Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Nov 13;13(11):802.
doi: 10.3390/toxins13110802.

Ochratoxin A Induces Steatosis via PPARγ-CD36 Axis

Qian-Wen Zheng  1   2 , Xu-Fen Ding  1 , Hui-Jun Cao  1 , Qian-Zhi Ni  1 , Bing Zhu  1 , Ning Ma  1 , Feng-Kun Zhang  1 , Yi-Kang Wang  1 , Sheng Xu  1 , Tian-Wei Chen  1 , Ji Xia  1 , Xiao-Song Qiu  1   2 , Dian-Zhen Yu  1 , Dong Xie  1   2   3 , Jing-Jing Li  1
Affiliations

Ochratoxin A Induces Steatosis via PPARγ-CD36 Axis

Qian-Wen Zheng et al. Toxins (Basel). .

Abstract

Ochratoxin A(OTA) is considered to be one of the most important contaminants of food and feed worldwide. The liver is one of key target organs for OTA to exert its toxic effects. Due to current lifestyle and diet, nonalcoholic fatty liver disease (NAFLD) has been the most common liver disease. To examine the potential effect of OTA on hepatic lipid metabolism and NAFLD, C57BL/6 male mice received 1 mg/kg OTA by gavage daily. Compared with controls, OTA increased lipid deposition and TG accumulation in mouse livers. In vitro OTA treatment also promoted lipid droplets accumulation in primary hepatocytes and HepG2 cells. Mechanistically, OTA prevented PPARγ degradation by reducing the interaction between PPARγ and its E3 ligase SIAH2, which led to activation of PPARγ signaling pathway. Furthermore, downregulation or inhibition of CD36, a known of PPARγ, alleviated OTA-induced lipid droplets deposition and TG accumulation. Therefore, OTA induces hepatic steatosis via PPARγ-CD36 axis, suggesting that OTA has an impact on liver lipid metabolism and may contribute to the development of metabolic diseases.

Keywords: OTA; PPAR; fatty liver disease; lipid metabolism.

PubMed Disclaimer

Conflict of interest statement

None of the authors participated in this study have anything to disclose regarding conflict of interest.

