MOLECULAR BASIS OF INFLAMMATION AND INSULIN RESISTANCE IN OBESITY ... · resistÊncia À insulina resistÊncia À insulina

MOLECULAR BASIS OF INFLAMMATION

AND INSULIN RESISTANCE IN OBESITY:

CONTRIBUTION OF THE GUT MICROBIOTA.

Mario J A Saad

FCM-UNICAMP

INSULIN

RESISTANCE

Hypothalamus

GIT Muscle

SpleenAdiposeBone Marrow

INSULIN RESISTANCE IN DIO

Liver

IRS1/2

PO4

Ras/Raf

MEK

ERK

PI 3

kinase

insulin

PO4PP

PIP2 PIP3

PP

PPP

PDK

AKTPO4

PO4

PO4

FoxO1

GSK3

aPKC

Glucose Uptake

Gliconeogenesis

Gene Transcription

Protein Synthesis

Cell Growth,

Differentiation

Glycogen

Synthesis

PO4

PO4

PO4

SOS

Grb2

Shc

INSULIN SIGNALING

0

100

150

200

250

300

350

C

*

(nm

ol/g

/min

)

50

HFD

MOLECULAR MECHANISMS OF INSULIN

RESISTANCE IN HIGH FAT DIET – 3 DAYS

Glucose uptake

0

20

40

60

80

100

120

C

*

(%)

pIRS-1

0

20

40

60

80

100

120

C

*

(%)

pAKT

MUSCLE

HFD HFD

MOLECULAR MECHANISMS OF INSULIN RESISTANCE –

MODULATORS OF INSULIN SIGNALING

PI 3

Kinase

S - Sa

S-S

b

ATP

insulin

S-S

PPPO4

IKKb

JNK

IL6 – TNFa – FFA - AII

HIGH FAT DIET

PO4

PO4

PO4 PO4

PO4

P

P

INSULIN

RESISTANCE

IRS1/2

PO4

LPS

0

0,5

1,0

2,5

1,5

2,0

(EU

/mL)

HFD 3days

*

LPS CIRCULATING LEVELS IN MICE ON HFD - 3 DAYS

Control



PI 3

Kinase

S - Sa

S-S

b

ATP

insulin

S-S

PPPO4

IKKb

JNK

HIGH FAT DIET

LPS

PO4

PO4

PO4 PO4

PO4

P

P

INSULIN

RESISTANCE

IRS1/2

PO4

Duodenum101-103 cfu/ml

Stomach101-103 cfu/ml

Jejunum104-107 cfu/ml

Cólon1011-1012 cfu/ml

MICROBIOTA IN THE GASTROINTESTINAL TRACT

Another bacteria

Firmicutes

Firmicutes

Bacteroidetes

Bacteroidetes

LEAN

Another bacteria

OBESE

PROPORTION OF FIRMICUTES AND BACTEROIDETES IN

LEAN AND OBESE MICE

PHENOTYPE

GENOTYPE MICROBIOTA ENVIROMENT

PHENOTYPE

GENOTYPE MICROBIOTA ENVIROMENT

MECHANISMS BY WHICH MICROBIOTA CAN

INFLUENCE OBESITY

LIVER


INFLUENCE OBESITY

ADIPOSE HYPOTHAL. MUSCLE

pAKT

0

20

40

100

60

80

- ++ -

GF

(%)

CONTROL

*

LIVER

EFFECT OF HIGH FAT DIET -3DAYS ON INSULIN SIGNALING

IN LIVER GERM-FREE AND CONTROL MICE

pIRS-1

0

20

40

100

60

80

- ++ -

GF

(%)

CONTROL

*

pJNK

0

20

40

100

60

80

GF

(%)

CONTROL

pIKK

0

20

40

100

60

80

GF

(%)

CONTROL

pIRS-1 Ser307

0

20

40

100

60

80

GF

(%)

CONTROL

**

*

Liver

EFFECT OF HIGH FAT DIET -3 DAYS ON INSULIN SIGNALING

IN LIVER GERM-FREE AND CONTROL MICE

LPS

0

0,5

1,0

2,5

1,5

2,0

GF

(EU

/mL)

CONTROL

*

SCFA- Acetate

0

1.0

2,0

5.0

3.0

4,0

GF

(mm

ol/

L)

CONTROL

*

EFFECT OF HIGH FAT DIET -3 DAYS ON LPS AND SCFAON GERM-FREE AND CONTROL MICE

CLAUDIN

OCCLUDIN

ACTIN

ZO-1

MIOSIN(%

)

