Figure 2.

miR-185 Is Abundant in LX-2 Cells but Downregulated in Activated LX-2 Cells

The human hepatic stellate cell (HSC) cell line LX-2 was treated with 5.0 ng/mL transforming growth factor β-1 (TGF-β1) for 0, 3, 6, 12, or 24 hr or with 2.5, 5.0, or 7.5 ng/mL TGF-β1 for 24.0 hr. (A) mRNA expression of key genes involved in the activation of HSC α-SMA, COL1A1, COL1A2, and COL3A1 was analyzed by real-time PCR. All genes increased in a time-dependent manner. mRNA levels of Ras homolog enriched in brain (RHEB) and rapamycin-insensitive companion of mammalian target of rapamycin (RICTOR) were assessed by real-time PCR. When HSCs were activated by TGF-β1 with a different dose (B) or time (C), RHEB and RICTOR were upregulated in mRNA levels. (D) Protein expressions of α-SMA, COL I, COL III, RHEB, and RICTOR was determined by western blotting; GAPDH or β-actin was used as a loading control. When HSCs were activated, all genes were upregulated in protein levels. miR-185-5p expression was reduced in activated HSCs in a dose-dependent manner (E) or time-dependent manner (F) examined by real-time PCR (n = 3, *p < 0 0.05, **p < 0.01). Data represent mean ± SEM.