Figures

Figure 1
Figure 1
Effects of OTA on lipid accumulation in HepG2 cells. HepG2 cells are treated with OTA at the concentrations of 5, 10 and 15 μM for 24 h, then intracellular lipid droplets are labeled with BODIPY (a) and TG contents are determined (c). HepG2 cells are treated with OTA at concentrations of 10 μM for 6, 12, 24 and 48 h, then intracellular lipid droplets are labeled with BODIPY (b) and TG contents are determined (d). Data shown as the mean ± S.E.M. * p < 0.05 and ** p < 0.01, n = 6 biological replicates.
Figure 2
Figure 2
Effects of OTA on lipid accumulation in primary hepatocytes. (a) Primary hepatocytes are treated with OTA at the concentrations of 5, 10 and 15 μM for 24 h, then intracellular lipid droplets are labeled with BODIPY (a) and TG contents are determined (c). Primary hepatocytes are treated with OTA at concentrations of 10 μM for 6, 12, 24 and 48 h, then intracellular lipid droplets are labeled with BODIPY (b) and TG contents are determined (d). Data shown as the mean ± S.E.M. * p < 0.05, n = 6 biological replicates.
Figure 3
Figure 3
OTA induces simple steatosis in vivo. Mice are treated with OTA or NaHCO3 for 12 weeks (n = 12 in each group). Effects of OTA on mouse: body weight (a), liver weight (b), representative images of livers (c), H&E and Oil Red O staining of liver sections of control and OTA-treated mice (d), liver TG contents and serum TG levels (e), serum ALT levels and AST levels (f). Data shown as the mean ± S.E.M. * p < 0.05 and ** p < 0.01, ns means no significant difference.
Figure 4
Figure 4
Reprogramming of hepatic gene expression after OTA treatment. (a) Heatmap for clustering analysis of genes in the livers of control and OTA-treated mice. (b) KEGG analysis showing top 15 enriched pathways in OTA-treated livers with p < 0.05. (c) Results of GSEA showed PPAR signaling pathway were differentially enriched upon OTA treatment. (d) Heatmaps of hepatic RNA-seq raw gene counts for PPAR signaling pathway. (e) Examination of top upregulated genes associated with lipid metabolism in livers of control and OTA-treated mice (n = 12 in each group) by qPCR. (f) Examination of top upregulated genes associated with lipid metabolism in HepG2 cells by qPCR (n = 6 biological replicates). Data shown as the mean ± S.E.M. * p < 0.05, ** p < 0.01 and *** p < 0.001.
Figure 5
Figure 5
OTA induces hepatic steatosis through upregulation and activation of PPARγ. (a) Western blot analysis of PPARγ expression in livers of control and OTA-treated mice. (b) Immunohistochemical staining of PPARγ in liver tissues of control and OTA-treated mice. (c) PPARγ protein expression in control and OTA-treated HepG2 cells. (d) Western blot analysis showing effect of OTA on PPARγ expression in cytosolic and nuclear fractions of HepG2 cells. (e) Expression and localization of PPARγ under indicated treatment is examined by immunofluorescence. Nuclei were stained with DAPI, and (f) intracellular lipid droplets are labeled with BODIPY.
Figure 6
Figure 6
OTA regulates PPARγ by post-translational modification. (a) PPARγ mRNA expression in livers of control and OTA-treated mice (n = 12 in each group). (b) Influence of OTA on the ubiquitination of PPARγ is examined in HEK293T and HepG2 cells. (c) PPARγ expression after OTA and cycloheximide (CHX) treatment. (d) Western blot analysis of SIAH2 expression in livers of control and OTA-treated mice. (e) Endogenous interaction between PPARγ and SIAH2 is examined by immunoprecipitation with SIAH2 antibody in control and OTA-treated HepG2 cells. (f) HepG2 cells are treated with OTA at the concentrations of 5, 10 and 15 μM for 24 h, and endogenous interaction between PPARγ and SIAH2 is examined by immunoprecipitation with SIAH2 antibody. Data shown as the mean ± S.E.M. ns means no significant difference.
Figure 7
Figure 7
OTA-induced hepatic steatosis is CD36-dependent. (a) Western blot analysis of CD36 expression in HepG2 cells under indicated treatment. (b) Western blot analysis of CD36 expression in livers of control and OTA-treated mice. (c) Immunohistochemical staining of CD36 in liver tissues of control and OTA-treated mice. (d) Knockdown efficiency of CD36 in HepG2 cells. (e) BODIPY staining of lipid droplets in control and CD36-knockdown HepG2 cells treated with DMSO and OTA. (f) Left, TG contents in control and CD36-knockdown HepG2 cells treated with DMSO and OTA. Right, TG contents in HepG2 cells under indicated treatment (n = 6 biological replicates). Data shown as the mean ± S.E.M. * p <0.05 vs. shControl DMSO treatment; and # p < 0.05 vs. shControl OTA treatment.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Ringot D., Chango A., Schneider Y.-J., Larondelle Y. Toxicokinetics and toxicodynamics of ochratoxin A, an update. Chem. Biol. Interact. 2006;159:18–46. doi: 10.1016/j.cbi.2005.10.106. - DOI - PubMed
    1. Hajok I., Kowalska A., Piekut A., Ćwieląg-Drabek M. A risk assessment of dietary exposure to ochratoxin A for the Polish population. Food Chem. 2019;284:264–269. doi: 10.1016/j.foodchem.2019.01.101. - DOI - PubMed
    1. Clarke R., Connolly L., Frizzell C., Elliott C.T. Challenging conventional risk assessment with respect to human exposure to multiple food contaminants in food: A case study using maize. Toxicol. Lett. 2015;238:54–64. doi: 10.1016/j.toxlet.2015.07.006. - DOI - PubMed
    1. Bui-Klimke T.R., Wu F. Ochratoxin A and Human Health Risk: A Review of the Evidence. Crit. Rev. Food Sci. Nutr. 2015;55:1860–1869. doi: 10.1080/10408398.2012.724480. - DOI - PMC - PubMed
    1. Damiano S., Iovane V., Squillacioti C., Mirabella N., Prisco F., Ariano A., Amenta M., Giordano A., Florio S., Ciarcia R. Red orange and lemon extract prevents the renal toxicity induced by ochratoxin A in rats. J. Cell. Physiol. 2020;235:5386–5393. doi: 10.1002/jcp.29425. - DOI - PubMed

Publication types

MeSH terms