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Pre- and post-sexual maturity liver-specific ERα knockout does not impact hepatic mitochondrial function or steatosis but does alter gene expression

ABSTRACT: Compared to males, premenopausal women and female rodents are protected against hepatic steatosis and present with higher functioning mitochondria (greater hepatic mitochondrial respiration and reduced H2O2 emission). Despite evidence that estrogen action mediates female protection against steatosis, mechanisms remain unknown. Here we validated a mouse model with inducible reduction of liver ERα (LERKO) via AAV Cre. We phenotyped the liver health and mitochondrial function of LERKO mice (n=10-12 per group) on a short-term high-fat diet (HFD), and then tested if timing of LERKO induction at 2 timepoints (sexually immature: 4 wks old (n=11 per group) vs. sexually mature: 8-10 wks old (n=8 per group)) would impact HFD-induced outcomes. We opted for an inducible LERKO model due to known estrogen-mediated developmental programming, and report both receptor and tissue specificity with our model. Control mice were ERαfl/fl receiving AAV with GFP only. Results show that there were no differences in body weight/composition or hepatic steatosis in LERKO mice with either short-term (4 wk) or chronic (8 wk) high-fat feeding. Similarly, LERKO genotype nor timing of LERKO induction (pre vs post sexual maturity) did not alter hepatic mitochondrial O2 and H2O2 flux, coupling, or OXPHOS protein. Transcriptomic analysis showed that hepatic gene expression in LERKO was significantly influenced by developmental stage. Together, these studies suggest that hepatic ERα is not required in female protection against HFD-induced hepatic steatosis nor does it mediate sexual dimorphism in liver mitochondria function.

ORGANISM(S): Mus musculus

PROVIDER: GSE228773 | GEO | 2023/05/24

REPOSITORIES: GEO

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Project description:Individuals with hepatic steatosis often display several metabolic abnormalities including insulin resistance and muscle atrophy. Previously, we found that hepatic steatosis results in an altered hepatokine secretion profile, thereby inducing skeletal muscle insulin resistance via inter-organ crosstalk. In this study, we aimed to investigate whether the altered secretion profile in the state of hepatic steatosis also induces skeletal muscle atrophy via effects on muscle protein turnover. To investigate this, eight-week-old male C57BL/6J mice were fed a chow (4.5% fat) or a high-fat diet (HFD; 45% fat) for 12 weeks to induce hepatic steatosis, after which the livers were excised and cut into ~200 µm slices. Slices were cultured to collect secretion products (conditioned medium; CM). Differentiated L6-GLUT4myc myotubes were incubated with chow or HFD CM to measure glucose uptake. Differentiated C2C12 myotubes were incubated with chow or HFD CM to measure protein synthesis and breakdown, and gene expression via RNA sequencing. Furthermore, proteomics analysis was performed in chow and HFD CM. It was found that HFD CM caused insulin resistance in L6-GLUT4myc myotubes compared with chow CM, as indicated by a blunted insulin-stimulated increase in glucose uptake. Furthermore, protein breakdown was increased in C2C12 cells incubated with HFD CM, while there was no effect on protein synthesis. RNA profiling of C2C12 cells indicated that 197 genes were differentially expressed after incubation with HFD CM, compared with chow CM, and pathway analysis showed that pathways related to anatomical structure and function were enriched. Proteomic analysis of the CM showed that 32 proteins were differentially expressed in HFD CM compared with chow CM. Pathway enrichment analysis indicated that these proteins had important functions with respect to insulin-like Growth Factor transport and uptake, and affect post-translational processes, including protein folding, protein secretion and protein phosphorylation. In conclusion, the results of this study support the hypothesis that secretion products from the liver contribute to the development of muscle atrophy in individuals with hepatic steatosis.

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Pre- and post-sexual maturity liver-specific ERα knockout does not impact hepatic mitochondrial function or steatosis but does alter gene expression

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