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Reduction of circulating PCSK9 and LDL-C levels by liver-specific knockdown of HNF1? in normolipidemic mice.

ABSTRACT: The transcription factors hepatic nuclear factor (HNF)1? and HNF1? can bind to the HNF1 site on the proprotein convertase subtilisin/kexin type 9 (PCSK9) promoter to activate transcription in HepG2 cells. However, it is unknown whether one or both HNF1 factors are obligatory for transactivating hepatic PCSK9 gene expression in vivo. We developed shRNA adenoviral constructs (Ad-shHNF1? and Ad-shHNF1?) to examine the effects of knockdown of HNF1? or HNF1? on PCSK9 expression and its consequent impact on LDL receptor (LDLR) protein levels in cultured hepatic cells and liver tissue. We demonstrated that infection with Ad-shHNF1?, but not Ad-shHNF1?, markedly reduced PCSK9 mRNA expression in HepG2 cells with a concomitant increase in LDLR protein abundance. Injecting Ad-shHNF1? in mice fed a normal diet significantly (? 50%) reduced liver mRNA expression and serum concentration of PCSK9 with a concomitant increase (? 1.9-fold) in hepatic LDLR protein abundance. Furthermore, we observed a modest but significant reduction in circulating LDL cholesterol after knockdown of HNF1? in these normolipidemic mice. Consistent with the observation that knockdown of HNF1? did not affect PCSK9 mRNA or protein expression in cultured hepatic cells, Ad-shHNF1? infection in mice resulted in no change in the hepatic mRNA expression or serum content of PCSK9. Altogether, our study demonstrates that HNF1?, but not HNF1?, is the primary positive regulator of PCSK9 transcription in mouse liver.

SUBMITTER: Shende VR

PROVIDER: S-EPMC4373738 | biostudies-literature | 2015 Apr

REPOSITORIES: biostudies-literature

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Reduction of circulating PCSK9 and LDL-C levels by liver-specific knockdown of HNF1α in normolipidemic mice.

Shende Vikram Ravindra VR Wu Minhao M Singh Amar Bahadur AB Dong Bin B Kan Chin Fung Kelvin CF Liu Jingwen J

Journal of lipid research 20150204 4

The transcription factors hepatic nuclear factor (HNF)1α and HNF1β can bind to the HNF1 site on the proprotein convertase subtilisin/kexin type 9 (PCSK9) promoter to activate transcription in HepG2 cells. However, it is unknown whether one or both HNF1 factors are obligatory for transactivating hepatic PCSK9 gene expression in vivo. We developed shRNA adenoviral constructs (Ad-shHNF1α and Ad-shHNF1β) to examine the effects of knockdown of HNF1α or HNF1β on PCSK9 expression and its consequent imp ...[more]

PMID: 25652089

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Project description:Proprotein convertase subtilisin/kexin type 9 (PCSK9) can promote the degradation of low-density lipoprotein (LDL) receptor (LDLR), leading to hypercholesterolemia and myocardial dysfunction. The intracellular regulatory mechanism by which the natural polyphenol pterostilbene modulates the PCSK9/LDLR signaling pathway in cardiomyocytes has not been evaluated. We conducted Western blotting, flow cytometry, immunofluorescence staining, and mean fluorescence intensity analyses of pterostilbene-treated mouse HL-1 cardiomyocytes. Pterostilbene did not alter cardiomyocyte viability. Compared to the control group, treatment with both 2.5 and 5 μM pterostilbene significantly increased the LDLR protein expression accompanied by increased uptake of LDL. The expression of the mature PCSK9 was significantly suppressed at the protein and mRNA level by the treatment with both 2.5 and 5 μM pterostilbene, respectively, compared to the control. Furthermore, 2.5 and 5 μM pterostilbene treatment resulted in a significant reduction in the protein hepatic nuclear factor 1α (HNF1α)/histone deacetylase 2 (HDAC2) ratio and sterol regulatory element-binding protein-2 (SREBP2)/HDAC2 ratio. The expression of both hypoxia-inducible factor-1 α (HIF1α) and nuclear factor erythroid 2-related factor 2 (Nrf2) at the protein level was also suppressed. Pterostilbene as compared to short hairpin RNA against SREBP2 induced a higher protein expression of LDLR and lower nuclear accumulation of HNF1α and SREBP2. In addition, pterostilbene reduced PCSK9/SREBP2 interaction and mRNA expression by increasing the expression of hsa-miR-335 and hsa-miR-6825, which, in turn, increased LDLR mRNA expression. In cardiomyocytes, pterostilbene dose-dependently decreases and increases the protein and mRNA expression of PCSK9 and LDLR, respectively, by suppressing four transcription factors, HNF1α, SREBP2, HIF1α, and Nrf2, and enhancing the expression of hsa-miR-335 and hsa-miR-6825, which suppress PCSK9/SREBP2 interaction.

Dataset Information

Reduction of circulating PCSK9 and LDL-C levels by liver-specific knockdown of HNF1? in normolipidemic mice.

Publications

Reduction of circulating PCSK9 and LDL-C levels by liver-specific knockdown of HNF1α in normolipidemic mice.

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