RECEPTORS AND THE ACTIVATION OF
mTOR, AND SIMI, SIMI CAN BIND
AND USE THIS EXPRESSION, WHICH
CAN PROMOTE GLYCOLYSIS, IN THE
MEAN TIME IN THE SAME CELLS BUT
IN A COMPLETELY UNCOUPLED WAY,
TASK CAN PROMOTE ACTIVATION, CAN
DECREASE INFLAMMATION, AND SO
YOU HAVE KINE OF A BALANCE WITH
INCREASED COAGULATION AND
INCREASED INFLAMMATION BUT THE
INCREASED TBLI COLSIS, IN VIVO,
OKAY, YEAH, THE INCREASED
GLYCOLYSIS, WILL LEAD TO AN
INCREASE COAGULATION THAT WOULD
LEAD TO THE TEAGHT OF THE MICE
DURING SEPSIS, SO WE JUST
PUBLISHED THIS PAPER IN THE
SIGNALING A KOWPY MONTHS AGO, SO
IF YOU'RE INTERESTED TO READ THE
FULL STORY, YOU ARE WELCOME TO
DO SO.
ASK THEN I'M GOING TO SWITCH TO
THE SECOND PART OF MY TALK,
WHICH IS ABOUT THE REGULATION OF
SMAD 3 DURING INFLAMMATION.
AND SO, JUST TO MENTION SO THIS
IS UNPUBLISHED DAILY BASIS THEA
SO PLEASE DO NOT SHARE.
SO IF I COME BACK TO THE FIRST
SLIDE I SHOWED YOU DURING THE
INTRODUCTION, TGF BETA IS
EXTREMELY WELL KNOWN TO INDUCE
PHOSPHORYLATION, VERY WELL
KNOWN, BUT WHAT IS HAPPENING IN
THE CONTEXT IS ESPECIALLY
INFLAMMATION, AND HOW SMAD 3 CAN
BE REGULATED IN THIS CONTEXT
REMAINS PRETTY MUCH LARGELY
UNKNOWN.
AND SO WE STUDIED BY DOING VERY
BASIC EXPERIMENTS IN THE LAB,
INVITRORY HARVESTED MACROPHAGES
AND WE GAVE THEM LPS TO ACTIVATE
THEM AND SO SURPRISINGLY TO US,
IF YOU GIVE LPS TO MACROPHAGES,
IT CAN ENHANCE THE
PHOSPHORYLATION OF SMAD 3 IN THE
C-TERMINAL LEVEL.
THIS HAPPENS, THIS STARTS AT 4
HOURS, PEAKS AT 6 AND COMES DOWN
AFTER 24 HOURS.
SO DEMRONITRATION THAT LPS CAN
INDUCE PHOSPHORYLATION, IN THE
C-TERMINAL DOMAIN AND SO, WHAT
WE NOTICED IS THAT THIS ONLY
HAPPENS AT 4 HOURS AND THAT
BEFORE, SO SUGGESTING THAT IT'S
NOT A DIRECT SIGNALING BUT IT'S
A SECONDARY RESPONSE AND THAT
YOU MIGHT NEED NEW PROTEIN, TO
INDUCE THIS PHOSPHORYLATION, SO
WE TRY TO INVESTIGATE THE LITTLE
BIT MECHANISM BY WHICH THEY
INDUCE PHOSPHORYLATION, SO FIRST
WE PLOKED THE TRANSLATION OF THE
PROTEIN AND WE USED THE SIGHTY
HEXAMIDHERE AND IF YOU OBSERVE
THE ABILITY FORTHE REGULATION IS
COMPLETELY PROHIBITED WITH
[INDISCERNIBLE].
YOU NEED NEW PROTEIN TO INDUCE
SMAD 3 PHOSPHORYLATION, SO WE
SCREEN FOR SOME TGF BETA MEMBERS
BY QPC R, THE THEY ARE AFFECTED
BY LPS TREATMENT AT EARLY TIME
POINT, THE TGF BETA RESOAPT OR2
IS A LITTLE BIT DECREASED AND
THE LEVEL OF TGF BETA MRNA
INCREASE AT 6 HOURS, THEN IT DID
NOT FIT WITH THE KINETIC BECAUSE
WE OBSERVE PHOSPHORYLATION AT 4
HOURS ALREADY AND WE COULD
NONAPOPTOTIC THE FIND TGF BETA
IN THE SUPERINATE EPT OF THOSE
MACROPHAGES.
SO THEN WE USE THE DEFICIENT
MACROPHAGES COMING FROM THE
MICE, AND AS YOU CAN OBSERVE
HERE IN THE WILD-TYPE
MACROPHAGE, IT CAN INDUCE VERY
WELL AND THEN WITH THE DEFESHT
CELLS, LPS CAN DO IT EVEN MORE
WHILE TGF BETA EFFECT IS
DRAMATICALLY ABROGATED SO DEMON
TRAITING THAT THE INDUCTION OF
SMAD 3 PHOSPHORYLATION BOY LPS
IS COMPLETELY INDEPENDENT OF TE
SIGNALING PATHWAY.
