Learn more about GAN rodent diet

The AMLN diet with a trans-fat source or GAN with palm oil are named for the companies where the model characterization occurred. These types of diets are used to exacerbate hepatic inflammation and fibrosis.

Prior to the 2018 FDA ban on trans-fats, diets containing hydrogenated vegetable oil were used to reliably induce metabolic and liver histopathological changes in rodents.  Following 2018, modifications to fat sources had to be made.

Within the MASH literature, the AMLN or GAN diets are described as purified rodent diets with 22% fructose, 2% cholesterol, and 40% of kcal either predominately from palm oil (GAN) or trans-fat containing hydrogenated vegetable oil that is very comparable to what we had prior to 2018 (AMLN).

Examples:

• TD.200591 40% Kcal Fat (Palm Oil, 2% Chol, 20% Kcal/Fruct)
• TD.180939 40% Kcal Fat (HVO, 2% Chol, 20% Kcal/Fruct)

Research use and timeline to achieve phenotype:

Diet formulations of these patterns have reported hepatic steatosis, systemic insulin resistance, and obesity after 12 weeks of feeding an AMLN diet.  Others have reported the GAN diet produced steatohepatitis and fibrosis after 20 weeks of feeding. While not required, the AMLN/GAN model often pairs a glucose/fructose solution to the drinking water.

References:

See MASH rodent diet references

FPC (Fat - Palmitate - Cholesterol) are popular MASH diets that include features of both the “Western” and American Lifestyle-Induced Obesity Syndrome (ALIOS) model diets to achieve both metabolic and hepatic MASH features within an accelerated time frame. FPC diets intentionally contain a lower methionine and choline content to exacerbate these phenotypes. FPC diets are high fat (~52% kcal from fat) and include milkfat fat, palmitic acid and hydrogenated vegetable shortening (source of trans-fats) with high sucrose (~34% by weight) and 1.25% cholesterol.

Examples:

• TD.160785.PWD 52 kcal/Fat Diet (C16:0, HVO, AMF, Choline/Met)
• TD.190142^* 52 kcal/Fat Diet (C16:0, HVO, AMF, Choline/Met)

* a version of TD.160785.PWD with regular casein that can be pelleted

Research use and timeline to achieve phenotype:

Increased chemokines, steatosis, lobular inflammation, liver fibrosis, glucose intolerance, increased liver weight, and body weight gain has been reported after 16 weeks of feeding.  While not required, the FPC model often pairs a glucose/fructose solution to the drinking water.

References:

See MASH rodent diet references

Western or fast-food style diets fed to induce MASH with metabolic syndrome contain 40 - 45% kcal from milkfat (a fat source high in palmitate) with added cholesterol (primarily 1.25%) and high sucrose (>30%). Dietary palmitate and cholesterol have both previously been associated with the progression from simple steatosis to MASH.

Examples:

• TD.120528 42% Kcal/Fat Diet (Increased Sucrose, 1.25% Chol.)
• TD.96121 21% MF, 1.25% Chol. Diet
• TD.88137*  Adjusted Calories Diet (42% from fat)
  
  

* primarily for MASLD; for MASH-like phenotype expression, feed greater than 36 weeks 

Research use and timeline to achieve phenotype:

Obesity with metabolic syndrome and inflammation has been achieved after 12 weeks of feeding. Following 24 weeks, significant steatosis, moderate hepatic ballooning and bridging fibrosis have been reported. While not required, the Western model often pairs a glucose/fructose solution to the drinking water.

References:

See MASH rodent diet references

Expression of MASH phenotypes in a shorter time frame, without metabolic syndrome or obesity can be achieved with a methionine and choline devoid (MCD) approach. MCD diets are amino acid defined rodent diets deficient in methionine and choline, with high sucrose (>40% by weight) and ~10% corn oil by weight. The polyunsaturated fat in corn oil promotes hepatic lipid oxidation.

Methionine and choline deficiency decreases fat oxidation and export of fat from the liver. Dietary sucrose is necessary for hepatic lipid accumulation and oxidation. Expect that animals will lose weight (up to 40%) on this type of diet, so careful monitoring is recommended.

A variation of this MCD diet contains 60% kcal from fat and 0.1% methionine to mitigate weight loss commonly seen when feeding an MCD diet, while still driving more severe MASH phenotypes.

Examples:

• TD.90262* Methionine/Choline Deficient Diet
• TD.240263 CDAA Diet 0.1% Met (60/Fat, Lard, O)

*Modifications of our MCD diets with adjusted methionine, choline, and/or fat levels are available upon request.

Research use and timeline to achieve phenotype:

Steatosis, increased serum alanine aminotransferase (ALT), inflammation, and hepatic fat oxidation has been observed within 3 weeks of feeding our MCD diets with fibrosis development reported after 6 weeks of feeding.

Insulin-resistance, inflammation, increased ALT and TG, upregulated cytokine production, fibrosis, and hepatic ballooning have been reported within 8 weeks of feeding a high fat CDAA diet.

References:

See MASH rodent diet references

The following references describe MASH phenotype expression in Sprague-Dawley and GEM rat models.

Carreres, Lydie, et al. "Modeling diet-induced NAFLD and NASH in rats: a comprehensive review." Biomedicines 9.4 (2021): 378.

Zou, Yuhong, et al. "High-fat emulsion-induced rat model of nonalcoholic steatohepatitis." Life sciences 79.11 (2006): 1100-1107.

Aldiss, Peter, et al. "FGF21 deletion mildly exacerbates hepatic dysfunction in MASH diet and alcohol fed rats." bioRxiv (2024): 2024-03.  

Zhu, An-Qi, et al. "Transcriptomic insights into the lipotoxicity of high-fat high-fructose diet in rat and mouse." The Journal of Nutritional Biochemistry 128 (2024): 109626.

Janevski, Mile, et al. "The effect of cocoa supplementation on hepatic steatosis, reactive oxygen species and LFABP in a rat model of NASH." Comparative Hepatology 10 (2011): 1-13.

Dietary models of MASH continue to evolve with the goal of more accurately recapitulating both the metabolic and hepatic symptoms of human disease. Commonly researchers are studying the synergistic effects of various MASH dietary features to accelerate progression of the model and severity of liver disease.

A Teklad nutritionist can work with you to formulate new diets in order to investigate novel dietary models of MASH. Contact a nutritionist at askanutritionist@inotiv.com for a diet consultation.

The choice of control diets is dependent on the specific research goal. Many researchers choose to compare their MASH diet-fed animals to animals fed a natural ingredient, grain-based diet (also referred to as standard diet or chow). These diets differ in the source and level of nutrients as well as in the presence of non-nutritive factors.

Depending on what your main comparisons are, it may be suitable to have a grain-based diet as your control/reference group. However, making such comparisons limits inferences to dietary patterns versus a specific dietary component. In some cases, such as those studies feeding amino acid defined diets like the MCD model, a matched control diet is recommended given the very different formulations and protein sources of grain-based diets.

When making inferences about specific nutrients within the diet, an ingredient matched, low fat control diet may be necessary. There are many options with different levels and types of fat in addition to different types of carbohydrate ranging from sucrose (highly refined and digestible) to corn starch (refined, but more complex) to resistant starch (refined, but not fully digestible).

A very basic purified control diet would be AIN-93M TD.94048 or AIN-93G TD.94045 . AIN-93 diets have a moderate amount of sucrose at ~10% with fat from soybean oil providing a healthy fatty acid profile. Learn more about AIN diet formulas.

Contact a nutritionist for an additional information and control diet recommendations.