« Predisposition de la réponse à l’agression »

D Payen, MD, Ph Ddpayen1234@orange.fr

Interface INSERM-SFAR-SRLF

Inflammation et métabolismesont étroitement intriqués…

Argument phylogénique et « anatomique »

the Drosophila fat body

incorporates the mammalian homologues of

the liver and the haematopoietic and immune

Systems.

Function:sensing energy and nutrient

availability,

coordinates the appropriate

metabolic and survival responses

site of coordination of pathogen

responses with metabolic status.

Therapeutic targets at the interface between metabolic and inflammatory pathways

Argument génétique constitutif

The constitutive view

Topic:Studies on SNP’s associations with

innate immune response.

3 examples of leading pathways:– TNF- promoter variants– TLR4 haplotypes– HLA-DB variants

Strategy: known prot gene SNPs association

Genetic Polymorphisms and Severe Sepsis

Meningococcemia; Severe sepsis

Meningococcemia; Severe sepsis

PAI-1

FactorV Leiden

Severe SepsisViral Pneumonia

IL-1 locusIL-4

MeningococcemiaSeptic Shock; Cerebral MalariaSevere SepsisSevere Sepsis, MeningococcemiaSevere sepsis

TNF locus

IL-18IL-10IL-6

Meningococcemia; Pneumococcemia

FCRII Receptor

Meningococcemia, Pneumococcemia

Severe sepsis

Mannose Binding Lectin

Gene

Gram negative/positive Septic

ShockLegionnaire’s DiseaseSeptic Shock

Toll-Like Receptor 4/2

Toll-Like Receptor 5

CD14

Susceptibility and/or Outcome

Limitations of gene by gene approachLimitations of gene by gene approach

Hope in development of genome wide association methods (600 000 variants studied in one shot!)

Hope in development of genome wide association methods (600 000 variants studied in one shot!)

Clark et al. Intensive Care Med 2006 32:1706-1712

Argument génomique

• Gene expression in whole blood leukocytes determined before and at 2, 4, 6, 9 and 24 h after the i.v LPS to 4 HV, compared to 4 additional subjects without LPS.

• Only 3% of the genes modified their expression

• Essentially under-expression human blood leukocyte response to acute systemic inflammation the transient dysregulation of leukocyte bioenergetics &modulation of translational machinery

Argument neuro-hormonal

Insulin-receptor signalling interfaces with inflammatory signalling at the level of insulin-receptor substrates through activation of serine kinases.

(b) ACTH responders

P value for log rank test: 0.9370

0.25

0.50

0.75

1.00

0 5 10 15 20 25 30

placebo

steroid

P value for log rank test: 0.8500

0.25

0.50

0.75

1.00

0 5 10 15 20 25 30

(a) ACTH non-responders

placebo

steroid

P value for log rank test: 0.813

0

0.25

0.50

0.75

1.00

0 5 10 15 20 25 30

day

(c) All patients

placebo

steroid

Figure 2 - Kaplan Meier Curves for 28 day all-cause mortality in (a) ACTH nonresponders (b) ACTH

responders and (c) all patients

Figure 3 - Kaplan Meier Curves for time to reversal of shock in (a) ACTH

nonresponders (b) ACTH responders and (c) all patients

placebo

steroid

P value for log rank test: <0.001

0

0.25

0.50

0.75

1.00

0 5 10 15 20 25 30

day

(c) All patients

P value for log rank test: 0.0380

0.25

0.50

0.75

1.00

0 5 10 15 20 25 30

placebosteroid

(a) ACTH non-responders(b) ACTH responders

P value for log rank test: <0.0010

0.25

0.50

0.75

1.00

0 5 10 15 20 25 30

placebosteroid

Sub-paper from Corticus data base

Our article: shock reversal & outcome

QS: Why success in shock reversal did not improve outcome?

Within the first five daysIncidence of shock reversal according to treatment arm

P<0.0001

0 1 2 3 4 5 6

0.0

0.2

0.4

0.6

0.8

1.0

Days after randomization

Cu

mu

lati

ve i

nci

den

ce

HydrocortisonePlacebo

Within the first five days

Incidence of death prior to shock reversal according to treatment arm

0 1 2 3 4 5 6

0.0

0.2

0.4

0.6

0.8

1.0

Days after randomization

Cu

mu

lativ

e in

cid

en

ce

HydrocortisonePlacebo

P= 0.28

Argument biochimique

Free energy used by cells respiration or glycolysis

Alternatively, reactions can be centred around ATP.

