Renel Denervation By Noninvasive Dual-Frequency Focusedultrasound System In Canine Model

PART ? DEVELOPMENT AND FUNCTIONAL DETECTION OF NONINVASIVE DUAL-FREQUENCY FOCUSED ULTRASOUND SYSTEMObjective: To cooperate and develop the noninvasive dual-frequency focused ultrasound system,and to detect its acoustic characteristics.Observing its working status in the mode of the same frequency and different frequency,as well as understanding the basic function of this ultrasonic system.In addition,the working state of this system was detected by tissues in vitro,the formation of biological focal region under different acoustic parameter was observed,providing a platform for animal experiments.Methods: The ultrasound system is mainly composed of well focused dual-frequency focused ultrasound transducer,precise frequency and phase controlling ultrasound signal generation system,stepping three-dimensional motion controlling system and high-quality ultrasound imaging system.The radiative force method was used to detect the electro-acoustic conversion ratio of transducer,the distribution of sound field was observed by the needle hydrophone,and the formation of biological focal region and the coordination ability of all components were studied by isolated bovine liver.Finally,the positions of virtual focus and actual focus were verified through the guidance of ultrasound imaging and software system,which can be a foundation for accurate determination of target in the following experiments.Results: The ratio of electro-acoustic conversion of ultrasound transducer under the same frequency mode was up to 50%.The actual electrical output power and the electric-acoustic conversion ratio decreased slightly after using the difference frequency mode.A low frequency envelope was detected at the focal point by the hydrophone,and the distribution of the acoustic field showed that the morphology of focal region shaped "spindle",with the longitudinal axis of 15mm×2mm×2mm.Coagulative necrosis region were largest and in the same frequency mode using isolated bovine liver.With the change of frequency difference,the morphology of biological focal region changed from "oval" to "long spindle",and the coagulative necrosis region tended to decrease gradually.Through the guidance of ultrasound imaging system and software controlling system,the position of actual focus and the virtual focus that on imaging system was the same.The ultrasound system can work effectively and stably after the above detection.Conclusion: According to the results of detection,the noninvasive dual-frequency focused ultrasound system can achieve seamless switching between the same frequency and the difference frequency mode,and the frequency can be adjusted within a certain range.The size and shape of focus about ultrasound transducer are good,and its electric-acoustic conversion ratio is high.Meanwhile,the 3d motion controlling system has high precision,and the ultrasound imaging system can provide high-quality imaging and doppler flow imaging,this ultrasound system can control the position of focus under the visualization software to ensure the accuracy of experiment.This noninvasive dual-frequency focused ultrasound system provides a powerful guarantee for animal experiments.PART ? RENAL SYMPATHETIC NERVE STIMULATION BY NONINVASIVEDUAL-FREQUENCYFOCUSEDULTRAS OUNDSYSTEM INCANINEObjective: Renal sympathetic nerve stimulation bydifference frequency model of noninvasive dual-frequency focused ultrasound system.Based on the change of blood pressure,heart failure and hematology,determiningthe distribution ofpositive stimulation,proving the efficacy of renal sympathetic nerve stimulationthat mediated by difference frequency ultrasound,further to provide a new method to determine target of RDN.Methods: A total of 8 adult healthy beagles were included in this study.Real-time electrocardiogram and invasive arterial blood pressure monitoring were performed.Color Doppler flow imaging(CDFI)was obtained along the long axis of renal artery using noninvasive dual-frequency focused ultrasound system.Virtual focus was moved to both sides of the proximal,middle and distal segments of the renal artery,point-by-point stimulation was performed with 50 Wacoustic power,?f=10K,and duration 6s×5 acoustic parameters.The distance of each point was 2mm on the same section of renal arteryuntil renal artery CDFI disappeared.The change of blood pressure,heart rate and renal function were recorded before,during and after stimulation 10 s,20s and 30 s,and histological examination was performed.Results: Positive points were systolic blood pressure increased equal or over 5mm Hg during stimulation,and negative points without obvious change.Both systolic and diastolic blood pressure were elevated from baseline at positive points during stimulation(P<0.05),then gradually decreased after stimulation,and recovered to the baseline within 30 s.There was no change in systolic or diastolic blood pressure in the negative stimulus.There was no significant change in heart rate before and after the experiment(P>0.05).Positive points were observed in a total of 7 dogs that located in the proximal ventral renal artery.The level of NE was higher than baseline,there were no renal vascular and nerves injury in gross specimens and histological examination.Conclusion: The difference frequency mode of self-developed noninvasive dual-frequency focused ultrasound system for RNS,which realized effective renal sympathetic nerve stimulation without causing renal vascular and nerve injury.This method of stimulating nerves by the mechanical effect of difference frequency ultrasound that provided a new strategy to determine the specific position of sympathetic nerve,and provided a new direction to determine the effective target for RDN.PART ? RDN BY NONINVASIVEDUAL-FREQUENCYFOCUSED ULTRASOUNDSYSTEM IN CANINEObjective: In this part,we used noninvasive dual-frequency focused ultrasound system to stimulate nerves with the mode of difference frequency,and high-intensity acoustic energy to ablate nerves by the same frequency mode,then re-stimulation by the mode of difference frequency.Through the method of stimulation,ablation and re-stimulation could determine target and ablation endpoint,realizing RDN effectively.Methods: Twenty-eight healthy adult beagles were randomly divided into stimulation+ablation group(n=8),random ablation group(n=8),simple stimulation group(n=8)and the sham control group(n=4).Under the guidance of CDFI,the two sides of the proximal,middle and distal renal arteries,as well as their ventral and dorsal parts were stimulated with the acoustic power of 50 W,?f=10K,duration of 5s×6 in stimulation+ablation group and simple stimulation group.The acoustic power of 350 W and duration of 3s×2 was used to ablate positive points in stimulation+ ablation group.Then this point was re-stimulated with the same acoustic stimulation parameters.The process was repeated by stimulation-ablation and re-stimulation until the positive points were ablated without the increase of blood pressure under stimulation.In the randomized ablation group,random ablation was performed with acoustic power of 350 W and time of 3s along renal artery without stimulation.In simple stimulation group,the sympathetic nerve was stimulated by ultrasound without ablation.The sham control group received the above procedures but no ultrasonic energy release.Changes in blood pressure,NE of renal tissue,renal function,gross specimens and histological examination on baseline and day 28 were recorded.Compared with the baseline,the systolic and diastolic blood pressuredecreased by-16±5.4mm Hg and-10±4.4mm Hgin stimulation+ablation group respectively on day 28 after surgery(P<0.05).The systolic and diastolic blood pressures decreased by-8±3.7 and-5±3.4mm Hgrespectivelyin random ablation group(P<0.05).There was not significant decline of blood pressure in simple stimulation group and sham control group(P>0.05).Compared with other three groups,the systolic blood pressure decreased significantly in stimulation+ablation group on day 28 after surgery,with the lowest level of NE of renal tissue(P<0.05).No obviously change on renal function were observed in the four groups before and after surgery(P>0.05).The peripheral adipose tissue of the renal artery was slightly reduced after ablation,thenerves around renal arteries were necrosis with varying degrees by histological examination.Conclusion: The strategy of difference frequency stimulation,same frequency ablation and difference frequency re-stimulation for RDN using noninvasive dual-frequency focused ultrasound system,which could reduce blood pressure significantly.Meanwhile,compared with random ablation,this method could reduce systolic blood pressure and improve therapeutic effects obviously.This combination of acoustic stimulation and ablation realized safe and effective ablation,providing a new technical method for the treatment of refractory hypertension by RDN.