Pristimerin Attenuates Fibrogenesis and Inflammation Through TIPE1 ‐Regulated P2X7r Inflammatory Signaling Cascades in Hepatic Fibrosis

Pristimerin (Pris), a natural triterpenoid extracted from Celastrus orbiculatus Thunb. and Gymnosporia acuminata Hook. f., has various pharmacological activities, including anti-inflammatory and anti-tumor effects. Our study explored the influence of Pris on fibrogenesis and inflammatory modulation via the tumor necrosis factor-alpha-induced protein 8-like 1 (TIPE1)-P2X7r axis in hepatic fibrosis. We induced a model of hepatic fibrosis in mice through intraperitoneal administration of Thioacetamide (TAA). To elucidate the pathways associated with fibrosis, RNA sequencing was performed. HSC-T6 cells were stimulated with transforming growth factor-β (TGF-β) while LX-2 cells were treated with siRNA targeting TIPE1. Bone marrow-derived macrophages (BMDMs) were exposed to a combination of lipopolysaccharide and adenosine triphosphate (LPS/ATP) along with Pris to produce conditioned medium. RNA sequencing data indicated significant alterations in pathways involving TIPE1, NOD-like receptors, and neutrophil extracellular traps (NETs) in the fibrosis context. Pris decreased accumulation of extracellular matrix (ECM) and the epithelial-mesenchymal transition (EMT) in the fibrotic liver. Pris significantly reduced the secretion of inflammatory mediators, NETs, and fibrosis markers. Pris activated TIPE1 while suppressing P2X7r and NLRP3 (NOD-like receptor family pyrin domain-containing protein 3) expression in activated HSCs and in TAA-challenged mice. Furthermore, Pris diminished ECM expression and reversed macrophage-induced activation of HSCs, supporting previous findings. The absence of TIPE1 impaired Pris's ability to inhibit P2X7r in LX-2 cells. Additionally, while LPS/ATP stimulation upregulates NLRP3, GSDMD, and IL-23 in mouse BMDMs, Pris exerts a notable inhibitory effect on these pyroptosis factors. Thus, our study demonstrates that Pris alleviates hepatic fibrosis by regulating the TIPE1-P2X7r axis. This finding not only positions Pris as a promising therapeutic candidate but, more importantly, highlights the critical role of the TIPE1-P2X7r axis in hepatic fibrosis, integrating purinergic P2X7r signaling, NLRP3 inflammasome activation, macrophage-HSC crosstalk, and NETs formation into an integrated mechanistic network.