Download Role of Liver In Triglyceride Homeostasis

Role of Liver In Triglyceride
Homeostasis
Larry L. Swift, Ph.D.
Department of Pathology
Vanderbilt University School of Medicine
Metabolic Cycle of Fatty Acids and
Triglycerides
VLDLTG
Fatty Acids
• Aliphatic carboxylic acids usually having an even number
of carbon atoms
• Chains may be saturated or unsaturated
• Position of double bonds described in relation to
carboxy-terminus ( ) or methyl carbon ( or n)
Common Fatty Acids
Fatty Acid
C:DB, position
Stearic
18:0
Oleic
18:1 ω9
Linoleic
18:2 ω6
α-Linolenic
18:3 ω3
γ-Linolenic
18:3 ω6
Arachidonic
20:4 ω6
Eicosapentaenoic 20:5 ω3
Structure
Fatty Acid Biosynthesis
• Takes place in the cytosol
• Acetyl CoA is substrate; palmitate (C16:0) is
product
• Fatty acids are synthesized by a multi-enzyme
complex, fatty acid synthase (FAS)
• Elongation beyond C16:0 and desaturation are
catalyzed by enzymes associated with the
endoplasmic reticulum
Amino Acids
Glucose
Fatty acids
Pyruvate
Pyruvate
Denhydrgenase
Cytosol
Acetyl CoA
Fatty Acids
Mitochondria
Fatty Acid Biosynthesis
•First committed step of fatty acid synthesis pathway
•Acetyl CoA Carboxylase (ACC) is major regulatory
site of fatty acid synthesis
Fatty Acid Biosynthesis
Final product
is palmitic acid,
C16:0
Fatty Acid Synthase
Fatty Acid Elongation / Desaturation
Palmitate (16:0)
elongation
Mammals
and plants
Stearate (18:0)
desaturation
Oleate
18:1 (∆9 or 9)
desaturation
* Plants
only
*
Other PUFAs
Longer saturated
fatty acids
*
Linoleate
18:2 (∆9,12 or 6)
-linoleate
18:3 (∆9,12,15 or 3)
Palmitoleate
16:1 (∆9 or 7)
-linoleate
18:3 (∆6,9,12 or 6)
elongation
Eicosatrienoate
20:3 (∆8,11,14 or 6)
desaturation
Arachidonate
20:4 (∆5,8,11,14 or 6)
Synthesis of Polyunsaturated Fatty Acids
Regulation of Fatty Acid Biosynthesis
Acetyl CoA Carboxylase
• Allosteric regulation – citrate activates and
palmitoyl CoA, stearoyl CoA and arachidyl CoA
inhibit ACC
• Hormonal regulation – glucagon inactivates and
insulin activates ACC via
phosphorylation/dephosphorylation of the
enzyme
Regulation of ACC by Phosphorylation
• Inactivation (starved
state)
– glucagon increases
cAMP
– activates protein
kinase A
– inactivates ACC
• Activation (fed state)
– insulin induces
protein phosphatase and activates
ACC
Regulation of Lipid Biosynthesis via SREBP
Horton et al., J. Clin. Invest. 109: 1125, 2002.
SREBP1c
• Enhances transcription of genes required for FA
synthesis
• Responsive genes include:
–
–
–
–
–
–
–
ATP citrate lyase
Acetyl-CoA carboxylase
Fatty acid synthase
Fatty acid elongase
Stearoyl-CoA desaturase
Glycerol-3-phosphate acyltransferase
Genes required to generate NADPH
Insulin
• Stimulates hepatic FA synthesis during
carbohydrate excess
• Increases SREBP-1c mRNA in liver
• Induction of target genes blocked if dominant
negative form of SREBP-1c is expressed
• Incubating primary hepatocytes with glucagon
decreases mRNAs for SREBP-1c and its associated
target genes
What happens to these newly
synthesized fatty acids?
