The effect of aldafermin expressing-Escherichia coli Nissle 1917 along with dietary change on visceral adipose tissue in MASLD mouse model

Abstract

Background: Visceral adipose tissue (VAT) accumulation in obesity has been implicated as a key factor in the development of metabolic dysfunction-associated steatotic liver disease (MASLD). Apart from lifestyle change interventions, there is no effective therapy against MASLD. In this study, the effect of a novel microbial therapy along with dietary change on VAT and VAT-liver crosstalk was evaluated in a MASLD mouse model.

Methods: MASLD was induced by feeding eighteen C57BL/6J male mice with the American Lifestyle-Induced Obesity diet for fourteen weeks. Subsequently, during the following seven weeks, all mice were switched to standard diet and the intervention group received single gelatine cubes containing 10⁹ CFU each of aldafermin-expressing Escherichia coli Nissle (EcNA, n = 6); while the control groups received either 10⁹ CFU/gelatine cube of non-modified Escherichia coli Nissle (EcN, n = 6) or gelatin cube with no treatment (CTRL, n = 6). The effect of EcNA on epididymal visceral adipose tissue (eVAT) morphology was evaluated by histology and the gene expression profile in eVAT and liver by RNA-sequencing analysis.

Results: After seven weeks of intervention, EcNA, when compared to CTRL group, induced smaller adipocytes (p-value = 0.0217 for diameter, p-value = 0.0386 for area). Gene Set Enrichment Analysis in eVAT showed significant upregulation of fatty acid metabolism (FDR-adjusted p-value = 0.001), oxidative phosphorylation (FDR-adjusted p-value < 2.2e-16), peroxisome (FDR-adjusted p-value = 0.0185), and thermogenesis (FDR-adjusted p-value = 0.0199) pathways when EcNA was compared to EcN group. In addition, the impact of EcNA in eVAT-liver gene expression crosstalk was underlined by the upregulation of Bcl6 and Cnst expression in both tissues when EcNA was compared to CTRL and EcN groups.

Conclusions: These results support the beneficial effects of EcNA, along with dietary change intervention, in obesity-associated MASLD. This microbial therapy could potentially boost the improvements induced by dietary change in eVAT metabolism and its crosstalk with the liver.

Fig. 1. EcNA intervention reduced the size of adipocytes in eVAT.

A Representative picture of H&E staining after seven weeks of intervention, n = 6 mice/group, scale bar 100 µm, 10× objective. Quantification of adipocyte by using Adiposoft [19] B diameter (µm) and C area (µm²), n = 6 mice/group, 6 acquisitions/mouse. Chart results were presented as average values, error bars denote SD. eVAT epididymal adipose tissue; H&E Haematoxylin and Eosin.

Fig. 2. eVAT and liver gene expression change in EcNA intervention group.

A Venn diagram showing the number of DEGs and B Volcano plot showing the proportion of up- (in red) and downregulated (in blue) genes with FDR-adjusted p-value < 0.05 in EcNA compared to either CTRL or EcN in both, in eVAT and the liver. eVAT epididymal visceral adipose tissue; DEGs differentially expressed genes.

Fig. 3. EcNA intervention induced changes in gene expression pathways in eVAT and liver.

Bar plots showing the upregulated (in blue) and downregulated (in orange) pathways in A EcNA vs EcN in eVAT, B EcNA vs CTRL in eVAT, C EcNA vs EcN in the liver and D EcNA vs CTRL in the liver. Pathways with FDR-adjusted p-value < 0.05 were considered significant. eVAT: epididymal visceral adipose tissue. Details of the pathways can be found in Supplementary Table 9.

Fig. 4. EcNA effect on shared gene expression patterns between eVAT and liver.

A Heatmaps showing the log2foldchange values (positive in red and negative in blue) of the overlapped DEGs with FDR-adjusted p-value < 0.05 between eVAT and the liver in EcNA compared to EcN or CTRL groups. B Bar plots showing the upregulated (in blue) and downregulated (in orange) pathways in EcNA vs CTRL in both eVAT and the liver. Pathways with FDR-adjusted p-value < 0.05 were considered significant. eVAT epididymal visceral adipose tissue; DEGs differentially expressed genes. Details of the pathways can be found in Supplementary Table 9.

Fig. 5. Correlation analysis showing the eVAT-liver gene expression crosstalk in EcNA intervention group.

A Diagram showing the analysis approach for Spearman correlation analysis. B heatmap showing the Spearman correlation analysis done using the FPKM of the 38 overlapped DEGs in both eVAT and liver in EcNA compared to CTRL. C Heatmap showing the Spearman correlation analysis done using the FPKM of the 19 overlapped DEGs in both eVAT and liver in EcNA compared to EcN. Positive correlation in red and negative correlation in blue. White asterisks identified notable correlations. FPKM Fragments Per Kilobase per Million mapped fragments; DEGs differentially expressed genes; eVAT epididymal visceral adipose tissue.