Yubin Ji

Juglone-induced apoptosis in human gastric cancer SGC-7901 cells via the mitochondrial pathway.

This study was designed to investigate the effect of juglone on the apoptosis of human gastric cancer SGC-7901 cells. The cytotoxic activity of juglone on SGC-7901 cells was tested by the sulforhodamine B (SRB) assay. The morphological changes in the cells were observed by transmission electron microscopy (TEM). The apoptotic rate, the level of reactive oxygen species (ROS), mitochondrial transmembrane potential and the expression of cytochrome c protein were detected by flow cytometry (FCM). The expression of Bcl-2 and Bax proteins were examined by Western blot. Caspase 3 activity was determined with a microplate reader. Our results were as follows: the GI(50) values for SGC-7901 cells were 36.51 ± 1.05 μmol/L (24h) and 25.37 ± 1.19 μmol/L (48 h). After 24h of exposure to juglone (5, 10, 15 and 20 μmol/L), the cells presented the typical morphological changes of apoptosis, and the rate of apoptosis was found to increase in a dose-dependent manner. After cells were treated with juglone at the same dose for 24h, the level of ROS was significantly higher, the expression of Bcl-2 was significantly down-regulated and the expression of Bax was significantly up-regulated compared to the control. The mitochondrial transmembrane potential was significantly lower, and the expression of the cytochrome c protein was significantly higher relative to the control. Caspase 3 was activated in a concentration-dependent manner. In conclusion, juglone can induce apoptosis in SGC-7901 cells through a mitochondrial pathway that seems to be mediated by the generation of ROS and a reduction in the Bcl-2/Bax ratio.

[Effect of Sargassum fusiforme polysaccharides on the complex mobility of erythrocytes in tumor-bearing organisms using high performance capillary electrophoresis].

This study analyzed the effect of Sargassum fusiforme polysaccharides (SFPS) on the complex mobility of erythrocytes in tumor-bearing organisms. A mouse tumor model was developed in which mice were intraperitoneally injected with high, medium, and low dosages of SFPS for 7 d. The erythrocytes were collected and prepared into suspensions, and the complex mobilities of cells were measured using high performance capillary electrophoresis (HPCE). Experimental conditions included the following: capillaries, 75 microm x 50 cm; buffer for electrophoresis, phosphate solution containing hydroxypropylmethyl cellulose (0.1 mol/L, pH 7.4); injection pressure, 3.448 kPa; injection time, 10 s; separation voltage, 20 kV; column temperature, 25 degrees C. The electroosmotic mobility was calculated to be 2.16 x 10(-4) cm2 x V(-1) s(-1). It was found that SFPS can decrease the migration time of erythrocytes in tumor-bearing organisms, which was 17.56 min for the control group, 13.96 min for the low dosage group, 12.90 min for the medium dosage group, 13.51 min for the high dosage group, and 11.51 min for the normal group, and increase its complex mobility, which was 1.06 x 10(-4) cm2 x V(-1) x s(-1) for the control group, 1.19 x 10(-4) cm2 x V(-1) x s(-1) for the low dosage group, 1.29 x 10(-4) cm2 x V(-1) x s(-1) for the medium dosage group, 1.23 x 10(-4) cm2 x V(-1) x s(-1) for the high dosage group, and 1.45 x 10(-4) cm2 x V(-1) x s(-1) for the normal group. Compared with the negative control group, all three dosage groups had significant differences (P < 0. 01) in the complex mobility of erythrocytes. It was found that SFPS can change the complex mobility of erythrocytes in tumor-bearing organisms with the tendency to the condition in normal organisms, which is possibly because SFPS can change the charges density on erythrocytes surface. It is believed that HPCE can be used as an auxiliary tool for determining the physiological state and functions of erythrocytes.

Alkaloids from beach spider lily (Hymenocallis littoralis) induce apoptosis of HepG-2 cells by the fas-signaling pathway.

