RESUMEN
Objective The active protein of traditional Chinese medicine has anti-tumor effect, and salvia miltiorrhiza is an important anti-tumor traditional Chinese medicine. Here, the effect of Salvia miltiorrhiza lectin protein (SMLP) on the apoptosis of gastric cancer cells was studied. Methods SMLP expressed and purified from prokaryotic cells was used to treat the gastric cancer cells SGC-7901. The experiment was divided into the control group (untreated) and the SMLP treatment group (final concentration of 10 μmol / L of SMLP was treated for 24 h). Real-time RT-PCR and Western blot were used to detect the changes of apoptosis gene expression. Flow cytometry and Hoechst staining were applied to detect the apoptotic status. Caspase-3 and Caspase-9 activity assay kits were used to detect the apoptotic level. Results The result of RT-PCR showed that the mRNA level of Bax in the SMLP treatment group was significantly higher than in the control group (1.00±0.12 vs 0.67±0.10）(P<0.05). After treatment with SMLP to gastric cancer cells, the activity and expression level of cleaved Caspase-3 and Caspase-9 were increased significantly compared with the control group (P<0.05). The cell nucleus in the control group was bigger and rounder, with smooth surface and uniform staining, whilst in the SMLP-treated group, the cell nucleus became deeper with pyknosis, representing typical morphological characteristics of apoptosis. The early apoptosis level in the control group was 6.55%, and the SMLP treatment group reached 10.18%, showing an increase in the level of apoptosis. Conclusion SMLP expressed and purified in vitro can promote the apoptosis of gastric cancer cells, which is of great significance for further revealing the function of plant lectin and investigating the anti-tumor effect on the protein of traditional Chinese medicine.
RESUMEN
Objective Gastric cancer is the most common cancer in the world. In China, Patients with gastric cancer are mostly treated with platinum-based chemotherapy. Programmed cell death 4 (PDCD4) was found as an important proapoptosis recently, the aim of the present study was to investigate the role of PDCD4 reversed the apoptosis induced by cisplatin in gastric cancer cell. The study will provide the target marker for treatment and diagnosis of cisplatin resistance in gastric cancer.Methods Stable transfection with pCMV-PDCD4 vector into human cisplatin resistance gastric cancer cell line-SGC7901/DDP; the cells were divided into control group, over-expression group, control with cisplatin group, over-expression with cisplatin group for following experiments. Hoechst dying with immunofluorescence and flow cytometry were used to measure the cell apoptosis in vitro; Real-time PCR was used to detect the mRNA expression levels of PDCD4, and the protein levels of PDCD4, pAK, pGSK3β, BCL-2 and Bak were detected by Western blot. The cells were divided into vector group, PDCD4 group, PDCD4 with activator group for detect the level of PARP(C) by Western blot.Results Compared with control group, the Results of real-time PCR and western blot were showed the level of PDCD4 was augmented in over-expression group (also in the over-expression with cisplatin group), which was indicated stable transfection with PDCD4 was successful. Immunofluorescence (with hoechst dying) and flow cytometry demonstrated that PDCD4 facilitated cell apoptosis exposed to cisplatin. PDCD4 overexpression attenuated the protein levels of pAkt, pGSK3β and BCL-2, but increased the protein levels of BAK. Furthermore, incubation with SC-79 (the activator of Akt) reversed cell apoptosis induced by PDCD4.Conclusion Overexpression of PDCD4 promotes the apoptosis induced by cisplatin through pAKT/pGSK3β pathway, which is favorable to reverse cisplatin resistance in gastric cancer.