Bioinformatics and experimental verification to explore the potential mechanism of ginsenoside Rg3 suppresses hepatocellular carcinoma progression.

Ginsenoside Rg3 is an extract from ginseng and has the activities of antitumor in a variety of carcinomas, making it a promising monomer of Chinese herbal medicine for tumor treatment. This study aimed to investigate the targets and mechanisms of action of Rg3 in hepatocellular carcinoma (HCC). The molecular structure of Rg3 was obtained, and 100 potential targets were identified using the SwissTargetPrediction database. Univariate Cox regression analysis on the TCGA-LIHC cohort identified 25 genes as candidate targets of Rg3 with significant prognostic relevance. Consensus clustering divided the TCGA-LIHC cohort into two subtypes: C1 and C2. The C2 subtype exhibited worse overall survival, and significant differences in the expression patterns of the 25 candidate genes were observed between the subtypes. Clinical characteristics also differed significantly between the C1 and C2 subtypes. Mutation analysis showed a higher mutation rate and specific gene mutations in the C2 subtype. Increased immune cell infiltration, including macrophages, Th1 cells, and Th17 cells, was observed in the C2 subtype. Here, an orthotopic murine HCC model was established using Hepa1-6 (murine liver carcinoma cells) in immunocompetent C57BL/6 mice. Rg3 was administered via intraperitoneal injection. Tumor volumes were measured using calipers, and the volume was calculated using the formula: width^2 × length × 0.5. This method, while traditional, has been validated and provides consistent and reliable data for assessing tumor progression and treatment efficacy. The function of Rg3on tumor growth and angiogenesis was evaluated in vitro and in vivo. Especially the role of Rg3 in regulating the myeloid-derived suppressor cells (MDSCs) recruitment and Kupffer cells function were investigated. In the present study we found that Rg3 administration suppressed tumor growth and angiogenesis in vivo. In the liver tissues of HCC-bearing mice, Kupffer cells expressed more co-inhibitory molecule CD274 and less co-stimulatory molecules CD86 and MHCII, whereas Rg3 treatment induced the restoration of Kupffer cells function. Rg3 also increased the efficiency of Kupffer cells as antigen-presenting cells. Furthermore, an increased MDSCs proportion in tumor tissues and surrounding parenchyma was detected in HCC-bearing livers, and this enhancement was blocked after Rg3 administration. In vitro, co-culture of Kupffer cells with MDSCs resulted in decreased CD86 expression and increased CD274 expression in Kupffer cells. Kupffer cells also produced decreased IL-6 and IL-18 level and upregulated IL-10 level after co-culture with MDSCs. This study provides insights into the potential targets and mechanisms of Rg3 in HCC and lays a foundation for personalized treatment strategies.