Hepatic transcriptome signatures in mice and humans with nonalcoholic fatty liver disease

Abstract

Background: Nonalcoholic fatty liver disease (NAFLD) is the main reason for cirrhosis and hepatocellular carcinoma. As a starting point for NAFLD, the treatment of nonalcoholic fatty liver (NAFL) is receiving increasing attention. Mice fed a high-fat diet (HFD) and hereditary leptin deficiency (ob/ob) mice are important NAFL animal models. However, the comparison of these mouse models with human NAFL is still unclear.

Methods: In this study, HFD-fed mice and ob/ob mice were used as NAFL animal models. Liver histopathological characteristics were compared, and liver transcriptome from both mouse models was performed using RNA sequencing (RNA-seq). RNA-seq data obtained from the livers of NAFL patients was downloaded from the GEO database. Global gene expression profiles in the livers were further analyzed using functional enrichment analysis and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway.

Results: Our results showed that the biochemical parameters of both mouse models and human NAFL were similar. Compared with HFD-fed mice, ob/ob mice were more similar in histologic appearance to NAFL patients. The liver transcriptome characteristics partly overlapped in mice and humans. Furthermore, in the NAFL pathway, most genes showed similar trends in mice and humans, thus demonstrating that both types of mice can be used as models for basic research on NAFL, considering the differences.

Conclusion: Our findings show that HFD-fed mice and ob/ob mice can mimic human NAFL partly in pathophysiological process. The comparative analysis of liver transcriptome profile in mouse models and human NAFL presented here provides insights into the molecular characteristics across these NAFL models.

Keywords: animal model; high-fat diet; nonalcoholic fatty liver disease; ob/ob mice; transcriptomics.

FIGURE 1

Basic parameters and histology in NAFL (nonalcoholic fatty liver) mice. (A) Body weight, (B) liver weight, and (C) percentage of body weight of C57BL/6J male mice fed on chow diet (CD) or high‐fat diet (HFD) for 8 weeks (n = 8 per group). (D) Body weight, (E) liver weight, and (F) percentage of body weight of C57BL/6J (wild type [WT]) or ob/ob mice at age 8 weeks (n = 5 per group). (G) Representative images of livers, H&E (hematoxylin and eosin) staining of livers, and Oil Red O staining of livers from mice fed 8 weeks on CD or HFD (n = 8 per group). Scale bar = 50 μm for H&E and Oil Red O staining. Arrows represent lipid droplets. Quantitative analysis of Oil Red O staining (n = 4 per group). (H) Representative images of livers, H&E staining of livers, and Oil Red O staining of livers from WT or ob/ob mice at age 8 weeks (n = 5 per group). Scale bar = 50 μm for H&E and Oil Red O staining. Arrows represent lipid droplets. Quantitative analysis of Oil Red O staining (n = 4 per group). All quantitative data for mice were presented as mean ± SEM (standard error of the mean). Statistical comparison between groups was performed using Student's t‐test. *p < 0.05, **p < 0.01, and ***p < 0.001.

FIGURE 2

Serum lipids and biochemical parameters in NAFL (nonalcoholic fatty liver) mice. (A) Serum total cholesterol (TC) and triglyceride (TG) levels in C57BL/6J male mice fed 8 weeks with CD (chow diet) or HFD (high‐fat diet) (n = 5–8 per group). (B) Serum TC and TG levels in WT (wild type) or ob/ob mice at age 8 weeks (n = 5 per group). (C) Serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels in mice fed 8 weeks with CD or HFD (n = 6–8 per group). (D) Serum AST and ALT levels in WT or ob/ob mice at age 8 weeks (n = 5 per group). All quantitative data for mice were presented as mean ± SEM (standard error of the mean). Statistical comparison between groups was performed using Student's t‐test. *p < 0.05, **p < 0.01, and ***p < 0.001.

FIGURE 3

Overview of hepatic gene expression profiles in CD (chow diet) versus HFD (high‐fat diet) mice, WT (wild type) versus ob/ob mice, and normal versus NAFL (nonalcoholic fatty liver) patients. (A) Numbers of DEGs (differential expression genes) in the livers of three groups. (B) Numbers of upregulated and downregulated genes in the livers of three groups. (C) Venn diagrams of DEGs. The cutoff for the differential expression is onefold. (D) The expression changes in 24 common DEGs in three groups. Volcano plots of mRNAs (messenger RNA) between (E) CD and HFD mice, (F) WT and ob/ob mice, and (G) normal people and NAFL patients. The plots were constructed by plotting –log10 adjusted p‐value on the y‐axis and log2 fold change on the x‐axis. Blue blots represent downregulated genes, red dots represent upregulated genes, and black blots represent mRNAs without significant difference.

FIGURE 4

Enrichment analysis of differences in two mouse models and NAFL (nonalcoholic fatty liver) patients. (A–C) The top 10 enriched GO (gene ontology) terms and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways of DEGs (differential expression genes). The left figure shows GO enrichment, and the right figure shows KEGG pathway analysis. (A) CD (chow diet) versus HFD (high‐fat diet), (B) WT (wild type) versus ob/ob mice, and (C) normal people versus NAFL patients.

FIGURE 5

Gene set analysis of differences in two mouse models and NAFL (nonalcoholic fatty liver) patients. Heat maps of log2 fold change for significantly regulated (A) lipid metabolism‐related genes, (B) fatty acid metabolism‐related genes, (C) glucose metabolism and insulin signaling‐related genes, (D) cholesterol metabolism‐related genes, (E) steroid metabolism‐related genes, (F) immunity‐related genes, and (G) monocyte recruitment and inflammation‐related genes in two mouse models and NAFL patients.

FIGURE 6

Comparison of differential expression response for NAFL (nonalcoholic fatty liver) pathway‐related genes in two mouse models and NAFL patients. Color scale of log2 fold changes: the lowest (blue) to the highest (red).