SO IT'S NOT TGF BETA, SO WHAT
COULD INDUCE SMAD 3
POSFORALATION IN RESPONSE TO LPS
AND WHAT WE NOTICE INDEED A VERY
EARLY TIME POINT AFTER LPS
TREATMENT, THE MACROPHAGE ARE
ACTIVATING A RECEPTOR, AND AS
WELL AS [INDISCERNIBLE] AND WE
ALSO INCREASED A PREDICTION IN
THE SUPERINATENT OF THE CELLS,
SPECIALLY TARTING AT 4 HOURS.
SO DEMONSTRATING THAT LPS CAN
PROMOTE THE INDUCTION OF THE
PATHWAY, AND THIS IS INTERESTING
BECAUSE IT'S BEEN DEMON TRAIT
INDEED ALL CELL TYPES TO BE ABLE
TO INDUCE SMAD 3
PHOSPHORYLATION, SO THAT COULD
BE THE TARGET WE'RE LOOKING FOR
AND SO WE'RE FORTUNATE TO BE
ABLE TO GET THE PLUCKS MICE FROM
DR. FLORRIA AT COLUMBIA
UNIVERSITY AND SO WE GENERATED
THE CREE MICE THAT HAS
DEFICIENCY OF ACR1 B IN
MACROPHAGES AND THEN AS CAN YOU
APPRECIATE HERE, IT'S VERY
DRAMATIC, LPS CAN INDUCE THE
PHOSPHORYLATION AND IT IS
COMPLETELY ABROGATED WHEN THE
ACTIVITY RECEPTOR IS DOWN.
SO DEMON TRAITING THAT SMAD 3
CAN INDUCE PHOSPHORYLATION IN A
DEPENDENT MANNER.
SO WE'RE TRYING TO UNDERSTAND A
BIT MORE THE SIGNALING PATHWAYS
BY WHICH IT HAPPENS, I WILL NOT
GO INTO ALL THE DETAILS ABOUT
YOU WE SHOW OF COURSE THAT IT
CAN WORK WITH THE CLR RECEPTOR,
SO IT'S NOT AFFECTED, IT'S NOT
IMPORTANT, BUT IT IS DEPENDENT
ON NYDATE, AND THE MYK KINASES,
SO NOW AGAIN, THEY ARE KINASES
SO THEY HAVE TRANSCRIPTION
FACTORS SO WE TRY TO LOOK FOR
THE TRAPGZ KRIPGZ FACTOR.
WE CHECK THEMMER, THEY'RE NOT
INVOLVED AND THEN WE TART
STUDYING TO BE IN AT THAT TIME
51st BECAUSE STAT 5 STUDIED TO
BE PHOSPHORYLATED 2 HOURS AFTER
LPS TREATMENT, SO THAT COULD FIT
OUR KENETICS, AND ALSO BECAUSE
AT THAT TIME 5 HAS SOME
POTENTIAL BINDING SITES INTO THE
ACTIVE PROMOTER AND IN RESPONSE
TO LPS, STAT 5 CAN BIND TO THE
ACTIVITY PROMOTER.
THE DEMONITRATION THAT LPS CAN
INHIBIT STAT 5 AND IF YOU USE
STAT 5 MACROPHAGES, THE ABILITY
OF LPS TO INDUCE ACTIVE
EXPRESSION IS DRAMATICALLY
REDUCED AND THE ABILITY TO
INDUCE SMAD 3 PHOSPHORYLATION IS
PRIMITIVE FOR REUSE, THIS CAN
INCREASE EXPRESSION AND SMAD
THROW PHOSPHORYLATION, FOR STAT
5 DEPENDENT PATHWAY.
SO, THIS IS THE MOLECULAR
MECHANISM, WHAT IS THE FUNCTION
OF THIS SMAD 3 PATHWAY.
SO IF YOU KNOCK DOWN THE
MACROPHAGES, YOU HAVE INCREASED
PRODUCTION OF IL6, VERY DRAMATIC
AND LARGELY DECREASED EXPRESSION
OF TGP BETA EYE WHICH IS AN
ANTIINFLAMMA ARE TOY FACTOR.
IF YOU DO THE SAME THING WITH
GUT MACROPHAGES YOU HAVE THE
SAME THING, SO IS THE WHYED THAT
YOU CAN HAVE NATURAL BRICK TO
INFLATION THAT IS PUT IN PLACE
BY THE CELLS TO ARK VOID THE
OVER ACTIVATED, THEY WOULD START
TO PRODUCE ACTIVITY A THAT
PRODUCE THE PHOSPHORYLATION BUT
WILL DOWN REGULATE THE
PHOSPHORYLATION, SO WE WANTED TO
UNDERSTAND A LITTLE BIT DEEPER
WHAT COULD BE THE MECHANISM BY
WHICH THESE CHANGE VS BEEN AND
SO WE PERFORM SOME RNASEQ ANINAL
SIS IN THE SMAD 3 MACROPHAGES
WITH THE LPS SO IT'S NOT
SURPRISING I SHOWED NUCLEOTIDES
RESPONSE TO LPS, THEY HAVE
INCREASED YEENS BELONGS TO
INFLAMMATION, IL6, CD64, AND SO
ON AND SO FORTH AND THEY HAVE
JEERPS RELATED TO ANTIINFLMATORY
FACTORS.
SO WE CAN SEE AT THE PATHWAY
LEVEL, BUT WHAT'S VERY
INTERESTING THAT IF YOU LOOK AT
THE DOWN REGUTED PATHWAY THIS
TIME, NOT UPREGULATED YOU CAN
OBSERVE THAT MANY OF THIS
PATHWAY, MOST OF THEM BELONG TO
METABOLISM.
