Non-alcoholic fatty liver disease (NAFLD) is a growing epidemic associated with key aspects of metabolic disease such as obesity and diabetes. The first stage of NAFLD is characterized by lipid accumulation in hepatocytes, but this can further progress into non-alcoholic steatohepatitis (NASH), fibrosis or cirrhosis, and hepatocellular carcinoma (HCC). A western diet, high in fats, sugars and cholesterol is linked to NAFLD development. Murine models are often used to experimentally study NAFLD, as they can display similar histopathological features as humans;however, there remains debate on which diet-induced model most appropriately and consistently mimics both human disease progression and pathogenesis. In this study, we performed a side-byside comparison of two popular diet models of murine NAFLD/NASH and associated HCC: a high fat diet supplemented with 30% fructose water (HFHF) and a western diet high in cholesterol (WDHC), comparing them to a common grain-based chow diet (GBD). Mice on both experimental diets developed liver steatosis, while WDHC-fed mice had greater levels of hepatic inflammation and fibrosis than HFHF-fed mice. In contrast, HFHF-fed mice were more obese and developed more severe metabolic syndrome, with less pronounced liver disease. Despite these differences, WDHC-fed and HFHF-fed mice had similar tumour burdens in a model of diet-potentiated liver cancer. Response to diet and resulting phenotypes were generally similar between sexes, albeit delayed in females. Notably, although metabolic and liver disease phenotypes are often thought to progress in parallel, this study shows that modest differences in diet can significantly uncouple glucose homeostasis and liver damage. In conclusion, long-term feeding of either HFHF or WDHC are reliable methods to induce NASH and diet-potentiated liver cancer in mice of both sexes; however, the choice of diet involves a trade-off between severity of metabolic syndrome and liver damage.
HCC Mouse CohortWild-type inbred C57BL/6N genetic background mice were housed in the same fashion as the NASH cohort mice described above. Mice were injected with 25 mg of diethylnitrosamine (DEN) per kg body mass at 2 weeks of age. Mice were fed ad libitum either the HFHF diet or the WDHC diet as described above. 14 males (n=6 on HFHF, n=8 on WDHC) and 14 females (n=7 on HFHF and n=7 on WDHC). Body mass was taken weekly, and mice were sacrificed after 24 weeks of feeding following a 3-hour fast. Samples were processed in a similar manner as the NASH cohort.
Blood CollectionA minimum concentration of 3.2 µL per 600 µL blood of Aprotinin from bovine lung (Sigma, MA, USA) was added to all blood samples (except those taken during the glucose tolerance test) to prevent protein degradation. Blood collected during the oral glucose tolerance test (OGTT) was collected in Microvette Ò 100µL K3 EDTA (Sarstedt, Germany). Blood was incubated at 4°C for 90 minutes, followed by an 8-minute centrifugation at 4000 rpm at 4°C. The top layer of serum was collected and stored at -80°C unti...