ABSTRACT
ObjectiveTo a nalyze the morphologic features of SMMC-7721 cannibalistic cells that induced by triazole Schiff base derivative(LH-37) in vitro. Methods The SMMC-7721 cells (1×10~4/ml)were cultured in the medium containing of 1×10~(-5) mol/L LH-37 for 24h,48h.The character of cells was detected by Papanicolaou and Wright′s Staining. Immunohistochemical method was used to observe the cleaved Caspase-3 positive cells. The ultrastructure of cannibalism cells was observed by JEM 100CX-II transmission electronic microscope. Results Microscopic analysis demonstrated the complete internalization of one cell within another. We noted that some cannibalistic cells in small aggregates appeared to be inside of large vacuoles, suggesting that they were internalized within a neighboring cell. The proportion of cannibalistic cells were increased after SMMC-7721 cells were cultured in the presence of LH-37 for 48 hours. The proportion of the cannibalistic cells in control and LH-37 group was 0.47% and 5.23% respectively . Many internalized cells were positive for cleaved caspase-3 staining . Ultrastructural analysis of engulfed cells from 24 hours exhibited evidence of live-cell internalization consistent with cannibalism, The most common fate for internalized cells was death after treatment with LH-37 for 48 hours, as evidenced by nuclear degradation and the eventual disappearance of some cells within the enveloping cell . Conclusion The data presented indicate that LH-37 can lead to an increase of cannibalism in human hepatocarcinoma cell in vitro.
ABSTRACT
Aim To study the effect of 3-pyridin-3-yl-4-[(4-hydroxy-3-methoxy-benzylidene)amino]-5-me-thylsulfanyl-4H-[1,2,4] triazole (LH-37) on induction of differentiation and apoptosis of hepatocarcinoma BEL-7402 cells. Methods BEL-7402 cells were cultured in RPMI 1640 and treated by LH-37 at different doses. The proliferation of the cells and the inhibition effect of LH-37 on the cell proliferation were examined by MTT assay. The reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the changes of alpha fetoprotein(?-FP)mRNA and albumin(Alb). The concentration of ?-FP in the cells was detected by ELISA assay. Cell morphology was observed by fluorescence microscope techniques. The protein expressions of active caspase-3 in BEL-7402 cells were observed by immunohistochemical staining. Western blot was used to assay caspase-3 and caspase-9. Colorimetric method was used to assay activity of superoxide dismutase (SOD) and catalase (CAT). The apoptotic cells were assayed by flow cytometry (FCM) with annexin V-FITC conjugated and propidium iodide (PI) staining. Results The cell proliferation was inhibited by LH-37 at 10 ?mol?L-1~1 mmol?L-1 in dose dependent manners. After treated with 10 ?mol?L-1 or 1.0 ?mol?L-1 LH-37, the mRNA and protein expression of ?-FP were significantly reduced but mRNA expression of Alb was significantly increased. Treatment of BEL-7402 cells with different LH-37 concentrations for 48 hours increased the percentage of active caspase-3 positive cells and protein expression of caspase-3 and caspase-9. More apoptosis features of cells were observed in LH-37 ( 100 ?mol?L-1) treatment groups than in control group. LH-37 markedly promoted the viability of SOD and decreased CAT in BEL-7402 cells. Counclusion LH-37 might inhibit proliferation and induce differentiation and apoptosis of human hepatocarcinoma BEL-7402 cells, which might be related to the cytotoxicity of the intracellular hydrogen peroxide (H2O2).