Bufalin suppresses hepatocellular carcinogenesis by targeting M2 macrophage-governed Wnt1/ß-catenin signaling.

BACKGROUND: The interplay of tumor-associated macrophages (TAMs) and tumor cells plays a key role in the development of hepatocellular carcinoma (HCC) and provides an important target for HCC therapy. The communication between them is still on the investigation. Bufalin, the active component derived from the traditional Chinese medicine (TCM) Chansu, has been evidenced to possess anti-HCC activity by directly suppressing tumor cells, while its immunomodulatory effect on the tumor microenvironment (TME) is unclear. PURPOSE: To explore the mechanism of M2 TAM-governed tumor cell proliferation and the inhibitory effect of bufalin on HCC growth by targeting M2 macrophages. METHODS: Morphology and marker proteins were detected to evaluate macrophage polarization via microscopy and flow cytometry. Cellular proliferation and malignant transformation of HCC cells cultured with macrophage conditioned medium (CM) or bufalin-primed M2-CM, were assessed by cell viability, colony formation and soft agar assays. Regulations of gene transcription and protein expression and release were determined by RT-qPCR, immunoblotting, immunoprecipitation, ELISA and immunofluorescence. Tumorigenicity upon bufalin treatment was verified in orthotopic and diethylnitrosamine-induced HCC mouse model. RESULTS: In this study, we first verified that M2 macrophages secreted Wnt1, which acted as a mediator to trigger ß-catenin activation in HCC cells, leading to cellular proliferation. Bufalin suppressed HCC cell proliferation and malignant transformation by inhibiting Wnt1 release in M2 macrophages, and dose-dependently inhibited HCC progression in mice. Mechanistically, bufalin specially targeted to block Wnt1 transcription, thus inactivating ß-catenin signaling cascade in HCC cells and leading to tumor regression in HCC mouse model. CONCLUSION: These results clearly reveal a novel potential of bufalin to suppress HCC through immunomodulation, and shed light on a new M2 macrophage-based modality of HCC immunotherapy, which additively enhances direct tumor-inhibitory efficacy of bufalin.

Study on the action mechanism of the Polygonum perfoliatum L. on non-alcoholic fatty liver disease, based on network pharmacology and experimental validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine (TCM) holds that non-alcoholic fatty liver disease (NAFLD) belong to the category of "thoracic fullness". Polygonum perfoliatum L. (PPL), a Chinese medicinal herb with the effect of treating thoracic fullness, was recorded in the ancient Chinese medicine book "Supplements to Compendium of Materia Medica". It has been used since ancient times to treat NAFLD. However, the underlying mechanism and active components of PPL against NAFLD remains unclear. AIM OF STUDY: To identify the main active components and the anti-NAFLD mechanism of PPL. MATERIALS AND METHODS: Network pharmacology, UPLC/QE-HFX analysis, and molecular docking were employed to determine the main bioactive compounds and key targets of PPL for the NAFLD treatment. This effect was further validated with administration of PPL (200 mg/kg and 400 mg/kg) to NAFLD model mice for 5 weeks. Systemic signs of obesity, biochemical parameters, and histological changes were characterized. Immunohistochemistry, western blot, and PCR analysis were conducted to elucidate the mechanistic pathways through which PPL exerts its effects. RESULTS: Network pharmacology revealed 77 crossover genes between the PPL and NAFLD. The kyoto encyclopedia of genes and genomes (KEGG) analysis show that PPL treat NAFLD mainly regulating glucose-lipid metabolism mediated by PI3K/AKT signal pathway. The Gene Ontology (GO) enrichment analysis show that PPL treat NAFLD mainly regulating inflammation mediated by cytokine-mediated signaling pathway. In accordance with the anticipated outcomes, administration of PPL in a dose-dependent manner effectively mitigated insulin resistance induced by a high-fat diet (HFD) by activating the PI3K/AKT signaling pathway. Histopathological evaluation corroborated the hepatoprotective effects of PPL against HFD-induced hepatic steatosis, as evidenced by the inhibition of de novo fatty acid synthesis and promotion of fatty acid ß-oxidation (FAO). Further research showed that PPL blocked cytokine production by inhibiting the NF-κB pathway, thereby reducing immune cell infiltration. Furthermore, five flavonoids from PPL, including quercetin, baicalein, galangin, apigenin, and genistein were identified as key compounds based on ingredient-target-pathway network analysis. Molecular docking show that these active compounds have favorable binding interactions with AKT1, PIK3R1, and MAPK1, further confirming the impact of PPL on the PI3K/AKT pathway. CONCLUSIONS: Through the combination of network pharmacology prediction and experimental validation, this work determined that therapeutic effect of PPL on NAFLD, and such protective effect is mediated by activating PI3K/AKT-mediated glucolipid metabolism pathway and hepatic NF-κB-mediated cytokine signaling pathway.

