metabolic stress, mitochondria and organ failure during ... · organ failure during critical...

Metabolic stress, mitochondria and organ failure during critical illness:underlying mechanisms revealing

therapeutic potential

Jan Gunst, MD, PhD

33rd Annual SIZ meeting

Vilvoorde, June 14, 2013

Metabolic interventions can affect outcome

Tight glycemic control with intensive insulin therapy

Early parenteral nutrition

Van den Berghe et al. NEJM 2001Van den Berghe et al. NEJM 2006 Vlasselaers et al. Lancet 2009Casaer et al. NEJM 2011

Study aims

Mechanisms of organ protection by preventing hyperglycemia with insulin

Detailed impact of early vs late PN on the kidney

Data published:

Kidney Int 2009;76(5):512-20

Part 1: Glycemic control vs insulin & renal damage

Normoinsulinemia/Normoglycemia NI/NGHyperinsulinemia/Normoglycemia HI/NG Normoinsulinemia/Hyperglycemia NI/HGHyperinsulinemia/Hyperglycemia HI/HG

InsulinemiaGlycemia

3 5 70Basel.

200

50

1 2 3 4 5 6 7Basel.

350

100500

450500550

150200250300

400

mg/d

l

Days Days

300

250

150

100

0

mU

/l

Ellger et al., Diabetes 2006

4-arm study:blood glucose and insulinemia regulated independently

Glucose vs insulin & kidney function

: p≤0.1, p≤0.05 versus control§, #

mg/d

l

0.0

0.5

1.0

1.5

2.0

2.5

3.0

#

§

Bars indicate mean s.e.m.

: p≤0.1, p≤0.05 between sick groups

Plasma creatinine

n 4 8 8 89

Glyc. NG NG HG HG

Ins. NI NI HIHI

Control Sick

Glucose vs insulin & kidney function

: p≤0.1, p≤0.05 versus control§, #

mg/d

l

0.0

0.5

1.0

1.5

2.0

2.5

3.0

#

§

Bars indicate mean s.e.m.

: p≤0.1, p≤0.05 between sick groups

Plasma creatinine

n 4 8 8 89

Glyc. NG NG HG HG

Ins. NI NI HIHI

Control Sick

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

0 5 10 15 20 25 30 35

Cre

atinin

e (

mg/d

l)Glucose (mmol/g cortex)

R=0.546P=0.0005

Mechanisms of organ protection

Mitochondrial functionTissue perfusion & DO2

r=-0.211p=0.24

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

-1 -.8 -.6 -.4 -.2 0 .2 .4 .6 .8 1

Cre

atinin

e (

mg/d

l)

log O2 delivery to cortex

U/g

cort

ex

Complex I

0

5

10

15

20

25

30

35

40

#

# #

n 4 8 8 89

Complex V

0

25

50

75

100

125

150

175

200

225

##

##

n 4 8 8 89

: p≤0.05 versus control# : p≤0.1, p≤0.05 between sick groups Bars indicate mean s.e.m.

Complex V (U/g cortex)

r=-0.609P<0.0001

0 50 100 150 200 250

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

Cre

atinin

e (

mg/d

l)

Complex I (U/g cortex)

r=-0.640P<0.0001

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

0 5 10 15 20 25 30 35 40

#

0.0

1.0

2.0

3.0

4.0

5.0

ml O

2/1

0g p

er

min

Cortical O2 delivery

n 8 8 8 89

HI/HGNI/HGHI/NGControl NI/NG

Conclusion Part 1Glycemic control versus Insulin & renal damage

Normoglycemia

Hyperinsulinemia

Renal protection

Protection againstmitochondrial damage

Glucose

Glucose-6-P

Glyceraldehyde-3-P + DHAP

Glycerate-1,3-biP

Pyruvate

L-lactateAcetyl-CoA

Krebs cycle-Respiratory chainFatty acids/TGKetone bodies

MethylglyoxalAGEs

Free adducts

D-Lactate

Glycogen

Glyoxalase IGlyoxalase II

Glyoxal

3-deoxyglucosone

Mechanisms of glucose toxicity

GAPDH

Mitochondrial dysfunction(Multiple) organ failure

Direct insult

Insufficient mitochondrial repair

Part 2: Role of intact mitochondrial repair in critical illness

Data published: J Clin Endocrinol Metab 2011;96(4):E633-645J Clin Endocrinol Metab 2012;97(1):E59-64Crit Care Med 2013; 41(1):182-94

Mitochondrial repair: autophagy

LC3-IIp62

ububub

SSub

p62ub

ubub

Sub

p62ub

ubub

Sub

Isolationmembrane

Autophagosome Autophagolysosome

Lysosome

LC3-I

LC3-I

p62

Ubiquitinated substrate

LC3-II

Boxes indicate median/IQR, whiskers interdecile range

: p≤0.05 versus control

Control (elective rectal surgery)

Conventional insulin therapy

Autophagosome

0

50

100

150

200

250

300

350

400

5 5n

Numberautophagic vacuoles

Human liver

LC3-I

p62

Ubiquitinated substrate

LC3-II


: p≤0.05 versus control

Control

Conventional insulin therapy

?

