Objective To analyze the chemical composition and characterize the polysaccharide structure of Shancigu (Cremastrae Pseudobulbus Pleiones Pseudobulbus) polysaccharide (CPP), and investigate its anti-hepatocellular carcinoma (HCC) efficacy and mechanism. Methods Total sugar and protein contents of CPP samples were determined using phenol-sulfuric acid method and Coomassie brilliant blue method, respectively. Structural characterization was performed by UV spectroscopy, infrared spectroscopy, scanning electron microscopy, PMP pre-column derivatization-HPLC and gel permeation chromatography. Male Balb/c mice were inoculated with H22 primary cells under the right forelimb axilla, mice were randomly divided intomodel, sorafenib (30 mg/kg) group and CPP low-, medium-. high-dose (200, 400, 800 mg/kg) groups, with 10 mice per group, another 10 normal mice were selected as the control group. Administration was performed for 16 consecutive days, tumor inhibition rates were calculated. Hematoxylin-eosin (HE) staining was used to observe histopathological changes in tumor tissues. Tumor tissues from model group and CPP medium-dose group were analyzed using transcriptomic sequencing to elucidate the mechanism of CPP’s anti-HCC activity. Key signaling targets and pathways of CPP against HCC were investigated through Kyoto encyclopedia of genes and genomes (KEGG) and gene ontology (GO) enrichment analysis. In vivo mechanism validation was performed using Western blotting, qRT-PCR and ELISA. Results CPP contained 95.19% total sugar and 0.33% protein. The structural characterization results showed that CPP was a pyran sugar ring connected by α-glycosidic bonds, irregularly coiled, composed of mannose and glucose (with a mass ratio of 2.388∶1), and had a weight average relative molecular weight of 1.27 × 10⁵. The animal experiment results showed that CPP low-, medium- and high-dose groups had tumor inhibition rates of 45.13%, 48.68% and 35.94% on H22 tumor bearing mouse models, respectively. The optimal dose for anti-HCC was 400 mg/kg. Histopathological sections revealed significant tumor tissue lesions in model group, with improvements observed across all treatment groups, particularly marked nuclear fragmentation in CPP medium-dose group. Transcriptomic analysis suggested CPP may exert its anti-HCC effects by modulating NOD like receptor family pyrin domain containing 3 (NLRP3) signaling pathway. Western blotting results showed that compared with model group, the expression levels of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), cystein-asparate protease-1 (Caspase-1) and gasdermin D (GSDMD) in tumor tissues of mice in CPP medium-dose group were significantly reduced (P < 0.05, 0.01, 0.001). ELISA results showed that compared with model group, the levels of interleukin-1β (IL-1β) and IL-18 in tumor tissues of CPP medium-dose group were significantly reduced (P < 0.001). Conclusion The anti-HCC effect of CPP may be related to the inhibition of NLRP3 inflammasome activation and blockade of Caspase-1/GSDMD mediated cell apoptosis pathway.

Cremastrae Pseudobulbus Pleiones Pseudobulbus polysaccharide  /  hepatocellular carcinoma  /  transcriptomics  /  pyroptosis  /  NLRP3 signaling pathway  /  mannose  /  glucose