MicroRNA expression profile in Lieber-DeCarli diet-induced alcoholic and methionine choline deficient diet-induced nonalcoholic steatohepatitis models in mice

Abstract

Background: Alcoholic and nonalcoholic steatohepatitis are leading causes of liver diseases worldwide. While of different etiology, these share common pathophysiological mechanisms and feature abnormal fat metabolism, inflammation and fibrosis. MicroRNAs (miRNA) are highly conserved noncoding RNAs that control gene expression at the post-transcriptional level either via the degradation of target mRNAs or the inhibition of translation. Each miRNA controls the expression of multiple targets; miRNAs have been linked to regulation of lipid metabolism and inflammation.

Methods: We fed Lieber-DeCarli alcohol or methionine-choline-deficient (MCD) diets to C57Bl6 and analyzed livers for histopathology, cytokines by ELISA, alanine aminotransferase (ALT) by biochemical assay, and microRNA profile by microarray.

Results: Both Lieber-DeCarli and MCD diets lead to development of liver steatosis, liver injury, indicated by increased ALT, and elevated levels of serum TNFalpha, suggesting that animal models portray the pathophysiological features of alcoholic and nonalcoholic fatty liver, respectively. We identified that Lieber-deCarli diet up-regulated 1% and down-regulated 1% of known miRNA; MCD diet up-regulated 3% and down-regulated 1% of known miRNA, compared to controls. Of miRNAs that changed expression levels, 5 miRNAs were common in alcoholic and nonalcoholic fatty livers: the expression of both miR-705 and miR-1224 was increased after Lieber-DeCarli or MCD diet feeding. In contrast, miR-182, miR-183, and miR-199a-3p were down-regulated in Lieber-deCarli feeding, while MCD diet lead to their up-regulation, compared to corresponding controls.

Conclusions: Our findings indicate etiology-specific changes in miRNA expression profile during steatohepatitis models, which opens new avenues for research in the pathophysiology of alcoholic and nonalcoholic fatty liver disease.

Figure 1

Lieber-deCarli and methionine-choline deficient (MCD) diet feeding feature liver steatosis, impaired fat metabolism, liver injury and pro-inflammatory activation. C57Bl6 mice were fed with Lieber-deCarli or mehionine-choline deficient diets to establish ASH and NASH models, respectively. Liver sections were stained with H&E and analyzed for morphology (A); serum levels of triglycerides (B), alanine aminotransferase (C), TNFα (D) and alcohol (E) were analyzed. In panel A, representative images out of n=6/group are shown. In panels B–E, data are shown as mean±SD from 6 mice/group; the * indicate p<0.05 compared to pair-fed controls; the # indicate p<0.05 compared to MCS controls.

Figure 2

Steatohepatitis modulates liver microRNA expression. Liver microRNA expression was analyzed using miRNA Microarray. Panels A and D show the percentage of microRNA that changed during feeding of Lieber-deCarli (A) and MCD (D) diets, compared to corresponding controls. Panels B and E show the heat-maps of microRNAs expression in Lieber-deCarli diet feeding at p<0.05 and p<0.01, respectively. Panels C and F show the expression of microRNAs in MCD diet feeding at p<0.05 and p<0.01, respectively. In panels B,C,E and F (shown n=3), the black color indicates no change, the red color represents up-regulation, and the green color indicates down-regulation of the gene expression; the intensity of the color correlates with the extent of changes.