Animal experiments were approved by the University Committee on Use and Care of Animals (UCUCA) of Southern Medical University. C57BL6/J (wild type, stock no. 005304), and Lgr5-eGFP-IRES-CreERT2 (Lgr5-GFP, stock no. 008875) mice were purchased from Jackson Laboratory Generation and genotyping of the Lgr5-GFP mice has been previously described16 (link).
For single CCL4 treatment, 8-week-old male mice received an i.p. injection of CCL4 (2 ml/kg, Sigma-Aldrich) dissolved in olive oil at a ratio of 1:4, or olive oil alone (2 ml/kg). Mice liver was harvested and analyzed at day 5.
For induction of hepatic fibrosis by CCL4, 8-week-old mice were i.p. injected with CCL4 (2 ml/kg, Sigma-Aldrich) dissolved in olive oil at a ratio of 1:4, or olive oil alone (2 ml/kg) twice a week for 6 weeks.
For Sirius Red staining, 5 µM sections were stained with Sirius Red (Sigma), and the results were quantified using Nikon Elements software. Six random fields from each section were analyzed at a final magnification of ×40. Five random sections from each mouse were analyzed, and the statistical data from 10 mice per group were obtained for comparison of the different treatment groups.
For single CCL4 treatment, 8-week-old male mice received an i.p. injection of CCL4 (2 ml/kg, Sigma-Aldrich) dissolved in olive oil at a ratio of 1:4, or olive oil alone (2 ml/kg). Mice liver was harvested and analyzed at day 5.
For induction of hepatic fibrosis by CCL4, 8-week-old mice were i.p. injected with CCL4 (2 ml/kg, Sigma-Aldrich) dissolved in olive oil at a ratio of 1:4, or olive oil alone (2 ml/kg) twice a week for 6 weeks.
For Sirius Red staining, 5 µM sections were stained with Sirius Red (Sigma), and the results were quantified using Nikon Elements software. Six random fields from each section were analyzed at a final magnification of ×40. Five random sections from each mouse were analyzed, and the statistical data from 10 mice per group were obtained for comparison of the different treatment groups.
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