Anti-Tumor Activities And Primary Mechanism Of Polysaccharides From Paecilomyces Hepiali Mycelium Against N-Diethylnitrosamine-Induced Hepatocellular Carcinoma In Mouse

Paecilomyces Hepiali,an endophytic fungus,is separated from Cordyceps sinensis.Its mycelia polysaccharides have various pharmacological activities,such as anti-tumor,immunomodulatory,antioxidant and anti-inflammatory,while little has known about its effect on the prevention and treatment of primary liver cancer.Hepatocellular carcinoma is one of the most common malignant tumors with high morbidity and mortality.The development of methods for its prevention and treatment has been of great interest.The objective of the present study was to investigate the antiproliferation activity and the antitumor mechanisms of polysaccharide(PHps1)purified from Paecilomyces Hepiali in human hepatoma cells(HLE cells)and in Balb/c mice with N-diethylnitrosamine(DEN)induced liver injury and hepatocellular carcinoma-like lesions.Four P.Hepiali polysaccharide-rich fractions(PH60,PH70,PH80 and PH90)were obtained following hot water and alcohol precipitation with the antioxidant activities of PH90>PH70>PH80>PH60.One polysaccharide fraction(PHps1)was further purified from PH90 by Saphadex G100 column chromatography.The total ash content of PHps1 was 1.3%.HPLC analysis showed that the main molecular weight of PHps1 was 10.8 k Da.No absorbance peaks at 260 nm and 280 nm were found indicating no containing of protein and nucleic acid.FT-IR spectrum analysis showed that PHps1 contained-OH,C-H,C=O,C-O-C,C-O-H groups and a pyranose ring.The inhibitory activity of PHps1 on human hepatoma cells(HLE cells)was determined by 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide(MTT)methods and cell colony formation assay.The results showed that PHps1 significantly inhibited the growth,proliferation and colony formation of HLE cells.When the concentration of PHps1 was 800 μg/ml,the proliferation inhibition rate of HLE cells reached 50.5% after 48 h and 77.06% after 72 h.The colony formation experiment showed that the 600 μg/ml and 800 μg/ml of PHps1 significantly inhibited colony formation of HLE cells.In vivo,after 15 weeks administration,the liver-weight and body weight ratio of mice treated with PHps1 significantly increased compared with the model group(P<0.01).PHps1 prevented liver cell necrosis,inhibited the higher expression of proliferating cell nuclear antigen and Ki-67 antigen induced by the DEN.Levels of alanine aminotransferase,aspartate aminotransferase,alkaline phosphatase were also significantly decreased in PHps1 treated group(P<0.05),indicating a significantly preventive and therapeutic effects of PHps1 on DEN-induced liver injury and primary liver cancer-like lesions in mice.Furthermore,the levels of inflammatory cytokines such as interleukin(IL)-6,IL-1β,IL-10,tumor necrosis factor-α,interferon-γ in liver were significantly decreased in PHps1 treated group(P<0.05).The contents of immunoglobulin A in spleen and immunoglobulin M in plasma and spleen were significantly increased(P<0.05).The antioxidant defense was also stimulated by reducing reactive oxygen species(ROS)levels,increasing total antioxidant capacity(T-AOC)and superoxide dismutase(SOD)activities(P<0.05).Moreover,PHps1 reduced the levels of phosphor-nuclear factor-κB(P<0.001),and enhanced the expression of heme oxygenase-1(P<0.05),superoxide dismutase 1(P<0.01),nuclear factor-erythroid 2-related factor-2(P<0.05)and catalase(P<0.01)in liver of DEN-injected mice,indicating a significant immunomodulatory,antioxidant and anti-inflammatory activities.In conclusion,the study indicats that P.Hepiali polysaccharides can significant prevention and treatment effects on DEN induced liver injury and hepatocellular carcinoma-like lesions,and its activity may be realized by regulating p-NF-κB and Nrf2/HO-1 related signaling pathways.