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Background spleen is the largest immune organ in the human body, has a variety of immune cells factor has an important anti-infective, anti-tumor function, but it is also the body's blood reservoir, filter blood, hematopoietic organs and destroy blood; study also found that the spleen close to the body's endocrine; variety of physiological functions of the gut plays an important role in the regulation of the liver. The traditional treatment of hypersplenism splenectomy, splenic artery partial embolization (PSE) and spleen radiofrequency ablation (RFA). Splenectomy is not only easily lead to decreased immunity, increased risk of infection, but also may increase plasma viscosity, improve the incidence of thromboembolic; partial embolization of splenic artery and spleen radiofrequency ablation induced risk of bleeding, and accompanied severe pain, fever and other reactions. The effects of ultrasonic cavitation ultrasound thermal effects outside of another physical effect; liquid microbubble ultrasound shock, expansion, contraction, and the implosion of a series of dynamic processes accompanied by transient high-temperature, high-pressure, shock wave, discharge and micro-jet energy release behavior. The microbubbles can reduce ultrasonic cavitation threshold, with a low-energy ultrasound excitation case, significant cavitation can occur, which can be caused by small vessel wall mechanical damage, bleeding, hematoma, and thrombosis. Research purposes known microbubbles enhanced ultrasonic cavitation could produce significant vascular damage effect. In this study, using a self-developed new pulsed ultrasound treatment device, combined with intravenous ultrasound contrast agent for the treatment of vascular occlusion, healthy New Zealand white rabbits spleen, the spleen visual comparison of before and after treatment target grayscale ultrasound imaging, quantitative analysis splenic parenchyma before and after the experiment angiography acoustic intensity, to understand the state of the blood supply and blood circulation;, pathological biopsy of the spleen vascular occlusion preliminary pathological mechanisms of biological effects and potential therapeutic role of spleen ultrasonic cavitation a preliminary study. Materials and methods Experimental animals and grouping healthy adult 36 New Zealand white rabbits, male or female, body weight 1.7-2.5kg provided by the Experimental Animal Center of Guangdong Province. Depending on whether the ultrasound irradiation, whether mediated microbubble destruction, with or without prothrombin involved 36 rabbits were randomly divided into four groups, A: simple micro-bubble group (of Microbubbles, MB);: simple ultrasound group (Ultrasound, US ); C: ultrasound micro-bubble group (Ultrasound and Microbubbles USMB); D: prothrombin ultrasound microbubble group (Prothrombin and Uultrasound and Microbubbles, PUSMB); each group of nine. PUSMB group immediately after I slow intravenous bolus prothrombin (20IU/kg) is pulsed ultrasound irradiation combined with intravenous microbubbles injected; The USMB groups using pulsed ultrasound irradiation combined with intravenous microbubbles injected; U.S. group with the same amount of physiological the the saline alternative microbubbles using pulsed ultrasound irradiation; the MB group after intravenous injection of microbubbles and ultrasound treatment head sham. Second, laboratory instruments and materials pulsed focused ultrasound therapy instrument, provide by the Second Affiliated Hospital of the Third Military Medical University, Department of Ultrasound, ultrasound frequency 831KHZ, pulse width adjustable 100-1000 microseconds peak sound pressure 4.6Mpa, the average sound 0.89 W/cm2. Fat fluorine significant lipid membranes microbubbles and ultrasound research system of the Second Affiliated Hospital of the Third Military Medical University, the core gas perfluoropropane, milky white curd-like appearance, the average particle size of 2 μm, 98% of which is less than 8μm microbubbles concentration of 4-9 × 109/ml. Experimental methods by ear vein injection of 2% sodium pentobarbital anesthetized animals, dose 1.5ml/kg establish the marginal ear vein channel. Anesthesia after rabbit supine fixed the experimental device autopsy table, gently pull the abdominal cavity after surgical exposure spleen around to saline gauze fixed. PUSMB group was given intravenously to Note 20IU/kg prothrombin immediately slow intravenous bolus injection of microbubbles a dose 0.1ml/kg of saline diluent, and contact irradiation 5min ultrasound treatment head vertically spleen light; USMB Group no prothrombin injection rest with PUSMB group the same; U.S. group to probe the vertical irradiation of ultrasound therapy 5min microbubbles with the same dose of saline alternative; the MB group after intravenous microbubbles same the USMB group, while ultrasound treatment of head fake according spleen 5min. 0min, 30min and 60min four time points before treatment and after treatment, spleen diagnostic ultrasound imaging and recording the second harmonic gray scale ultrasound contrast imaging. Complete contrast, playback dynamic image analysis software enable acoustic density QLAB analysis at each time point the the spleen substance of the region of interest (ROI) before irradiation and post-irradiation time intensity curve (TIC), the calculation of peak time (t) and The peak intensity (PI). Each group after treatment 0min randomly selected three rabbits were killed, the rest of the rabbits were killed after 60min after treatment, both for spleen irradiation area and its surrounding tissue, routine biopsy, HE staining, light microscopic examination of tissue The pathological changes. Ultrasound contrast analysis using PILLIPS company the IU22 color Doppler ultrasonic diagnostic apparatus, 9L high frequency linear array probe, frequency 5-9MHz, ultrasound contrast mode. First imaging using two-dimensional imaging longitudinal section of the spleen, adjust the zoom factor and image depth; then choose the best display section, switch to contrast the state to open a double-width mode, dual-site real-time image display, and one for the used to observe the contrast of the image, and the other one for the real-time image of the gray scale, used to monitor thus ensuring always in the