• ATP-producers include glycolysis & oxidative phosphorylation

• ATP-consumers: transport of cations & synthesis of macromolecules

energy consuming functions of immune cells

• Synthesis of macromolecule & ion transport; O2 is mainly used by mitochondria, non-mitochondrial O2 consumption is negligible

ADN/ARN synthesis

O2 is used for what type of functions? 

O2

ADP+Pi

ATP

H+

Actinomycine D

ATP synthase

Respiratoire Chain

NADPH oxydase

protein Synthesis

Na+, K+ ATPase

Ca2+ ATPase

Antimycine A

Cycloheximide

Inhibition

Buttgereit, Biochem J, 1995.

Ouabaïne

DPIChlorure de lanthanum

Global VO2: the different fractions

% O2

75

80

85

90

95

100

0 500 1000 1500 2000 2500 3000

Sec

Cellules

DPI

Cycloheximide

Ouabaïne

ActinomycineDChlorure de lanthanum

Antimycine A

78 ngatomes O2

Septic plasma  modifications similar to those observed in septic cellsPlasmatic factors +++ 

Effects septic plasma septique on baseline (V0) & stimulated reponse

Mitochondrial VO2 becomes more decoupled after incubation in septic plasma.

It cannot result from tissue hypoxia!!!!

MIntegration of stress-signalling mechanisms.

Le sucre et inflammation…

J Appl Physiol 1997

Exogenous glucose (NUTRITION)

FedFastedFasted+iP G

J Appl Physiol 1997

In conclusion, glucose metabolism interferes with hemodynamic, metabolic, and inflammatory responses to LPS. MAJOR ROLE OF EXOGENOUS GLUCOSE

Exogenous glucose (NUTRITION)

Fig. 4

O2 c

on

sum

pti

on

rat

e, n

gat

om

s O

2/m

in/1

07 c

ells

0

4

8

12

16

NG HG

VoVstim ionomycinVstim PMA p=0.003

A

Del

ta i

ncr

ease

in

O2 c

on

sum

pti

on

, %

0

50

100

150

200

NG HG

Delta ionomycinDelta PMA

p=0.004

B

Fig. 5

Vo

, n

gat

om

s O

2/m

in/1

07 ce

lls

HLA-DR, site number

0

2

4

6

8

10

0 10000 20000 30000 40000 50000

sepsis

HV

Del

ta i

ncr

ease

in

O

2 co

nsu

mp

tio

n,

%

HLA-DR, site number

0

50

100

150

200

0 10000 20000 30000 40000 50000

sepsis

HV

Del

ta i

ncr

ease

in

O

2 co

nsu

mp

tio

n,

%

-25

25

75

125

175

225

0 10000 20000 30000 40000 50000

sepsis

HV

HLA-DR, site number

r=0.44p=0.009

r=0.57p=0.0014

r=0.58p=0.049

A

B

C

Iono+PMA

ADP

Un example of acute inflammation aigue in human beings: hepatic  transplantation

Day 4 after OLT

Argument inflammatoire

Immunity triggers inflammation

Pathogen pathwayPAMPs or MAMPs“stranger model”Infection and sepsis

Cell and tissue lesions DAMPs“danger model” infection, trauma,

postop, burns

Kono et coll. Nat Rev Imm 2008

Cell death and inflammation.

- Necrotic cell death DAMPs receptors

on leukocytes + prod of pro-inflam cytokines

(IL‑1).

- Other molecules proteases; hydrolases

act on EC components + mediators

(complement fragments) or DAMPs prod of

pro-inflam cytokines by host cells.

- Pro-inflam mediators local vascular

endothelium ‘leaky’, attracts neutrophils

and monocytes/macrophages soluble

(antibody) and cellular defences in the

tissue

Donc, les facteurs de prédisposition

Outcome

ConstitutivePolymorphismsSNPs…

AcquiredPolymorphisms?Epigenetic…

Chronic disease Chronic

treatment

Poor or goodQuality of feeding

PharmacogenomicDrug induced geneExpression changes

Reversing shock ≠Improve outcome

Surviving shock increasesthe risks of disease

Proteins releasedAre changed byEnvironment (nitrosylated…)

Metabolic failureNot related to perfusion

Need forMarkers

D Payen 2010

Difficult to reverse but good reserve

Easy to reverse But high co-morbidity in elderly…

TISSUE RESERVE = Co-morbidity

AGECo-morbidity

Sev

erit

y S

core

INJURY

Severe Injury + good tissue reserveModerate injury + high Co-morbidity Similar severity score

Shock severity is characterized by scores…Outcome may then depend on injury but also

on acquired negative factors…

Survivors of hospitalization for community-acquired pneumonia are at increased risk of cardiovascular events, repeated infections, and death in the following months.

