Inhibition of adipose triglyceride lipase alleviates NASH in diabetic mice via impaired Pparasignalling favouring hydrophilic bile acid composition

Background:  Metabolic comorbidities such as diabetes, obesity, and metabolic syndrome play a crucial role in the pathogenesis and progression of metabolic dysfunction-associated steatotic liver disease (MASLD). Inhibition of adipose triglyceride lipase (ATGL/PNPLA2) activity in insulin resistance and MASLD is an attractive therapeutic target. This study evaluated the impact of Atglistatin-mediated ATGL inhibition on non-alcoholic steatohepatitis (NASH) development in diabetic and hyperlipidaemic mice.

Materials and methods:  Streptozotocin-injected male mice were fed an HFD to induce NASH. Atglistatin (ATGLi) was administered on week 4 for four weeks. We analysed liver histology, hepatic lipid content, immunohistochemistry, RNA sequencing, and serum biochemistry. Additionally, we treated Caco2 cells, human primary extrahepatic cholangiocyte organoids (ECO), and HepG2 ATGL KD cells with the human ATGL inhibitor, NG-497, and Ppara agonist WY14643.

Results:  Atglistatin reduced body weight (-2.54 g) despite a comparable food intake between the STAM and Atglistatin groups. The bodyweight reduction was due to fat (-40%) and liver mass (-23%) reduction. Atglistatin treatment significantly improved fasting blood sugar (-12%). In line with a trend for reduced ALT (-45%), AP (-25%), total acylglycerol, and cholesterol contents, there was a significant improvement in histological liver injury assessed by H&E staining and NAFLD score (-45%). The NAFLD score revealed improvement of steatosis (-60%) and inflammation (-35%) but not ballooning in the Atglistatin group. Immunochemistry showed that Atglistatin treatment significantly reduces Mac2-positive cells in the liver (-50%) and gonadal white adipose tissues (gWAT) (-60%) compared to the untreated STAM group, indicating a reduced infiltration of immune cells into the liver and gWAT. Mechanistically, the NGS data revealed that Atglistatin reduced Cpt1a, Acox1, and Ppara gene expression, suggesting impaired Ppara signalling that favours hydrophilic bile acids synthesis. In line, Cyp7a1, Cyp27a1, and Cyp2c70 were increased while Cyp8b1 was reduced. Accordingly, Intestinal lipid transporters in the Atglistatin mice, like Abca1 and Cd36, were reduced, consistent with the reduction of liver TAG species, mostly linoleic acids. Because of the lack of efficacy of Atglistatin in humans, we used the novel human-specific ATGL inhibitor NG-497 to validate the in vivo findings. The NG-497-treated Caco2 cells showed reduced ATGL, ABCA1, FATP5, and MTTP mRNA transcripts. PPARA signalling was impaired in ECO treated with NG-497 and HepG2 ATGL KD cells since PPARA, CPT1A, and ABCA1 were reduced, and CYP7A1 increased. Furthermore, gel-shift analysis showed reduced PPARA binding activity in Hepa 1c1c7 and HepG2 cells treated with the Atglistatin and NG-497 respectively. Luciferase analysis also revealed reduced Ppara transcriptional activity in response to the Atglistatin treatment in Hepa 1c1c7 cells.

Conclusions:  Impaired Ppara signalling translated into hydrophilic BA synthesis that interferes with dietary lipid absorption, improving the metabolic disturbances in the STAM model of NASH associated with diabetes. The validation with the human-specific ATGL inhibitor should open a new clinical avenue for MASLD clinical trial and treatment.

 

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Artikel online veröffentlicht:
05. Juni 2024