USP7 Regulates Macrophage Polarization via PKM2-Mediated Metabolic Reprogramming in Severe Acute Pancreatitis

Study Background and Research Question

Severe acute pancreatitis (SAP) is a critical inflammatory condition marked by high morbidity and mortality, often leading to systemic inflammatory response syndrome and multi-organ failure. Despite advances in understanding its pathogenesis, effective therapies to interrupt the disease course remain scarce. Macrophages, as central players in the immune response, exhibit distinct functional phenotypes: the pro-inflammatory (M1) and anti-inflammatory (M2) states. Early in SAP, M1 macrophages infiltrate pancreatic tissue, intensifying inflammation, whereas subsequent M2 macrophage accumulation helps resolve tissue damage. Deciphering mechanisms that control macrophage polarization in SAP could thus identify new therapeutic avenues.

In this context, the reference study (Wu et al., 2025) investigates the role of ubiquitin-specific protease 7 (USP7) in regulating macrophage polarization, focusing on its interaction with pyruvate kinase M2 (PKM2), a pivotal glycolytic enzyme implicated in immune cell metabolic programming.

Key Innovation from the Reference Study

The core innovation of this research is the identification of a USP7–PKM2 metabolic axis that governs macrophage polarization during SAP. By elucidating how USP7 regulates PKM2 stability and activity through deubiquitination, the study connects metabolic reprogramming with inflammatory phenotype switching in macrophages. This mechanistic insight extends the field’s understanding of immune metabolism beyond cancer cells, highlighting the relevance of PKM2 in inflammatory disorders such as SAP. Furthermore, the study provides experimental evidence that pharmacological PKM2 inhibition can modulate disease outcomes, pointing toward new therapeutic strategies.

Methods and Experimental Design Insights

Wu et al. employed a comprehensive suite of in vivo and in vitro methodologies:

• Animal Models: SAP was induced in mice, with or without genetic or pharmacological modulation of USP7 and PKM2.
• Histological Assessment: Pancreatic tissue was examined for inflammation and macrophage infiltration using immunostaining and histopathology.
• Flow Cytometry and Immunofluorescence: Macrophage phenotypes (M1 vs. M2) were quantified in pancreatic tissue and cultured cells.
• Biochemical Assays: Serum amylase and lipase levels were measured as indices of pancreatitis severity. Pro-inflammatory cytokine expression was assayed via RT-qPCR and ELISA.
• Metabolic Profiling: Seahorse extracellular flux analysis quantified glycolytic (ECAR) and oxidative (OCR) metabolism in macrophages.
• Protein Interaction Studies: Co-immunoprecipitation (Co-IP) and ubiquitination assays delineated USP7’s effect on PKM2 stability and localization.
• Pharmacological Intervention: A selective PKM2 inhibitor was administered to SAP mice to assess its impact on disease phenotype and rescue experiments in USP7-deficient contexts.

This multi-pronged approach ensured robust mechanistic dissection and translational relevance.

Core Findings and Why They Matter

The study reports several pivotal findings:

• USP7 Expression is Upregulated in SAP: Pancreatic macrophages from SAP mice exhibit increased USP7 levels, correlating with heightened inflammation (Wu et al., 2025).
• USP7 Promotes M1 Macrophage Polarization: Genetic knockdown of USP7 decreases M1 polarization, reduces pro-inflammatory cytokines, and alleviates SAP severity, shifting macrophages toward an M2 (anti-inflammatory) phenotype.
• USP7–PKM2 Interaction Regulates Macrophage Metabolism: USP7 stabilizes PKM2 via deubiquitination, promoting its nuclear translocation and supporting glycolytic reprogramming characteristic of M1 macrophages. Loss of USP7 increases PKM2 ubiquitination, diminishes its pro-inflammatory function, and skews metabolism toward oxidative phosphorylation, favoring M2 polarization.
• Pharmacological PKM2 Inhibition Modulates Disease Outcome: Administration of a pyruvate kinase M2 inhibitor partially reverses the anti-inflammatory and protective effects of USP7 knockdown, confirming that USP7’s influence on macrophage polarization is PKM2-dependent.

Collectively, these data highlight metabolic reprogramming—specifically, aerobic glycolysis disruption—as a linchpin of immune cell function in SAP. The findings suggest that targeting PKM2, either genetically or pharmacologically, could provide a novel route to modulate inflammation in acute pancreatitis.

Comparison with Existing Internal Articles

The mechanistic role of PKM2 in immune and cancer cell metabolism has been previously explored in oncology and immunometabolism literature, as reflected in several internal resources. For example, “Targeting Cancer and Immune Metabolism: Strategic Insight” situates PKM2 inhibitor (compound 3k) at the intersection of cancer biology and immune modulation, underscoring its utility for dissecting metabolic reprogramming underlying both tumor proliferation and immune cell polarization. Similarly, “Optimizing Cancer & Immunometabolic Assays” details how selective pyruvate kinase M2 inhibitors, including compound 3k, enable researchers to probe the functional consequences of glycolytic flux in macrophages and tumor cells.

What distinguishes the reference study is its application of these metabolic principles to acute inflammatory disease rather than cancer, directly implicating PKM2 in the pathogenesis of SAP and demonstrating the therapeutic potential of PKM2 inhibition in this new context. This cross-domain relevance supports the broader use of PKM2-targeted agents in inflammatory research beyond oncology.

Limitations and Transferability

While the study provides robust mechanistic evidence in murine SAP models and cultured macrophages, several limitations should be considered:

• Translational Uncertainty: Findings in mouse models may not fully recapitulate human SAP pathophysiology. Further validation in human tissues and clinical settings is necessary.
• Specificity of PKM2 Inhibition: Although a selective PKM2 inhibitor was used, off-target effects cannot be entirely excluded, and long-term metabolic consequences remain to be characterized.
• Complexity of Inflammatory Networks: Macrophage polarization is influenced by numerous microenvironmental cues; targeting PKM2 may have broader immunometabolic effects that warrant careful investigation.

Despite these caveats, the study’s insights advance the conceptual framework for metabolic intervention in inflammatory diseases and provide a foundation for further translational research.

Protocol Parameters

• SAP induction in mice: Follow established protocols for cerulein or L-arginine administration to induce acute pancreatitis, with careful monitoring of disease severity and animal welfare.
• Macrophage polarization assays: Isolate peritoneal or pancreatic macrophages; phenotype using flow cytometry for M1 (CD86, iNOS) and M2 (CD206, Arg1) markers.
• PKM2 inhibitor administration: In mouse models, oral dosing of a selective PKM2 inhibitor at 5 mg/kg every two days for 31 days was effective in preclinical cancer studies, as reported in the product information; adjust duration and endpoints based on SAP model requirements.
• Metabolic profiling: Use Seahorse XF analysis to measure ECAR and OCR in polarized macrophages, assessing glycolytic versus OXPHOS flux.
• Protein interaction studies: Employ Co-IP and ubiquitination assays to validate USP7–PKM2 interactions in macrophage lysates.

Research Support Resources

Researchers interested in probing PKM2’s role in macrophage metabolism or modeling metabolic interventions in SAP can consider using the PKM2 inhibitor (compound 3k) (SKU B8217), a potent and selective small molecule available from APExBIO. This inhibitor has demonstrated efficacy in preclinical studies for both cancer and immunometabolic research, providing a validated tool for disrupting PKM2-dependent glycolytic pathways. For further context on experimental design and translational considerations, internal articles such as “Targeting Cancer and Immune Metabolism” and “Optimizing Cancer & Immunometabolic Assays” provide additional workflow guidance.