TIGHT JUNCTIONS - ZO-1, CLAUDIN AND OCCLUDINE

CLAUDIN

OCCLUDIN

ACTIN

ZO-1

MIOSIN

ZO-1

0

20

40

60

80

100

120

GF CONTROL

*

(%)

ZO-1 EXPRESSION IN INTESTINE OF GERM-FREE MICE ON

HIGH FAT DIET

6 sem

WEIGHT

0

5

10

15

20

25

30

TOM

g

TOM TLM TOM TLM

14 sem

600

400

200

00 40 80 120

Time (min)

Blo

od

Glu

cose

mg

dl-1

)

TLM

GTT

METABOLIC CHARACTERISTICS OF LEAN MICE SUBMITTED TO

MICROBIOTA TRANSPLANTATION FROM OBESE MICE

*

LPS

0

0,2

0,4

0,6

0,8

1

1,2

TFO C

pAkt

0

20

40

60

80

100

120

TFO TFO

AntibCTFO

Antib

TFO CTFO

Antib

0

10

15

20

25

30

35

(mg

/kg

/min

)

5

40

Glucose Clamp

INSULIN SENSITIVITY IN LEAN IN MICE SUBMITTED TO

MICROBIOTA TRANSPLANTATION FROM OBESE MICE

EFEITO DE ANTIBIMTNZ+AMP+NEOM)

(EU

/ml)

UA



PI 3

Kinase

S - Sa

S-S

b

ATP

insulin

S-S

PPPO4

IKKb

JNK

HIGH FAT DIET

LPS

PO4

PO4

PO4 PO4

PO4

P

P

INSULIN

RESISTANCE

IRS1/2

PO4

INSULIN RESISTANCE IN HIV PATIENTS

INSULIN LPS

Indivíduo obeso

Flora

ComensalEpitélio Intestinal

Fígado Tec. Adiposo Hipotal Muscle

LPS LPS

Indivíduo obeso

Flora

Comensal

Fígado Adipose

tissue

OBESEIntestinal

Micorbiota

Liver

Short chain fatty acids:

ACETATE, BUTIRATE, PROP.

INSULIN

RESISTANCE


INFLUENCE OBESITY

0

2

4

6 *

AC

ETA

TO

(n

g/u

l)

ACETATE

CONT OB

CORRELATION BETWEEN SCFA LEVELS –

ACETATE- AND AMPK ACTIVITY IN OBESE MICE

0

20

40

60 *

AM

PK

AC

TIV

ITY

(a.u

.)

CONT OB

80

AMPK

EFFECT OF ACETATE ON AMPK ACTIVITY

0

30

60

90 *

UA

CONT 30min 60min

ACETATE GAVAGE IV

30min

*

pAMPK

PI 3

quinase

AKT

S - Sa

S-S

b ATP

insulina

EFFECT OF AMPK ON GLUCOSE AND LIPID METABOLISM

S-S

P

P

P

PPO4PO4

PO4

PO4

IRS1/2

AMPK

AGCC

ACETATO

GLUCOSE TRANSPORT

HEPATIC GLUCOSE OUTPUT

LIPID

OXIDATION

INSULIN RESISTANCAE AND LPS LEVELS IN

OVERWEIGHT AND OBESE HUMANS

0

1

2

3

*4

**

INSULIN

LPS

ADIPOSE TISSUE

MORPHOLOGY IMMUNOHISTOCHEMISTRY

EFECTO OF HIGH FAT DIET ON ADIPOSE

MORPHOLOGY AND MACROPHAGE INFILTRATION

Adipose Tissue - LEAN

Adipokines

(adiponectin, leptin)

M2 - Alternatively

Activated

Inflammatory Molecules

(TNFα, IL-6, iNOS)

Weight Gain

POLARIZATION OF MACROPHAGE IN ADIPOSE

TISSUE

Adipose Tissue - OBESE

M1 - Classicaly

Activated

M2ACETATE

M1

EFFECT OF ACETATE ON POLARIZATION OF

MACROPHAGE IN ADIPOSE TISSUE


(TNFα, IL-6, iNOS)

ACETATE

M2

EFFECT OF ACETATE ON M2 MACROPHAGE INFILTRATION IN

ADIPOSE TISSUE OF MICE ON HFD

Lean ObeseObese +

ACETATE

M2 macrophage

ACETATE

Adipose Tissue - LEAN

Adipokines

(adiponectin, leptin)

M2 - Alternatively

Activated


(TNFα, IL-6, iNOS)

