Background:
GVIA calcium-independent phospholipase A2 (iPLA2β) regulates homeostasis and remodeling
of phospholipid (PL). iPLA2β is one of modified genes of non-alcoholic fatty liver
disease (NAFLD) which leads to nonalcoholic steatohepatitis (NASH), end-stage liver
disease, and hepatocellular carcinoma. Feeding mice with methionine-choline-deficient
(MCD) diet provides a model of advanced NASH. As we have shown that iPLA2β regulates
bile acid (BA) homeostasis during autoimmune hepatitis (BBA,1852:1646 – 57,2015),
we here aim to explore whether iPLA2β deficiency could modulate MCD-induced NASH by
altering BA production and gene expression of BA synthesis and transporters.
Methods:
Female iPLA2β-null mice and age-matched wild-type (WT) were fed by MCD diet for 5
weeks. Liver and intestine samples were subjected to histology and immunohistochemistry
(IHC). BA profiles were analyzed by liquid chromatography mass spectrometry. Quantitative
RT-PCR and western blot were performed for gene expression analyses.
Results:
Compared with MCD-fed WT, MCD-fed iPLA2β-null mice were not protected from hepatic
steatosis, but showed an increase of hepatic stellate cell activation (α-smooth muscle
actin IHC) and biliary epithelial proliferation (cytokeratin 19 IHC). MCD feeding
of WT caused marked elevation of cholic acid, muricholic acid (MCA), tauromuricholic
acid (TMCA) in bile, portal vein, and inferior vena cava serum. Most of these BA species
were further increased by iPLA2β deficiency, and the increased BA in vena cava was
due to intestinal damage. Compared with MCD-fed WT, livers of MCD-fed iPLA2β-null
mice showed marked elevation of MCA and TMCA as well as BA classified as FXR antagonists.
Intestinal BAs were however not altered by iPLA2β deficiency. Exaggerated BA in MCD-fed
iPLA2β-null mice were accompanied by increased expression of hepatic BA synthesis
gene Cyp7a1 and decreased expression of bile salt export pump, which exports BA from
liver to bile. Furthermore, these MCD-fed mutant mice showed decreased expression
of farnesoid X-activated receptor (FXR) in liver as well as FXR and FGF15 in ileum.
As alterations of ER-stress may be responsible for the decrease of hepatic FXR, we
measured ER-stress markers. Compared to MCD-fed WT, livers of MCD-fed mutant mice
showed an increase of p-IRE1a, but a decrease of XBP-1 s, p-PERK, CHOP, and splicing
factor, Arg/Ser rich 3 expression.
Conclusion:
iPLA2β deficiency during MCD exacerbated BA within the enterohepatic circulation by
downregulating hepatic FXR and markedly altered ER stress markers. Hence, biliary
epithelial proliferation and hepatic stellate cell activation during iPLA2β and MCD
deficiency was likely a consequence of increased hepatic BA. Our results also highlight
an ER stress-FXR axis as a regulator of BA homeostasis, and this can be used as a
therapeutic target for inhibiting fibrosis in advanced NASH.