ESPECIALLY METABOLISM OF LIPIDS,
CARBOHYDRATES, METABOLISM AND SO
ON AND SO FORTH.
SO SUGGESTING THAT THERE COULD
BE LIKE A METABOLIC REGULATION
IN THE CELLS, AND WAWE COULD
OBSERVE NOTABLY, THE
MITOCHONDRIA METABOLISM COULD BE
THOSE METABOLISM, GENERATION,
FUNCTION, THEY ARE ALL
NONREGULATED, AND BY DOING FAST
STAINING FROM MIGHT O TRACKER
WHICH IS A WAY TO STAIN
MITOCHONDRIA, WE CAN PRESERVE
THAT AND WE HAVE DECREASED
MITOCHONDRIA LEVELS, AND THIS IS
EVEN MORE TRUE IN PRESENCE OF
MACROPHAGES AND WE HAVE THE SAME
RESULTS IN PRESENCE OF ACGR 1
BEING THE MACROPHAGE,
DEMONSTRATING THAT THE PATHWAY
SUPPORT THE BIODPEN SIS, THAT
COULD BE VERY IMPORTANT BECAUSE
MITOCHONDRIA IS A HUB TO CONTROL
METABOLISM, INFLAMMATION,
MACROPHAGES FUNCTION AND SO, WE
TRY TO GO A BIT FURTHER AND WHAT
YOU COULD OBSERVE IS THAT THOSE
CHANGES IN MITOCHONDRIA LEVELS
WILLING TO DECREASE LEVELS OF
ATP, WE HAVE LESS ATP INTO THE
CELLS AND DEMON TRAITING THIS
ACTIVITY A INDUCTION PATHWAY CAN
LEAD TO MITOCHONDRIA BIOGENESIS
AND GENERATION OF ATP, SO I
REALLY WANT TO EMPHASIZE THAT
THIS NOT LIKE EXTREME LEVELS OF
ATP, IT'S TRY TO MAINTAIN HOMEIO
STASE EXPIS HAVE NORMAL LEVELS
OF ATISSUING P WHICH NORMAL IN
THE CELLS TO FUNCTION.
SO, YOU HAVE DECREASED ATP, SO
DO THOSE LEVELS SUSTAIN
INFLAMMATION, SO WE TRY TO SEN
ILLEGALSEN BACK TO THE CELLS SO
WE USE THE ATP TO INDUCE
ACTIVATION OF COURSE, AND SO,
WHAT YOU COULD OBSERVE IN THE
MACROPHAGES IN PRESENCE OF LPS,
AND LEVELS OF ATP, CAN YOU HAVE
THE LEVEL OF EXPRESSION WHICH IS
AN ANTIINFLAMMATORY FACTOR
AGAIN, THIS IS TRUE IN
[INDISCERNIBLE] MACROPHAGES.
SO DEMONSTRATING AGAIN, HAVE YOU
THE PATHWAY, THIS ACTIVITY IN
THE PATHWAY THAT CAN LEAD TO
MITOCHONDRIA BIOGENESIS, NO MORE
PREDICTION THAT CAN PROMOTE THE
EXPANSION OF ANTIINFLAMMATORY
EFFECTS.
ATP CAN CONVERGE INTO ADP AND
AMP BY C13 AND 39, AND THEN ADEN
O SEEN CAN INDUCE RECEPTORS AND
E DUCE SIGNALING WITH KREB,
THAT'S OUR ANTIFLAMMATORY.
IS IT THE CASE IN OUR SETTINGS,
SO WE TRY TO INTERROGATE THAT,
AND SO THE FIRST THING WE
OBSERVE IS THAT LPS CAN
DRAMATICALLY ENHANCE THE
ACTIVATION OF CD73 AND THIS IS
DEPENDENT ON SMAD 3, SO THE
LEVELS ARE LARGELY DECREASED,
AND ON THE ACT VIN A EXPRESSION,
SO IT COMES BACK DOWN IN
[INDISCERNIBLE] KNOCK OUT
MACROPHAGES, SO DEMONITRATION
THAT THIS CAN PROMOTE THE
EXPRESSION OF CD7.
AND SO NOW WE HAVE
FUNCTIONALITY.
SO IF YOU RESORT BACK THE LEVELS
OF ATP AS I SHOWED YOU, YOU CAN
PROMOTE THIS AND IF YOU PLOK IT,
DOWN STREAM TRANSCRIPTION FACTOR
OF KREB, CAN YOU ABNORMALITIES
BRO IMAIT THE ATP EFFECT, THIS
IS TRUE IN THE GUT MACROPHAGES
AND THIS IS ALSO TRUE IN THE--WE
CAN BIND TO BELONGING TO
MITOCHONDRIA FUNCTION WHICH CAN
PROMOTE MITOCHONDRIA BIOGENESIS,
THE SYNTHASE OF ATP AT NORMAL
HOMEOSTATIC LEVELS WHICH CAN BE
CONVERTED JUST IN CD73 THAT CAN
CIRCLE BACK INTO THE CELLS,
ACTIVATE KREB AND IN THE WITH
SMAD 3 CAN WORK ON THE FOR
EXAMPLE.
