Caspase-1-mediated hepatic stellate cells activation in vivo and in vitro.

2026-03
Journal of ethnopharmacology 359

PubMed: 41443484
DOI: 10.1016/j.jep.2025.121066

Study Design

Population: chronic liver fibrosis mouse models and the human liver stellate cell line-LX2 cells stimulated by TGF-β1
Methods: Extracting Cs-4 polysaccharides using water extraction, alcohol precipitation, and weak anion-exchange chromatography; two models on chronic liver fibrosis were set up by intraperitoneal injection of 15% carbon tetrachloride (CCl4)-olive oil solution, and long-term methionine-choline-deficient (MCD) diet feeding with or without Cs-4 polysaccharides; LX2 cells were performed in vitro with varying the presence or absence of CSP-W administration

Ethnopharmacological relevance

Paecilomyces hepiali Chen (Cs-4), recognized as the classic deficiency-tonifiying medicine of traditional Chinese medicine (TCM), is the principal and most valuable medicinal species of Ophiocordyceps sinensis (known as Cordyceps sinensis (Berk.) Sacc., or 'Dong-Chong-Xia-Cao' in Chinese). According to the TCM theories, liver fibrosis results from 'deficiency of healthy qi' and 'accumulation due to yin and deficiency in liver and kidneys', and O. sinensis is traditionally believed to strengthen healthy qi, tonify yin and kidneys. And its efficacy in treating hepatic fibrosis has been widely proven by clinical applications over millennia and contemporary research. Despite extensive research and evidence supporting its beneficial effects on liver fibrosis, the primary active components and underlying mechanism of this substance remain unclear.

Aim of the study

Extracting Cs-4 polysaccharides using water extraction, alcohol precipitation, and weak anion-exchange chromatography to elucidate structural characteristics, and evaluate efficacy and potential mechanisms in the chronic liver fibrosis mouse models and in vitro studies.

Materials and methods

The structural characteristics of Cs-4 polysaccharides were elucidated and mainly characterized by FT-IR spectroscopy and monosaccharide composition. Subsequently, two models on chronic liver fibrosis were set up by intraperitoneal injection of 15 % carbon tetrachloride (CCl₄)-olive oil solution, and long-term methionine-choline-deficient (MCD) diet feeding with or without Cs-4 polysaccharides. Then, serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and hepatic histological evaluations were conducted to assess the effects of Cs-4 polysaccharides. Furthermore, α-smooth muscle actin(α-SMA), collagen type I (Col-Ⅰ), transforming growth factor-β1(TGF-β1), the NLR family, pyrin domain containing protein 3(NLRP3) and Caspase-1 expressions were examined using immunohistochemistry, Real-time quantitative PCR analysis (RT-qPCR) analysis and Western blot to investigate hepatic stellate cells (HSCs) activation and inflammatory responses. Additionally, the human liver stellate cell line-LX2 cells stimulated by TGF-β1 were performed in vitro with varying the presence or absence of CSP-W administration.

Results

CSP-W, neutral polysaccharides derived from Cs-4, is composed of Gal, Man, Xyl, Ara, Glc, Fuc, and Rha (molar ratio= 52.4: 35: 1.7: 6.8: 2.8: 0.5: 0.8). Additionally, CSP-W was observed to significantly reduce serum AST, ALT levels and liver histological injury in mice with chronic liver fibrosis induced by CCl₄ and an MCD diet. It also decreased collagen fiber deposition and HSCs activation in Sirius Red (SR) or Masson staining, along with a reduction in Col-Ⅰ, α-SMA expressions. Mechanistically, CSP-W lowered TGF-β1 levels, preventing Smad2/3 phosphorylation and activation, and markedly reduced NLRP3, Caspase-1, and IL-1β expressions in both CCl₄ and an MCD diet models. Furthermore, CSP-W administration in vitro in TGF-β1-stimulated LX2 cells resulted in the downregulation of markers associated with the TGF-β1/Smad signaling pathway and NLRP3/Caspase-1 signaling pathways.

Conclusions

As neutral polysaccharides, CSP-W exerted a notable curative impact on chronic liver fibrosis in mice induced by CCl₄ and MCD diet. The mechanism mainly hindered the activation of HSCs and diminished collagen fibers accumulation through downregulating the TGF-β1/Smad and NLRP3/Caspase-1 signaling pathways in vivo and in vitro. Our findings provided compelling evidence supporting the potential application of CSP-W for managing chronic liver fibrosis.

Research Insights

Supplement	Dose	Health Outcome	Effect Type	Effect Size	Source
Paecilomyces hepiali	—	Reduced Hepatic Inflammation Markers	Beneficial	Moderate	View source It also decreased serum AST, ALT levels and liver histological injury in mice with chronic liver fibrosis induced by CCl4 and an MCD diet.
Paecilomyces hepiali	—	Reduced Hepatic Stellate Cell Activation	Beneficial	Large	View source It also decreased collagen fiber deposition and HSCs activation... along with a reduction in Col-Ⅰ, α-SMA expressions.
Paecilomyces hepiali	—	Reduced Liver Fibrosis	Beneficial	Large	View source CSP-W exerted a notable curative impact on chronic liver fibrosis in mice induced by CCl4 and MCD diet.