• Depends on the cell and metabolic state
• Can be stored (but not as fatty acids) or utilized
for energy
• Synthesis of other lipid classes
Triglyceride
• Glycerol backbone with
3 fatty acids (FA)
• Highly concentrated
source of energy, easily
stored in cytosolic
droplets
• Before use as energy
source, it must undergo
lipolysis to release FA
Triglyceride Synthesis
Synthesis of Glycerol Phosphate in Liver
Glucose
Glycolysis
CH2OH Glycerol
CH2OH
CHOH
DHAP C=O
O
CH2-O-P-OO
ATP
ADP
NADH
Glycerol 3-phosphate
dehydrogenase
CH2OH
NAD+
CH2OH
HO-C-H
O
CH2-O-P-OO
L-Glycerol 3-Phosphate
Glycerol Kinase
Biosynthesis of Triglyceride
What Happens to TG Synthesized by the
Liver?
• Energy
• Export as VLDL
• Storage
Lipoprotein Model
Very Low Density Lipoproteins (VLDL)
• Spherical TG-rich particles 40-60 nm in diameter
• Synthesized by the liver and transport triglyceride to
periphery for storage and/or utilization
• Major structural protein is apo B
Apolipoprotein B
• Structural apoprotein for triglyceride-rich
lipoproteins
• Expressed in two forms: apoB100 & apoB48
• B100 is found on VLDL, IDL, and LDL
• B48 is found on chylomicrons
• ApoB100 and B48 synthesis are essential for VLDL
and chylomicron assembly
Apolipoprotein B
VLDL Assembly
• Initiated in endoplasmic reticulum (ER)
• Assembly and secretion are dependent on
the ability of the cell to synthesize apoB and
MTP
• Constitutive process modulated by a variety
of factors
Assembly of Triglyceride-Rich
Lipoproteins
G. Shelness, Wake Forest School of Medicine
Microsomal Triglyceride Transfer Protein
(MTP)
• A heterodimeric protein complex consisting of 97 kDa
subunit and protein disulfide isomerase (PDI)
• Transfers lipid from the ER membrane to the forming
lipoprotein in the ER lumen
• Absolute requirement for assembly of VLDL by the liver
• Inhibition of MTP leads to reduction in plasma
triglycerides and cholesterol
Regulation of VLDL Production
• Availability of functional MTP
• Apo B degradation
• Substrate availability
Functional MTP
• Inhibition of MTP leads to decreased VLDL
secretion and a reduction in plasma TGs
• Over-expression of MTP leads to increased VLDL
secretion
• Absence of MTP (abetalipoproteinemia) leads to
absence of TG-rich lipoproteins in plasma
ApoB Degradation
• Proteasome inhibitors block degradation of apoB
and ubiquitinated apoB accumulates within the
cell
• ALLN, a calpain inhibitor, inhibits degradation of
apoB, but does not increase apoB secretion
• Insulin regulates degradation of apoB through
PI3-kinase pathway
• Insulin resistance leads to decreased apoB
degradation and increased secretion
Assembly of Triglyceride-Rich
Lipoproteins
Davidson and Shelness, Ann. Rev. Nutr. 20: 169, 2000
Targets for Regulation of
VLDL Assembly
G.F. Gibbons, et al., Biochem Soc Trans 32: 59-64, 2004
Hepatic Lipid Mobilization
Gibbons et al., Biochim. Biophys. Acta 1483: 37, 2000
What is the fate of VLDL secreted
from the liver?
VLDL Metabolic Pathway
Chylomicron Metabolic Pathway
Adipocyte Lipid Metabolism
Sethi J.K. , Vidal-Puig A.J., J. Lipid Res. 48:1253-1262, 2007
Sources of Fatty Acids for Liver and
VLDL Triglycerides
Adiels, M. et al. Arterioscler Thromb Vasc Biol 28:1225-1236, 2008.
Why Does Liver Store TG?
