Damaging the membrane of Ehrlich ascitic tumor cells with focused ultrasound / 中华物理医学与康复杂志

Objective To study the damage focused ultrasound inflicts on the membrane permeability of Ehrlich ascitic tumor (EAC) cells and the relationship between changes in membrane permeability and focused ultra-sound exposure time. Methods The relative survival rate of tumor cells was examined at various intensities and dif-ferent exposure times using focused 2.2 MHz ultrasound. The uhrastructure changes were evaluated with a scanning electron microscope after different exposures. Membrane permeability was investigated by incorporating fluorescein isothiocyanate dextran (FD5OO) , and membrane damage was evaluated by measuring lactate dehydrogenase (LDH) release. Results Morphological observation showed there were numerous microvilli on the surface of un-exposed cells. When the cells had been irradiated with focused ultrasound for 30 s there was only a slight effect on the shape of the cells and the number of microvilli was slightly reduced. When the cells were exposed to ultrasound for 60 s, the surface of many cells became relatively smooth with no obvious microvilli, and several small craters were seen on the surfaces of cells where the cytoplasm seemed to have extruded through the membrane. The cell membrane was seri-ously damaged by sonoporation. The loading of FD500 in the unexposed cells was only 0.21%. When the cells had been sonicated with focused ultrasound for 30 s or 60 s, the loading of FD500 increased to 11.46% and 18.50% re-spectively. The released LDH activities in the 30 s group and 60 s group were 2.94±0.02 and 3.28±0.04 U/L, respectively. The activities of LDH increaased as the focused ultrasound exposure time was prolonged. Conclusion Focused ultrasound may damage the cell membrane permeability of EAC cells, and the damage increases as the expo-sure time is prolonged from 30 s to 60 s.

Effect of ultrasound activating hematoporphyrin on the activities of antioxidative enzymes in mouse hepatoma 22 / 生物医学工程学杂志

This investigation was made with regard to the influences of ultrasound combined with hematoporphyrin on the activities of antioxidative enzyme in ascites hepatoma 22 (H-22) tumor cells, and to a better understanding of the potential biological mechanism of sonodynamic therapy which involved the damage to cells. Combined with 100 microg/ml hematoporphyrin, high intensity focused ultrasound sonication at a frequency of 1.43 MHz and an intensity level of 2.0 W/cm2 was delivered to H-22 tumor cells for 1 min. The viability of cells was evaluated by typan-blue blue exclusion test. The intracellular reactive oxygen species (ROS) levels were determined by 2',7'-dichlorofluorescein diacetata (DCFH-DA). Enzymatic chemical methods were used to measure the activities of key antioxidative enzymes. The results indicated that the cell damage rate of ultrasound combined with hematoporphyrin was significantly higher than that of the treatment with ultrasound alone, and hematoporphyrin alone had no killing effect on H-22 cells. The level of ROS in cell suspension was significantly increased, and the key antioxidative enzyme activities were obviously decreased after treatment with the combined use of ultrasound and hematoporphyrin. We speculated that the decreased activities of key antioxidative enzymes in cells might be involved in mediating the killing effect on H22 cells in sonodynamic therapy.

The effect of focused ultrasound on the physicochemical properties of Sarcoma 180 cell membrane / 生物医学工程学杂志

This study was amied to detect the changes in the cell membrane of Sarcoma 180 (S180) cells induced by focused ultrasound and to probe the underlying mechanism. The viability of tumor cells was examined at various intensities and different treatment times by ultrasound at the frequency of 2.2MHz. Flow cytometry and fluorescence microscopy were used to detect the loading of fluorescein isothiocyanate dextran (FD500) which signifies the change of membrane permeability. The results showed that after the cells were treated by ultrasound, especially when irradiated for 60s, the number of fluorescent cell, which represented the transient change of membrane permeabilization with cell survival, increased significantly. Then the damage of cell membrane was evaluated by the measurement of lactate dehydrogenase (LDH) release which became more severe as the radiation time was increasing. The generation of lipid peroxidation was estimated using the Thibabituric Acid (TBA) method after irradiation. The results reveal that the instant cell damage effects induced by ultrasound may be related to the improved membrane lipid peroxidation levels post-treatment. The physicochemical properties of S180 cell membrane were changed by focused ultrasound. The findings also imply an exposure time-dependent pattern and suggest that the lipid peroxidation produced by acoustic cavitation may play important roles in these actions.