?

?

60

0

10

20

30

40

50

LC3-II/LC3-I ratio

18 18n

0

10

20

30

40

50

60

p62

Rela

tive p

rote

in level

18 18n0.20

0.25

0.30

0.35

Ubiquitin staining

12 18n

Inte

nsi

ty

Autophagosome

Human liver

Mitochondrial repair in vivo?

Human post mortem biopsies: relation with outcome?

Rabbit model of prolonged critical illness: survivors vs. non-survivors

Autophagy ~ Outcome

p62

Liver Kidney

LC3-II/LC3-I ratio

Sick-Non Survivor

Sick-Survivor

Healthy

0

1

2

3

4

0

1

2

3

4

5

6

7

8

9

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

3

4

5

6

7

8

9

10

11

12

ALT

last

day

(sqrt

-tra

nsf

orm

ed)

0.4 0.6 0.8 1.0 1.2 1.4 1.6 1.8 2.0 2.2

p62 liver (sqrt-transformed)

r=0.647p<0.0001

n 8 26 13 n 8 26 13

Boxes indicate median/IQR, whiskers interdecile range. Organs could not be sampled in 5/18 non-surviving animals.

, () : p≤0.05, 0.05<p≤0.1 vs control : p≤0.05, 0.05<p≤0.1 survivor vs non-survivor

, () : p≤0.05, 0.05<p≤0.1 vs control : p≤0.05, 0.05<p≤0.1 between sick groups

Boxes indicate median/IQR, whiskers interdecile range. Pearson correlation calculated after square root transformation of p62 and markers of organ damage

LIVER KIDNEY

3 days 7 days 3 days 7 days

0

1

2

3

4

5

6

0

1

2

3

4

5

6

7

0

1

2

3

4

0

1

2

3

4

()

p62 protein

Normo-glycemia

Hyper-glycemia

Control

Mitochondrial protection by preventing hyperglycemia (in part) explained by maintaining autophagy more efficient?

ALT last day (U/l)AST last day (U/l) Creatinine last day (mg/dl)

Complex V activ.(U/g liver)

Complex I activ.(U/g kidney)

Complex V activ.(U/g kidney)

Complex I activ.(U/g liver)

LIVER KIDNEY

3 days 7 days 3 days 7 days

0

1

2

3

4

5

6

0

1

2

3

4

5

6

7

0

1

2

3

4

0

1

2

3

4

0

1

2

3

4

5

6

7

0.6 0.8 1.0 1.2 1.4 1.6 1.8 2.04

6

8

10

12

14

16

0.6 0.8 1.0 1.2 1.4 1.6 1.8 2.0

0.6

0.8

1.0

1.2

1.4

1.6

1.8

2.0

0.8 1.0 1.2 1.4 1.6 1.8

0

2

4

6

8

0.0 0.5 1.0 1.5 2.00

10

20

30

40

50

60

70

0.0 0.5 1.0 1.5 2.0

2

3

4

5

6

7

8

9

0.5 1.0 1.5 2.0 2.5

n 7 8 8n 8 8 8 n 5 5 5n 5 5 5

()

()

2

3

4

5

6

7

8

9

0.5 1.0 1.5 2.0 2.5

p62 protein

, () : p≤0.05, 0.05<p≤0.1 vs control : p≤0.05, 0.05<p≤0.1 between sick groups

Boxes indicate median/IQR, whiskers interdecile range. Pearson correlation calculated after square root transformation of p62 and markers of organ damage

Correlation p62 protein (X-axis) with:

r=0.808p=0.0002

r=0.606p=0.01

r=0.562p=0.02

r=-0.692p=0.004

r=-0.671p=0.006

r=-0.611p=0.004

r=-0.605p=0.005

Mitochondrialdamage

Organdamage

Normo-glycemia

Hyper-glycemia

Control

Activation of autophagy

Sick-Rapamycin

Sick-Saline

Healthy


0.4

0.8

1.2

1.6

2.0

2.4

p62 kidney

n 4 6 60.5

0.6

0.7

0.8

0.9

1.0

1.1

1.2

Plasma creatinine

n 4 6 6

Complex V activity kidney

20

40

60

80

100

120

140

160

()

n 4 6 6

, () : p≤0.05, 0.05<p≤0.1 vs healthy : p≤0.05, 0.05<p≤0.1 between sick groups

Insufficient autophagy

Mitochondrial dysfunctionOrgan damage

Adverse outcome

Hyperglycemia (Parenteral) feeding1

Conclusion Part 2: Role of intact mitochondrial repair in critical illness

1Derde et al. Endocrinology 2012

Casaer et al. N Engl J Med 2011

Part 3: Detailed impact of early vs late PN on AKI

Data published:

