Chrono-metabolic regulation of hepatic fibrosis via NR1D1-mediated hepatic stellate cell activation

Table 1 Key circadian regulators in hepatic stellate cell activation and liver fibrosis

Circadian gene/regulator	Model/cell type	Intervention/condition	Main findings	Mechanistic insight	Ref.
NR1D1/Rev-erbα	Mouse liver, HSCs	Circadian disruption/CCl4	Dysregulated NR1D1/Rev-erbα; enhanced HSC activation	Loss of NR1D1 promotes TGF-β signaling and ECM deposition	[1,10,20,24]
BMAL1	HSCs, LX2 cells	TGF-β1 induction	Downregulated BMAL1; increased glycolysis and HSC activation	BMAL1 inhibits phenotypic transformation via IDH1/α-KG-mediated glycolysis	[23,38,40]
CLOCK, Per1-3	HSCs	CCl4-induced fibrosis	Reduced CLOCK gene expression	Disruption of CC contributes to HSC proliferation and collagen synthesis	[18,21,37]
REV-ERBα	HSCs, mice	Melatonin/SR9009	Upregulation prevented HSC activation	Modulates circadian clock and PPARα signaling	[18,39]

This table summarizes the primary circadian clock components involved in hepatic stellate cell activation and hepatic fibrosis, including experimental models, interventions, main findings, mechanistic insights, and supporting references. HSC: Hepatic stellate cell; CC: Circadian clock; TGF-β: Transforming growth factor-beta; ECM: Extracellular matrix.

Table 2 Experimental evidence linking NR1D1 to HIF-1α inhibition and ammonia-mediated hepatic stellate cell activation

Pathway/target	Model/cell type	Intervention	Main findings	Mechanistic insight	Ref.
NR1D1–HIF-1α	LX2, primary HSCs	CCl4-induced fibrosis/NR1D1 overexpression	NR1D1 restoration suppressed HIF-1α, reduced α-SMA and collagen	NR1D1 inhibits HIF-1α signaling, reduces ammonia-mediated HSC activation	[20,29]
Ammonia	LX2, human HSCs	NH4Cl/hyperammonemia	Increased proliferation, ROS, ER stress, α-SMA, PDGF-Rβ	Ammonia promotes HSC activation; toxicity reversed by NR1D1	[12,41]
HIF-1α	HSCs	Hypoxia/metabolic stress	Upregulated fibrogenic genes	HIF-1α drives ammonia-induced fibrogenesis	[19,29]

This table summarizes experimental studies investigating the NR1D1–HIF-1α–ammonia axis, including models, interventions, main findings, mechanistic insights, and supporting references. HSC: Hepatic stellate cell; ROS: Reactive oxygen species; ER: Endoplasmic reticulum; α-SMA: Alpha-smooth muscle actin; PDGF-Rβ: Platelet-derived growth factor receptor beta.

Table 3 Recent preclinical studies of Hedyotis diffusa and other bioactive compounds targeting the NR1D1-HIF1-ammonia axis

Compound/extract	Model	Intervention	Main findings	Mechanistic insight	Ref.
Hedyotis diffusa	CCl4/HF mice, LX2	Hedyotis diffusa extract/injection	Reduced α-SMA, collagen; restored NR1D1	Modulates NR1D1–HIF1–ammonia axis; normalizes urea cycle	[21,29]
Dihydroartemisinin	CCl4 mice, HSCs	Dihydroartemisinin treatment	Restored lipid droplets in HSCs; inhibited activation	NR1D1-mediated Rab7 ubiquitination regulates lipophagy	[32,51,52]
Ferulic acid	LX2 cells, SD rats	TGF-β1/CCl4	Inhibited α-SMA, collagen, p-Smad 2/3	Blocks TGF-β/Smad signaling	[31,47]
Eriocitrin	TAA mice, LX2 cells	Eriocitrin treatment	Reduced inflammasome activation and collagen deposition	PPARα-mediated NLRP1/NLRC4 pathway	[33]
HDW extract	CCl4 mice	HDW treatment	Reduced HSC activation; improved liver function	Modulates gut microbiota, FXR/SHP/CYP7A1 pathway; chrono-metabolic effects	[21]
FA11	CCl4 mice	FA11 treatment	Reduced α-SMA, collagen	Inhibits TGF-β1-induced HSC activation	[46]
Ghrelin	CCl4 mice	Ghrelin treatment	Decreased HSC proliferation, ECM deposition	Modulates HIF-1α and ROS pathways	[45]
Physalin D	HSCs	PD treatment	Reduced HSC activation	Blocks TGF-β/Smad and YAP signaling	[49]

This table summarizes interventions, experimental models, main findings, mechanistic insights, and references for natural and pharmacological compounds affecting the NR1D1–HIF-1α–ammonia pathway. HSC: Hepatic stellate cell; ECM: Extracellular matrix; ROS: Reactive oxygen species; TGF-β: Transforming growth factor-beta.

Table 4 Key controversies in NR1D1–HIF-1α–ammonia axis research

Controversy	Evidence/context	Knowledge gap	Ref.
Disease-specific applicability	Most mechanistic studies performed in CCl4-induced fibrosis or NASH/MASH models; relevance to ALD, drug-induced fibrosis, congenital fibrosis, or NAFLD unclear	Uncertain if NR1D1–HIF-1α–ammonia axis functions similarly across diverse etiologies; need multi-model validation	[12,41,53-55]
Single-target vs integrated chrono-metabolic therapy	NR1D1 modulation alone shows anti-fibrotic effects; bioactive compounds (e.g., dihydroartemisinin, notoginsenoside R1) regulate multiple HSC pathways including lipophagy, PPAR-γ/TGF-β	Whether single-node targeting is sufficient vs combinatorial strategies integrating NR1D1, ammonia-lowering therapy, bile acid modulation, and circadian-aligned dosing	[20,21,29,32,55,56]
Clinical translation challenges	Variability in circadian rhythms, fibrosis stage, etiology-specific metabolic alterations; existing models often do not reflect human heterogeneity	Optimal dosing schedules, patient stratification, and model selection; incorporation of patient-derived systems for translation	[4,16,21,30,57]

This table summarizes key issues, supporting evidence, and gaps in knowledge for the NR1D1–HIF-1α–ammonia axis, highlighting areas requiring further investigation. HSC: Hepatic stellate cell; NASH: Nonalcoholic steatohepatitis; MASH: Metabolic dysfunction-associated steatohepatitis; ALD: Alcoholic liver disease; NAFLD: Nonalcoholic fatty liver disease.