CD1 male mice (8 to 10 weeks old) were maintained in pathogen-free housing and cared for in accordance with the Universidad Autónoma Metropolitana and NIH Guide for the Care and Use of Laboratory Animals.
Twenty mice were randomly divided in two groups; in HC group 10 animals were fed with a high-cholesterol diet (HC, 2% cholesterol and 0.5% sodium cholate) for two days, as reported by Marí and coworkers [2 (link)]. Ten control animals were fed with a regular rodent Chow diet (Purina #5001). After the two days under HC and Chow diets, five animals were sacrificed.
Hepatocytes were isolated from the rest of the HC and Chow mice by the two-step collagenase perfusion, as we previously described [12 (link)]. The viability was >90% as assessed by trypan blue exclusion. Hepatocytes were seeded at 2.13 × 105 cells per cm2 either in Lab-Tek chambered slides or in 10 cm dishes (Nalge, Nunc) in the Ham's F-12/Dulbecco's modified Eagle's basal hepatocyte growth medium supplemented with 10% fetal bovine serum. After 4 h of cells attachment, media were replaced by a serum-free basal hepatocyte growth medium. In the following day, cells were treated with 50 ng/mL HGF.
Twenty mice were randomly divided in two groups; in HC group 10 animals were fed with a high-cholesterol diet (HC, 2% cholesterol and 0.5% sodium cholate) for two days, as reported by Marí and coworkers [2 (link)]. Ten control animals were fed with a regular rodent Chow diet (Purina #5001). After the two days under HC and Chow diets, five animals were sacrificed.
Hepatocytes were isolated from the rest of the HC and Chow mice by the two-step collagenase perfusion, as we previously described [12 (link)]. The viability was >90% as assessed by trypan blue exclusion. Hepatocytes were seeded at 2.13 × 105 cells per cm2 either in Lab-Tek chambered slides or in 10 cm dishes (Nalge, Nunc) in the Ham's F-12/Dulbecco's modified Eagle's basal hepatocyte growth medium supplemented with 10% fetal bovine serum. After 4 h of cells attachment, media were replaced by a serum-free basal hepatocyte growth medium. In the following day, cells were treated with 50 ng/mL HGF.
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