The Killing Efect Of Focused Ultrasound Combined With Hematoporphyrin On S180

Malignant tumor is one of the most deadly diseases for human beings .About one of ten deaths is due to cancer, according to the numbers announced by the world heath organization.At present ,there are many modalities being used for cancer therapy including radiotherapy ,chemotherapy,surgicaltherapy and so on.While in the ultrasound antitumor fields there concerns about high intensity focused ultrasound (HIFU)surgery,ultrasound thermal treatment and sonodynamic therapy(SDT).At the end of last century,Japanese Umemura firstly put forward sonodynamic therapy ,this therapy is based on preferential and long time retention of a sonosensitizing drug in tumor tissues and subsequent activation of the drug by ultrasound .ultrasound can penetrate deeply into organic tissues without any harm and can be focused into a small region of tumor to activate a sonosensitizing drug .The ability to enhance drug cytoxicity with ultrasound can be localized on a pathological site with minimal damage to peripheral healthy tissues,which might be a valuable clinical asset.Recently ,many scholars at home and abroad have done many researches in this field and have obtained some primary results ,but we haven't got the definite mechanism about SDT.Based on earlier results,this paper has done some work about national scientific funds-the cytological molecular biological mechanism of ultrasound activating HpD.This expriment studied the killing effect of ultrasound combined with HpD on S180 ascitic tumor cells and solid tumor cells at the frequency of 2.2MHz. The results can be explained as follows:1. With the fluorescence photometer ,we detected the maximal excition spectrum(402nm) and emission spectrum(596nm) of HpD.determinded HpD concentration in S180 tumor cells to choose the hightest concentration time (45min),and established the distribution of HpD in tumor bearing mouse to get the best time for exposure to ultrasound that probably be between 24-72h.2. At different ultrasound intensity and different exposure time ,cell livability decreased as the intensity and time increased.3. After establishing the parameters of ultrasound, we chose the fixed intensity(4W/cm2) to evaluate morphologic changes after different treatments using alight microscope(LM),a scanning electron microscope(SEM),and a transmission electron microscope(TEM).The experiment results showed: ultrasound alone showed a slight antitumor effect,while ultrasound plus HpD showed a significant synthetic antitumor effect which can destroyed the microstrure and ultrastrure of tumor cells,especially in the cell membrane and the organelle membrane.we also studied the cell apoptosis phenomena induced by ultrasound activating HpD with AV-PI staining method.These results suggested that there might be two kinds of cell killing methods-inducing cell apoptosis or maybe sdirectly destroying cell ultrastrure.4. Using immunocytochemistry, we detected the protein expression changes about apoptosis associated genes bcl-2 and caspase-3 after treatment in vitro.Bcl-2 protein expression degraded in ultrasound plus HpD group compared with other three groups in which the expression was changed a little. We also analyzed the significant differences(p<0.05 ).Caspase-3 expression was similar in control and HpD groups,While the expression increased in ultrasound alone and ultrasound plus Hpd groups.5. With the method of TBA chromotest, we detected the influence of ultrasound combined with HpD on the degree of lipid overoxidation of tumor cells,and found the quantity of lipid overoxidation in ultrasound combined with HpD higher than ultrasound and HpD alone.6. In this study, ultrasonically induced cell damage enhanced by HpD was significantly reduced by histidine and mannital, but not by superoxide dismutase (SOD). Possible mechanism of ultrasound activating HpD derived hydroxyl radicals as a mediator for sonodynamically induced cell damage.7. The killing effect of different ultrasound intensity on S180 solid tumors indicated 5w/cm2 maybe the threshold intensity to produce antitumor effect.8. Antitumor effects were estimated by measuring tumor size and tumor weight. The tumor size inhibition of ultrasound combined with HpD was almost twice than ultrasound alone, and the tumor weight inhibition of them is respectively 33.09% and 20.67%.9. After exposure to ultrasound at the frequency of 2.2MHz and the intensity of 5W/cm2 in vivo,the tumor cell structure was observed by LM and TEM.The results showed: ultrasound alone had some antitumor effect in vivo which would become more serious as the time delayed, and the synthetic antitumor effect of ultrasound in the presence of HpD was more significant at the same experiment.