Accumulation of succinate in the blood is a potential early indicator of metabolic dysfunction-associated steatotic liver disease (MASLD).

There is currently a need to identify new biomarkers to diagnose metabolic dysfunction-associated steatotic liver disease (MASLD). Here, using C57BL6N male mice fed a high-fat diet (HFD), we provide evidence that extracellular succinate buildup is a sex-dependent diagnostic marker for MASLD. Male mice fed the HFD for 2-weeks developed simple steatosis, which was associated with the plasma buildup of succinate to 50 μM. Feeding the mice this diet for up to 7 weeks advanced the condition to MASLD, resulted in cardiac fibrosis, and the further increased plasma succinate to 100 μM. Using Huh-7 hepatoma cells as a model, we found fructose overload increased the concentration of succinate in the culture media, and this was associated with mitochondrial dysfunction and the hyper production of mitochondrial hydrogen peroxide (mtH2O2). HepG2 hepatocellular blastoma cells subjected to fructose overload in culture also accumulated succinate in the media. Treatment of the Huh-7 and HepG2 cells exposed to fructose with ursodeoxycholic acid (UDCA) or its taurine-conjugated form, TUDCA, which are known to elicit protective hepatocellular effects by inducing antioxidant defenses, strongly inhibited succinate build up by preserving mitochondrial function and preventing H2O2 hyper-production. Finally, using our glutaredoxin-2 (Glrx2-/-) gene knockout mouse model on a C57BL6N background, we found deleting the Glrx2 gene in male mice completely abrogated the accumulation of succinate, cis-aconitate, and itaconate in plasma. Importantly, wild-type (Wt) or Glrx2-/-female littermates did not accumulate any of these metabolites in plasma when fed the HFD, which coincided with MASLD resistance. Collectively, our findings show succinate accumulates rapidly in the extracellular milieu in our mouse model for MASLD and cell culture models for hepatic lipotoxicity. These findings suggest the applicability of succinate as a biomarker of early MASLD particularly among males and especially in pediatric populations.