J Am Soc Nephrol 2013; 24(6):995-1005

Incidence of AKI

Early PN

n (%)

Late PN

n (%)p

AKI (any) 568 (24.9) 565 (24.6) 0.8

AKI Stage 1 219 (9.6) 197 (8.6) 0.2

AKI Stage 2 99 (4.3) 107 (4.7) 0.6

AKI Stage 3 250 (11.0) 261 (11.4) 0.7

AKI stage 1 and 2 were defined as peak creatinine 1.5-2x, respectively 1.5-2x baselinevalue. AKI stage 3 was defined as preak creatiinine 2x baseline value OR Creatinine >4mg/dl (and 0.5 mg/dl rise) OR new renal replacement therapy

Recovery from AKI

Early PN Late PN p

AKI stage 1

Days with AKI in ICU1 1 (1-2) 2 (1-3) 0.4

AKI stage 2


AKI stage 3


1 Data show median (interquartile range) for ICU survivors only

Recovery from AKI

Early PN Late PN p

AKI stage 1


Alive and AKI-free at hospital discharge, n (%) 168 (76.7) 148 (75.1) 0.7

AKI stage 2



AKI stage 3



1 Data show median (interquartile range) for ICU survivors only

2312Early PN

0.0

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

Plasma creatinine

d1 d3 d5 d7 d9 d11 d13

1Excluded: dialyzed patients (n=428) and patients with missing samples on more than 2 consecutive days (n=584)

Bar graphs indicate mean and 95% CI

Late PN

Early PN

0.001<p≤0.01; # p≤0.001 between sick groups

Plasma urea

0

20

40

60

80

100

No. in ICU

1438 975 736 601 517 4322328 1399 913 655 524 436 365Late PN

#

## # ##

d1 d3 d5 d7 d9 d11 d13

Urea/creatinine ratio

Creatinine clearance1

0

20

40

60

80

100

120

#

##

######

0

20

40

60

80

100

d1 d3 d5 d7 d9 d11 d13

d1 d3 d5 d7 d9 d11 d13

Nitrogen loss and balance over time in ICU

Nitrogen loss (g) Nitrogen balance (g)

d1 d3 d5 d7 d9 d11 d13 d1 d3 d5 d7 d9 d11 d13

Excluded: dialyzed patients (n=428) and patients with missing samples on more than 2 consecutive days (n=584)Bar graphs indicate mean and 95% CI

No. in ICU

1801 1003 622 436 336 276 220

1827 989 598 402 298 234 191

Early

LatePN

1801 1003 622 436 336 276 220

1827 989 598 402 298 234 191

Early

LatePN

0.001<p≤0.01; # p≤0.001 between sick groups

#

#

#####

#

# ## # #

# # #

0.0

2.5

5.0

7.5

10.0

12.5

15.0

17.5

20.0

22.5

-16

-14

-12

-10

-8

-6

-4

-2

0

Late PN

Early PN

63% of extra nitrogen administration net wasted!

Conclusion Part 3Early versus late PN & AKI

Early PN:

No major impact on incidence and recovery of AKIprolonged stage 2 AKI?

Inefficient to reverse the negative nitrogen balanceIncreased ureagenesis

prolonged duration of renal replacement therapy? (as supported by multiple regression-data not shown)

Summary

Future perspectives

Open perspectives for therapies that

activate autophagy in critical illness,

to stimulate damage removal,

especially in combination with therapies that are able to

effectively suppress excessive catabolism of healthy, lean tissue

Acknowledgements

Department and Laboratory of Intensive Care Medicine

Greet Van den Berghe

Ilse Vanhorebeek Miet Schetz

Eric-Jan Ververs Annelies Aertgeerts Inge Derese

Sarah Derde Andy Wauters Pieter Wouters

Michaël Casaer Greet Hermans Fabian Güiza

Björn Ellger Yves Debaveye Magaly Boussemaere

all colleagues from the Laboratory of Intensive Care Medicine, KU Leuven

clinical staff and research nurses Intensive Care Medicine, University Hosp. Leuven

External collaborators

Wim Martinet (Univ. Antwerp) Jasperina Dubois (Jessa Hosp. Hasselt)

Christophe Magnes (Univ. Graz) Kathleen Claes (KU Leuven)

Evelyne Lerut (KU Leuven)

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