Liver samples were fixed with 4% (w/v) paraformaldehyde, embedded in paraffin, cut into 4-μm-thick sections, and stained with hematoxylin and eosin (H&E) for morphological investigations. H&E staining was carried out after deparaffination as previously described 51. Frozen 8-μm-thick sections of liver tissues were stained with Oil Red O (ORO) to detect neutral lipids 52. Cells were washed three times with PBS, fixed with 10% paraformaldehyde for 30 min and then stained with 0.6% Oil Red O for 10min. After hematoxylin staining, cells were observed by microscope (Carl Zeiss, Jena, Germany).

Immunofluorescent staining of liver tissues was performed as we reported 53. Briefly, liver tissues were frozen in optimum cutting temperature compound, cut at a thickness of 5 μm, fixed with acetone and methanol (1:1) for 30 minutes, then ruptured in Triton X-100 for 30 minutes at room temperature. After the non-specific binding sites were blocked with 0.2% BSA for 60 minutes at 4 °C, the sections were incubated with primary antibodies at 4 °C overnight. After washing extensively in PBS, with Alexa Fluor 488- or rhodamine-conjugated secondary antibodies at room temperature for 60 minutes. The tissue sections were then mounted with DAPI, observed under a scanning microscope (Carl Zeiss, Jena, Germany) and analyzed by Image-Pro-Plus 6.0 software (Media Cybernetics, Silver Spring, MD, USA). The following primary antibodies were used for immunofluorescence: mouse anti-albumin (1:200 dilution, Abcam, Cambridge, UK) and rabbit anti-Prkab1 (1:200 dilution, Proteintech Group, Chicago, USA).

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