| Background and ObjectivesHigh Intensity Focused Ultrasound(HIFU) as a new micro-invasive and non-invasive technology of treatment of tumors, the past decade treatment has been in clinical use for liver cancer, breast cancer, pancreatic cancer, prostate cancer and other malignant tumors, for benign tumor such as uterine fibroids, as well as non-neoplastic disease such as adenomyosis, etc. The technology has been more mature, how improve their therapeutic efficacy and safety has become a research hotspot. The clinical application of ablation of tumors at once,use more continuous wave of high intensity focused ultrasound(CHIFU), mainly to thermal effects, but complications of HIFU to clinical treatment of the most common thermal damage. Pulse wave high intensity focused ultrasound(PHIFU) output power intermittently, primarily destroy tissue by cavitation, mechanical effects and other non-thermal effects, but to the larger tumor, the treatment time is too long. Effective synergistic manner to reduce treatment time, which the ultrasound contrast agent in the majority. Nanoscale ultrasound contrast agents compared with the conventional micron-sized ultrasound contrast agents, better stability, more penetrating, can cross the blood pool to reach the organization, is one of the hot research.In this experiment, normal rabbit liver tissue for the study, changes in liver tissue acoustic environment by Nanobubbles(NBs). Explore the feasibility of synergistic of NBs joint with PHIFU, and the impact on BRF of continuous wave and pulsed wave of HIFU. Then set up rabbit liver VX2 tumor model, further separately treat liver cancer use continuous wave HIFU joint with Nanobubbles and pulsed wave HIFU joint with Nanobubbles,then compare the safety and efficacy of these two therapy methods.Materials and Methods1. Preparation of Nanobubbles0.25 mg polylactic acid / glycolic acid dissolved in dichloromethane, was added 100 ul PFH under ice, sound array instruments sound array 2min, join in 5% polyvinyl alcohol, high-speed disperser homogenizer homogeneous 4min, get double emulsion. Stir to volatile the dichloromethane, washed with distilled water, after centrifugation,then get PFH / PLGA microspheres. Detect physical and chemical properties.2. The impact of Nanobubbles joint with CHIFU and PHIFU irradiation of normal rabbit liver tissue32 healthy rabbits were randomly divided into A, B, C, D 4 group, successively were group pure CHIFU, group CHIFU joint with nanobubbles, group simple PHIFU, group PHIFU joint nano synergist(PHIFU groups duty cycle are 50%), n = 8, were HIFU point irradiation, the changes of the temperature at whole focus area were observed,then after irradiation measuring damage irradiation zone, calculate the volume of necrotic injured area after irradiation.3. Nano synergist ablation on rabbit liver VX2 tumor of CHIFU and PHIFURabbit liver VX2 tumor model by embedding laparotomy. 41 rabbits were planted. Take one randomly to observe modeling situations. The remaining 40 were randomly divided into A, B, C, D 4 group, followed by pure CHIFU group, CHIFU joint with nanobubbles group, simple PHIFU group, PHIFU joint with nanobubbles group(Duty cycle of PHIFU group were 50%). Each group has 10,HIFU were irradiated through ultrasound monitoring. Recording irradiation time, total irradiation energy and adverse reactions of animals during irradiation. After irradiation take 5 rabbits in each group to observe damage, measure lesion volume and calculate the energy efficiency factor. The remaining 5 rabbits in each group continue to raise two weeks, observe the growth of rabbits, then after two weeks These rabbits were anatomicaled, observe outcome of the situation of tumor.4. Compare nanobubbles joint with CHIFU and PHIFU ablation of rabbit liver VX2 tumor,obsever cell apoptosis and proliferationRabbit liver VX2 tumor model by embedding laparotomy. 41 rabbits were planted. Take one randomly to observe modeling situations. The remaining 20 were randomly divided into A, B, C, D 4 group were pure CHIFU group, CHIFU joint nanobubbles group, simple PHIFU group, PHIFU joint nanobubbles group(group PHIFU duty cycle is 50 %), 5 in per group, Then these were irradiated by HIFU by B-mode ultrasound monitoring. After 24 hours, take the edge of necrotic tissue at the outer 0.5cm, fixed, embedded, sliced, detection kit according to the prompts. Light microscope to calculate AI and PI.Result1. Prepare nanospheres of PLGA parcel of PFH successfully. The average particle diameter is 146 nm.2. Temperature: CHIFU Joint nanobubbles group higher than CHIFU irradiation, PHIFU joint nanobubbles group higher than PHIFU irradiation; CHIFU Joint nanobubbles group was higher than the irradiation PHIFU joint nanobubbles group(P <0.05).3. Coagulation necrosis Volume: CHIFU Joint nanobubbles group is bigger than simply CHIFU group, PHIFU joint nanobubbles group is larger than simply PHIFU group; CHIFU Joint nanobubbles group is larger than PHIFU Joint synergist group(P <0.05).4. Energy efficiency factor: CHIFU Joint synergist group less than pure CHIFU group, PHIFU joint synergist is less than simple PHIFU group, PHIFU Joint synergist group less than CHIFU with synergist group(P <0.05).5. Complications: CHIFU Joint Nano synergist group and pure CHIFU group there are different degrees of skin redness, simple PHIFU group had three cases of skin irritation, PHIFU joint nano synergist group had one case of skin irritation.6. The outcome: each group can effectively control tumor metastasis. CHIFU Joint synergist group and CHIFU group all show gray coagulation necrosis, PHIFU joint synergist group has 4 patients necrosis liquefaction. PHIFU simple irradiation group has 2 cases of necrosis liquefaction.7. Immunohistochemistry: CHIFU Joint Nano synergist group and PHIFU joint synergist group with the corresponding pure CHIFU irradiation and PHIFU irradiation, the apoptotic index were higher, lower proliferation index(P <0.05). Compare PHIFU Joint Nano synergist and CHIFU joint synergist group,expression of PCNA and apoptosis index nearly homogeneous(P> 0.05).Conclusion1. PHIFU joint Nano synergist radiation than simply PHIFU irradiation has bigger coagulation necrosis and higher target temperature, PFH / PLGA polymer nanoparticles as PHIFU enhancers are significantly effective.2. Whether or PHIFU CHIFU joint with nano synergist,United lesion volume was more than simple HIFU irradiation, nano synergist can enhance CHIFU and PHIFU. PHIFU joint Nano synergist than CHIFU joint synergist has lower thermal deposition, PHIFU joint Nano synergist safer than CHIFU, the specific mechanism remains to be further studied.3. Nano boosters for higher duty cycle of PHIFU to enhance the thermal effect based, collaborative Cavitation efficiency.4. PHIFU combined with nano synergist therapy tumor efficiency, more safety, outcomes shortened.5. Nano synergist joint CHIFU and PHIFU could induce apoptosis and inhibit the proliferation of new cells of residual tumor tissue. |
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