The document presents research on the secretion and regulation of apolipoprotein B48 (ApoB48) by primary hamster intestinal enterocytes. Studies show increased intracellular stability and oversecretion of ApoB48 from fructose-fed hamster enterocytes. The results suggest that hyperinsulinemia and increased de novo lipogenesis in the intestine may enhance ApoB48 secretion and contribute to elevated triglyceride levels.

1. Secretion and Regulation of ApoB48 by
Primary Hamster Intestinal EnterocytesIntestinal Enterocytes
B C
LabeledApoB48(%control)
LabeledApoB48(%control)
0
20
40
60
80
100
120
Total Cells Media
Control
MG132
P < 0.05
0
20
40
60
80
100
120
Total Cells Media
Control
Oleate
P = 0.01
Cells Media
0 45 90 45 90
A
Chase Time (min)

2. Cells Media
Fructose-Fed
Chow-Fed
Chase Time (Min)
0 45 90 45 90
D
Chase Time (Min) Chase Time (Min)
100
100
0
10
20
30
40
50
60
70
80
90
0 20 40 60 80
Total apoB 48
LabeledApoB48(%of0Time)
LabeledApoB48(%of0Time)
p= 0.003 p= 0.001
Secreted apoB 48
0
10
20
30
40
50
60
70
80
90
100
0 20 40 60 80 100
Chow-Fed
Fructose-Fed
E F
Ex vivo evidence for oversecretion of intestinal apoB48
& Enhanced intracellular apoB48 stability in fructose-
fed hamster enterocytes

3. In Vivo Production of TG & Intestinal ApoB48
in the Fasting State
Time (min)
55 60 65 70 75 80 85
Sf>400TGconcentration
(mmol/l)
0
1
2
3
4
Sf >400 after Triton WR1339
Time (min)
55 60 65 70 75 80 85
Sf>400ApoB48concentration
(µg/ml)
140
160
180
200
220
240
260
280
Time (min)
55 60 65 70 75 80 85
Sf100-400TGconcentration
(mmol/l)
0
1
2
3
Sf 100-400 after Triton WR1339
Time (min)
55 60 65 70 75 80 85
Sf100-400ApoB48concentration
(µg/ml)
90
100
110
120
130
140
150
160
170
Triglyceridesecretion
0.0
0.2
0.4
0.6
0 1 2 3
ApoB48secretion
0
4
8
12
16
Triglyceridesecretion
0.0
0.1
0.2
0.3
ApoB48secretion
0
2
4
6p = 0.002 p = 0.01
p = 0.14
p = 0.59
A B
DC

4. In Vivo Production of TG & Intestinal ApoB48
in the Postprandial State
Time (min)
55 60 65 70 75 80 85
Triglyceridelevel(mmol/l)
0.0
0.1
0.2
0.3
0.4
0.5
0.6
Time (min)
55 60 65 70 75 80 85
ApoB48level(µg/ml)
80
90
100
110
120
130
140
150
Time (min)
55 60 65 70 75 80 85
Triglyceridelevel(mmol/l)
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
Time (min)
55 60 65 70 75 80 85
ApoB48level(µg/ml)
50
60
70
80
90
100
Triglyceridesecretion(µmol/min)
0.00
0.01
0.02
0.03
0.04
0.05
0.06
0.07
Col 48: 0.07
Col 48: 0.06
Triglyceridesecretion(µmol/min)
0.00
0.02
0.04
0.06
0.08
0.10
Col 50: 0.08
Col 50: 0.09
0 1 2 3
ApoB48secretion(µg/min)
0
1
2
3
4
5
6
7
8
Col 52: 2.65
Col 52: 7.46
ApoB48secretion(µg/min)
0
1
2
3
4
5
6
7
8
Col 54: 3.85
Col 54: 7.26
Sf 100-400 after Triton WR1339
Sf >400 after Triton WR1339
A B
C D
p = 0.3 p = 0.003
p = 0.8 p = 0.007

5. 0
20
40
60
80
100
120
140
160
Fructose-Fed
MTPProteinMass
(%ofControl)
Chow-Fed Fructose-Fed
0
20
40
60
80
100
120
0 0.5 1 1.5 2 2.5 3 3.5
MTP inhibitor (µM)
ApoB48Secretion(%ofcontrol)
Fructose-fed
Chow-fed
∗
∗ ∗ ∗
Chow-Fed
p=0.006D
E
LabeledCholesterylEster
(%ofControl)
p= 0.002
0
50
100
150
200
250
Cellular
p=0.001
Media
B
LabeledCholesterol
(%ofControl)
Chow-Fed
Fructose-Fed
0
100
200
300
400
500
600
700
800
Cellular
p=0.0018
p=0.0013A
Media
LabeledTriglyceride
(%ofControl)
Cellular
0
20
40
60
80
100
120
140
160
180
p=0.04
p=0.003
Media
C
Ex Vivo Evidence for Intestinal Overproduction of
Lipoprotein Lipids and Increase MTP mass/activity

