Immuno-metabolism of liver resident macrophages in non-alcoholic fatty liver disease
Introduction: Non-alcoholic steatohepatitis (NASH), the progressive form of fatty liver
disease, is associated with hepatic inflammation and liver fibrosis. Typically, serum levels of
adiponectin, a potent anti-inflammatory cytokine, are reduced in NASH. “Immunometabolism” is an emerging field of research that investigates the co-regulation of metabolic
and inflammatory pathways within immune cells. Chronic exposure of the immune system to
particular nutrients can lead to metabolically triggered inflammation (meta-inflammation). In
this context tissue resident macrophages play a pivotal role in monitoring tissue function and
restoring metabolic homeostasis. We explored how the intrinsic metabolic state of liver
resident macrophages (Kupffer cells) change in NASH.
Materials and Methods: Male C57BL/6 adiponectin knockout (ADN) and wild type mice
were fed either normal chow (NC) or a high cholesterol (HC) diet for 12 weeks. At the end of
this period, serum and hepatic cholesterol were measured. Liver tissues were examined by
haematoxylin and eosin (H&E), and by CD68 immunostaining. For in vitro analyses of
immuno-metabolism, rat Kupffer cells were isolated and cultured in various concentrations of
glucose and treated with LPS +/- adiponectin, and palmitate. qPCR and western blots were
performed to determine mRNA expression and to study changes in signaling molecules in
liver tissue and kupffer cells.
Results: When fed the high cholesterol diet, both genotypes had similar increases in average
liver/total body weight ratio and reduced fat pad weight. Biochemistry and histology showed
that KO mice fed the HC (ADN-HC) diet had significantly increased ALT levels, and
elevated hepatic total (148 vs. 101 mg/dl, p<0.05), and free cholesterol (114 vs. 80 mg/dl,
p<0.05). ADN-HC livers had increased gene expression for TNFα (11-fold, p<0.05) CD68
(6-fold, p<0.05), IL-1β (1.4-fold, p<0.05), CCL19 (6.0-fold, p<0.01), liver pyruvate kinase
(L-PK), (WT-HC 6.7-fold, ADN-HC 5-fold, p<0.05), and 1.8-fold increase in expression of
RelB by western blot. In preliminary in vitro experiments, we find that treatment of Kupffer
cells with LPS leads to increased glucose uptake. We detected incremental increases in LPS
mediated activation of the NFkB pathway in macrophages cultured without glucose, low
glucose (5mM) and high glucose (25mM) media. Adiponectin increased glucose uptake and
adding adiponectin to the media increased NFkB activation, whereas palmitate reduced the
uptake of glucose and suppressed activation of NFkB pathways.
Conclusion: In vivo, adiponectin KO mice fed the high cholesterol diet had severe
inflammation, with glycolytic pathway activation. Likewise, exposure to glucose modified
the inflammatory status of Kupffer cells. These data suggest that Kupffer cell driven metainflammation plays an important role in hepatic immunometabolism and NASH
pathogenesis.