Chenfeng Ji

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.

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.

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

Analysis of apoptosis mechanism in SGC-7901 induced by CSBE with flow cytometry and confocal laser scanning techenology

Flow cytometry is a laser based, biophysical technology employed in cell counting, sorting, biomarker detection and protein engineering, by suspending cells in a stream of fluid and passing them by an electronic detection apparatus. Confocal laser scanning microscopy is a technique for obtaining high-resolution optical images with depth selectivity. The flow cytometry and confocal laser scanning microscopy were used to study the changes of initiator, effector and marker in process of mitochondrial apoptosis pathway induced by CSBE. In addition, the regulation genes of apoptosis were surveyed with western blotting and flow cytometry. As a result, the mechanism of apoptosis induced by CSBE was to increase the expression of pro-apoptotic protein Bax, decrease the expression of anti-apoptotic protein Bcl-2, which induced mitochondrial PTP channel open, decreased mitochondrial membrane potential, led to release of mitochondrial Cyt-c and caused activation of caspase-3. To sum up, the mitochondrial apoptosis pathway of SGC-7901 induced by CSBE was revealed by the flow cytometry and confocal laser scanning microscopy.

Effects of Juglone on ROS Production and Mitochondrial Transmembrane Potential in SGC-7901 Cells

To investigate the effects of juglone on ROS production and mitochondrial transmembrane potential (ΔΨm) in SGC-7901 cells in vitro, further to elucidate the apoptosis mechanism. The apoptosis morphological changes of SGC-7901 cells were observed using fluorescence microscope. The level of ROS and ΔΨ<inf>m</inf> of cells were detected by FCM. The change of [Ca²⁺]<inf>i</inf> in cells stained with Fluo-3/AM was observed by LCSM. Hoechst 33258 staining assay showed thick granulated fluorescence in condensed nuclei and cytoplasm, and the number of apoptotic bodies was increased with increasing of juglone concentration. The level of ROS at the dose of 5, 10, 15 and 20μM of juglone was 34.83±1.45%, 42.43±1.36%, 53.73±1.38% and 68.67±1.33%, respectively, which were significantly higher than that in control group (P≪0.01). The concentration of Ca²⁺ in cells exposed to juglone for 24 h was increased significantly in a dose dependent manner compared with control group (P≪0.01). ΔΨm of treatment groups was 85.53±1.82%, 53.57±2.48%, 46.33±1.46%, 36.43±2.64%, respectively, which were significantly lower than that in control group (P≪0.01).Therefore, juglone can increase intracellular level of ROS and concentration of Ca²⁺, decrease ΔΨm and induce the cells apoptosis.

The Effect of Sargassum fusiforme Polysaccharides on the Functioning of Erythrocyte Membrane of S180 Mice by Affecting Ca²+ Channels

To analyze the effect of Sargassum fusiforme polysaccharides (SFPS) on the functioning of erythrocyte membrane of S180 mice by affecting Ca2+ channels. A tumor mouse model was developed, and the mice were treated with high, medium, and low doses of SFPS. Changes in [Ca2+] in the erythrocytes in the tumor-bearing mice were measured using laser confocal scanning; changes in the content of sialic acid and the acitivities of ATPase were meausred using relevant test kits; changes in the level of erythrocyte membrane potential were analyzed using FCM; and changes in the complex mobility of erythrocytes were measured using HPCE. SFPS lowered the intracelluar [Ca2+] of erythrocytes in tumor-bearing mice, increased the content of sialic acid on surface of erythrocyte membrane, increased the acitivities of Na+,K+-ATPase and Ca2+, Mg2+-ATPase, and raised the level of erythrocyte membrane potential and the complex mobility of erythrocytes in elelctrophoresis. By affecting the Ca2+ channels, SFPS can increase the content of sialic acid on the surface of erythrocyte membrane. The sialic acid combines with Ca2+ to increase the activities of Na+,K+-ATPase and Ca2+, Mg2+-ATPase, resulting in a rise in the membrane potential of the erythrocytes and an increase in the complex mobility of the erythrocytes in electrophoresis, thus restoring many physiological and biochemical functions of the erythrocyte membrane. This may be one of the anti-tumor mechanisms of SFPS.

