Diet Induced NASH B6

Taconic is the first and only vendor to offer a diet-induced NASH model off the shelf

Taconic Biosciences maintains an inventory of C57BL/6NTac male mice conditioned on a modified Amylin liver NASH (AMLN) diet. Diet # D09100310i (source Research Diets) contains 40 kcal% fat, 20 kcal% fructose and 2% cholesterol and is an irradiated diet. C57BL/6NTac males are put on this diet at 6 weeks of age and housed at reduced density. Control males are housed in the same location, also at reduced density, and are fed NIH-31M diet. Control males can also be generated using a low-fat purified diet upon request.

Phenotypic characterization of the NASH B6 model is ongoing, but literature reports of B6 mice fed similar diets report that mice become obese, get fatty liver and develop liver inflammation and fibrosis after 26+ weeks on diet, with inter-animal variability observed for development of the inflammation and fibrosis phenotype.

NASH B6 mice in transit for an extended period of time (more than 48 hours by ground or any air transit) can lose 10-20% of their body weight and may require extended acclimation (two or more weeks). Weight loss induced by transit may delay disease phenotype relative to animals which have remained in the same facility for the entire conditioning period. Additionally, phenotypic penetrance of certain NASH phenotypes is incomplete, so animals conditioned on diet for the same length of time will vary in disease phenotype. Taconic recommends a pilot study to determine the appropriate number of mice to order for a specific study type.

NOTE: The NASH diet D09100310i may take several weeks to receive after ordering from Research Diets. Please ensure you understand timeline of diet receipt in your facility prior to ordering mice. Taconic cannot provide the NASH diet beyond the amount the mice are shipped with in the TTC for use in transit.

Figure 1: Comparison between C57BL/6NTac mice placed on D09100310 diet (NASH B6NTac) or kept on chow diet (Control B6NTac) from 5 weeks of age. A) Liver weight as a percentage of total body weight. B) Liver hydroxyproline content. C) Liver triglycerides. D) Serum alanine aminotransferase levels (ALT). For each time point, n=8 for NASH B6NTac and n=4 for Control B6NTac. Different individual animals were used for each time point (i.e. data is not longitudinal by animal). * and ** indicate statistical significance between NASH and control animals. Two-way ANOVA with multiple comparisons, with p-value<0.05* or p-value<0.01** Data provided by an anonymous pharmaceutical company.

Figure 2: Histopathology for C57BL/6NTac mice placed on D09100310 diet (NASH B6NTac) or kept on chow diet (Control B6NTac) from 5 weeks of age. Animals were on diet for 26 weeks (top set) or 38 weeks (bottom set). Picrosirius red (PSR) staining illustrates collagen I and III fibers, hematoxylin and eosin (H&E) staining illustrates steatosis. Two control and two NASH animals are shown for each time point, with PSR and H&E shown for the same individual animal. Different individual animals were used for each time point (i.e. data is not longitudinal by animal). Data provided by an anonymous pharmaceutical company.

Necropsy results for 10 male C57BL/6NTac mice at 16 weeks on diet D09100310i

Gross exam: Mice were bright, alert, and responsive. Haircoats were oily, but no wounds or dermatitis were observed. Obesity was present and all mice had a body condition score of 5/5.

General findings: Following euthanasia via CO2 asphyxiation, mice were weighed and dissected via a midline incision from prepuce to manubrium. All internal abdominal and thoracic organs were examined grossly. All mice had marked adipose tissue in the mesentery, retroperitoneal fat pads, and caudal subcutaneous fat pads. Liver in all mice was grossly pale to tan, smooth, and uniformly enlarged (see Fig 3). Liver was removed intact and weighed. Liver was then transected into 1 cm sagittal sections for examination of cut surface. No masses were identified on serosal or cut surfaces of the liver.

Summary: Moderate diffuse hepatomegaly with steatosis.

Figure 3: Liver of C57BL/6NTac mouse on NASH diet D09100310i for 16 weeks
displays moderate diffuse hepatomegaly with steatosis.

Body weight and liver weight for Diet Induced NASH B6 mice on diet for 16 weeks

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Necropsy results for 10 male C57BL/6NTac mice at 27 weeks on diet D09100310i

Gross exam: All 10 mice were bright, alert, and responsive. Fur coat condition was visibly oily, but no wounds or dermatitis observed. Obesity was present in all, with adipose tissue plentiful. All animals had a body condition score of 5/5.

General findings: Following euthanasia via CO2 asphyxiation, mice were weighed and dissected via a bilateral paramedial incisions from the midline. All internal abdominal and thoracic organs were examined grossly. All animals had marked mesentary and subcutaneous adipose tissue. Liver was removed intact (including gallbladder) and weighed. Gross findings include Male M000671 found to have an approximately 1.0 mm lesion, flat, circular, flush to the medial lobe, milky white in coloration, located on the lower right side of the medial lobe (Fig 5). All other internal other abdominal and thoracic organs appeared unremarkable in all animals examined. Liver was then transected into 1 cm sagittal sections for examination of cut surface. No masses were identified on serosal or cut surfaces of the liver.

Figure 4: Liver of C57BL/6NTac mouse on NASH diet D09100310i for 27 weeks
displays moderate diffuse hepatomegaly with steatosis.

Figure 5: Liver of C57BL/6NTac mouse on NASH diet D09100310i for 27 weeks
displays moderate diffuse hepatomegaly with steatosis. This mouse
(Male M000671) displayed a flat lesion flush to the medial lobe.

Body weight and liver weight for Diet Induced NASH B6 mice on diet for 27 weeks and age-matched B6 controls