But the cause is unknown…Early and long-lasting mechanisms?But the cause is unknown…Early and long-lasting mechanisms?

Viral co-infections and tissue damageViral co-infections and tissue damage

Crit Care Med 2009; 37:1850-1857

242 ICU patients, 39 CMV infections

ICU mortality: 54% in CMV patients vs 37% in others (p=0.082)

In-hospital mortality: 59% vs 41% (p=0.058)

ICU LOS: 32d vs 12d (p<0.001)

Length of MV: 27d vs 10d

At least one bacterial nosoc inf: 69% vs 33% (p<0.001)

242 ICU patients, 39 CMV infections

ICU mortality: 54% in CMV patients vs 37% in others (p=0.082)

In-hospital mortality: 59% vs 41% (p=0.058)

ICU LOS: 32d vs 12d (p<0.001)

Length of MV: 27d vs 10d

At least one bacterial nosoc inf: 69% vs 33% (p<0.001)

En conclusion

• Métabolisme et inflammation sont intriqués• Les raisons et les preuves sont multiples• Les facteurs enzymatiques, les substrats, les organelles, les facteurs nucléaires interférent pour assurer et moduler la réponse inflammatoire

• Le traitement et les biomarqueurs métaboliques et inflammatoires constituent un espoir diagnostique et thérapeutique

Attendons les données futures….

Mitochondrial oxygen consumption - oxidative phosphorilation (> 90 %).

Electron transport chain Saraste M. Science, 1999.

Migration

Cytokinesis

Phagocytosis

Antigen processing

Antigen presentation

Activation

Effector functions

ATP

Active transport of molecules and ions

Synthesis of macromolecules

(RNA/DNA/protein synthesis)

Specific immune functions

General housekeeping functions

ENERGY CONSUMING FUNCTIONS OF IMMUNE CELLS

Buttgereit F. Immunology Today, 2000.

LPS injection: an example of ACUTE INFLAMMATION

…

Calgranulines: an example of DAMPs molecule

May be a prognostic marker…

deadalive

p<0,0001

0

5

10

15

20

25

S10

0A8/

A9

pg/m

l

n=61 n=50

Plasma S100 A8/A9 complex D0SOFA D0

2

4

6

8

10

12

14

16

18

20

NS (p<0,09)

Gobal population n = 111

Payen D, Lukazsewicz AC et al. (in review)(Patented in December 2008)

*p=<0.0001, test de Mann WhitneyCinétique des “alive”: D0-D28, p<0.0001, test de Friedman, n=34

D0-D14, p<0.0001, test de Friedman, n=41D0-D7, p=0.0005, test de Friedman, n=57

Cinétique des “dead”: D0-D14,NS, test de Friedman, n=8D0-D7, p=0.0010, test de Friedman, n=15

* *

(41)

0

2

4

6

8

10

12

14

16

18

20

22

24

26

28

30

Pla

sma

S10

0A8/

A9

(g

/ml)

D0 D1 D7 D14 D28

dead

alive

(15)(8)

*

*

(43) (46)(59)

(59)(60)

(50)

Figure: trend over time of S100A8/A9 complex, according to outcome, in plasma. Results expressed in g/ml. Effectives in brackets.

S100A8/A9 in patients without shock at D0N=8 survivorsN=8 non survivors

Mann whitney test, p=0.0008

0

4

8

12

16

20

24

28

D0

dead

alive

Pla

sma

S1

00

A8

/A9

(

g/m

l)

p=0.0008

Metabolic alterations: Singer et al. Lancet 2004, Belikova et al. CCM 2007

Impairment of capacity of repair?

Exple: autophagy – to sustain metabolism during nutrient

deprivation

– to prevent the accumulation of damaged, toxic proteins and organelles during stress

– Implicated in anti-microbial defense

inhibits cell death by necrosis

Metabolic failure and cell recoveryMetabolic failure and cell recovery

HO

ME

OS

TA

SIS

ST

RE

SS

AD

AP

TA

TIO

N

Provision of nutrientsduring catabolism

Generation of ATPin stressed cells

Signals forheterophagic removalof apoptotic cells

Degradation ofmisfolded proteins

Removal of surplusor damage organelles

Genomic stability

Levine et al. Cell 2008 132:27-42

Autophagy is impaired in:

degenerative disease, heart disease, aging, cancer, dysimmunity…

Autophagy is impaired in:

degenerative disease, heart disease, aging, cancer, dysimmunity…

Levine et al. Cell 2008 132:27-42

Autophagy in innate and adaptive immunityAutophagy in innate and adaptive immunity