Secretion and Regulation of ApoB48 by

1
Secretion and Regulation of ApoB48 by Primary
Hamster Intestinal Enterocytes
Media
A
Cells
Chase Time (min)
45
90
0
45
90
B
C
P lt 0.05
P 0.01
Control
Control
MG132
Oleate
Labeled ApoB48 ( control)
Labeled ApoB48 ( control)
Media
Total
Cells
Total
Cells
Media
2
Ex vivo evidence for oversecretion of intestinal
apoB48 Enhanced intracellular apoB48 stability
in fructose-fed hamster enterocytes
Media
Cells
D
Fructose-Fed
Chow-Fed
Chase Time (Min)
0
45
90
45
90
E
F
Total apoB 48
Secreted apoB 48
100
90
80
Labeled ApoB 48( of 0 Time)
70
p 0.003
Labeled ApoB 48( of 0 Time)
p 0.001
60
Chow-Fed
50
Fructose-Fed
40
30
20
10
0
0
20
40
60
80
0
20
40
60
80
100
100
Chase Time (Min)
Chase Time (Min)
3
In Vivo Production of TG Intestinal ApoB48 in
the Fasting State
4
In Vivo Production of TG Intestinal ApoB48 in
the Postprandial State
5
Ex Vivo Evidence for Intestinal Overproduction of
Lipoprotein Lipids and Increase MTP mass/activity
p0.04
Labeled Cholesterol ( of Control)
p0.0013
A
C
B
p0.003
800
Labeled Triglyceride ( of Control)
Labeled Cholesteryl Ester ( of Control)
p 0.002
250
p0.001
700
Chow-Fed
200
600
Fructose-Fed
500
p0.0018
150
400
300
100
200
50
100
0
0
Media
Media
Media
Cellular
Cellular
Cellular
E
Fructose-fed Chow-fed
Chow-Fed
Fructose-Fed
?
D
?
p0.006
?
?
160
ApoB 48 Secretion ( of control)
140
120
100
MTP Protein Mass ( of Control)
80
60
40
20
0
0.5
1
1.5
2
2.5
3
3.5
0
MTP inhibitor (?M)
Fructose-Fed
Chow-Fed
6
Density/Size Distribution of Intestinal
Lipoproteins Evidence for Increased Number Size
A
B
Chow-fed
p 0.06
80
plt0.026
Fructose-Fed
70
60
50
plt0.024
of Total Labeled APoB48
40
of Total Labeled ApoB48
plt0.05
30
20
plt0.03
10
0
Large CM
Small CM
VLDL
LDL
HDL
Particleslt 1.006
Particles gt 1.006
140
Fasting
Postprandial
C
D
plt0.01
120
plt0.0007
P0.0001
P0.008
100
80
Labeled ApoB48 ( of Fructose-fed)
Labeled ApoB48 ( of Fructose-fed)
60
40
40
20
0
0
Total apoB48
CM apoB48
Total-apoB48
CM apoB48
7
Increased De Novo Lipogenesis in FF
Enterocytes Evidence that ApoB48 Secretion is
Linked to the Rate of De Novo Lipogenesis
A
p 0.002
Fatty Acid Synthesis
Labeled Fatty Acid ( as control)
Labeled Lipid ( as control)
Sensitivity of TG FA Secretion to Cerulenin
Cholesterol
Fatty Acid
Triglyceride
0
2
4
6
8
10
12
14
16
Chow-fed
Fructose-fed
Cerulenin ?g/ml
C
Labeled Secreted ApoB-48 ( as control)
Sensitivity of ApoB48 Secretion to Cerulenin
0
2
4
6
8
10
12
14
16
Cerulenin ?g/ml
8
Acute Fructose Feeding or Exposure Does NOT
Affect Intestinal ApoB48 Secretion
Two-day Fructose Feeding
In Vitro Incubation of Enterocytes with Fructose
120
120
100
100
80
80
Labeled ApoB-48 ( of control)
Labeled ApoB-48 ( of control)
60
60
40
40
20
20
0
0
Chow-fed
Fructose-fed (2 days)
Control
Fructose (3mM )
450
P lt 0.01
P lt 0.0001
400
350
14C Fructose Incorporation into TG
Cholesterol
300
Labeled cholesterol
Labeled Triglyceride
250
200
150
100
50
0
hepatocytes
enterocytes
hepatocytes
enterocytes
9
Mechanisms of Intestinal Lipoprotein
Overproduction in Insulin Resistant/ Hyperinsulin
emic States
TG dietary
monoglyceride
Apical Surface
Intestinal Enterocyte
FA
De Novo (FA)
FABP
TG
Glucose

FABP-FA
FA
FAS ACC
ER
TG
TG
SREBP1c
MTP
PL
Golgi
PKB (Akt)
TG
CE
PI3-K
ApoB48
SOS
IRS
Proteasome
p110
Grb-2
Degradation
SHP-2
NCK
Basolateral Surface
PTK
PTK
ApoB48 Lipoprotein Particles
Insulin (??)
10
Acknowledgements
CollaboratorsGary Lewis- TorontoAndre
Carpentier- SherbrookeSven Olof Olofsson -
Sweden Janet Sparks - Rochester, NY Raphael
Cheung Windsor Michel Tremblay Montreal Denny
Trinh - Toronto
Laboratory Group Changiz Taghibiglou Mehran
Haidari Steven Van Iderstine Wei QuiTaryne
Chong Farhana Mahboob Biao ChenLeyla
MangalogluLouisa Pontrelli Fariborz Rashid Rita
Kohen Debbie Rudy
Funding Heart Stroke Foundation of
Ontario Canadian Institutes of Health
Research NSERC Hospital for Sick
Children Merck-Frosst
pfizer GlaxoSmithKline BBDC, University of
Toronto