Understanding NASH: the 'silent' liver disease

The so-called “silent” liver disease of NASH ranks third for the leading causes of liver transplants and our global obesity epidemic has only increased the prevalence of NASH. Without an approved NASH treatment, the healthcare industry must explore therapeutic strategies for this unmet medical need.

Animal models of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) have proven helpful in understanding disease progression, developing new treatment options and improving diagnostic techniques. To produce rodent models of NAFLD and NASH, diet manipulation is a useful tool. When choosing dietary features to induce NAFLD pathologies, researchers should consider the animal model, time frame and desired disease outcome.

Dietary methods to induce NAFLD/NASH in rodents can be split into two categories:

1) Feed rodents for longer periods of time to induce obesity, metabolic syndrome and mild NASH

2) Feed a nutrient deficient diet to induce hepatic features of severe NASH in a relatively short period of time, without inducing metabolic symptoms such as obesity and insulin resistance

Exploring these dietary methods in more detail will help you to make informed decisions about the therapeutic strategies available.

Inducing obesity, metabolic syndrome and mild NAFLD/NASH

Western/fast food style diets with milkfat and cholesterol are high in palmitate and sucrose. Dietary palmitate and cholesterol have previously been associated with the progression from simple steatosis to NASH. These diets can induce obesity, metabolic syndrome and simple steatosis within nine weeks of feeding. Increased hepatic inflammation has been observed after 12 weeks of feeding. NASH typically requires longer feeding, with fibrosis developing within nine months and late stage fibrosis after 14 to 20 months.

With the American Lifestyle-Induced Obesity Syndrome (ALIOS) model, an “American fast food” diet is offered, which is high in trans-fats and sugar. Trans-fats from hydrogenated vegetable shortening are associated with increased insulin resistance and hepatic inflammation in rodent NASH models. The ALIOS model develops obesity with insulin resistance, elevated ALT levels and steatosis within 16 weeks. Increased inflammation and early development of fibrosis has been observed as early as 6 months. After 12 months, mice exhibited severe steatosis with fibrosis and inflammation, and 50% of mice developed hepatic neoplasms.

Understanding the FPC diet: fructose, palmitate, cholesterol and trans-fat

These diets include Western and ALIOS model diets to achieve both metabolic and hepatic NASH features within an accelerated time frame. These diets have lower methionine and choline content than typical rodent diets, and are high in sucrose. The high milkfat in these diets provide SFA and palmitic acid. Hydrogenated vegetable shortening provides trans-fats. Male mice developed insulin resistance and NAFLD with inflammation, hepatocyte death and fibrosis within 16 weeks of feeding.

In all of the above diets, a glucose/fructose solution can be added to the drinking water, which may further promote NASH development.

Working with high fat diets

These diets typically contain 40 to 60 percent kcal from fat without supplemented cholesterol or cholate. The degree of NASH pathology is limited or mild and varies depending on the animal model, length of feeding and dietary components. For more information, see our Diet Induced Obesity page.

Inducing more severe hepatic NAFLD/NASH

To induce more severe hepatic NAFLD/NASH without obesity or metabolic syndrome, researchers have some options with:

1) Atherogenic diets high in fat, cholesterol and cholate

2) methionine/choline deficient (MCD) diets

Atherogenic diets high in fat, cholesterol and cholate have added cholesterol (1 – 1.25%) and cholate to help induce NASH. This diet option includes purified “Western” style diets with increased cholesterol and cholate and also hybrid diets, which are a mix of a natural ingredient mouse diet in a 3:1 ratio with a concentrated purified diet. The hybrid diet is associated with increased gallstone formation and liver damage as compared to similar purified diets. Atherogenic diets induce varied degrees of NASH with increased hepatic inflammation, with early fibrosis observed after ten weeks of feeding.

Methionine/choline deficient (MCD) diets are amino acid defined rodent diets deficient in methionine and choline, which decreases fat oxidation and export of fat from the liver. In addition, these diets are high in sucrose, with about 10% corn oil by weight. Steatosis, increased serum alanine aminotransferase, inflammation and hepatic fat oxidation has been observed within three weeks, with fibrosis development after six weeks. This diet does not produce metabolic syndrome (which is an aspect of NASH in human models), and is associated with progressive weight loss (up to 40%).

Control diets

Many researchers compare their NAFLD/NASH diet-fed animals to animals fed a natural ingredient, grain-based diet, depending on the research goals. You can chat with a nutritionist and obtain additional information and control diet recommendations.

Careful planning and preparation are critical when producing high quality hybridomas and mAbs. By working with an experienced partner, researchers can eliminate many of the common pitfalls while also saving time and money.

The future of NAFLD/NASH diets

Dietary NAFLD/NASH models continue to evolve. Currently, diets which induce obesity, metabolic syndrome and mild NASH require long term feeding (4 to 12 months) to induce mild fibrosis; often add a glucose/fructose solution to the drinking water and promote sedentary behavior by removing overhead cage feeders. Diets to induce more severe hepatic NASH often do not recapitulate metabolic symptoms associated with NASH such as obesity or insulin resistance and are commonly fed for 3 to 12 weeks to induce hepatic inflammation and early fibrosis.

As research continues to expand, new diets can be formulated to investigate emerging dietary models of NAFLD/NASH and help address this urgent unmet medical need.

Download our mini-paper on NAFLD/NASH studies in rodents to learn more about dietary factors.