SO THIS IS THE MECHANISMS
INVITRO, NOW WHAT IS HAPPENING
IN VIVO IS THE PATHWAY IS STILL
ACTIVE, AS IT PLAYS, SO WE CAME
BACK TO OUR LPS, WE WE INYEBED
LPS INTO THE MICE, WE ISOLATED
THE MACROPHAGES 6 HOURS AFTER
THE INJECTION OF THE DISEASE AND
WE LOOKEDDA THE THIS PATHWAY AND
IT WAS ACTIVE SO AGAIN, I'M
GOING TO SHOW YOU THE LPS DATA
BUT WE HAVE SAME DATA WITH THE
SEPSIS MODEL WHERE WE HAVE REAL
INFECTION ONGOING.
AND SO IN THOSE MICE IN INCREASE
THE PHOSPHORYLATION IN RESPONSE
TO LPS, IN THE SEPSIS MODEL, SO
IT'S PHOSPHORYLATED, AND THEN,
THIS IS HAPPENING MANICALLY ON
MACROPHAGES, SO WOE HAVE
INCREASED FOCUSED ONS FORALATION
OF MACROPHAGES BUT NOT THAT MUCH
INTO THE NUTRIFILL OR A LITTLE
BIT WITH THE B-EABT CELLS AND
THEN, IS IT DEPENDENT ON TGF
BETA OR ACT VIN, WHAT WE COULD
OBSERVE AS CAN YOU OBSERVE IN
THIS GRAPH, SO IN WILD-TYPE,
MICE, THE MACROPAGES DO NOT
REALLY HAVE ANY SMAD 3
PHOSPHORYLATION WHICH MAKES
SENSE IN RESPONSE OF LPS, THIS
IS PLURIBU CURVE, INCREASED BUT
IN THE TGF BETA MICE, THEY CAN
STILL INCREASE THE
PHOSPHORYLATION AND THERE IS
ABSOLUTELY NO DIFFERENCE.
SO DEMON TRAITING THAT LPS CAN
WORK IN VIVO DURING SEPSIS AND
SO IS IT DEPENDENT ON THE
ACTIVITY LEVEL AFTER LPS
INJECTION INTO THE MICE 6 HOURS
AFTER INFECTION.
DEMON TRAITING LPS CAN REDUCE
SMAD 3 PATHWAY, SO WHAT IS THE
ROLE OF THIS PATHWAY IN TERMS OF
INFLAMMATION AND SURVIVAL, IF
YOU INDUCE SEPSIS IN THIS CREE
MY, HAVE YOU DECREASED SURVIVAL,
AND THIS IS LINKED TO INCREASED
LEVELS OF INFLAMMATION.
CERTAINLY--CERTAINLY
SPECIALENTIAL LOAMACYY -I WELL,
6, YOU HAVE SO, A QUESTION THAT
WE WANTED TO UNDERSTAND IS OKAY,
IN IS TRUE IN THE CONTEXT OF
LPS, WHICH IS A BACTERIAL LIGAND
AND A TNF LIGAND BUT WHAT ABOUT
OTHER TOLL-LIKE RECEPTORS THAT
ARE ABLE TO MEDIATE THIS EFFECT,
WHAT ABOUT THE LIAISON GAPPEDS,
AND ALSO WHAT ABOUT ENDOSTUDIES
OF MULTIPLE ENDOCRINAL RECEPTORS
SO TO TRY TO ANSWER THOSE
QUESTIONS, WE FIRST USED THE
EPROTEIN FROM [INDISCERNIBLE]
THAT HAS BEEN DEMONSTRATED HERE,
AND BE ABLE TO INFLAMMATION, AND
WE ALSO USE TLR7 LIGAND WHICH IS
MORE IN THE CONTEXT OF
AUTOIMMUNITY.
AND SO, SHOULD SHOULD GIVE
PROTEIN TO MACROPHAGES INVITRO,
YOU CAN INCREASE THE EXPRESSION
OF ACTIN A AND YOU CAN ALSO
INCREASE THE PHOSPHOROUS 3
REGULATION.
THE SAME THING HAPPENS IN
RESPONSE TO, YOU HAVE INCREASED
IN ACTIVITY A, AND HAVE YOU
INCREASED SMAD 3 PHOSPHORYLATION
WHICH IS COMPLETELY DEPENDENT ON
AND DEMONSTRATING THOSE 2
LIGANDS CAN INDUCE ACTIVE A AND
PHOSPHORYLATION AND THAT IT'S A
GENERAL PHENOMENON THAT SEEMS TO
BE OCCURRING.
SO WE DID QUITE A BIT OF WORK ON
THE PROTEIN MODEL IN VIVO AND
INVITRO.
I WILL NOT SHOW THOSE DATA TODAY
AND I WILL FOCUS MORE ON THE
IMMUNITY PART AND THE
[INDISCERNIBLE] AND SO CAN THE
WHAD CAN BE APPLIED ON THE BACK
SKIN OF THE MICE FOR 6
CONSECUTIVE DAYS AND THAT CAN
INDUCE [INDISCERNIBLE] DISEASE
EMPLOY SO WE DID THE SAME THING,
WE IMAGED FOR INFLAMMATION, WE
DID IT FOR 6 HOURS AND WE USE IT
FOR THE ACTIVATION OF THIS
[INDISCERNIBLE] PATHWAY.
