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Sodium Butyrate increases hepatic lipid accumulation

Hyun jae Sung, Dong Yeol Kim, Jiwon Park, Sang-min kim, Quynh Nguyen Thi Nhu, Duong Tran Thi Thuv, An Bui Ngan, Inn-Oc Han
Program in Biomedical Science and Engineering (BMSE) Inha University, 100 Inha Ro, Michuholgu, Incheon 22212, South Korea

Abstract 3.5
3
Short Chain Fatty Acids (SCFAs) including butyrate are produced from dietary fiber by gut microbiota. 2.5
Butyrate has been shown that reduce lipid accumulation in human hepatocytes in high fat conditions. 2
However, it still remains unclear the mechanism by which butyrate regulates hepatic lipid accumulation. In ORO/Protein 1.5
the current study, we newly discovered that sodium butyrate (NaB) induces lipid accumulation in Hep G2 0.5 1
cells in dose dependent manner under normal glucose conditions. Other types of SCFA such as acetate 0
5 10
1
0
1
5 10 0
uM μM μM μM μM μM μM μM
or propionate did not induce lipid accumulation in Hep G2 cells. Since NaB is a histone deacetylase 5.5 mM 33 mM
(HDAC) inhibitor, we examined the effect of another well known HDAC inhibitor, Trichostatin A (TSA) for
lipid accumulation in Hep G2 cells. TSA also induced lipid accumulation in Hep G2 cells in dose Figure 3. Effects of serial concentration of trichostatin A (TSA) on fat accumulation in HepG2 cell
dependent manner. Next we examined in vivo effect of NaB using adult Zebrafish model. NaB resulted in
NaB TSA 4
lipid accumulation in the liver of zebrafish under the normal diet condition. Both NaB and TSA increased
3.5
protein and mRNA expression of sterol regulatory element-binding protein 1c (SREBP-1c) and fatty acid
0 mM 2 mM 4 mM 0 μM 5 μM 10 μM 3
synthase (FAS), a key markers related to fat accumulation, in Hep G2 cells in a dose dependent manner. 2.5
Our study collectively indicates that NaB increased hepatic lipid accumulation under normal diet H3 H3K9ac / H3 (fold) 2
conditions potentially through HDAC pathway 1.5 1
0.5
H3K9ac 0
Control 2 mM 4 mM Control 5 μM 10 μM
NaB TSA
Materials and Methods Figure 4. HDAC inhibitors increase Histone protein acetylation level in HepG2 cell
.
● HepG2 cell ORO staining ● Zebrafish NaB IP injection NaB TSA NaB TSA
Western Blot RT-PCR
0 mM 2 mM 4 mM 0 μM 5 μM 10 μM 0 mM 2 mM 4 mM 0 μM 5 μM 10 μM
β-actin β-actin
SREBP-1c SREBP-1c
FAS
4.5 1.6 1
NaB (1 mg/g, 2.5 mg/g)
4 1.4 0.9
3.5 3 1.2 0.8
SREBP-1c/ β-actin (fold) 1.5 1 SREBP-1c/ β-actin (fold) 0.6 0.4 0.4 0.3
Results 2.5 2 0.8 1 FAS/ β-actin (fold) FAS/ β-actin (fold) 0.7 0.6 0.5
0 0.5 0.2 0 0.2 0.1 0
Control 2 mM 4 mM Control 5 uM 10 uM 0 mM 2 mM 4 mM 0 μM 5 μM 10 μM Control 2 mM 4 mM Control 5 uM 10 uM
2.5 2 NaB TSA NaB TSA NaB TSA
ORO/Protein 1.5 1 Figure 5. HDAC inhibitors increase SREBP-1c,FAS mRNA and protein level in HepG2 cell
0.5
0
0 1 2 4 0 1 2 4 14
NaB mM mM mM mM mM mM mM mM
12
Glucose 5.5 mM 33 mM 10
Figure 1. Effects of serial concentration of sodium butyrate (NaB) on fat accumulation in HepG2 cell ORO area (%) 8 6
2 4
0
Control 1 g/kg 2.5 g/kg 5 g/kg
2.5 2 Figure 6. Effects of serial concentration of sodium butyrate (NaB) on fat accumulation in zebra fish
ORO/Protein 1.5 1
0.5
0 Conclusion
0 0.1 1 2 0 0.1 1 2
NaP mM mM mM mM mM mM mM mM
Glucose 5.5 mM 33 mM
- NaB induces lipid accumulation in HepG2
cells and zebrafish liver.
- NaB induced fat accumulation by acting
Sodium butyrate
as HDACi, not as SCFA.
1.6
1.4 1.2 - It was confirmed that SREBP-1c, a
ORO/Protein 0.8 0.6 0.4 1 NaB and TSA were treated.
lepogenesis marker, was affected when
0 0.2 lipogenesis marker - Further studies are needed to determine
0 0.1 1 2 0 0.1 1 2 (SREBP-1c, FAS, SCD-1…) which protein transcription was affected
NaA mM mM mM mM mM mM mM mM transcription↑ by NaB as HDACi.
Glucose 5.5 mM 33 mM
Figure 2. Effects of serial concentration of sodium propionate (NaP) & sodium acetate (NaA)
on fat accumulation in HepG2 cell
Hepatic lipid accumulation ↑