6. Fructose-Fed
Chow-fed
0
10
20
30
40
50
60
70
80
Large
CM
Small
CM
VLDL LDL HDL
p<0.05
p= 0.06
p<0.03
%ofTotalLabeledApoB48 A
0
20
40
60
80
100
120
140
CM apoB48 Total-apoB48
LabeledApoB48
(%ofFructose-fed)
p<0.0007
p<0.01C
0
10
20
30
40
50
60
70
80
Particles< 1.006 Particles > 1.006
%ofTotalLabeledAPoB48
p<0.024
p<0.026
B
0
40
20
60
80
100
120
CM apoB48 Total apoB48
LabeledApoB48
(%ofFructose-fed)
P=0.008 P=0.0001
DFasting Postprandial
Density/Size Distribution of Intestinal Lipoproteins
Evidence for Increased Number & Size

7. Increased De Novo Lipogenesis in FF Enterocytes
Evidence that ApoB48 Secretion is Linked to
the Rate of De Novo Lipogenesis
0
10
20
30
40
50
60
70
80
90
100
110
0 2 4 6 8 10 12 14 16
Cholesterol
Fatty Acid
Triglyceride
LabeledLipid(%ascontrol)
Cerulenin µg/ml
0
50
100
150
200
250
300
350
Chow-fed Fructose-fed
LabeledFattyAcid(%ascontrol)
0
10
20
30
40
50
60
70
80
90
100
110
0 2 4 6 8 10 12 14 16
Cerulenin µg/ml
LabeledSecretedApoB-48(%ascontrol)
A
C
p= 0.002
Sensitivity of ApoB48 Secretion
to Cerulenin
Fatty Acid
Synthesis
Sensitivity of TG & FA Secretion
to Cerulenin

8. 20
60
80
100
0
40
120
Chow-fed Fructose-fed
(2 days)
0
20
40
60
80
100
120
Control Fructose
(3mM )
0
200
400
600
800
1000
enterocytes hepatocytes
P < 0.01
LabeledApoB-48(%ofcontrol)
LabeledApoB-48(%ofcontrol)
Labeledcholesterol
0
50
100
150
200
250
300
350
400
450
enterocytes hepatocytes
LabeledTriglyceride
P < 0.0001
Acute Fructose Feeding or Exposure Does NOT Affect
Intestinal ApoB48 Secretion
[14
C] Fructose
Incorporation
into TG &
Cholesterol
Two-day Fructose
Feeding In Vitro Incubation
of Enterocytes with
Fructose

9. Mechanisms of Intestinal Lipoprotein
Overproduction in Insulin Resistant/
Hyperinsulinemic States
Apical Surface
Basolateral Surface
FA
monoglyceride
FABP-FA
FABP
TG
TG
TG
ApoB48
PL
CE
De Novo (FA)
Insulin (??)
TGdietary
SOS
Grb-2
IRS
PI3-K
p110
SHP-2
NCK
PTKPTK
β β
α α
IR
PKB (Akt)
SREBP1c
FAS & ACC
Glucose
+
FA
MTP
ERER
GolgiGolgi
Intestinal
Enterocyte
TG
ApoB48 Lipoprotein
Particles
p85Proteasome
Degradation

10. Acknowledgements
Laboratory Group:
Changiz Taghibiglou
Mehran Haidari
Steven Van Iderstine
Wei Qui
Taryne Chong
Farhana Mahboob
Biao Chen
Leyla Mangaloglu
Louisa Pontrelli
Fariborz Rashid
Rita Kohen
Debbie Rudy
Collaborators:
Gary Lewis- Toronto
Andre Carpentier- Sherbrooke
Sven Olof Olofsson - Sweden
Janet Sparks - Rochester, NY
Raphael Cheung – Windsor
Michel Tremblay – Montreal
Denny Trinh - Toronto
Funding:
Heart & Stroke Foundation of Ontario
Canadian Institutes of Health Research
NSERC Hospital for Sick Children
Merck-Frosst pfizer
GlaxoSmithKline
BBDC, University of Toronto