Alkaloids are the most extensively featured compounds of natural anti-tumor herbs, which have attracted much attention in pharmaceutical research. In our previous studies, a mixture of major three alkaloid components (5, 6-dihydrobicolorine, 7-deoxy-trans-dihydronarciclasine, littoraline) from Hymenocallis littoralis were extracted, analyzed and designated as AHL. In this paper, AHL extracts were added to human liver hepatocellular cells HepG-2, human gastric cancer cell SGC-7901, human breast adenocarcinoma cell MCF-7 and human umbilical vein endothelial cell EVC-304, to screen one or more AHL-sensitive tumor cell. Among these cells, HepG-2 was the most sensitive to AHL treatment, a very low dose (0.8μg/ml) significantly inhibiting proliferation . The non- tumor cell EVC-304, however, was not apparently affected. Effect of AHL on HepG-2 cells was then explored. We found that the AHL could cause HepG-2 cycle arrest at G2/M checkpoint, induce apoptosis, and interrupt polymerization of microtubules. In addition, expression of two cell cycle-regulated proteins, CyclinB1 and CDK1, was up-regulated upon AHL treatment. Up-regulation of the Fas, Fas ligand, Caspase-8 and Caspase-3 was observed as well, which might imply roles for the Fas/FsaL signaling pathway in the AHL-induced apoptosis of HepG-2 cells.

Human gastric cancer cell line SGC-7901 apoptosis induced by SFPS-B2 via a mitochondrial-mediated pathway

This study was to investigate the effect of Sargassum fusiforme polysaccharides (SFPS-B2) on the proliferation and apoptosis of human gastric cancer cell line SGC-7901. Cells were treated with different concentrations of SFPS-B2. MTT and flow cytometry (FCM) assays were performed to evaluate the effect of SFPS-B2 on the cell growth and apoptosis. Inverted fluorescent microscope was used to observe cell morphology. Laser scanning confocal microscope (LSCM) was used to analyze intracellular calcium ion concentration, mitochondrion permeability transition pore (MPTP) and mitochondrial membrane potential (MMP). Spectrophotometer was applied to quantify the activity of Caspase-9 and Caspase-3. FCM was used to determine the expressions of Bcl-2, Bax and cytochrome C. It was shown that SFPS-B2 inhibited the growth of SGC-7901. After the treatment for 72 h, the cell apoptosis morphology was obvious, which showed that cell protuberance and apoptotic body appeared, and the cytoplasm was concentrated; the apoptotic peak appeared and the apoptotic rate increased in a dose-dependent manner. After the treatment for 24 h, SFPS-B2 activated intracellular MPTP and decreased MMP. It also increased the activity of Caspase-9 and Caspase-3, down-regulated the expression of Bcl-2 and up-regulated the expression of Bax, induced the release of Cyt-C. SFPS-B2 induced SGC-7901 apoptosis through a mitochondrial-mediated pathway, suggesting it may be an agent for cancer treatment.

Schedule-Dependent Effects of Kappa-Selenocarrageenan in Combination with Epirubicin on Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) has a relatively higher incidence in many countries of Asia. Globally, HCC has a high fatality rate and short survival. Epirubicin, a doxorubicin analogue, may be administered alone or in combination with other agents to treat primary liver cancer and metastatic diseases. However, the toxic effects of epirubicin to normal tissues and cells have been one of the major obstacles to successful cancer chemotherapy. Here, we investigated the effects of epirubicin in combination with kappa-selenocarrageenan on mice with H22 implanted tumors and HepG-2 cell proliferation, immune organ index, morphology, cell cycle and related protein expressions in vivo and in vitro with sequential drug exposure. The inhibitory rate of tumor growth in vivo was calculated. Drug sensitivity was measured by MTT assay, and the Kings principle was used to evaluate the interaction of drug combination. Morphological changes were observed by fluorescent microscopy. Cell cycle changes were analyzed by flow cytometry. Expression of cyclin A, Cdc25A and Cdk2 were detected by Western blotting. In vivo results demonstrated that the inhibitory rate of EPI combined with KSC was higher than that of KSC or EPI alone, and the Q value indicated an additive effect. In addition, KSC could significantly raise the thymus and spleen indices of mice with H22 implanted tumors. In the drug sensitivity assay in vitro, exposure to KSC and EPI simultaneously was more effective than exposure sequentially in HepG-2 cells, while exposure to KSC prior to EPI was more effective than exposure to EPI prior to KSC. Q values showed an additive effect in the simultaneous group and antagonistic effects in the sequential groups. Morphological analysis showed similar results to the drug sensitivity assay. Cell cycle analysis revealed that exposure to KSC or EPI alone arrested the cells in S phase in HepG-2 cells, exposure to KSC and EPI simultaneously caused accumulation in the S phase, an effect caused by either KSC or EPI. Expression of cyclin A, Cdc25A and Cdk2 protein was down-regulated following exposure to KSC and EPI alone or in combination, exposure to KSC and EPI simultaneously resulting in the lowest values. Taken together, our findings suggest that KSC in combination with EPI might have potential as a new therapeutic regimen against HCC.