SO IF YOU APPLY [INDISCERNIBLE]
ON THE MICE AFTER THE FIRST
APPLICATION, HAVE YOU INCRACIED
ACTIVE EXPRESSION, AND YOU HAVE
INCREASED LEVEL OF SMAD 3
PHOSPHORYLATION AS CAN YOU
APPRECIATE HERE, DEMONSTRATE
THANKSGIVING PATHWAY IS INDUCED.
SO NOW WHAT IS THE FUNCTION OF
THIS PATHWAY, IF WE USE
[INDISCERNIBLE] CREE MICE AND
APPLY ON THE BACK OF THE MICE,
SO, THOSE MICE HAVE A HIGHER
[INDISCERNIBLE] WHICH MEANS THEY
ARE MORE SICK, AND WE COULD
OBSERVE THAT WE HAVE MORE
MACROPHAGES IN THE SKIN AND
THOSE MACROPHAGES PRODUCE MORE
IL6 SO THEY'RE MORE
INFLAMMATORY.
AND NOW THEY BECOME MORE
INFLAMMATORY, THEY WILL DRIVE
THE GAMMA T-CELLS AND THEY WILL
STOP PRODUCING IL15
INTERFERON-GAMMA ANDILE L22.
AND DEMON TRAIT THANKSGIVING
ACTIVE A AS THE PATHWAY CAN
CONTROL THE MACROPHAGES BEHAVIOR
THAT CAN BE PROINFLAMMATORY AND
CONTROL THE ACTIVATION OF THE
GAMMA T-CELLS DURING SORTING
SIGNALS AYA SIS.
AND WEEE INYEBED ACTIVEIN A INTO
THE INTRA DERMALLY INTO THE SKIN
OF THE MICE AND SO IF YOU INJECT
ACTIVEIN INTO THE MICE, THIS IS
DEPENDENT ON SMAD 3, SO THE
EFFECTIVE IT IN THERE IS
[INDISCERNIBLE].
AND IN THOSE TREATED MICE WE
OBSERVED A DECREASED LEVEL OF
MACROPHAGES AND THIS IS
DEPENDENT ON SMAD 3, AND OF
COURSE WE HAVE DECREASE
EXPRESSION OF IL17 AND GAMMA AND
THIS IS AGAIN COMPLETELY
REVERTED TO MORE LEVELS IN SMAD
3 KNOCK OUT DEFESHT MICE.
SO DEMONSTRATING THIS ACTIVE A
PATHWAY CAN CONTROL INDUCED THE
[INDISCERNIBLE] AND BE A BREAK
FOR INPOLICEMANNATION AND IPT
GREATERUNITY AND ACCEPTIS.
SO FINALLY IS THIS PATHWAY
ACTIVE IN HUMAN IN SO WE HOPE TO
HAVE MORE DATA ON THIS AS WELL.
SO WE CAME BACK TO THE DATA FROM
THE FIRST PROJECT AND WE
OBSERVED THIS PATHWAY TO BE
ACTIVE IF PATIENTS THAT HAVE
SEPSIS, DEVELOP SEPSIS,
INCREASED LEVEL OF ACTIVITY A,
3, RECEPTOR IN STAT 5, IN
MACROPHAGES AND IN COVID-19, YOU
ALSO HAVE INCREASED EXPRESSION
OF THOSE YEENS IN COVID-19
PATIENTS.
DEMONSTRATING THIS ACTIVITY OF
THE PATHWAY COULD ALSO BE ACTIVE
IN PATIENT'S THAT DEVELOP SEPSIS
AND COVID-19.
SO TO CONCLUDE THIS PART OF THE
TALK, WHAT I SHOWED YOU IF
THAT'S LPS, PROTEIN, AND
[INDISCERNIBLE], SIGNALS WITH
THEIR TLRs AND CAN INDUCE STAT
5 ACTIVATION, SO I DID NOT SHOW
IT TO YOU FOR A PROTEIN, BUT
BASICALLY THEY HAVE THE SAME
THING IN PATHWAY THROUGH MYD88
STAT PATHWAY AND AT THAT TIME 5,
AND THEN STAT 5 CAN BIND TO THE
PROMOTER OF ACTIVEIN A, CAN
PRODUCE THE PRODUCTION OF
ACTIVIN A, BUT WE BELIEVE IT'S
HAPPENING IN OTHER CONTEXT, IT
CAN INDUCE PHOSPHORYLATION, SMAD
3 CAN BIND TO SELF-PROMOTERS AND
REGULATE INFLAMMATION WITH
METABOLISM DURING SEPSIS, SUCH
AS IN COVID-19.
SO WHAT ARE THE FUTURE
DIRECTIONS OF THIS WORK?
SO 1 OF THE DIRECTION WHICH IS
RELATED TO THE FIRST PROJECT IS
THAT WE STUDIED THE ROLE OF
MACROPHAGES IN THE ROLE OF
RESPONSE TO TGF BETA AND
INFECTION ANDS VERY HIGH LEVELS
OF INFLAMMATION BUT WHAT
HAPPENING IF YOU HAVE HIGH
LEVELS OF TGF BETA AND LOW GRADE
OF INFLAMMATION, ESPECIALLY THE
RULE OF TUMOR MICROENVIRONMENT.
