Protease Inhibitor VR23 Alleviates Severe Acute Pancreatitis with High Trypsin Synthesis in Mice

Abstract

Background: Acute pancreatitis (AP), characterized by pancreatic autodigestion and systemic inflammation, remains a life-threatening condition with a high mortality rate in severe cases. Cholinergic signaling through muscarinic receptors (mAChRs) regulates pancreatic exocrine secretion, yet the role of chronic cholinergic hyperactivation in AP progression is poorly understood. Protease inhibitors like VR23, a potent trypsin-selective compound, hold therapeutic potential but lack evaluation in AP models.

Objective: This study aimed to investigate the effects of prolonged cholinergic stimulation on AP severity and evaluate the efficacy of VR23 in mitigating pancreatic injury.

Methods: Male C57BL/6 mice were pretreated with intragastric carbachol (5 mg/kg, twice daily for 7 days) to mimic chronic cholinergic hyperactivation, followed by intraperitoneal caerulein (100 μg/kg, 10 hourly doses) to induce AP. VR23 (30 mg/kg) was administered intraperitoneally 12 h and 3 h before caerulein. Pancreatic injury was assessed via histopathology (HE-staining), serum amylase/lipase activity, western blot (trypsin/amylase), RT-qPCR (TNF-α, IL-1β, IL-6, IL-18), MPO immunohistochemistry (neutrophil infiltration), HMGB1 immunohistochemistry (necrosis) and TUNEL staining (apoptosis).

Results: Long-term carbachol stimulation induced acinar cell hypertrophy and elevated intracellular trypsin synthesis, exacerbating caerulein-induced AP with increased pancreas-to-body weight ratio, histopathological scores, serum amylase/lipase, and pro-inflammatory cytokines. Neutrophil infiltration and apoptosis were significantly amplified. VR23 pretreatment attenuated pancreatic injury, reducing histopathological scores, serum enzymes, inflammatory cytokines, neutrophil infiltration, and apoptosis.

Conclusion: Long-term chronic cholinergic stimulation exacerbates AP by enhancing trypsin synthesis, inflammatory responses, and apoptosis. VR23 demonstrates therapeutic efficacy by inhibiting protease activity and modulating inflammation, highlighting its potential as a targeted treatment for AP.

Keywords: Acute pancreatitis; Carbachol; Inflammatory response; Muscarinic receptor; Protease inhibitor.