Effect of Asparagus Polysaccharides on Migration Time of Erythrocytes in Tumor-Bearing Organisms Using High Performance Capillary Electrophoresis

This study analyzed the effect of Asparagus polysaccharides on the migration time of erythrocytes in tumor- bearing organisms. A mouse tumor model was developed in which mice were intraperitoneally injected with high, medium, and low dosages of Asparagus polysaccharides for 7 d. The erythrocytes were collected and prepared into suspensions, and the migration time of cells were measured using high performance capillary electrophoresis (HPCE). Experimental conditions included the following: capillaries, 75 mum x 50 cm; buffer for electrophoresis, phosphate solution containing hydroxypropylmethyl cellulose (0.1 mol/L, pH 7.4); injection pressure, 3.448 kPa; injection time, 10 s; separation voltage, 20 kV; column temperature, 25degC. It was found that Asparagus polysaccharides can decrease the migration time of erythrocytes in tumor-bearing organisms, which was 17.20 min for the control group, 14.33 min for the low-dosage group, 11.95 min for the medium-dosage group, 13.03 min for the high-dosage group, and 11.42 min for the normal group. Compared with the negative control group, all three dosage groups were significantly different (P < 0.01). It was also found that Asparagus polysaccharides can change the migration time of erythrocytes in tumor-bearing organisms with the tendency to the condition in normal organisms, which is possibly because Asparagus polysaccharides can change the charges density on erythrocytes surface.

Study on human promyelocytic leukemia HL-60 cells apoptosis induced by fucosterol

In this study, we investigated the effect of fucosterol on HL-60 and the molecular mechanism. HL-60 Cells were treated with fucosterol, and 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) method was used to study fucosterol anti-tumor activity. Morphology of HL-60 cells was observed. Flow cytometry (FCM) was employed to detect the cell cycle. Laser scanning confocal microscope (LSCM) was used to analyze mitochondrial membrane potential (MMP) and the expressions of Fas, FasL, Fadd and Caspase-8. Western blot was performed to analyze the expressions of Cyt-C, Pro-Caspase-9 and Pro-Caspase-3. Caspase activity kits were used to determine the activity of Caspase-9, Caspase-8 and Caspase-3. The results showed fucosterol could inhibit the growth of HL-60 cells, and the cell cycle was arrested at G2/M phase. HL-60 cells showed obvious apoptosis morphology. After being treated with fucosterol for 24 h, HL-60 cells decreased MMP, induced Cyt-C release and Caspase-9, Caspase-3 activation. Fucosterol also increased the protein expression of Fas, FasL, Fadd and Caspase-8. Moreover, the activity of Caspase-9, Caspase-8 and Caspase-3 was increased significantly. In conclusion, Fucosterol can induce HL-60 cells apoptosis, suggesting that it may be a potent agent for cancer prevention and treatment.

Chelidonine induces mitotic slippage and apoptotic-like death in SGC-7901 human gastric carcinoma cells