ONE POINT WHICH I THINK IS
EXTREMELY IMPORTANT, COULD WE
TARGET THE TGF BETAACIS DURING
SEPSIS IN HUMANS, SO OF COURSE,
THERE IS SOME WORK REQUIRED THAT
WE--THAT WOULD REQUIRE TO
DEVELOP SOME ANTIBODIES AND
INHIBITORS, BUT WE HAVE ALL
TOOLS AVAILABLE AND INHIBITORS
AND ACTIVATION THAT ARE
AVAILABLE ALREADY AND THERE IS
NOW AND DRIVE THIS, WE WILL HEAR
THE GOOD COAGULATION, AND IT'S
BEEN TARGETED WE WOULD LIKE TO
COLLABORATE AND BE MORE THAN
HAPPY.
AND THEN IA LITTLE BIT ABOUT THE
PROGECS I SHOW YOU DURING
SEPSIS, ACTIN A IS A GOOD GUY
BUT WE WANT TO TRY TO UNDERSTAND
WHAT CONTEXT THIS IS MORE
IMPORTANT THAN TGF BETA, WE
BELIEVE IN THE EXAMPLE OF THE
TUMOR MICROENVIRONMENT, IT COULD
BE MORE IMPORTANT THAN ACTIN A
BUT IT DEPENDS ON ALSO THE
ACTIVATION OF THOSE PROTEINS AND
MANY DIFFERENT FACTORS AND HOW
ARE THEY DIFREPRESENTIALLY
REGULATE, SPECIALLY WE FOCUS ON
SMAD 3 ACTIVATE BOOED I BOTH BUT
THEY HAVE DIFFERENT SIGNALING
PATHWAY, LIKE mTOR AND
SIGNALING FOR TGF BETA SO WHICH
PATHWAY IS MORE IMPORTANT AND
FINALLY WE COULD USE ACT VIN A
TO FIGHT INFECTIONS IN HUMAN AND
TO DECREASE INFLAMMATION AND
INDUCE A RETURN TO HOMEOSTASIS.
SO THE LAST SLIDE BUT OF COURSE
ALSO THE MOST IMPORTANT.
SO I WILL LIKE TO THANK
EVERYBODY WHO HELPED ME FOR THE
STUDIES, MY MENTOR DR. CHEN, AN
AMAZING MENTOR THESE PAST 5
YEARS, AS WELL AS EVERYBODY IN
THE LAB WHO IN THE PICTURE WHO
HELPED ME PERFORM SO MANY SPRPTS
AS WELL AS [INDISCERNIBLE], I
WOULD ALSO LIKE TO THANK ALL OUR
COLLABORATORS ESPECIALLY INSIDE
NIH, FOR THE RNASEQ ANALYSIS AND
DR. CHEN YRK AO, AND
[INDISCERNIBLE] FOR THE HELP IN
ATTACK SEQ AND CHIP ANALYSIS,
DR. LARRION AND DR. BEWLY, AND
DR. SU FOR PROVIDING THE ACR1 B
FLUX MICE AND I WOULD LIKE TO
THANK YOU FOR YOUR ATTENTION AND
IF YOU HAVE ANY QUESTIONS, THE
FLOOR IS YOURS.
[ APPLAUSE ]
>> THANK YOU EMPLOY WE HAVE
QUESTIONS COMING ALREADY SO I
WILL HOLD MINE BACK.
>> INTERESTING TALK, I HAVE A
FEW QUESTIONS REGARDING THE
FIRST PART EMPLOY ANDIME SO CURE
QUOWS WHAT IS GOING TO HAPPEN IF
YOU TAKE THE MACROPHAGE THAT THE
TGF BETA DEFESHT RECEPTOR AND
YOU STIMULATE THEM WITHOUT THE
[INDISCERNIBLE], THEY WILL
REGULATE GLYCOLYSIS OR WHAT DO
THEY DO?
>> YES, SO WE DIDN'T GO THAT
MUCH FURTHER ON THAT, BUT IT
LOOKS LIKE, YES.
AND BAGGED ON THAT, MY QUESTION
IS WHAT DO THOSE MACROPHAGE DO
BECAUSE YOU REGULATE DPLI COLSIS
AND THEY WILL PRODUCE
INTERLEUKIN 1 BETA BY ACTIVATING
INFLAMMASOME OR WHAT DO THEY
PROTIEWS?
>> WE DIDN'T SEE THAT MUCH IL1
BETA CHANGES ESPECIALLY IN THE
INPOLICEMANNASOME ACTIVATION,
AND PROINFLAMMATORYITE O KINES,
AND HAVE YOU CHECKED BY CHANCE
WHEN YOU KNOCK DOWN, I DON'T
KNOW IF YOU DO NOT TGFPAYS BETA
TO THE MACROPHAGE OR SOMETHING,
IN THE FAIG O SITTIC CAPACITY OF
THE CHANGES WHEN YOUACIVATE THEM
IN THE PRESENCE OF THE TGF BETA
OR NOT BECAUSE YOU DON'T KNOW
WHEN YOU PUT THE TGF BETA INTO
MACROPHAGES THEY ARE BECOMING
mTOR MACROPHAGES NSO TO ANSWER,
THE FIRST THING WE DID NOT TRY
TO DO THAT BUT IT'S A VERY GOOD
POINT.
