Ultrasound Activated Hematoporphyrin Destruction Of H22 Ascites Tumor Cell Apoptosis Induced By,

Malignant tumor is one of the diseases, which directly threat to human beings, there are thousands upon thousands of people under the affliction of this diseases every year. At present, the treatment of tumor especially malignant tumor is the most important investigative task in home and abroad. There are many modalities being used for cancer therapy including radiotherapy, chemotherapy, operation and so on, but these methods exist disadvantages each other, and of these, radiotherapy and chemotherapy have significant side effects on human. In 1978, American scholar Doughtery firstly present PDT, compared with traditional cancer therapy, which is a new idea on cancer therapy without wound and side effect. But due to incapable penetrability of beam, the method is mainly used to cure surface tumor and has difficulty in curing deep and terminal tumor.Ultrasonic wave with the frequency of 20kHz upwards is mechanical wave; ultrasonic wave has quietly strong penetrability and is provided with some direction, especially focused ultrasound can be focused in a small region and can penetrate deeply in tissues, then locally activate a preloaded sonosensitizer. In 1989, Japonic scholar Umemura et al firstly demonstrated that ultrasound can activate certain hematoporphyrin and thereby induce significant anti-tumor effects. SDT is based on the local activation of a systemically administered hematoporphyrin which showed preferential and long-term retention in tumor tissues, by ultrasonic exposure. At present SDT has more potential and promising method for cancer treatment.Basing on the international development and earlier experiment results, this paper has done some work about the National Natural Science Foundation of China (Grant No.39870240 and No. 30270383). The sonodynamical effect was investigated on hepatoma 22 cells exposed to the combination of 100μg/mL hemaporphyrin (Hp) and focused ultrasound at the frequency of 1.43MHz, and the present results can be explained as follows:1. The intact fractions of H-22 cells in the presence of 100-μg/mL Hp for up to 60 seconds of exposure at an ultrasonic intensity of 2 W/cm~2 were examined. The results showed no apparent difference between the Hp group and the control group. After 60 seconds of exposure, the viability of cells was reduced to 63.7% in the presence of 100-ug/mL Hp but only 87.3% without Hp. It suggested that ultrasound combined hemaporphyrin showed synergistic killing effect on hepatoma 22.2. The changes of ultrastructure on H22 cells were observed by scanning electron microscope and transmission electron microscope after SDT treatments. Under the scanning electron microscope, we found that ultrasound alone had some effects on the surface of H22 cells, but with the same ultrasound intensity, the H22 cells showed significantly damaged in ultrasound combined Hp group, and the cell damage in ultrasound plus Hp group was more serious than ultrasound group as time passed. Under the transmission electron microscope, we observed that mitochondria and membrane systems were the sensitive site to be destroyed in ultrasound group and ultrasound plus Hp group, and the damage became more and more significant as time delayed. Moreover, the apoptosis was observed in ultrasound combined with Hp group after 3h treatment.3. Physiological and biochemical analysis: after designed treatment, we found there were significant changes of apoptosis-relative proteins, including Caspase-3, Caspase-8 and Cytochrome C, suggested that the death process might also exist in both endogenous and exogenous apoptosis pathway.4. The generation of oxygen free radicals in cell suspensions was immediately detected after treatment using a fluorescence microplate reader. Results suggested that: Hp had synergistic effects with ultrasound and significantly increased the ROS content compared with the other three groups5. Colorimetry and enzymatic chemical methods were used to measure the lipid peroxidation levels and activities of key antioxidant enzymes in H22 cell after treatment, and found that the malondialdehyde content was remarkably enhanced, and antioxidative enzyme activities were obviously decreased at different level compared with the other three groups.