TREATMENT OF INSULIN RESISTANCE -

MODULATE THE POLARIZATION OF MACROPHAGE

Adipose Tissue - OBESE

M1 - Classicaly

Activated

0

2

4

6

*

Perc

enta

ge

-%

Akkermansia muciniphila

8

CONT DH DH+MET

EFFECT OF METFORMINE ON THE EXPRESSION OF

DE AKKERMANSIA MUCINIPHILA IN COLON

0

2

4

6

*

Perc

enta

ge

-%

Akkermansia muciniphila

8

CONT CLOZAPINE OLANZAPINE

EFFECT OF ANTIPSICOTHIC DRUGS ON THE

EXPRESSION OF DE AKKERMANSIA MUCINIPHILA

*

IN GUT WE TRUST

Bruno Geloneze

Candida Parisi

Kleber Franchini

Lauro Kubota

Lício Velloso

Maria Helena M Lima

Patrícia Prada

Rui Curi

Sr. Luiz Janeri

Sr. Jósimo Pinheiro

AGRADECIMENTOS

Alexandre Gabarra Oliveira

Andrea Cariccili

Angélica Souza

Bruno de Melo Carvalho

Dioze Guadagnine

Fernanda Pace

Francine Mitzetneir

Kelly Lima Calixto

Natalia Tobar

Renata Bagarolli

Tiago Araújo

RESIST À INSULINA (HOMA) E NÍVEIS DE LPS EM

ADOLESCENTES COM SOBREPESO E OBESOS

0

1

2

3

*4

**

HOMA

25

20

40

35

30

15

10

5

01974 1980 1986 1992 1998 2004

Prevalence of obesity

Prevalence of ascaridiasis

%

Year

Cont. do apetite

AKT

HIPOTÁLAMO

Prod. de glicose

AKT

FÍGADO

Capt de glicose

AKT

MÚSCULO

ADIPOSO

Infiltração de

macrófagos

VASOS

eNOS

RESISTÊNCIA À

INSULINA

RESISTÊNCIA À INSULINA – PAPEL DO TGI E ADIPOSO

TNFa

IL6

TGI

LPS

Flora

INFILTRAÇÃO DE CÉLULAS NO ESTROMA

VASCULAR DO TECIDO ADIPOSO

Macrófago M2

Macrófago M1

Neutrófilo

Eosinófilo

CÉLS NO ESTROMA VASCULAR DO ADIPOSO

Tec. Adiposo - MAGRO

Adipocinas

(adiponectina, leptina)

M2 - Ativado

Alternativamente

M1 - Ativado

Classicamente

Tec. Adiposo - OBESO

Moléculas pro-inflamatórias

(TNFα, IL-6, iNOS)

Ganho de Peso

TRANSFORMAÇÃO DE MACRÓFAGOS

REGENERATIVOS-M2 EM INFLAMATÓRIOS-M1.

Indivíduo obeso

Flora

Comensal

Ácidos graxos de cadeia curta:

ACETATO, BUTIRATO, PROP.

Epitélio Intestinal

Fígado Tec. Adiposo Hipotál Músculo

MECANISMOS QUE LIGAM FLORA INTESTINAL E

RESISTÊNCIA À INSULINA – OBESIDADE

Indivíduo obeso

Flora

Comensal

Fígado Tec. Adiposo

Indivíduo obeso

Flora

Comensal

Fígado

RESISTÊNCIA

À INSULINA

Indivíduo obeso

Flora

Comensal







TregLPS

Indivíduo obeso

Flora

Comensal


Indivíduo obeso

Flora

Comensal

Fígado

Treg

INFILTRAÇÃO DE CÉLULAS NO ESTROMA

VASCULAR DO TECIDO ADIPOSO

Macrófago M2

Macrófago M1

Neutrófilo

Cel T (Treg)

CÉLS NO ESTROMA VASCULAR DO ADIPOSO


M2 - Ativado

Alternativamente

M1 - Ativado

Classicamente



INFLAMATÓRIOS-M1 EM REGENERATIVOS-M2.

AGCC

AGCC


INFLAMATÓRIOS –M1 EM REGENERATIVOS-M2

M2

Regenerativo M1

Inflamatório

TregAcetato

M2Treg

INFECÇÃO PELO STRONGYLOIDES VENEZUELENSIS A

INFILTRAÇÃO DE M2 NO TECIDO ADIPOSO DE OBESOS

CONTROLE OBESO OBESO + SV

MGL1 – macrófago M2

EFEITO DA INFILTRAÇÃO DE CÉLS Treg NA

DIFERENCIAÇÃO DO MACRÓFAGO EM M2 OU M1

M2Treg

M1Treg

ESTUDOS DE METAGENOMA DE MICROBIOTA EM DM2

145 F 345 M e F

N, IG e DM2 N e DM2

Bactérias produtoras de butirato: Roseburia e F Praunitzi


Proteobactéria –

LPS

Bactérias - stress oxidativo

Karlsson et al. Qi et al.