WE DID NOT LOOK AT PHAGOCYTOSIS,
SO WE CAN TELL YOU THERE ARE NO
DIFFERENCE IN THE LEVEL IN THE
BLOOD OF THE MICE BUT LOOKING AT
PHAGOCYTOSIS WOULD BE VERY
IMPORTANT.
I FORGET WHAT I WANTED TO SAY
FOR A SECOND PART.
WHAT WAS THE END OF THE
QUESTION.
>> THE FAIG O SITTIC CAPACITY
DIFFERENT BETWEEN THE USE
STIMULATOR [INDISCERNIBLE]
TGF BETA?
>> YEAH, SO WE DID NOT LOOK AT
THAT, WE KNOW THAT IN CONTEXT OF
THE SIGNIFYITOSEIS THAT WE DID
BUT NOT IN CONTEXT, BUT IN VIVO
THERE'S NO DIFFERENCE IN LEVELS
OF INFECTION OF THOSE INTO THE
BLOOD OF THE MICE.
IN, OKAY, THANK YOU NYOU'RE
WELCOME.
NI HAVE A QUESTION ABOUT THE
FITTER PART OFIOURE TALK AS
WELL, VERY NICE TALK, SO,
COAGULATION, FACTOR 13 IS THE
END STAGE WOULD ACTUALLY TABLIZE
THE CLOT, AND LPS I'M SURE
INDUCES CLOTTING FACTORS FOR
EXAMPLE, INDUCES ALL RECOLLECT
FACTORS AND COMPLEMENT
ACTIVATION, ET CETERA, DO YOU
THINK TGF BETAS THOSE LESMS, 7,
8, 9, WHATEVER YOU WANT TO CALL
THEM, THE BLOOD CLOTTING IS GOOD
AND HOW IS THIS STACK FACTOR THE
MOST IMPORTANT AS OPPOSE TO
CHANGING LEVELS INCREASES OF
OTHER BLOOD CLOTTING LEVELS
AFTER SEPSIS.
>> THAT'S A VERY GOOD QUESTION
ACTUALLY, UNFORTUNATELY WE WERE
NOT ABLE TO GO FURTHER INTO THE
COAGULATION PART SO WE WOULD
HAVE LOVED TO HAVE THOSE MYSELF
AND WITH CREE AND PROVE THAT F13
IS THE MOST IMPORTANT BUT WE
WERE NOT ABLE TO GO THAT FAR SO
YEAH, THE THINK OF THAT BECAUSE
WE INJECTED TGF BETA AND BLOCKED
TGF BETA WE BELIEVE THAT THERE
MIGHT BE OTHER CELLS AFFECTED BY
TGF BETA LIKE [INDISCERNIBLE] OR
OTHERS THAT PLAY A ROLE IN
COAGULATION NDID YOU SEE
CLOTTING IN THE ANIMALS?
WHERE DOES IT OCCUR?
IN SEPSIS WHERE DO YOU SEE A LOT
IN.
>> IN THE BLOOD IF YOU TAKE THE
BLOOD, THE COAGULATION IS MUCH
FASTER.
>> OKAY, THANK YOU NYOU'RE
WELCOME.
>> WE CAN LET TRAINEES GO FIRST,
RIGHT?
, TRAINEES FIRST.
>> SO, WHEN YOU HAVE CYTOKINE
STORM TOXIC SHOCK CONNECK WIDE
SEPSIS FOR EXAMPLE OR OTHER
SITUATIONS THERE'S OFTEN IL6
INVOLVED AND TREATMENT WITH
[INDISCERNIBLE] WHICH IS AN IL6
INHIBITOR CAN BE EFFECTIVE AND
WE KNOW THAT TGF BETA CAN
COLLABORATE WITH IL6 TO INDUCE
TH15 AND ILPANE PRODUCTION, SO,
HOW DOES ALL THAT FIT INTO YOUR
SEPSIS MODEL WHERE I DON'T
RECALL YOUR MENTIONING EITHER
IL6 OR IL15 EXCEPT LATER IN THE
CONTEXT OF PSORIASIS, YOU
MENTIONED IL17.
SMRKS GOOD QUESTION, SO THE
ACTUALLY THE LEVELS OF HAVE NOT
CHANGED IN MACROPHAGES OR IN THE
BLOOD OF THOSE MICE.
THEY DON'T SEEM TO BE AFFECTED,
WE COULD NOT SEE ANY TH17
GENERATION BUT IT'S PRETTY EARLY
ON.
AND THEY'RE LIKE 18 HOURS AND
THEN, LIKE EVERYBODY THAT HAS TO
DIE, DIES BY 48 HOURS, SO IT'S
PROBABLY STILL A LITTLE BIT
EARLY TO SEE DRAMATIC TH17
RESPONSE, SO PROBABLY WHY WE
DIDN'T SEE IT NOW IN LONGER
INFECS, YEAH, PROBABLY I THINK
IT COULD PLAY A ROLE, YEAH,
DEFINITELY, YEAH.
>> OKAY, THANK YOU.
>> THANK YOU, THAT WAS A NICE
TALK.
I HAVE A QUESTION ABOUT YOUR
FIRST PART MORE CONCEPT WISE
BECAUSE YOU MENTIONED ABOUT
SEPSIS IN THE COVID INFECTION,
DO YOU THINK IT'S LIKE HOW CAN I
SAY THE REAL--I DON'T FINISH
IT'S THE RIGHT WORD BUT REAL
SETTING OF TGF BETA AND THOSE
LIKE SIGNALING PATHWAYS INVOLVED
TO SAY IN A SETTING OF COVID-19
SITUATION?
