Fig 1: Effects of anti-HIV drugs on expression of selective factors related to hepatic metabolism or secretion of bile acid or bilirubin. (A) Western blots of proteins from HepG2; Ctrl, non-senescent HepG2; Senescent, senescent HepG2; DMSO, dimethyl sulfoxide as vehicle control; RTV, ritonavir; LPV, lopinavir; RTV + LPV, ritonavir plus lopinavir; OPTP1B1 and OATPIB3, organic anion transporter polypeptide member 1B1 and 1B3; GAPDH, glyceraldehyde 3-phosphate dehydrogenase as an internal control; UGT1A1, UDP-glucuronosyltransferase (UGT) 1A1; (B–D), RT-PCR of mRNAs from HepG2; CYP3A4, CYP2D6, and CYP7A1: cytochrome P450 3A4, 2D6, and 7A1, respectively; Green dots indicate drug treatments; **, p < 0.01; ***, p < 0.005; (E) Enzyme activities of UGT from non-senescent PHHs (Ctrl) or senescent PHHs (Senescent); *, p < 0.05; **, p < 0.01 compared to Ct of the same cell group; δ, p < 0.05 and δδ, p < 0.01 compared to the same treatment between non-senescent and senescent PHHs.
Fig 2: Impact of miR-491-3p on UGT1A1-mediated xenobiotic metabolism in human liver cells, as examined with the probe NHPN. Hep3B (A) and HepG2 (B) were treated with 20 nM of RNA for 72 hours and then incubated with 10 μM NHPN for 30 minutes. After live cell images were acquired, the fluorescent intensities of NHPN glucuronides were measured with a plate reader (excitation at 370 nm and emission at 520 nm) and then normalized to total protein concentrations. Values are mean ± SD (N = 3 biological replicates per group). ∗∗P < .01; and ns, not significant; as compared with control RNA (Student’s t-test).
Fig 3: Recombinant miR-491-3p alters UGT1A1-mediated estradiol 3-glucuronidation capacity in Hep3B cells. (A) Schematic illustration of estradiol 3- and 17-glucuronidation catalyzed by different UGT isoforms. (B) Levels of glucuronides produced from estradiol in cell lysates after transfected with miR-491-3p and control RNA for 72 hours, as quantified by LC-MS/MS method. Values are mean ± SD (N = 4 biological replicates per group). ∗∗∗P < .001; and ns, not statistically significant as compared with the control RNA group (Student’s t-test).
Fig 4: BioRNA/miR-491-3p modulates UGT1A1 protein levels in human Hep3B (A) and HepG2 (B) cells. IFC studies confirmed UGT1A1 (green) localization on the ER (red), whose intensities were obviously reduced by recombinant miR-491-3p molecules (20 nM for 72 hours) in both Hep3B (A) and HepG2 (B) cells. Scale bar, 100 μm.
Supplier Page from Abcam for UGT Activity Assay / Ligand Screening Kit (Fluorometric)