DROGAS QUE MODULAM FLORA

NOVAS ASSOCIAÇÕES E PERPECTIVAS NO

ESTUDO DA MICROBIOTA

METFORMINA

ANTIPSICOTICOS ATÍPICOS – Clozapina - Olanzapina

DROGAS MODULADAS PELA FLORADIGOXINA

DOENÇAS CRÔNICAS –

Possível associaçao

OSTEOPOROSE – Massa Óssea

HIPERTENSÃO ARTERIAL

25

20

40

35

30

15

10

5

01974 1980 1986 1992 1998 2004

OBESITY

ASCARIDIASIS

%

YEARS

PREVALENCE OF ASCARIDIASIS AND

OBESITY IN CHILDREN IN BRAZIL

EFFECT OF ACUTE EXERCISE ON TLR4 ACTIVATION AND

INSULIN SIGNALING IN MUSCLE OF MICE ON HFD

C 16h HFD 2h

HFD+EXE

0

20

120

AU

40

*80

60

100

TLR4-MyD88

Muscle

C 16h HFD 2h

HFD+EXE

0

20

120

Un

ida

de

s a

rbit

rári

as

40 *

80

60

100

#

#

pAKT

0

4

12

L/m

in-1

8

Basal

Exercise

EFFECT OF ACUTE EXERCISE ON BLOOD FLOW IN

DIFFERENT ORGANS

LPS

0

0,25

0,5

0,75

1,0

1,25

C

*

HFD

(EU

/mL)

HFD + EXE

2h 16h

EFFECT OF ACUTE EXERCISE ON CIRCULATING

LPS

Indivíduo obeso

Flora

Comensal







Indivíduo obeso

Flora

Comensal


Indivíduo obeso

Flora

Comensal

Fígado

RESISTÊNCIA

À INSULINA

0

10

15

20

25

30

35

CONT

*(m

g/k

g/m

in)

5

STRG

EFFECT OS STRONGYLOIDES VENEZUELENSIS INFECTION ON

INSULIN SENSITIVITY IN MICE ON HFD

40

CONT

*

STRG

CHOW HFD

Glucose Clamp

0

30

40

50

60

70

80

CONT

*

(%)

20

STRG

90

CONT

*

STRG

LPS

0

10

15

20

25

30

35

CONT

*(EU

/L)

5

STRG

40

CONT

*

STRG

% Firmicutes

(lactobacilus)

CHOW HFD CHOW HFD

EFFECT OF STRONGYLOIDES VENEZUELENSIS INFECTION ON

INTESTINAL MICROBIOTA AND ON LPS CIRCULATING LEVELS

HL

HL+

ANTC

pAMPK

0

20

40

60

80

100

120

HLC

0

20

40

60

80

100

120

HL+

ANT

pAMPK

FÍGADO MÚSCULO

ATIVAÇÃO DA AMPK EM FÍGADO E MÚSCULO DE

ANIIMAIS OBESOS TRATADOS COM ANTIBIÓTICOS

Alexandre G Oliveira

Andréa Moro Caricilli

Angélica Camacho

Bruno de Melo Carvalho

Dioze Guadagnini

Francine Mittestainer

Fernanda Pace

Kelly Lima Calixto

Natalia Tobar

Patty Karoll Picardi

Renata Bagarolli

Tiago Araujo

José B C Carvallheira

Eduardo Ropelle

Lício A Velloso

A C Boschero

Rui Curi

Patrícia O Prada

CNPq – FAPESP

Instituto Nacional de

Obesidade e Diabetes

ACKNOWLEDGEMENTS

MODULATION OF GUT MICROBIOTA

MODULAÇÃO DA FLORA INTESTINAL

DIETA PROBIÓTICOS

MICROBIOTA

TRANSPLANTE

DE FLORAANTIBIÓTICOS

MODULATION OF GUT MICROBIOTA BY PROBIOTICS

COMPOSITION OF PROBIOTIC

Lactobacillus rhamnosus,

Lactobacillus acidophillus

Bifidobacterium bifidum.

