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| Study on the effect of apoptosis and autophagy of pine pollen on human hepatocellular carcinoma SK-Hep-1 cells |
| LI Beilin DENG Yibin LUO Yanhong |
| Youjiang Medical University for Nationalities, Guangxi Zhuang Autonomous Region, Baise 533000, China |
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Abstract Objective To observe the effect of pine pollen on apoptosis and autophagy of human hepatocellular carcinoma SK-Hep-1 cells, and to explore the anti-tumor effect and mechanism of pine pollen, so as to provide experimental basis for the clinical application of pine pollen. Methods SK-Hep-1 cells were cultured in vitro and randomly divided into 4 groups: the control group, the pine pollen low-dose group (30 mg/mL), the pine pollen medium-dose group (45 mg/mL), and the pine pollen high-dose group (60 mg/mL). The inhibition rate of SK-Hep-1 cells was determined by CCK-8 method after pine pollen intervention at different concentrations at different time (12, 24, 48 and 72 h). After pollen treatment for 48 h, cell morphology was observed by phase contrast microscopy and Hoechst33258 fluorescence staining. The apoptosis rate was observed by flow cytometry and mRNA expression levels of autophagy were detected by RT-PCR. Results CCK-8 assay showed that pine pollen could significantly inhibit SK-Hep-1 cell proliferation in time and dose dependence. Phase contrast microscopy showed that pine pollen could inhibit cell proliferation. Hoechst33258 fluorescence staining showed that, compared with the control group, the pine pollen low-dose, medium-dose and high-dose groups showed obvious apoptotic changes in cell morphology. Annexin V-FITC/PI double staining results showed that the apoptosis rates in the pine pollen low-dose, medium-dose and high-dose groups were significantly increased compared with the control group, with statistically significant differences (P ＜ 0.05 or P ＜ 0.01). RT-PCR analysis showed that, compared with the control group, intervention of pine pollen intervention after 48 h, the expression of Beclin-1, LC3-Ⅱwas up-regulated obviously in the pine pollen low-dose, medium-dose and high-dose groups, but P62 expression was down-regulated, with statistically significant differences (P ＜ 0.05 or P ＜ 0.01). Conclusion Pine pollen can effectively inhibit the proliferation of human hepatocellular carcinoma SK-Hep-1 cells, promote their apoptosis, and induce their autophagy. Pine pollen may promote apoptosis of hepatocellular carcinoma cells by inducing autophagy.
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