same sections of; extraction \ear vein after saline diluted into 5ml fast bolus is completed (3-5s within), then red with 2ml of saline injection; injection of contrast agent at the same time start the ultrasonic diagnostic apparatus built-real-time continuous observation spleen slice perfusion and echo intensity, records the pulse focused ultrasound treatment before and after treatment at 0min, 30min and 60min Targets secondary harmonic gray scale ultrasound contrast image; to enable the QLAB acoustic density analysis software, select the square sampling box, about the size of 1.5 × 1.5mm, in the irradiated area in the spleen (spleen exposed portions of the central region, the the ROI edge at the upper and lower envelope central), to avoid developing large vessels, drawn automatically by the instrument comes into - the intensity curve (TIC) to calculate the peak time (t) and the peak intensity (PI). General observation, pathological examination of irradiated spleen appearance irradiation damage; drawn irradiated area and the surrounding uninjured tissue fixed in formalin solution, HE staining, pathological changes observed by light microscopy organizations, including thrombosis, of microthrombi perivascular tissue cells change. Six statistical processing of each group of data are the mean ± standard deviation (X ± S) said, the data in the same group before and after treatment compared using a paired t-test was used to compare between different groups into groups mean comparison t test or t ' Inspection and testing are P lt; 0.05 with statistical significance, and all data are used SPSS16.0 software. 36 rabbits completed the experimental process. Gross pathology observed before treatment in each group the spleen was uniform bright red. After treatment, the the the prothrombin joint ultrasound microbubble treatment group, ultrasound microbubble treatment group spleen irradiation area was dark red, non-irradiated area slightly dimmed, the irradiated area with non-irradiated areas will have a clearer boundary; The simple ultrasound group and simply microbubble group spleen as a whole slightly darkened, no obvious boundaries between the irradiated area (false according to district) and normal tissue. Two, the ultrasound contrast the results of (a) visual observation: before treatment shows contrast agent quickly into the spleen tissue, spleen overall was uniform enhanced perfusion good filling defect, no significant difference in visual analysis. 0min, PUSMB group, USMB group after treatment slowly visible contrast agent into the irradiated area by the splenic artery, the region enhanced delay, reduce the degree of enhancement, showed the low perfusion and negative developing area, with surrounding normal spleen tissue boundaries. clear results reproducible; MB group and the U.S. group contrast agent is still rapidly into the irradiated area was homogeneous enhancement, well perfused, with no difference before treatment. 30min, 60min, PUSMB group irradiated area after treatment enhanced still delayed perfusion enhancements continue to reduce the defect area increases; the USMB groups irradiated area perfusion defects reduced perfusion strength has been restored; MB group and the U.S. group did not significantly change . (B) acoustic density (AD) Quantitative analysis: Before treatment, each group of TIC curve rising branch steep after treatment PUSMB group, the USMB group TIC curve rising branch gentle, peak intensity (PI) value from before treatment to 22.2 ± treatment 0.48,21.7 ± 0.50 down to 0min 13.8 ± 1.92,12.4 ± 1.89. Peak time (t) values ??were extended from before treatment to 8.2 ± 0.54,7.9 ± 0.64 after treatment 0min of 15.6 ± 1.37,10.8 ± 1.56. Treatment after 30min, 60min observation peak time, PUSMB group USMB group, with 0min similar PI values ??declining PUSMB group, the USMB group PI values ??increased to varying degrees. US groups before and after treatment of the MB group at each time point TIC curve no significant changes, no statistically significant difference in time to peak and peak intensity. Three the pathological examination MB Group: Treatment 0min and treatment after 60min visible the splenic sinus uniform size, see within a few red blood cells; splenic cord structure is clear, and no obvious bleeding, cell degeneration, edema, and thrombosis. U.S. group: after treatment 0min and after treatment 60min visible splenic sinus slightly expansion, but still uniform size, splenic cord structure clearly, we can see a few red blood oozing, no bleeding, cell degeneration, edema, and thrombosis. USMB group: after treatment at 0min visible splenic sinus different degrees of expansion within see more red blood oozing; the splenic cord compression structure is not clear, with varying degrees of perivascular tissue edema, thrombosis seen in the small blood vessels; treatment 60min, visible splenic sinus dilation slightly reduced, there is still more red blood cells oozing still visible in some of the small blood vessels thrombosis; the splenic cord compression a little relief, the structure looming on the horizon, the perivascular tissue cells with varying degrees of edema. 0min, splenic sinus PUSMB group: after treatment significant expansion, which see a large number of red blood cells siltation the splenic cord compression, the structure is not obvious, vascular endothelial cell damage, visible small blood vessels thrombosis; 60min, splenic sinus after treatment remained significant expansion, in addition to micro thrombosis seen in part sinusoids, slightly larger intravascular visible thrombosis; the perivascular tissue cells with varying degrees of edema. Conclusions of a microbubble enhanced ultrasound cavitation effect can be blocked by normal rabbit spleen tissue blood flow showed perfusion significantly delay, reduce, and the emergence of the defect area, but as time goes on, there may be varying degrees of recovery perfusion prothrombin enhance its blocking effect. Microbubble enhanced ultrasound cavitation effect blocking the main mechanism of the splenic blood flow may be damage to the small blood vessels, causing vascular congestion, thrombosis, and a small hematoma, thus choking off the blood circulation. Prothrombin to accelerate and consolidate thrombosis, and enhance the blocking effect.
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