Science - USA (2020-02-07)

cytokine, which is consistent with similar macro-
phage infiltration in Flox and LAKO mice. Like-
wise,LAKOmiceshowednodifferenceingrowth
factorPdgfaand receptorPdgfraexpression

or matrix remodeling genes tissue inhibitor

matrix metalloproteinase 1 (Timp1) and dis-

coidin domain receptor tyrosine kinase 2 (Ddr2)

(Fig. 1P).

We also fed mice with the Amylin (AMLN)

diet, which is used to mimic human NASH in

preclinical studies ( 2 , 17 ). AMLN also repressed

AMPK activation in C57BL/6J mice (fig. S2A).

Zhaoet al.,Science 367 , 652–660 (2020) 7 February 2020 2of9

A

D

I

O

B

E

J L

P

C

FGH

K

M N

ND CD-HFD

pAMPKαThr^172

AMPKα

Flox LAKO

0

5

10

15

20

25

30

Body weight (g)

Flox LAKO

0

100

200

300

400

ALT (U/L)

• 
  

Flox LAKO

100

120

140

160

180

200

ALP (U/L)

• 
  

Flox LAKO

0

1

2

3

Liver weight (g)

Flox LAKO

100

150

200

250

300

TG (mg/g liver)

Flox LAKO

0

100

200

300

400

500

AST (U/L)

• 
  

H&E F4/80 Sirius Red

Flox

LAKO

Flox

LAKO

TUNEL DAPI Merge

Green pMLKL

Blue DAPI

Flox LAKO

0

500

1000

1500

Hydroxyproline

(ug/g liver)

• 
  

Flox LAKO

0

5

10

15

Fibrosis (% area)

• 
  

Flox LAKO

0

200

400

600

TUNEL

+ cell/field

• 
  

Prkaa1 Prkaa2

0.0

0.5

1.0

1.5

mRNA (A.U.)

*

*

Flox

LAKO

Tnfa Ccl2 Ccr2 Il1b Adgre1

0.0

0.5

1.0

1.5

2.0

2.5

mRNA (A.U.)

Flox

LAKO

Casp3 Casp8Ripk1 Ripk3

0.0

0.5

1.0

1.5

2.0

mRNA (A.U.)

Flox

LAKO

Tgfb Timp1 Col1a1 Col3a1 Acta2 Pdgfa Pdgfb Pdgfra Ddr2

0

1

2

3

4

5

mRNA (A.U.)

• 
  

• 
  

• 
  
  
  
  • 
    
  
  
  
  

Flox

LAKO

Fig. 1. Liver-specific AMPK knockout exaggerates liver damage in NASH.
(A) Expression ofPrkaa1andPrkaa2in liver;n= 8 to 9 mice. A.U., arbitrary
units. (B) Immunoblot of liver lysate from ND or CD-HFD–fed mice;n= 3 mice.
(CtoP) Flox and LAKO mice fed CD-HFD for 11 weeks. (C) Body weight. (D) liver
weight. (E) Liver TG. [(F) to (H)] Serum ALT (F), AST (G), ALP (H);n=8to
9 mice. (I) Liver sections stained TUNEL (red) and 4′,6-diamidino-2-phenylindole
(DAPI) (blue) or pMLKL (Green). Scale bar, 50mm. (J) Quantification of TUNEL-

positive nuclei per field in (I);n= 7 mice. (K) H&E, F4/80, and Sirius red staining

of liver sections. Scale bar, 100mm;n= 7 mice. (L) Quantification of fibrosis area

(percent of total area) shown in (K);n= 7 mice. (M) Liver hydroxyproline;n=8to

9 mice. (N) Expression ofTnfa,Ccl2,Ccr2,Il1b,andAdgre1in liver;n=8to

9 mice. (O) Expression ofCasp3,Casp8,Ripk1,andRipk3in liver;n=8to9mice.

(P) Expression ofTgfb,Timp1,Col1a1,Col3a1,Acta2,Pdgfa,Pdgfb,Pdgfra,and

Ddr2in liver;n= 8 to 9 mice. Mean ± SEM; *P< 0.05, Student’sunpairedttest.

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