Integrative study reveals NR1D1 mediates Hedyotis diffusa’s antifibrosis via hypoxia inducible factor-1/ammonia axis

Abstract

BACKGROUND

Reversal of hepatic fibrosis (HF) represents a potential cure for chronic liver pathologies; however, clinically approved agents targeting this process remain scarce. Emerging evidence from traditional Chinese medicine (TCM) suggests that Hedyotis diffusa (HD), a botanical agent related to TCM principles of liver pathogenesis, may exert therapeutic effects against fibrotic liver damage. Despite its historical use, the molecular mechanisms underlying its antifibrotic properties and regulatory pathways require systematic elucidation.

AIM

To elucidate the efficacy and potential mechanism of HD against HF and to explore potential therapeutic targets.

METHODS

Liquid chromatograph mass spectrometer revealed six bioactive components of HD injection (HDI) that enter the blood and liver. Network pharmacology using these components predicted related signaling pathways. A HF mouse model was induced by administration of 10% carbon tetrachloride for 8 weeks to validate the efficacy of HDI. Integrated Gene Expression Omnibus (GEO) mining and liver proteomics revealed the antifibrotic mechanism of HD, which was confirmed via target gene interference to elucidate upstream-downstream regulatory relationships.

RESULTS

Network pharmacology analysis suggests that HDI may ameliorate HF through the modulation of circadian rhythm, urea metabolism, and hypoxia inducible factor-1 (HIF-1) signaling. GEO data mining and hepatic proteomic profiling in a fibrotic mouse model confirmed the close associations between disease progression and dysregulation of these pathways. HDI intervention significantly restored expression of the circadian regulator NR1D1. Further mechanistic investigations revealed NR1D1 as an upstream regulator of HIF-1 signaling, urea cycle function, and ammonia metabolism. In vitro experiments demonstrated that ammonium chloride-induced ammonia accumulation promoted LX2 cell activation, which is potentially associated with mitochondrial dysfunction.

CONCLUSION

HD demonstrates unequivocal efficacy in combating HF, potentially by modulating HIF-1 and the urea cycle through its influence on circadian rhythm genes, with NR1D1 as a prominent representative target.

Keywords: Hedyotis diffusa; Hepatic fibrosis; NR1D1; Hypoxia inducible factor-1; Urea cycle; Ammonia metabolism

Core Tip: This study reveals a novel antifibrotic mechanism of Hedyotis diffusa, demonstrating that its bioactive components alleviate hepatic fibrosis by restoring expression of the circadian rhythm gene NR1D1 expression. NR1D1 acts as an upstream regulator, inhibiting hypoxia inducible factor-1 signaling and restoring urea cycle function, thereby reducing hepatic ammonia accumulation and interrupting the vicious cycle of ammonia-induced hepatic stellate cell activation. These findings position circadian rhythm-metabolism interplay as a promising therapeutic axis for liver fibrosis treatment.