Up-regulation of selenoprotein P and HIP/PAP mRNAs in hepatocytes by intermittent hypoxia via down-regulation of miR-203

Figures

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  Fig. 1. The mRNA levels of SELENOP (A), ANGPTL6 (B), SHBG (C), FGF21 (D), LECT2 (E), LIPASIN (F), and AHSG (G) in HepG2 cells treated by normoxia or IH for 24 h. The levels of the hepatokine mRNAs were measured by real-time RT-PCR using β-actin as an endogenous control. Data is expressed as means ± SE for each group (n=4). The statistical analyses were performed using Student's t-test.

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  Fig. 2. Concentrations of SELENOP in HepG2 cell culture medium were measured by ELISA. HepG2 cells were treated by normoxia or IH conditions for 24 h. Data is expressed as means ± SE for each group (n=4).

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  Fig. 3. The mRNA levels of REG Iα (A), REG Iβ (B), REG III (C), HIP/PAP (D), and REG IV (E) in HepG2 cells treated by normoxia or IH for 24 h were measured by real-time RT-PCR. Data is expressed as means ± SE for each group (n=4). The statistical analyses were performed using Student's t-test.

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  Fig. 4. Effects of IH on cellular proliferation of HepG2 cells. They were exposed to normoxia or IH conditions for 24 h, and cellular proliferation was measured by WST-8 assay. Data is expressed as means ± SE for each group (n=12). The statistical analyses were performed using Student's t-test.

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  Fig. 5. Effects of siRNA transfection on cell proliferation. After transfection of siRNA against REG Iα or HIP/PAP into HepG2 cells, cells were exposed to normoxia or IH conditions for 24 h, and cellular proliferation was measured by WST-8 assay. Data is expressed as means ± SE for each group (n=12). The statistical analyses were performed using Student's t-test.

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  Fig. 6. Luciferase assays in HepG2 cells. Reporter plasmids prepared by inserting the promoter fragments of SELENOP (−2989~+10) and HIP/PAP (−4030~+27) upstream of a firefly luciferase reporter gene in the pGL4.17 vector were transfected into HepG2 cells. After cells were exposed to 24 h of either IH or normoxia, the cells were lysed, and promoter activities of SELENOP (A) and HIP/PAP (B) were measured. All data is represented as the mean ± SE of the samples (n=3). The statistical analyses were performed using Student's t-test.

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  Fig. 7. The mRNA levels of miR-203 (A), DROSHA (B), DICER (C), MCPIP1 (D), and TP63 (E) in HepG2 cells treated by normoxia or IH for 24 h. The levels of mRNAs were measured by real-time RT-PCR using β-actin as an endogenous control. Data is expressed as means ± SE for each group (n=4). The statistical analyses were performed using Student's t-test.

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  Fig. 8. Effects of miR-203 inhibitor transfection on SELENOP and HIP/PAP expression. The miR-203 inhibitor and non-specific control RNA (miR-203 inhibitor NC) were introduced into HepG2 cells just before normoxia or IH exposure. The expression of SELENOP and HIP/PAP mRNA was measured by real-time RT-PCR using β-actin as an endogenous control. The figure represents (A) SELENOP mRNA expression in miR-203 inhibitor NC-introduced cells, (B) SELENOP mRNA expression in miR-203 inhibitor-introduced cells, (C) HIP/PAP mRNA expression in miR-203 inhibitor NC-introduced cells and (D) HIP/PAP mRNA expression in miR-203 inhibitor-introduced cells. Data is expressed as means ± SE for each group (n=4). The statistical analyses were performed using Student's t-test.