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Effect of miRNA-10b in regulating cellular steatosis level by targeting PPAR-alpha expression, a novel mechanism for the pathogenesis of NAFLD.
J Gastroenterol Hepatol. 2010 Jan; 25(1):156-63.JG

Abstract

BACKGROUND AND AIM

Accumulating evidence supports the effects of miRNA in lipid metabolism, providing a potential linkage between certain miRNA and non-alcoholic fatty liver disease (NAFLD). We aimed to investigate the miRNA expression pattern in a steatotic L02 cell model and explore the function of certain miRNA target pairs.

METHODS

The cell model was established by culturing L02 cells with a high concentration of free fatty acid. Micro-array and stem-loop reverse transcription polymerase chain reaction (RT-PCR) were utilized to detect dysregulated miRNA, whereas computational algorithms were used for target prediction. Real time RT-PCR, Western blot, luciferase activity measurement, and other techniques were employed for target verification.

RESULTS

Seventeen upregulated and 15 downregulated miRNA were found in steatotic L02 cells, while miRNA-10b was proven to regulate the steatosis level. Peroxisome proliferator-activated receptor-alpha (PPAR-alpha) was also found to participate in steatosis, as its protein level was decreased in steatotic L02 cells and its overexpression by transfection into the PPAR-alpha-pcDNA 3.1 vector could partially alleviate steatosis. We further found that PPAR-alpha is the direct target of miRNA-10b as it showed significantly changed protein expression, but a relatively unchanged mRNA level in steatotic L02 cells transfected with pre-miRNA-10b and anti-miRNA-10b. Moreover, the action of miRNA-10b on PPAR-alpha depends on the presence of a single miRNA-10b binding site, as the activity of a luciferase reporter carrying the mutant PPAR-alpha 3'-untranslated region was not reduced by the expression of miRNA-10b.

CONCLUSION

The established miRNA profile of the steatotic L02 cell model and the novel effect of miRNA-10b in regulating hepatocyte steatosis may provide a new explanation of the pathogenesis of NAFLD.

Authors+Show Affiliations

Department of Infectious disease, First Affiliated Hospital, Medical School, Zhejiang University, Zhejiang, China.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

19780876

Citation

Zheng, Lin, et al. "Effect of miRNA-10b in Regulating Cellular Steatosis Level By Targeting PPAR-alpha Expression, a Novel Mechanism for the Pathogenesis of NAFLD." Journal of Gastroenterology and Hepatology, vol. 25, no. 1, 2010, pp. 156-63.
Zheng L, Lv GC, Sheng J, et al. Effect of miRNA-10b in regulating cellular steatosis level by targeting PPAR-alpha expression, a novel mechanism for the pathogenesis of NAFLD. J Gastroenterol Hepatol. 2010;25(1):156-63.
Zheng, L., Lv, G. C., Sheng, J., & Yang, Y. D. (2010). Effect of miRNA-10b in regulating cellular steatosis level by targeting PPAR-alpha expression, a novel mechanism for the pathogenesis of NAFLD. Journal of Gastroenterology and Hepatology, 25(1), 156-63. https://doi.org/10.1111/j.1440-1746.2009.05949.x
Zheng L, et al. Effect of miRNA-10b in Regulating Cellular Steatosis Level By Targeting PPAR-alpha Expression, a Novel Mechanism for the Pathogenesis of NAFLD. J Gastroenterol Hepatol. 2010;25(1):156-63. PubMed PMID: 19780876.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Effect of miRNA-10b in regulating cellular steatosis level by targeting PPAR-alpha expression, a novel mechanism for the pathogenesis of NAFLD. AU - Zheng,Lin, AU - Lv,Guo-cai, AU - Sheng,Jifang, AU - Yang,Yi-da, Y1 - 2009/09/25/ PY - 2009/9/29/entrez PY - 2009/9/29/pubmed PY - 2010/5/11/medline SP - 156 EP - 63 JF - Journal of gastroenterology and hepatology JO - J Gastroenterol Hepatol VL - 25 IS - 1 N2 - BACKGROUND AND AIM: Accumulating evidence supports the effects of miRNA in lipid metabolism, providing a potential linkage between certain miRNA and non-alcoholic fatty liver disease (NAFLD). We aimed to investigate the miRNA expression pattern in a steatotic L02 cell model and explore the function of certain miRNA target pairs. METHODS: The cell model was established by culturing L02 cells with a high concentration of free fatty acid. Micro-array and stem-loop reverse transcription polymerase chain reaction (RT-PCR) were utilized to detect dysregulated miRNA, whereas computational algorithms were used for target prediction. Real time RT-PCR, Western blot, luciferase activity measurement, and other techniques were employed for target verification. RESULTS: Seventeen upregulated and 15 downregulated miRNA were found in steatotic L02 cells, while miRNA-10b was proven to regulate the steatosis level. Peroxisome proliferator-activated receptor-alpha (PPAR-alpha) was also found to participate in steatosis, as its protein level was decreased in steatotic L02 cells and its overexpression by transfection into the PPAR-alpha-pcDNA 3.1 vector could partially alleviate steatosis. We further found that PPAR-alpha is the direct target of miRNA-10b as it showed significantly changed protein expression, but a relatively unchanged mRNA level in steatotic L02 cells transfected with pre-miRNA-10b and anti-miRNA-10b. Moreover, the action of miRNA-10b on PPAR-alpha depends on the presence of a single miRNA-10b binding site, as the activity of a luciferase reporter carrying the mutant PPAR-alpha 3'-untranslated region was not reduced by the expression of miRNA-10b. CONCLUSION: The established miRNA profile of the steatotic L02 cell model and the novel effect of miRNA-10b in regulating hepatocyte steatosis may provide a new explanation of the pathogenesis of NAFLD. SN - 1440-1746 UR - https://www.unboundmedicine.com/medline/citation/19780876/Effect_of_miRNA_10b_in_regulating_cellular_steatosis_level_by_targeting_PPAR_alpha_expression_a_novel_mechanism_for_the_pathogenesis_of_NAFLD_ L2 - https://doi.org/10.1111/j.1440-1746.2009.05949.x DB - PRIME DP - Unbound Medicine ER -