ABSTRACT
To investigate the effect of the total flavonoids in Scutellaria barbata(TF-SB) against autophagy in tumor cells in vivo, and further determine whether the mechanism is correlated with the PI3K/AKT/mTOR pathway, which lead to autophagy-induced tumor cell death. Melanoma-bearing mice were prepared and divided into control group, rapamycin group (Rap 1.5 mg•kg⁻¹), and high, middle and low-dose TF-SB (200, 100, 50 mg•kg⁻¹) groups. The groups were given drugs once a day for successively 11 days. The inhibitory effect of TF-SB on the growth of melanoma was determined by measuring tumor volume and tumor inhibition rate. TUNEL method was used to detect the apoptosis of tumor cells to further verify the antitumor activity of TF-SB. The protein expressions of LC3-Ⅰ and LC3-Ⅱ were detected by Western blot, and the relative expression of LC3-Ⅱ was calculated based on LC3-Ⅱ/LC3-Ⅰ. In addition, the effect of TF-SB on autophagy of tumor cells, the underlying molecular mechanism of TF-SB in inducing autophagy and PI3K/AKT/mTOR pathway marker protein phosphorylation were also studied. According to the results, TF-SB effectively inhibited melanoma growth in mice, reduced tumor volume, increased the tumor inhibition rate, and significantly increased tumor cell apoptosis index and the ratio of LC3-Ⅱ/LC3-I (P<0.05, P<0.01 or P<0.001). The protein expressions of p-PI3K, p-AKT and p-mTOR were also suppressed dramatically compared with those in control group (P<0.05, P<0.01 or P<0.001). In conclusion, the total flavonoids in S. barbata could inhibit the growth of melanoma in vivo by inducing autophagy and apoptosis of tumor cells, which may be correlated with suppression of PI3K/AKT/mTOR pathway.
ABSTRACT
This paper was aimed to investigate the relationship between autophagy and NLRP3 inflammasome activation by studying the effect oftotal flavonoids in Scutellaria barbata (TF-SB) on autophagy in tumor cells and NLRP3 inflammasome, and to provide experimental evidence for further study of the anti-tumor mechanism of TF-SB. Mielanoma models were established by inoculating B16-F1 cell line to mice, and then were randomly divided into 5 groups (n=10 in each group): model control, positive control control(Rap, 1.5 mg•kg⁻¹), and TF-SB low, middle and high groups (50, 100 and 200 mg•kg⁻¹). Meanwhile, healthy C57BL/6J mice were used as normal control group (n=10). The drugs were given once daily for 2 weeks consecutively. Thirty minutes after last treatment, the determinations at endpoint were performed; pathological changes of tumor tissue were evaluated by using HE staining; protein expressions of LC3-II/LC3-I or NLRP3inflammasome/caspase-1/IL-1β and IL-18 in tumor tissues were detected by using Western-blot; and serum levels of IL-1β and IL-18 were detected by using Elisa kit. The results showed that the tumor cells in model group showed obvious atypia and malignant proliferation; the invasion of tumor tissue was significantly reduced, the tumor necrosis area was significantly increased, and the inflammatory reaction was significantly alleviated in positive control group and various TF-SB groups. As compared with model control group, LC3-II/LC3-I was significantly increased, while NLRP3/caspase-1/IL-1βand IL-18 protein expressions were significantly decreased in positive control group and TF-SB groups. Serum IL-1β and IL-18 levels in model control group were found higher than those in control group (P<0.001), but they were significantly lowered in positive control group and TF-SB groups (P<0.05, P<0.01 or P<0.001). Taken together, total flavonoids in S. barbata could effectively alter the tumor growth micro-environment by inhibiting the expression of NLRP3 inflammasome, and its anti-tumor effect may be associated with the induction of tumor cell autophagy.