Albumin cre alb cre mice

To generate mice with a conditional knockout allele of TRIM31, the TRIM31^Flox/Flox mice with C57BL/6 N background were constructed using CRISPR/Cas9-regulated genome engineering system. The exon 4/5 of TRIM31 was selected as conditional knockout region (cKO). Briefly, the selected exons of TRIM31 were flanked by two loxP sites, and therefore two single guide RNAs (gRNA1# and gRNA2#) targeting TRIM31 introns were designed. The targeting vector containing TRIM31 exon 4/5 flanked by two loxP sites and the two homology arms was used as the template. The targeting vector, gRNA1# and gRNA2#, and together with Cas9 were co-injected into fertilized eggs for cKO mouse production. The obtained mice, which had exon 4/5 flanked by two loxP sites on one allele, were used to establish TRIM31^Flox/Flox mice. Hepatocyte-specific TRIM31 deletion (TRIM31^Hep-cKO) mice were produced by mating TRIM31^Flox/Flox mice with albumin-Cre (Alb-Cre) mice (Jackson Laboratory, Bar Harbor, Maine, USA). A simple schematic diagram has been indicated in Supplementary Fig. S13A. TRIM31^Flox/Flox (Flox) mice littermates were used in the work as controls for the obtained TRIM31^Hep-cKO mice. All TRIM31^Flox/Flox and TRIM31^Hep-cKO mice were then randomly divided into three subgroups, including the Oil/Veh group (n = 15), CCl₄/Veh group (n = 20), and CCl₄/Mul-H (high dosage 60 mg/kg; n = 15) group.

miR-34a^fl/fl mice, albumin-Cre (Alb-Cre) mice, and C57BL/6J mice were purchased from the Jackson Laboratory (Bar Harbor, Maine, USA) [4 (link)]. miR-34a^fl/fl mice were crossed with Alb-Cre mice to generate germline hepatocyte-specific miR-34a^−/− (miR-34a^gHep−/−) mice and control littermates (miR-34a^fl/fl mice). AAV8-TBG-Null or AAV8-TBG-Cre was i.v. injected into the miR-34a^fl/fl mice to generate control mice (miR-34a^fl/fl mice) or adult-onset hepatocyte-specific miR-34a^−/− (miR-34a^Hep−/−) mice, respectively. Locked nucleic acids (LNA) against scramble sequences (LNA-Scr) or miR-34a (LNA-miR-34a) were synthesized by Qiagen and i.p. injected into the mice at a dosage of 10 mg/kg/week. All of the mice were housed in a temperature- and humidity-controlled room with a 12-h light/12-h dark cycle under pathogen-free conditions. The high-fat/cholesterol/fructose (HFCF) diet contained 40% fat/0.2% cholesterol (AIN-76A Western diet from TestDiet) and 4.2% fructose (in drinking water). Unless otherwise stated, 2-month-old male mice were used and fed an HFCF diet for 16 weeks. The mice were fasted for 5–6 h during the light cycle prior to euthanasia. All the of animal experiments were approved by the Institutional Animal Care and Use Committee at Northeast Ohio Medical University (NEOMED).