Nr4a2

The following primary antibodies were used: DJ-1 (Santa Cruz Biotechnology), RUNX1 (Santa Cruz Biotechnology), Lamp1 (Santa Cruz Biotechnology), LC3 (MBL International Corporation), p62 (MBL International Corporation), PARP (Cell Signaling Technology), PRKAA/AMPK (Cell Signaling Technology), phosphor-PRKAA/AMPK (Cell Signaling Technology), Beclin1 (Cell Signaling Technology), ATG5 (Cell Signaling Technology), PCNA (abcam), NFIL3 (Santa Cruz Biotechnology), ETS1 (Santa Cruz Biotechnology), FOXA1 (Santa Cruz Biotechnology), C-JUN (Santa Cruz Biotechnology), EGR3 (Santa Cruz Biotechnology), ATF3 (Santa Cruz Biotechnology), MAFF3 (Santa Cruz Biotechnology), BHLHE40 (Santa Cruz Biotechnology), NR4A2 (Santa Cruz Biotechnology). Ciclopirox olamine, Rotenone, APO, NDGA, NAC, Glucose, Oligomycin, 2-DG, CQ, Bafilomycin A1, E64D and PepA were purchased from Sigma.

Hepatic tissues were embedded in paraffin and sectioned. The sections were incubated in 3% H2O2 in methanol and nonspecific binding was blocked with 10% normal goat serum. The sections were incubated with primary antibody overnight, washed and incubated with secondary antibody for 60 minutes. Antigen–antibody complexes were visualized using DAB kits (GeneTech, Shanghai, China). Immunofluorescence was performed as described previously33 (link). The following primary antibodies were used: α-smooth muscle actin (α-SMA) (Santa Cruz), NR4A2 (Santa Cruz), Alexa Fluor^® 546 IgG (Life Technologies), Alexa Fluor^® 488 IgG (H + L) (Life Technologies). Image scanning analysis system (Image-Pro Plus) was used to analyze the changes in integrated optical density (IOD) of NR4A2 and α-SMA in immunofluorescence images.

Repagermanium (lot. 006316 A, purity > 99.9%), the polymer of THGP, was produced at the Hakodate plant of the Asai Germanium Research Institute Co., Ltd. (Hokkaido, Japan), and Bis(2-carboxyethylgermanium(IV) sesquioxide) was purchased from Sigma-Aldrich (Missouri, USA). Potassium bromide (KBr) and D₂O were purchased from Kanto Chemical, Co., Inc (Tokyo, Japan). NHDFs and a Fibroblast Growth Medium 2 Kit were purchased from Takara Bio (Shiga, Japan). Xanthine oxidase (XOD from buttermilk) was purchased from Oriental Yeast Co., Ltd. (Tokyo, Japan). Hypoxanthine (HPX), hydrogen peroxide and 4% paraformaldehyde phosphate buffer solution were purchased from Wako Pure Chemical Industrial Co., Ltd. (Osaka, Japan). Cellstain-PI solution and Hoechst 33342 solution were purchased from Dojindo Laboratories Co., (Kumamoto, Japan). A complete tablets EASY pack was purchased from Roche Diagnostics GmbH (Mannheim, Germany). NR4A2 and β-actin antibodies were purchased from Santa Cruz Biotechnology (California, USA) and Abcam (Cambridge, England). Secondary antibodies, Goat Anti-Mouse IgG H&L (HRP) and Donkey Anti-Goat IgG H&L (HRP) were purchased from Abcam. All reagents used in this study were of special or molecular biological research grade.