An Integrated Strategy for Investigating Antioxidants from Ribes himalense Royle ex Decne and Their Potential Target Proteins.

Natural products have been used extensively around the world for many years as therapeutic, prophylactic, and health-promotive agents. Ribes himalense Royle ex Decne, a plant used in traditional Tibetan medicine, has been demonstrated to have significant antioxidant and anti-inflammatory properties. However, the material basis of its medicinal effects has not been sufficiently explored. In this study, we established an integrated strategy by online HPLC-1,1-diphenyl-2-picrylhydrazyl, medium-pressure liquid chromatography, and HPLC to achieve online detection and separation of antioxidants in Ribes himalense extracts. Finally, four antioxidants with quercetin as the parent nucleus were obtained, namely, Quercetin-3-O-ß-D-glucopyranoside-7-O-α-L-rhamnopyranoside, Quercetin-3-O-ß-D-xylopyranosyl(1-2)-ß-D-glucopyranoside, Quercetin-3-O-ß-D-glucopyranoside, and Quercetin-3-O-ß-D-galactoside. Until now, the four antioxidants in Ribes himalense have not been reported in other literatures. Meanwhile, the free-radical-scavenging ability of them was evaluated by DPPH assay, and potential antioxidant target proteins were explored using molecular docking. In conclusion, this research provides insights into the active compounds in Ribes himalense which will facilitate the advancement of deeper studies on it. Moreover, such an integrated chromatographic strategy could be a strong driver for more efficient and scientific use of other natural products in the food and pharmaceutical industries.

Risk assessment of genotoxic and carcinogenic alkenylbenzenes in botanical containing products present on the Chinese market.

In the present study, a risk assessment of plant food supplements (PFS), traditional Chinese medicines (TCM) and herbal teas containing alkenylbenzenes was performed using the Margin of Exposure (MOE) approach. The levels of alkenylbenzenes in botanical preparations collected on the Chinese market were quantified and the combined estimated daily intake (EDI) was determined using dose additivity. The combined EDI values obtained assuming equal potency of all alkenylbenzenes detected in the PFS, TCM and herbal teas were 0.3 to 14.3, 0.05 to 539.4 and 0.04 to 42.5 µg/kg bw/day, respectively. Calculating combined EDI values taking into account the toxic equivalency (TEQ) approach, the values for PFS, TCM and herbal teas were 0.3 to 7.7, 0.05 to 278.0 and 0.02 to 16.5 µg estragole equivalents/kg bw/day, respectively. The MOE values resulting from consumption of these PFS, TCM and one cup of herbal tea per day during life-time were generally lower than 10 000, suggesting a potential priority for risk management. For short-term exposure such as two weeks consumption, applying Haber's rule, only one TCM 6 () still had an MOE value below 10 000. It is concluded that selected consumption of Chinese botanical preparations raise a concern because of exposure to alkenylbenzenes, especially when exposure is for longer periods of time.

Anti-microbial screening of endophytic fungi from Hypericum perforatum Linn.

Anti-microbial properties of 21 endophytic fungal strains from Hypericum perforatum Linn. were evaluated against three human pathogens, Staphyloccocus aureus, Escherichia coli and Rhodotorula glutinis, and two phytopathogens, Rhizoctonia cerealis and Pyricularia grisea. The results indicated that the ethyl acetate extracts of endophytic fermentation broth had stronger anti-microbial activities than their fermentation broth. And the inhibitory effect of the endophytic extracts on human pathogens was better than those on phytopathogens. Among these endophytic fungi, strains GYLQ-10, GYLQ-24 and GYLQ-22 respectively showed the strongest activities against S. aureu, E. coli, R. glutinis. GYLQ-14 and GYLQ-22 exhibited the most pronounced effect on P. Grisea while both GYLQ-06 and GYLQ-08 had the strongest anti-microbial activities against R. cerealis. Till now, this study is the first report on the isolation of endophytic fungi from H. perforatum Linn. and their anti-microbial evaluation.