Obese diet-induced mouse models of nonalcoholic steatohepatitis-tracking disease by liver biopsy.
OBJECTIVE
To characterize development of diet-induced nonalcoholic steatohepatitis (NASH) by performing liver biopsy in wild-type and genetically obese mice.


METHODS
Male wild-type C57BL/6J (C57) mice (DIO-NASH) and male Lep(ob) /Lep(ob) (ob/ob) mice (ob/ob-NASH) were maintained on a diet high in trans-fat (40%), fructose (22%) and cholesterol (2%) for 26 and 12 wk, respectively. A normal chow diet served as control in C57 mice (lean chow) and ob/ob mice (ob/ob chow). After the diet-induction period, mice were liver biopsied and a blinded histological assessment of steatosis and fibrosis was conducted. Mice were then stratified into groups counterbalanced for steatosis score and fibrosis stage and continued on diet and to receive daily PO dosing of vehicle for 8 wk. Global gene expression in liver tissue was assessed by RNA sequencing and bioinformatics. Metabolic parameters, plasma liver enzymes and lipids (total cholesterol, triglycerides) as well as hepatic lipids and collagen content were measured by biochemical analysis. Non-alcoholic fatty liver disease activity score (NAS) (steatosis/inflammation/ballooning degeneration) and fibrosis were scored. Steatosis and fibrosis were also quantified using percent fractional area.


RESULTS
Diet-induction for 26 and 12 wk in DIO-NASH and ob/ob-NASH mice, respectively, elicited progressive metabolic perturbations characterized by increased adiposity, total cholesterol and elevated plasma liver enzymes. The diet also induced clear histological features of NASH including hepatosteatosis and fibrosis. Overall, the metabolic NASH phenotype was more pronounced in ob/ob-NASH vs DIO-NASH mice. During the eight week repeated vehicle dosing period, the metabolic phenotype was sustained in DIO-NASH and ob/ob-NASH mice in conjunction with hepatomegaly and increased hepatic lipids and collagen accumulation. Histopathological scoring demonstrated significantly increased NAS of DIO-NASH mice (0 vs 4.7 ± 0.4, P < 0.001 compared to lean chow) and ob/ob-NASH mice (2.4 ± 0.3 vs 6.3 ± 0.2, P < 0.001 compared to ob/ob chow), respectively. Furthermore, fibrosis stage was significantly elevated for DIO-NASH mice (0 vs 1.2 ± 0.2, P < 0.05 compared to lean chow) and ob/ob NASH (0.1 ± 0.1 vs 3.0 ± 0.2, P < 0.001 compared to ob/ob chow). Notably, fibrosis stage was significantly (P < 0.001) increased in ob/ob-NASH mice, when compared to DIO-NASH mice.


CONCLUSIONS
These data introduce the obese diet-induced DIO-NASH and ob/ob-NASH mouse models with biopsy-confirmed individual disease staging as a preclinical platform for evaluation of novel NASH therapeutics.
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Maria Nicoline Baandrup
MN
Kristiansen
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Sanne Skovgård
SS
Veidal
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Kristoffer Tobias Gustav
KT
Rigbolt
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Kirstine Sloth
KS
Tølbøl
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Jonathan David
JD
Roth
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Jacob
J
Jelsing
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Niels
N
Vrang
Maria Nicoline Baandrup Kristiansen, Sanne Skovgård Veidal, Kristoffer Tobias Gustav Rigbolt, Kirstine Sloth Tølbøl, Jacob Jelsing, Niels Vrang, Michael Feigh, Gubra Aps, 2970 Hørsholm, Denmark.
Michael
M
Feigh
Diet-induced obesity; Fibrosis; Liver biopsy; Nonalcoholic fatty liver disease; Nonalcoholic steatohepatitis