The killing effect of focused ultrsound activating protoporphyrin IX on S180 cells / 生物医学工程学杂志

The killing effect on S180 cells was studied using the combination of protoporphyrin IX and focused ultrasound at the frequency of 2.2 MHz and different intensities. Cell viability was determined by trypan blue exclusion test, morphology changes were evaluated by means of scanning electron microscopy and transmission electron microscopy after ultrasonic exposure. The results indicated that protoporphyrin IX(PPIX) alone showed no significant effect on S180 cells when compared with that of control group. Ultrasound alone and ultrasound combined with PPIX groups showed some anti-tumor effect, which became more noticeable as the ultrasound intensity and PPIX concentration increased, and when the concentration of PPIX increased to 120 microM, the ultrasound combined with PPIX exerted a more significant anti-tumor effect than did the ultrasound alone in the same experiment.

Killing effect on S180 by focused ultrasound activating Protoporphyrin Ⅸ / 基础医学与临床

Objective To study the cell killing effect on isolated sarcoma 180 cells by ultrasound activating Protoporphyrin IX and to explore its biological mechanism.Methods The sonodynamical effect was investigated on S180 tumor cells exposed to the combination of 120 mol/L protoporphyrin Ⅸ (PPⅨ) and focused ultrasound at the frequency of 2.2 MHz and an intensity of 3W/cm2. The livability of cells was evaluated by trypan blue staining. Scanning electron microscope (SEM) observation of the surface of cells was performed to evaluate the morphological changes induced by ultrasonic irradiation. The generation of oxygen free radicals in cell suspensions was immediately detected after treatment by the active oxygen detection kit. Oxidative stress was assessed by measuring the activities of key antioxidant enzymes (ie, Superoxide dismutase[SOD], Glutathione peroxidase [GSH-PX], Catalase [CAT]) in S180 cells after SDT.Results The cell damage rate of ultrasound combined with PPIX was significantly higher than that treated with ultrasound alone only, and PPIX alone had no killing effect on S180 cells. Enzymatic chemical methods showed the content of MDA significantly increased after treatment, while the activities of key antioxidant enzymes in tumor cells all decreased at different levels, and was associated to the generation of oxygen free radicals in cell suspension after treatment. Conclusion Oxygen free radical may play an important role inimproving the membrane lipid peroxidation, decreasing the activities of key antioxidant enzymes in cells, and the biological mechanism might be involved in mediating the killing effect of S180 cells in SDT.

Establishment of plantlet regeneration system of Rubus idaeus / 中草药

Object\ To establish a plantlet regeneration system of Rubus idaeus L for the purpose to obtain a large number of high quality seedling in a short time Methods\ Stem apex and part of the stem were used as the explant and the optimal culture media and conditions were selected by orthogonal design Results\ An optimum culture medium for the induction of callus, adventitious bud and root was obtained which can be carried out in the laboratory with comparative ease and good repeatability Conclusion\ A basic medium + BA 0 2 mg/L+NAA 1 0～1 5 mg/L was most suitable for the induction of callus; a medium + BA 1 mg/L+NAA 0 1～0 2 mg/L+GA 3 6～8 mg/L+CH 300 mg/L was good for the induction of bud; a medium +BA 1 mg/L+NAA 0 1 mg/L+GA 3 2 mg/L was suitable for propagation of the bud; and the basic medium+IBA 0 1 mg/L+NAA 0 5 mg/L was good for the induction of root