Effect of Arca subcrenata Lischke anticancer protein on cell proliferation and apoptosis of human myeloid leukemia K562 cells
10.3867/j.issn.1000-3002.2016.03.007
- VernacularTitle:毛蚶抗癌活性蛋白对K562细胞增殖和凋亡的影响
- Author:
Chen ZHAO
;
Ying FU
;
Huaying LIN
;
Zhenxiao SUN
- Publication Type:Journal Article
- Keywords:
Arca subcrenata anticancer protein;
K562 cells;
apoptosis;
cell cycle;
caspase 3
- From:
Chinese Journal of Pharmacology and Toxicology
2016;30(3):221-228
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE To investigate the cytotoxic activity of Arca subcrenata Lischke anticancer protein(ASAP)constituents on human myeloid leukemia K562 cells in vitro and analyze its anticancer mechanisms. METHODS ASAP was extracted by low temperature water and ammonium sulfate precipitation. Protein concentration of ASAP was detected by Bradford method. Morphological changes of cultured K562 cells treated with ASAP were observed under the inverted phase-contrast micro?scope. The cell and nucleus changes were analyzed by Giemsa staining. The cytotoxicity of ASAP on K562 cells was detected by MTT assay. Flow cytometry was used to detect apoptosis and cell cycle of K562 cells treated with ASAP. The expression of apoptosis and cell cycle related proteins procaspase 3, caspase 3,P53 and programmed cell death 4(PDCD4)were analyzed by Western blotting. RESULTS ASAP exhibited significant cytotoxic effect on K562 cells in a time- and concentration-dependent manner. The concentration-effect correlation coefficient of ASAP 50,100 and 200 mg · L-1 on K562 cells for 24, 48 and 72 h was 0.851,0.8977 and 0.8997,respectively. Under an optical microscope,K562 cells showed cytomorphosis,or nuclear fragmentation after treatment with ASAP 200 mg · L-1 for 48 h. Flow cytometry analysis and Giemsa staining assay indicated that apoptotic cells increased and G2/M phase cells accumulated significantly with the increase of ASAP concentration. After treatment with ASAP 200 mg · L-1 for 48 h,the early and late apoptosis cell rate increased to(32.8 ± 0.1)%and(31.2 ± 2.2)%vs control group(3.7 ± 1.1)% and (9.9 ± 0.8)%(P<0.01),respectively,and the G2/M phase cells increased to (55.2 ± 1.7)% vs (15.3 ± 0.8)% in control group(P<0.01). After treatment with ASAP 200 mg · L-1 for 0-40 h,the expression of apoptotic protein procaspase 3 was down-regulated and its active form caspase 3 was significantly up-regulated at 32 h,while PDCD4 and P53 protein expression was down-regulated significantly in 0-40 h. CONCLUSION Apoptosis and cell cycle arrest induced in G2/M phase may account for ASAP cytotoxic activity to K562 cells. K562 cell apoptosis induced by ASAP depends on caspase 3 signal pathway. Down-regulated expression of PDCD4 and P53 proteins may be related to K562 cell apoptosis and cell cycle arrest in G2/M phase by ASAP.