The aim of the present study was to investigate the effect of chelidonine on mitotic slippage and apoptotic-like death in SGC-7901 human gastric cancer cells. The MTT assay was performed to detect the antiproliferative effect of chelidonine. Following treatment with chelidonine (10 µmol/l), the ultrastructure changes in SGC-7901, MCF-7 and HepG2 cells were observed by transmission electron microscopy. The effects of chelidonine on G2/M phase arrest and apoptosis of SGC-7901 cells were determined by flow cytometry. Indirect immunofluorescence assay and laser scanning confocal microscopy (LSCM) were used to detect the phosphorylation level of histone H3 (Ser10) and microtubule formation was detected using LSCM following immunofluorescent labeling. Subsequent to treatment with chelidonine (10 µmol/l), expression levels of mitotic slippage-associated proteins, including BUB1 mitotic checkpoint serine/threonine kinase B (BubR1), cyclin-dependent kinase 1 (Cdk1) and cyclin B1, and apoptosis-associated protein, caspase-3 were examined by western blotting at 24, 48 and 72 h. The half maximal inhibitory concentration of chelidonine was 23.13 µmol/l over 48 h and chelidonine induced G2/M phase arrest of cells. The phosphorylation of histone H3 at Ser10 was significantly increased following treatment with chelidonine for 24 h, indicating that chelidonine arrested the SGC-7901 cells in the M phase. Chelidonine inhibited microtubule polymerization, destroyed microtubule structures and induced cell cycle arrest in the M phase. Giant cells were observed with multiple micronuclei of varying sizes, which indicated that following a prolonged arrest in the M phase, the cells underwent mitotic catastrophe. Western blotting demonstrated that the protein expression levels of BubR1, cyclin B1 and Cdk1 decreased significantly between 48 and 72 h. Low expression levels of BubR1 and inactivation of the cyclin B1-Cdk1 complex results in the cells being arrested at mitosis and leads to mitotic slippage. In addition, apoptotic morphological changes in multinucleated cells were observed, the apoptosis rates increased gradually with administration of chelidonine in a time-dependent manner and the protein levels of caspase-3 increased significantly between 24 and 72 h. Thus, chelidonine induces mitotic slippage, and apoptotic-like death occurs in SGC-7901 cells undergoing mitotic catastrophe. Gastric cancer is a common malignancy, and ranks second in overall cancer-associated mortalities worldwide. The present study demonstrated that chelidonine induces M phase arrest and mitotic slippage of SGC-7901 human gastric carcinoma cells via downregulating the expression of BubR1, Cdk1 and cyclin B1 proteins. With the prolongation of chelidonine treatment, the giant cells with multiple micronuclei underwent mitotic slippage and were maintained in the G1 phase and did not survive. A number of multinucleated cells underwent apoptosis via a caspase-dependent signaling pathway. The current study proposes that chelidonine induces mitotic slippage and apoptotic-like death of SGC-7901 cells.

Study on the Screening for Chloronitrobenzene: Degrading Bacteria and Degradation of Chloronitrobenzene

With sewage plant aeration tank activated sludge as mixed bacteria source, Nitrobenzene concentration gradient of the chlorine used to increase the aerobic bacterial degradation of oscillating bottle method of domestication and enrichment culture. Separation and purification out of one can right-chloronitrobenzene as the sole carbon, nitrogen and energy of bacteria. Through physiological and biochemical reactions characteristic morphology and 16S rDNA sequence analysis, to determine the strain is Bacillus subtilis.The degradation kinetics analysis showed that the degradation of single strain 4CNB kinetic characteristics consistent with features.

Effect of Asparagus saponins on HepG2 Apoptosis and Mitochondrial Membrane Potential and ROS Level

To study the effect of Saponins of asparagus on HepG2 apoptosis, reactive oxygen species (ROS) and mitochondrial membrane potential (Δψm) levels. Saponins of asparagus with different concentration was treated with HepG2 at different time, MTT assay was used to detect inhibitory rate, fluorescence staining was used to observe apoptosis morphology, flow cytometry was used to detect apoptosis rate and cell cycle, also ROS and Δψm were measured by flow cytometry. Results showed Saponins of asparagus inhibited cell proliferation; the IC50 on HepG2 was 172.3 μg/mL. Apoptosis morphology was observed by fluorescence microscope, some of the cells had changed from irregular shapes into round cells, the nuclei had decreased in size, the number of nucleoli had decreased; the chromatin had condensed, with the cytoplasm becoming more concentrated. The cell cycle of HepG2 was arrested at S phase, G2/M phase percent decreased. After 72h the treated group appeared apoptosis peak, and apoptosis rate with high dose group 30.94±1.74%;After 48h the ROS in treated group increased with high dose group 77.7±4.5%; and Δψm decreased with high dose group 77.8±1.9% From the above it can be seen that Asparagus saponins have a relatively strong inhibitory effect on HepG2 cells, and the mechanism involved has to do with their role in increasing the production of intracellular ROS, which leads to the lowering of mitochondrial membrane potential, thus initiating the apoptosis mechanism that induces apoptosis in HepG2 cells. I. INTRODUCTION Asparagus officinalis Linn is an herbaceous perennial plant of the Liliaceae family. Modern scientific research has shown that Asparagus officinalis can act to prevent the proliferation of cancer cells, and is effective for treating many cancers, including lymphoma, carcinoma of the bladder, pulmonary carcinoma, hepatocarcinoma, gastric carcinoma, and skin carcinoma(1,2), with saponin as one of its major anti-tumor agents. In the present experiment, modern biotechnology was used to explore the effect of the