DO YOU THINK THIS CYTOKINE STORM
MAY BE RELATED TO THOSE TGF BETA
SECRETION OR?
I JUST WANT TO SEE YOUR--
>> SO YOU'RE ASKING WHETHER YOU
HAVE CYTOKINE STORM IN RESPONSE
TO TGF BETA?
>> RIGHT, SO IT LEADS TO THE
SEPSIS.
>> EXACTLY.
SO THE FIRST THING THAT WE
INVITRO IF YOU GET THE PROTEIN
FROM COVID-19, HAVE YOU THE SAME
THING, CAN YOU ACTIVATE TBLI
COLSIS BUT YOU CAN DECREASE
INFLAMMATION IN RESPONSE TO
TGF BETA SO THAT ANSWERED PART
OF YOUR QUESTION, THE
SECURITIZATION. THING, SO WE
LOOK AT INFLAMMATION AFTER ALL
THE WORKING ANS THERE IS NO MORE
SCIENCE OF INNAMES--NAMESSATION,
THE BRAIN, LUNGS, THE KIDNEY,
THE LIVER, WHEREVER WE LOOKED,
WE LOOKED AT CYTOKINES, THE
MACROPHAGES ARE STILL
ANTIINFLAMMATORY IN RESPONSE TO
TGF BETA SO WE REALLY BELIEVE
THAT IT'S NOT RELATED TO
CYTOKINE STORM BUT TISSUE
RELEVANCE AND DISCOAGULATION IS
DIFFERENT AND DRIVING THE MICE
BUT THE CYTOKINE STORM.
>> THANK YOU.
>> THANKS THAT WAS REALLY GREAT.
REALLY ENJOYED IT, SO I HAVE A
QUESTION ABOUT MRI COLSIS DATA
WHICH I THOUGHT WAS REALLY
CONVIPSING BUT I WANT TO THROW
OUT ANOTHER POSSIBILITY WHICH IS
JUST THAT METABOLISM IN GENERAL
IS NEEDED SO HAVE YOU TRIED
TREATING YOUR MICE FOR EXAMPLE
WITH A FLUTA MIN OLYMPIC SIS
INHIBITOR LIKE WITH DON OR
BPTSAND DO YOU GET THE SAME OR
DO YOU GET TBLI COLSIS.
>> GOOD QUESTION AGAIN, WE DID
NOT TRY TO BLOCK IT, WE TRY TO
BLOCK THE TCR SICK SEAL AND THEN
WE HAVE THE SAME EFFECT BUT WE
DID NOT GET THE DPLIEWTA MINE
BUT--
RIGHT.
BUT IF WHEN YOU TOUCH THE TCA
TIKELE YOU'RE GETTING THE SAME
EFFECT.
IT MAY BE THAT IT'S METABOLISM
IN GENERAL THAT'S REQUIRED AND
IT DOESN'T HAVE TO BE DPLI
COLSIS PER SE, BUT IT CAN BE ALL
OF THESE THINGS FITTING IN,.
>> I MEAN I CAN GIVE YOU
PROTOCOLS FOR DON TREATMENT FOR
YOUR MICE IF YOU'RE INTERESTED.
>> THAT WOULD BE NICE, YES.
THAT COULD BE POSSIBLE.
THAT COULD BE POSSIBLE, THAT
THEY JUST NEED THE ENERGY,
ENOUGH ENERGY TO FEED THIS
PATHWAY AND DISCOAGULATION,
THAT'S A POSSIBILITY.
BUT WE DID NOT CHECK ON DPLIEWTA
MINE IN PARTICULAR.
>> IT WOULD BE INTERESTING ALSO
TO SEE IF YOU HAVE SWITCHES
BETWEEN SOME OF THE METABOLIC
PATHWAYS AND THEN JUST REALLY
QUICKLY BECAUSE YOU'RE ATP DATA
WAS REALLY INTERESTING, DO YOU
KNOW IF IF YOU HAVE AN
UPREGULATION OF ANY OF THE
NUCLEOTIDE TRANSPORTERS IF ARE
YOURSELF, LIKE ANTI1 OR 2,
BECAUSE IT HAS TO BE GETTING
INTO YOURSELF SOMEHOW SO HOW IS
THAT AFFECTING BECAUSE IT SEEMS
LIKE YOU HAVE A FEEDBACK
MECHANISM.
>> SO DISCIPLINARY NOT LOOK AT
THOSE ONCE BUT I WILL DEFINITELY
DO IT BECAUSE THAT COULD BE A
MECHANISM.
THAT COULD BE A MECHANISM.
SO WHAT I CAN SAY THAT FOR
EXAMPLE, THE LEVEL OF NTP 1, IS
ACTUALLY UPREGULATED IN KNOCKOUT
MICE SO IT'S SOMEHOW SPECIFIC ON
CD73, BUT WE DID NOT LOOK AT
OTHER TRANSPORTERS.
RIGHT, BECAUSE IT CAN BE A
CD973AXIS BUT IT CAN ALSO BE A
TRANSPORTER ITSELF.
CORRECT.
THANK YOU VERY MUCH.
GREAT.
[ APPLAUSE ]