Unraveling Hepatitis B-Induced Liver Fibrosis: The Critical Role of Pyruvate, PPARα, and Reactive Oxygen Species

Highlight

• HBV infection elevates pyruvate levels, enhancing reactive oxygen species (ROS) production and promoting liver fibrosis.
• Decrease in peroxisome proliferator-activated receptor α (PPARα) expression plays a key role in HBV-induced oxidative stress and fibrogenesis.
• Activation of PPARα or inhibition of ROS can reverse HBV and pyruvate-induced fibrosis, identifying potential antifibrotic therapeutic targets.
• These findings are supported across in vitro cell cultures, animal HBV models, and analyses of patient sera.

Study Background

Chronic hepatitis B (CHB) remains a significant global health burden, affecting over 250 million individuals worldwide and representing a leading cause of liver cirrhosis and hepatocellular carcinoma. Liver fibrosis, a wound-healing response characterized by excessive extracellular matrix deposition, is central to disease progression in CHB patients. Despite effective antiviral therapies, fibrosis and cirrhosis continue to develop in many individuals, underscoring an unmet need for elucidating molecular fibrogenic pathways directly connected to HBV infection and metabolic alterations. Previous observations revealed pyruvate, a key metabolite in cellular energy metabolism, enhances HBV replication. However, mechanistic insights linking pyruvate metabolism to HBV-induced fibrogenesis are lacking, limiting targeted therapeutic options for fibrosis in CHB. This study hypothesizes that HBV-induced liver fibrosis is mediated via a pyruvate-driven pathway causing inhibition of PPARα, a nuclear receptor regulating lipid metabolism and oxidative stress, subsequently triggering ROS generation and fibrogenic signaling.

Study Design

The study employed a comprehensive experimental setup integrating molecular cell culture models, animal HBV models, and clinical sample analyses:

1. Cell Models: HepAD38 cells (HBV-expressing hepatocytes), NTCP-HepG2 cells (HBV-infected human hepatocytes), primary human hepatocytes, and LX2 hepatic stellate cells (HSCs) were studied in isolated and co-culture environments to simulate hepatocyte-HSC interactions underpinning fibrosis.

2. Interventions: Pyruvate supplementation, PPARα agonists/antagonists, and ROS inhibitors were applied to dissect pathway components.

3. Animal Models: HBV carrier mice, HBV-transgenic (HBV-Tg) mice, and mice with humanized livers were utilized to confirm in vivo relevance.

4. Ex Vivo: Human precision-cut liver slices (PCLS) from patients provided translational insight into HBV-related fibrogenesis.

5. Clinical Correlation: Serum analyses from CHB patients quantified pyruvate and fibrosis-related biomarkers.

Endpoints included expression levels of profibrotic genes (TGF-β1, TIMP-1, COL1A1, α-SMA), ROS generation assays, PPARα protein expression, and histological fibrosis markers.

Key Findings

The investigation revealed a multifaceted mechanism of HBV-induced fibrosis involving metabolic and oxidative pathways:

1. Elevated Pyruvate and Fibrosis Markers: HBV infection significantly increased intracellular pyruvate levels and upregulated profibrotic gene expression in hepatocytes and patient sera. Pyruvate was independently capable of enhancing fibrogenic gene expression.

2. ROS Generation and PPARα Suppression: HBV infection and pyruvate supplementation led to elevated ROS production while concomitantly downregulating PPARα expression in hepatocytes. ROS is a known driver of hepatic stellate cell activation and ECM accumulation.

3. Modulation of PPARα Activity Alters Fibrogenesis: Pharmacologic activation of PPARα reduced ROS and profibrogenic gene expression induced by HBV and pyruvate, whereas PPARα antagonism or gene knockdown augmented these fibrotic responses.

4. ROS Function Downstream of PPARα: Application of ROS inhibitors effectively blocked liver fibrosis markers enhanced by HBV-conditioned media, pyruvate supplementation, or PPARα suppression, confirming ROS as a critical fibrogenic mediator downstream of PPARα inhibition.

5. Validation Across Models: These molecular insights were consistently demonstrated in cell culture, HBV-Tg and humanized liver mice, and human liver slice models, supporting physiological and clinical relevance.

Expert Commentary

This study provides compelling evidence for a novel metabolic-oxidative axis driving HBV-induced liver fibrosis via pyruvate-dependent ROS generation linked to PPARα suppression. The finding that pyruvate amplifies fibrogenic signaling while inhibiting PPARα aligns with emerging concepts of metabolic reprogramming regulating fibrosis progression. PPARα’s role in modulating oxidative stress and lipid metabolism is well established, and its downregulation by HBV unveils a critical pathway exploitable for therapy. Importantly, PPARα agonists and ROS scavengers reversed fibrogenic changes, unveiling practical translational avenues.

Nevertheless, the precise molecular mechanisms by which HBV regulates pyruvate metabolism and suppresses PPARα warrant further elucidation, including potential epigenetic or transcriptional regulators. Also, while preclinical models support these observations, clinical trials are essential to validate efficacy and safety of targeting pyruvate metabolism or PPARα pathways in CHB patients with liver fibrosis.

Conclusion

Chronic HBV infection promotes liver fibrosis through a pyruvate-dependent mechanism that suppresses PPARα expression, resulting in increased ROS production and activation of profibrotic pathways. Targeting pyruvate metabolism and restoring PPARα activity represent promising strategies to mitigate oxidative stress-mediated liver fibrosis in CHB. These insights expand our understanding of HBV pathogenesis beyond viral replication to include metabolic and redox alterations, offering new therapeutic targets to address the clinical burden of HBV-associated liver disease.

Funding and Clinical Trials

The reported study was supported by multiple institutional and governmental grants acknowledged in the original publication. No specific clinical trial registration was noted.

References

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2. Gaggini M, et al. Metabolic dysfunction and oxidative stress in chronic liver diseases. Nat Rev Gastroenterol Hepatol. 2020;17(8):437-452.
3. Puri P, et al. Hepatic oxidative stress and mitochondrial dysfunction in liver fibrosis. Am J Pathol. 2019;189(5):960–972.
4. Lefebvre P, et al. Role of PPARs in liver pathophysiology and fibrosis: From molecular mechanisms to therapeutic perspectives. Prog Lipid Res. 2021;83:101099.