6x108 UFC de cada cepa

0

5

10

15

*

Glu

co

se U

tilizati

on

(mg

/kg

/min

)

Hyperinsulinemic

Euglycemic Clamp

20

CONT HFD HFD+PB

EFFECT OF PROBIOTICS ON INSULIN SENSITIVITY

IN MICE ON HFDC

HF HF+PBC

pJNK

0

20

40

60

80

100

120

0

200

600

1200

AU

800

*

1000

400

- + - ++ -

Cont

Insulin

HF HF+ PB

pAKT

AU

EFFECT OF PROBIOTICS ON INSULIN SIGNALINGIN

MUSCLE OF MICE ON HFD

BACTEROIDETES

EFFECT OF PROBIOTICS ON THE MICROBIOTA OF

MICE ON HFD

FIRMICUTIS

ESTUDOS DE METAGENOMA DE MICROBIOTA EM DM2

145 F 345 M e F

N, IG e DM2 N e DM2



Proteobactéria –

LPS

Bactérias - stress oxidativo

Karlsson et al. Qi et al.

ER STRESS AND INSULIN RESISTANCE

PO4

PO4

INSULIN

RESISTANCEIRE1ATF6PERK

IKKb, JNK

iNOS

PO4

IRS1/2

PP

IRS1/2PI 3

kinase

NO

0

5

10

15

*

Glu

co

se f

lux (

mg

/kg

/min

)

Glucose Clamp

20

CONT OB OB+PB

EFFECT OF PROBIOTICS ON INSULIN SENSITIVITY

IN OBESE MICE

0

10

15

20

25

30

35

CONT

*(m

g/k

g/m

in)

5

STRG

40

CONT

*

STRG

LEAN OBESE

Glucose Clamp

EFFECT OF STRONDILOGIDIASIS ON MICROBIOTA AND ON

INSULIN SENSITIVITY IN OBESE MICE

0

30

40

50

60

70

80

CONT

*

(%)

20

STRG

EFFECT OF STRONDILOGIDIASIS ON MICROBIOTA AND ON

SERUM LPS IN LEAN AND OBESE MICE

90

CONT

*

STRG

LEAN OBESE

SERUM LPS

0

10

15

20

25

30

35

CONT

*(EU

/L)

5

STRG

40

CONT

*

STRG

LEAN OBESE

% Firmicutes

(lactobacilus)

0

1

2

4

*

3

INSULIN

(ng/dL)

HFD HFD+PB0

50

100

200

*150

GLUCOSE

(mg/dL)

HFD HFD+PB0

10

15

25 *

20

GLP1

(pmol/L)

HFD HFD+PB

EFFECT OF PROBIOTICS ON PLASMA GLUCOSE

HORMONAL PARAMETERS IN MICE ON HFD

CLAUDIN

OCCLUDIN

ACTIN

ZO-1

MIOSIN

ZO-1

EFFECT OF PROBIOTICS ON ZO-1 EXPRESSION

IN INTESTINE OF OBESE MICE

OB OB+PBC

0

20

40

60

80

100

120

0

10

15

20

25

30

35

(mg

/kg

/min

)

5

EFFECT OF HIGH FAT DIET ON INSULIN SENSITIVITY

AND SIGNALING IN MUSCLE OF TLR4 -/- MICE

40

*

CONT TLR4-/-

CONT TLR4-/-Insulin

pJNK

GLUCOSE UPTAKE

CONT TLR4-/-

IRS1

ser307

pAKT

CHOW HFD

HIGH FAT DIET

LPS - FFA

MYD88

MAL/TIRAP

IRAK

TRAF6

TAK1

JNK IKK / IkB / NFkB

TNFa IL-6 iNOS

TLR4

LPS SIGNALING – TLR4

INTERACTION BETWEEN INNATE IMMUNE

SYSTEM AND INSULIN RESISTANCE

DIET AND

PATHOGENS

INNATE IMMUNE SYSTEM

INSULIN

RESISTANCE

0

50

100

150

200

250

300

C HFD

EFFECT OF HFD ON THE INTESTINAL MICROBIOTA

AND CIRCULATING LEVELS OF LPS

0

5

10

15

20

25

C HFD

Gli

ce

mia

mg

/dl

A U

LPS levels

Plasma Glucose

Bac

teró

ides

65%

Bifidobactérias

20%

Fu

so

ba

cte

rias

12

%

Clo

strídio

s3%

0

20

40

60

80

100

C HFD

Bo

dy w

eig

ht

(g)

5m HFD

Documents

MOLECULAR BASIS OF INFLAMMATION AND INSULIN RESISTANCE IN OBESITY ... · resistÊncia À insulina resistÊncia À insulina