• TG accumulates when plasma NEFA exceeds hepatic
disposal via secretion and oxidation
• Neutralizes potential toxicity of fatty acids released
from adipose tissue
• Hepatic TG secretion in post-absorptive state exceeds
hepatic NEFA esterification, suggesting utilization of
hepatic TG stores
Insulin – A Key Regulator of
Hepatic Lipid Metabolism
•
•
•
•
•
Fatty acid flux to the liver
Hepatic de novo lipogenesis
MTP synthesis
ApoB degradation
Stimulates production of lipoprotein
lipase
• Fatty acid oxidation
De Novo Lipogenesis
• Insulin stimulates SREBP-1c expression and
increases cleavage of SREBP-1c to its mature form
which activates production of lipogenic enzymes
• Hepatic IR exists in the pathway regulating
gluconeogenesis, but this does not affect insulin’s
ability to stimulate lipogenesis
• ChREBP is also up-regulated by insulin and works
in synergy with SREBP-1c to promote lipogenesis
MTP Synthesis
• Mttp gene expression is negatively regulated by insulin in
part by MAPK pathway
• Suppression of MTP by insulin also occurs via Forkhead
box 01 (Fox01)
• Insulin induces phosphorylation of Fox01 leading to its
exclusion from the nucleus, resulting in inhibition of MTP
expression
• Fox01 gain of function is associated with enhanced MTP
expression
• Insulin resistance leads to increased MTP expression
Insulin Signaling through Fox01
Kamagate et al. J. Clin. Invest. 118: 234702364, 2008
Degradation of ApoB
• Insulin targets apoB for degradation via a PI3kinase pathway
• Acute increases in insulin decrease hepatic VLDL
and apoB secretion via a post-ER, non-proteasomal
process
• Insulin resistance leads to decreased apoB
degradation and may contribute to overproduction
of VLDL
Hepatic VLDL Assembly and Secretion
Sparks and Sparks, J. Clin. Invest. 118: 2012-2015, 2008
Substrate Sources Driving Assembly of
ApoB-Lipoproteins in IR
Ginsberg et al., Arch. Med. Res. 36: 232, 2005
Insulin, VLDL Secretion, and Fatty Liver
Ginsberg, et al., Endocrin. Metab.
Clinics 35: 491, 2006
Summary
• De novo lipogenesis and the regulation of fatty
acid synthesis
• Sources of fatty acids for liver TG biosynthesis
• Secretion of hepatic TG with VLDL and the fate
of TG-rich particles
• Insulin resistance and its impact on hepatic TG
homeostasis
Effects of Insulin Resistance on Hepatic
TG Homeostasis
• Increased fatty acid flux from adipose tissue to
the liver
• Increased hepatic uptake of VLDL, IDL, and
chylomicron remnants
• Increased de novo lipogenesis
• Decreased degradation of apoB
• Increased activity of MTP
• Overproduction of VLDL
• Hepatic steatosis
Questions?
Increased Fatty Acid Flux from Adipose
Tissue to the Liver
• Insulin stimulates adipocyte fatty acid uptake
and inhibits fatty acid release (HSL) leading to
decreased plasma NEFA
• Insulin resistance leads to increased
mobilization of fatty acids from adipose tissue
and increased plasma NEFA
De Novo Lipogenesis
• Hepatic insulin resistance leads to upregulation of
sterol regulatory element-binding protein-1
(SREBP-1c) and activates lipogenic enzymes
• Carbohydrate response element-binding protein
(ChREBP) regulates expression of key glucoseresponsive genes of lipogenesis
• Synergistic action of SREBP-1c and ChREBP directs
conversion of excess glucose to fatty acids and
enhances esterification
Substrate Sources Driving Assembly of
ApoB-Lipoproteins in IR
Ginsberg et al., Arch. Med. Res. 36: 232, 2005
Endocrine Reviews 20(5): 649-88
Changes in Lipoprotein Metabolism in the
Metabolic Syndrome
Adiels, M. et al. Arterioscler Thromb Vasc Biol 2008;28:1225-1236
Metabolic Syndrome
• Abdominal obesity
• Elevated blood pressure
• Insulin resistance
• Prothrombotic state
• Proinflammatory state
• Atherogenic dyslipidemia
Metabolic Syndrome Dyslipidemia