Analysis of SGC-7901 Apoptosis Induced by CSBE and Regulating Gene Expression with Laser Scanning Microscopy and Immunofluorescence Flow Cytometry Technique

The purpose of this article was to observe interaction between drug and cancer cell with fluorescence probes and molecular imaging technology. In order to study the effect of cell growth and apoptosis morphology of Human gastric adenocarcinoma SGC-7901 induced by n-butanol extract from Capparis spionosa L. (CSBE), laser scanning microscopy was employed to observe apoptosis imaging. In addition, the protein expression and mRNA expression of its regulating gene Bcl-2 and Bax were detected. Laser scanning microscope imaging showed cells that have bound Annexin V-FITC were in early period of apoptosis, which will show green staining in the plasma membrane. The cells that have lost membrane integrity will show red staining (PI) throughout the nucleus and a halo of green staining (FITC) on the cell surface (plasma membrane). The necrosis cells were stained by PI only. In the western blot test, CSBE could decrease the expression of Bcl-2 protein and the rate of inhibition increased as the concentration of drug rised. The immunofluorescence flow cytometry technique found that CSBE could increase the expression of Bax protein in a dose-dependent manner. Thus the rate of Bcl-2/Bax was reduced. The result of RT-PCR showed that CSBE could down-regulate the mRNA expression of Bcl-2 gene, and up-regulate the mRNA expression of Bax gene, which meant that it was by regulating gene transcription level. Eventually the apoptosis of human gastric cancer SGC-7901 cell is induced through mitochondrial signal transduction pathway.

Analysis of changes of Ca 2+ concentration and mitochondrial membrane potential in HepG-2 induced by CSBE by Laser Scanning Confocal Technology and Flow Cytometry

To discuss the mechanism of the apoptosis induced by n-butanol extract from Capparis spionosa L. (CSBE) in human heptocarcinoma cell Line HepG-2, Laser Scanning Confocal Technology and Flow Cytometry were used to investigat the changes of mitochondrial membrane potential and Ca2+ levels in HepG-2 cells. The changes of mitochondria membrane potential induced by CSBE on the HepG-2 cells were studied by flowcytometry. Intracellular Ca2+ levels of both the treated and untreated HepG-2 cells with CSBE were measured by laser confocal microscope. The mitochondria membrane potential of HepG-2 cells decreased in various degrees under CSBE. In addition, the intracellular Ca2+ level increased in the middle and high dose groups, resulting in an imbalance. CSBE can decrease the mitochondria membrane potential, open MPTP channel and cause the overload of intracellular Ca2+ level, which may induce the apoptosis of HepG-2 cell. These instructions give you basic guidelines for preparing camera-ready papers for conference proceedings.

Analysis of SGC-7901 apoptosis induced by CSBE and regulating gene expression with laser scanning microscopy and immunofluorescence flow cytometry technique

The purpose of this article was to observe interaction between drug and cancer cell with fluorescence probes and molecular imaging technology. In order to study the effect of cell growth and apoptosis morphology of Human gastric adenocarcinoma SGC-7901 induced by n-butanol extract from Capparis spionosa L. (CSBE), laser scanning microscopy was employed to observe apoptosis imaging. In addition, the protein expression and mRNA expression of its regulating gene Bcl-2 and Bax were detected. Laser scanning microscope imaging showed cells that have bound Annexin V-FITC were in early period of apoptosis, which will show green staining in the plasma membrane. The cells that have lost membrane integrity will show red staining (PI) throughout the nucleus and a halo of green staining (FITC) on the cell surface (plasma membrane). The necrosis cells were stained by PI only. In the western blot test, CSBE could decrease the expression of Bcl-2 protein and the rate of inhibition increased as the concentration of drug rised. The immunofluorescence flow cytometry technique found that CSBE could increase the expression of Bax protein in a dose-dependent manner. Thus the rate of Bcl-2/Bax was reduced. The result of RT-PCR showed that CSBE could down-regulate the mRNA expression of Bcl-2 gene, and up-regulate the mRNA expression of Bax gene, which meant that it was by regulating gene transcription level. Eventually the apoptosis of human gastric cancer SGC-7901 cell is induced through mitochondrial signal transduction pathway.