ABSTRACT
Objective:To study the effect of extract of livistona chinensis on the proliferation and apoptosis of bladder cancer cells and the related mechanism.Methods:T24 cells were cultured in medium with the final concentration of 0, 25, 50 and 100 mg/L livistona chinensis extract, respectively. And then they were divided into control group and low, medium and high dose groups. The cell survival rate was detected by cell counting kit 8 (CCK-8). Colony formation assay was used to detect the number of cell clones. Apoptosis was detected by flow cytometry. Western blot was used to detect the expression of related proteins. The Syk overexpression vector plasmid and its negative control were transfected into T24 cells. After transfection, the cells were treated with 100 mg/L livistona chinensis. The cell survival rate, colony formation number and apoptosis rate were detected by the above method. The bladder cancer model nude mice were treated with different concentrations of livistona chinensis extract. Under the microscope, the expression of protein was detected by immunohistochemical staining of bladder tissue.Results:Compared with the control group, the survival rate of T24 cells in the low, medium and high dose livistona chinensis extract groups were significantly decreased [(88.50±3.65)%, (70.58±2.47)%, (48.90±2.37)% vs. (98. 25±4.26)%], and the number of clone formation decreased significantly [(101. 33±3.40), (84.00±2.94), (60.00±2.16) vs. (121.33±4.64) ], and the apoptosis rate was significantly increased [(11.45± 0.59)%, (17.71±0.64)%, (21.33±0.83)% vs. (7. 86±0.43)%]. The expression level of Ki-67 protein was significantly decreased, while the expression levels of Caspase3 and Syk protein were significantly increased in a concentration dependent manner ( P < 0.05). The cell survival rate of pcDNA3.1-Syk group was significantly lower than that of pcDNA3.1 group [(63.87±2.53)% vs. (98. 45±3.54)%], the number of clone formation decreased significantly [(74. 33±2.87) vs. (121.33±3.68)], and the apoptosis rate was significantly increased [(18.39±0.63)% vs. (7.89± 0.45)%] (all P<0.05). The cell survival rate in the high-dose group of livistona chinensis+ pcDNA3.1-Syk was significantly lower than that in the high-dose group of livistona chinenisi+ pcDNA3.1 group [ (29.80±1.63)% vs.(49.33±2.76)% ], the number of clone formation decreased significantly [(33.00±2.94) vs. (59.67±3.30) ], and the apoptosis rate was significantly increased [(26.93±0.68)% vs. (21.25±0.78)% ]( P<0.05). The experimental results of nude mice of bladder cancer model showed that the tumor volume of transplanted bladder cancer nude mice in the control group and the low, medium, and high dose livistona chinensis extract groups were (1 209.75±64.37), (1 006.31±40.49), (530.58±42.87), (267.58±16.73)mm 3, respectively, the weight of the transplanted tumor were (0.36±0.08), (0.30±0.04), (0.26±0.03), (0.18±0.06)g, and the differences between the two groups were statistically significant ( P <0.05). Immunohistochemical staining results showed that the expression of Sky and Caspase3 was increased and the expression of Ki-67 was decreased in the middle and high dose groups compared with that in the control group. Conclusion:Extract of livistona chinensis can inhibit the proliferation and promote apoptosis of bladder cancer cells by up regulating Syk expression.
ABSTRACT
Background Coal workers' pneumoconiosis (CWP) is a serious occupational disease. Whether ferroptosis, a form of necrotic regulated cell death, is involved in coal dust induced mouse models of CWP needs further survey. Objective This experiment is designed to elucidate the role of ferroptosis in the formation of CWP induced by coal dust in mice. Methods C57BL/6J mice were randomly assigned to a saline group or a CWP group, with eight mice in each group. The mice were treated with 0.1 mL normal saline or 0.1 mL coal dust suspensions (50g·L-1) via intra-tracheal instillation. HE staining and Masson staining were used to show lung injury and lung fibrosis. Iron concentration in mouse lung tissues was measured using iron assay kit. Lipid peroxidation was estimated in lung tissues by malondialdehyde (MDA) concentration and immunofluorescence intensity, and the ratio of glutathione (GSH) to L-glutathione oxidized (GSSG). Western blotting and real-time fluorescence-based quantitative PCR were used to test protein and mRNA expression levels of glutathione peroxidase 4 (GPX4) and ferritin in mice. Results Coal dust injured pulmonary structure, thickened alveolar wall, and caused collagen deposition and infiltration of inflammatory cells in the CWP group. The iron concentration in the CWP group [(10.75 ± 5.42) mg·L−1] was higher than that in the saline group [(1.14 ± 0.37) mg·L−1] (P < 0.01). The MDA concentration in the CWP group [(37.32 ± 12.18) μmol·L−1] was higher than that in the saline group [(18.70 ± 8.22) μmol·L−1] (P <0.01). The immunofluorescence intensity of MDA in the CWP group was stronger than that in the saline group. The GSH/GSSG ratio decreased in CWP treated mice (1.50 ± 1.70) compared with the normal saline treated ones (4.95 ± 2.86) (P < 0.01). Compared with the saline group (38.84 ± 15.61 for GPX4, 225.90 ± 54.34 for ferritin), the relative expression levels of GPX4 and ferritin mRNA in the CWP group were downregulated (14.29 ± 7.21 for GPX4, 106.70 ± 36.70 for ferritin) (P < 0.01). Compared with the saline group (1.47 ± 0.54 for GPX4, 1.73 ± 0.34 for ferritin), the relative expression levels of GPX4 and ferritin protein in the CWP group were also downregulated (0.92 ± 0.22 for GPX4, 0.97 ± 0.09 for ferritin) (P < 0.05). Conclusion Ferroptosis may be involved in the formation of coal workers' pneumoconiosis induced by coal dust in mice.