• Elevated plasma triglycerides
• Decreased HDL cholesterol
• Normal levels of LDL cholesterol carried in small
dense particles
• Lipoprotein alterations contribute to increased risk for
CHD
Lecture Outline
• Lipoproteins, structure, apoproteins,
characteristics
• Source of triglyceride for lipoproteins
• Assembly of triglyceride-rich lipoproteins
• Triglyceride-rich lipoprotein catabolism
• Effects of insulin resistance on triglyceride-rich
lipoprotein production
• VLDL secretion and fatty liver
Hormonal control of adipocyte lipolysis
Biochem. Soc. Trans. (2003) 31, 1120-1124
Apolipoproteins
Lipoprotein
Class
Tissue Source
MW
liver and intestine
30K
17K
45K
intestine
liver
240K
512K
apoAI
apoAII
apoA-IV
HDL
HDL
HDL
apoB48
apoB100
chylomicrons
VLDL, LDL
apoCI
apoCII
apoCIII
HDL
HDL and VLDL
HDL and VLDL
liver
6.6K
8.9K
8.8K
apoE
HDL, LDL, VLDL
liver
34K
Apolipoprotein B
• Structural apoprotein for triglyceride-rich
lipoproteins
• Expressed in two forms: apoB100 & apoB48
• B100 is found on VLDL, IDL, and LDL
• B48 is found on chylomicrons
• ApoB100 and B48 synthesis are essential for
VLDL and chylomicron assembly
Increased MTP Activity
• Mttp gene has insulin response element in the
promoter
• In human liver cells Mttp gene expression is
negatively regulated by insulin through the MAPK
cascade
• Increased MTP mRNA is associated with enhanced
synthesis of VLDL in wild-type animals and in animal
models of insulin resistance
Increased MTP Activity
• Forkhead box 01 (Fox01) has been shown to mediate
inhibitory action of insulin on target gene expression
• Fox01 stimulates hepatic MTP expression; the effect
is counteracted by insulin
• Fox01 gain-of-function is associated with enhanced
MTP expression, augmented hepatic VLDL
production, and elevated plasma TG levels in Fox01
transgenic mice
Essential Fatty Acids (EFA)
• Fatty acids that cannot be synthesized and
must be obtained from the diet
• C18:2 -6 - linoleic acid
• C18:3 -3 - -linolenic acid
• These fatty acids are starting point for
synthesizing longer and more highly
unsaturated fatty acids
Arachidonic Acid
• C20:4 ω6
COOH
CH3
• A precursor for prostaglandins, leukotrienes,
and thromboxanes
Common Fatty Acids
• C14:0 - myristic acid
• C16:0 - palmitic acid
• C18:0 - stearic acid
• C18:1 9 - oleic acid
• C18:2 6 - linoleic acid*
• C18:3 3 - -linolenic acid*; 6 - -linolenic acid
• C20:4 6 - arachidonic acid
• C20:5 3 - eicosapentaenoic acid
• C22:5 3 - docosapentaenoic acid
• C22:6 3 - docosahexaenoic acid
Source of Acetyl CoA
• Derived primarily from pyruvate via
pyruvate dehydrogenase in mitochondria
• Transported into cytosol as citrate, then
regenerated by ATP citrate lyase
• Available for malonyl CoA formation and
fatty acid synthesis
O
R1-C-S-CoA
CH2OH
HO-C-H
O
CH2-O-P-OO
Acyl Transferase
O
R2-C-S-CoA
Acyl Transferase
L-Glycerol 3-Phosphate
CH2-OOCR1
R2COO-C-H
O
CH2-O-P-OO
Phosphatidic Acid
Addition of Fatty Acids to
Form Triglycerides
CH2-OOCR1
R2COO-C-H
O
R3-C-S-CoA
DGAT
Phosphatidate
phosphohydrolase
CH2-OOCR1
R2COO-C-H
CH2OH
CH2OOCR3
Triacylglycerol
Diacylglycerol
Plasma Lipoprotein Classes
Chylomicrons
• TG-rich particles 75 to 200 nm in diameter
• Assembled by enterocytes to transport
dietary fat to periphery for storage or
utilization
• Present in postprandial plasma
• ApoB-48 is the sole B apoprotein (humans
and rodents)
Negative Stain of Lipoproteins
Chylomicrons
LDL
VLDL
HDL
Hepatic TG Storage
• Human liver can store 5-45 mol/g under
normal conditions (6.7-60.3 g/1.5 kg liver)
• Hepatic TG in house musk shrew can
exceed 100 mol/g during starvation
• Livers of mice that overexpress SREBP1a contain >250 mol/g
• Liver accomodates TG in cytosolic
droplets
Adipocyte Lipid Metabolism
Sethi J.K. , Vidal-Puig A.J., J. Lipid Res. 48:1253-1262, 2007