Reactive Oxygen Species Induces Lipid Droplet Accumulation in HepG2 Cells by Increasing Perilipin 2 Expression
Abstract
:1. Introduction
2. Results
2.1. Exogenous Addition of Hydrogen Peroxide Promotes the Formation of Lipid Droplets in Cells
2.2. PLIN2 Expression Level Increased after Treatment by Hydrogen Peroxide
2.3. PLIN2 Overexpression Promotes Lipid Droplets Formation in Cells
2.4. PLIN2 Interference Inhibits Hydrogen Peroxide-Induced Lipid Droplet Accumulation
2.5. Hydrogen Peroxide Treatment Promotes the Formation of Lipid Droplets through Up-Regulating PLIN2
2.6. Exogenous Addition of Hydrogen Peroxide Promotes the Formation of Lipid Droplets in Murine Hepatocytes
2.7. Lipid Droplet-Related Proteins Enhanced Expression in Steatotic Liver Tissue
2.8. PLIN2 Modulates Lipid Droplet Formation in Cells via PPAR (Peroxisome Proliferator-Activated Receptor) and CREBBP (CREB Binding Protein) Signaling Pathways
3. Discussion and Conclusions
4. Methods and Materials
4.1. Animals and Cell Lines
4.2. Antibodies
4.3. Plasmid DNA Construction
4.4. Cell Culture and Transfection
4.5. Hydrogen Peroxide Treatment
4.6. Hematoxylin-Eosin Staining and Oil Red O Staining of Histological Sections
4.7. Lipid Droplets Marking and Observation
4.8. Western Blot and Real-Time PCR
4.9. Immunofluorescence Assay
4.10. Intraperitoneal Injection of Hydrogen Peroxide in Mice
4.11. Immunohistochemistry Assay
4.12. Bioinformatics and Data Analysis
4.13. Statistical Analyses
Authors Contribution
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
ROS | reactive oxygen species |
NAFLD | non-alcoholic fatty liver disease |
NASH | non-alcoholic steatohepatitis |
PLIN1 | perilipin1 |
PLIN2 | perilipin2 |
PLIN3 | perilipin3 |
PLIN4 | perilipin4 |
PLIN5 | perilipin5 |
FSP27/CIDEC | cell death-inducing DFFA-like effector c |
SREBF1 | sterol regulatory element binding transcription factor 1 |
SREBP1c | sterol regulatory element binding protein 1c |
PPAR | peroxisome proliferator activated receptor alpha |
RXRA | retinoid X receptor alpha |
CREB | cAMP responsive element binding protein |
CREBBP | CREB binding protein |
SOD1 | superoxide dismutase 1 |
JNK/MAPK8 | mitogen-activated protein kinase 8 |
AMPK | adenosine 5‘-monophosphate (AMP)-activated protein kinase |
OA | oleic acid |
CITED2 | Cbp/P300 interacting transactivator with Glu/Asp rich carboxy-terminal domain 2 |
NCOA1 | nuclear receptor coactivator 1 |
NCOA2 | nuclear receptor coactivator 2 |
HIF1 | hypoxia inducible factor 1 |
HIF1AN | hypoxia inducible factor 1 alpha subunit inhibitor |
TP53 | tumor protein p53 |
ARNT | aryl hydrocarbon receptor nuclear translocator |
VEGFA | vascular endothelial growth factor A |
EPAS1 | endothelial PAS domain protein 1 |
AR | androgen receptor |
MED1 | mediator complex subunit 1 |
FABP1 | fatty acid binding protein 1 |
HELZ2 | helicase with zinc finger 2 |
NR3C | nuclear receptor subfamily 3 group C member |
SEIPIN/BSCL2 | seipin lipid droplet biogenesis associated |
FITM1 | fat storage inducing transmembrane protein 1 |
FITM2 | fat storage inducing transmembrane protein 2 |
ATGL/PNPLA2 | patatin like phospholipase domain containing 2 |
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Gene Symbol | Primer Sequence 5′-3′ | Product Size (bp) | Accession |
---|---|---|---|
PLIN1 (perilipin1) | Forward: TGGGTGGTGTGGCACATAC | 143 | XM_005254934.4 |
Reverse: CCTCCCCTTGGTTGAGGAGA | |||
PLIN2 (perilipin2) | Forward: TTGCAGTTGCCAATACCTATGC | 148 | XM_017014259.2 |
Reverse: CCAGTCACAGTAGTCGTCACA | |||
PLIN3 (perilipin3) | Forward: GCCCAAGAGATGGTGTCTAGC | 119 | NM_005817.4 |
Reverse: CCGGTCACTACGGACTTTGT | |||
PLIN4 (perilipin4) | Forward: GGAGCTGCAACCTTCGGAAA | 131 | NM_001080400.1 |
Reverse: GGACCACTCCCTTAGCCAC | |||
PLIN5 (perilipin5) | Forward: AAGGCCCTGAAGTGGGTTC | 192 | NM_001013706.2 |
Reverse: GCATGTGGTCTATCAGCTCCA | |||
SREBP1 (sterol regulatory element binding protein 1c) | Forward: ACAGTGACTTCCCTGGCCTAT | 222 | XM_024450893.1 |
Reverse: CATGGACGGGTACATCTTCAA | |||
FSP27 | Forward: ATTGATGTGGCCCGTGTAACG | 122 | NM_001199623.1 |
Reverse: CAGCAGTGCAGATCATAGGAAA | |||
ATGL | Forward: ATGGTGGCATTTCAGACAACC | 89 | NM_020376.3 |
Reverse: CGGACAGATGTCACTCTCGC | |||
CREBBP | Forward: CGGCTCTAGTATCAACCCAGG | 237 | NM_004380.2 |
Reverse: TTTTGTGCTTGCGGATTCAGT | |||
PPARα | Forward: TTCGCAATCCATCGGCGAG | 146 | NM_005036.5 |
Reverse: CCACAGGATAAGTCACCGAGG | |||
GAPDH | Forward: CTGGGCTACACTGAGCACC | 101 | NM_002046.6 |
Reverse: AAGTGGTCGTTGAGGGCAATG |
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Jin, Y.; Tan, Y.; Chen, L.; Liu, Y.; Ren, Z. Reactive Oxygen Species Induces Lipid Droplet Accumulation in HepG2 Cells by Increasing Perilipin 2 Expression. Int. J. Mol. Sci. 2018, 19, 3445. https://doi.org/10.3390/ijms19113445
Jin Y, Tan Y, Chen L, Liu Y, Ren Z. Reactive Oxygen Species Induces Lipid Droplet Accumulation in HepG2 Cells by Increasing Perilipin 2 Expression. International Journal of Molecular Sciences. 2018; 19(11):3445. https://doi.org/10.3390/ijms19113445
Chicago/Turabian StyleJin, Yi, Yanjie Tan, Lupeng Chen, Yan Liu, and Zhuqing Ren. 2018. "Reactive Oxygen Species Induces Lipid Droplet Accumulation in HepG2 Cells by Increasing Perilipin 2 Expression" International Journal of Molecular Sciences 19, no. 11: 3445. https://doi.org/10.3390/ijms19113445