HMGB1-mediated macrophage polarization to M2 phenotype promotes ureteral stricture: therapeutic potential of HMGB1 inhibitors.

Background: Ureteral stricture is a common condition characterized by fibrotic narrowing of the ureter, often resulting in obstruction and impaired renal function. The condition is typically associated with chronic inflammation, epithelial injury, and abnormal wound healing responses, leading to excessive deposition of extracellular matrix proteins. High Mobility Group Box 1 (HMGB1), a damage-associated molecular pattern (DAMP) protein, plays a key role in promoting inflammation and fibrosis in various tissues. However, its specific role in the pathogenesis of ureteral stricture remains unclear. Glycyrrhizin, a natural HMGB1 inhibitor, has shown therapeutic potential in alleviating inflammation and fibrosis in preclinical studies of other fibrotic diseases. This study explores the involvement of HMGB1 in ureteral fibrosis and evaluates the efficacy of glycyrrhizin as a potential therapeutic agent.

Patients and methods: Ureteral tissue samples were collected from seven patients with ureteral stricture, and a rabbit model of ureteral stricture was established using thermal injury to the left ureter. HMGB1 expression in both human and rabbit tissues was analyzed via Western blot. SV-HUC-1 cells were subjected to three distinct injury models: UVB irradiation, radiotherapy, and chemotherapy with cisplatin, to simulate ureteral damage and assess HMGB1 secretion and localization. THP-1 macrophages were treated with recombinant HMGB1 or conditioned medium from injured SV-HUC-1 cells, and macrophage polarization was evaluated using flow cytometry, with IL-4 stimulation serving as a positive control. The role of macrophage polarization in fibroblast activation was investigated using conditioned medium to treat MRC-5 fibroblasts, followed by Western blot and qPCR to measure the expression of activation markers. The impact of the TGF-β inhibitor SB-431542 on fibroblast activation was also assessed. Glycyrrhizin's therapeutic effects and toxicity were evaluated in vitro through biochemical assays and in vivo by histological analysis in rabbit models.

Results: HMGB1 levels were significantly elevated in fibrotic segments of ureteral tissues from both patients and rabbit models. In SV-HUC-1 cells, injury induced HMGB1 translocation from the nucleus to the cytoplasm and extracellular medium. Conditioned medium from injured SV-HUC-1 cells and direct exposure of THP-1 macrophages to recombinant HMGB1 protein both stimulated M2 polarization of macrophages, which secreted TGF-β and activated fibroblasts to upregulate α-SMA, FAP, FN1, and COL1A1. In contrast, direct exposure of fibroblasts to recombinant HMGB1 showed minimal effects on these markers, suggesting an indirect mechanism mediated by macrophages. Treatment with the TGF-β inhibitor SB-431542 effectively suppressed fibroblast activation induced by HMGB1-conditioned medium from macrophages, confirming the critical role of TGF-β in this pathway. Glycyrrhizin effectively inhibited HMGB1-induced macrophage polarization and fibroblast activation in vitro and improved ureteral stricture and hydronephrosis in the rabbit model without detectable toxicity.

Conclusion: HMGB1 contributes to ureteral stricture by promoting macrophage polarization and fibroblast activation, leading to extracellular matrix accumulation and fibrosis. Glycyrrhizin effectively inhibits these processes and alleviates ureteral stricture, demonstrating its potential as a therapeutic agent for ureteral fibrosis. These findings provide a basis for further exploration of HMGB1-targeted therapies.