The Effects Of Ultrasound Microbubble Contrast Agents Upon Rabbit Testicular Complete Torsion-detorsion Under The Diagnostic Ultrasound Radiation

ObjectiveTo explore the ultrasound contrast agent’s (Sonovue) effects upon rabbit testis which wereinjured after the acute unilateral complete torsion-detorsion and its related mechanisms, on thecondition of the diagnostic ultrasound radiation with different doses and different imagingmodes.Method70healthy New Zealand white adult rabbits were randomly divided into10groups, with7in each group. Materials for groups (S1, A1, B1, C1, D1), were based on the immediately afterischemia reperfusion, while groups (S2, A2, B2, C2, D2) were after24hours. Ligate the lefttestis and block the blood flow completely for3hours. Removing the ligation when it reachedthe ischemic time. Reperfusion of groups(S1, S2) were administered without Sonovue,groups(A1,A2) were administered by using a small-dose(0.1ml/kg) Sonovue in conventionalimaging group, groups(B1,B2) were administered by using high-dose(1ml/kg) Sonovue inblasting imaging group, groups(C1,C2) were administered by using small-dose(0.1ml/kg)Sonovue in blasting imaging group, and groups(D1,D2) were administered by usinghigh-dose(1ml/kg) Sonovue in blasting imaging group. Before taking samples, eachexperimental group should have a bilateral testicular contrast-enhanced ultrasound. Thenanalyzed the changes of each group’s Contrast-enhanced ultrasound(CEUS) parameters,including speed parameters(β), time to peak intensity (TTP), peak and base intensitydifference(PBD), and half-time of the descending peak intensity (DT/2). Materials for groups(S1, A1, B1, C1, D1,) were based on the immediately after reperfusion. Cut the rabbitoperative-lateral testis as the materials to observe the testicular specimen by and large and thepathological changes (HE stain) of he tissue sections under the light microscope, to conduct contents of MDA and SOD of testicular tissue, to examine expressions of Caspase-3inspermatogenic epithelial cells by Westernbolt, to detect the cell apoptosis index (AI) by Tunel.While groups (S2, A2, B2, C2, D2) needed do the same job after24hours on the condition ofstandard breeding. Make a comparison study among these groups.Results1．CEUS parameters of operative-lateral testes showed no significance in all group beforeoperation(P＞0.05). In groups which were administered immediately by small-dose andhigh-does Sonovue after removing ligation, CEUS parameters β showed no significance（P＞0.05）,AT shortened and DT/2lengthened with significance (P＜0.05)，PBD ascended withsignificance (P＜0.05). In group D2, the TTP and DT/2are significantly prolonged(P<0.05),PBD is significantly descended(P<0.05), and the time-intensity curve manifested that the peakwas lower and delayed.2．Compared with groups（S1、A1、B1、C1、D1）, in groups （S2、A2、B2、C2、D2）Content of SOD was lower and MDA was higher significantly (P<0.05). SOD in C2waslower slightly than in group S2, and MDA in group C2increased with no significance (P>0.05). Compared with other groups, SOD in D2decreased, and MDA increased with significance(P<0.05).3．Under light microscope, the Johnen’s score showed no significance (P＞0.05) in groups(S1, A1, B1, C1, D1,). The Johnen’s scores in groups（S2、A2、B2、C2、D2）were lower thanin groups (S1, A1, B1, C1, D1,). The Johnen’s scores in groups A2, B2, C2showed nodifference with group S2(P>0.05), while were lower compared with group D2withsignificance (P<0.05).In groups S1, A1, B1, slight angiectasis could be seen in the operatedtestis, and hemorrhage and thrombosis could not be seen apparently. While in groups (C1, D1),angiectasis, congestion and thrombus in some part of small vessels could be seen. Under lightmicroscope and electron microscope, spermatogenic cells impeded seriously in differentiationand maturation.4．Tunel apoptotic index detection and Western blot analysis showed that there were nosignificance of spermatogenic cells’ apoptosis in groups（S1, A1, B1, C1, D1）(P＞0.05). AIand histone expression in groups (S2, A2, B2, C2, D2) was higher than in groups (S1, A1, B1, C1, D1) with significance (P＜0.05). AI, Bax and caspase-3in group C2increased slightlythan in groups (S2, A2, B2C2), while showed no significance (P＞0.05). Compared withothers groups, AI, Bax and caspase-3in group D2increased with significance (P＜0.05).Conclusion①Color Doppler ultrasound can make an accurate diagnosis on acute unilateralcomplete torsion and CEUS time-intensity curves can indirectly reflect the situations oftesticular microcirculation after detorsion. It can guide the further treatment;②Under the diagnostic ultrasound radiation, when conventional imaging and low-doesimaging blasted, they did not aggravate the ischemia testicular. While for high-dose, itaggravated;③in vivo, microbubble increased the sound cavitation effect, which affected thecapillary wall and impeded the microcirculation and damaged the parenchyma cell. Relatedmechanisms had the potential to cause capillary injury and aggravate further ischemia andhypoxia, through cavitation. Through increasing Oxidative Stress, it also can aggravateischemic testicular damage, such as raising a large number of neutrophil infiltration andincreasing the pro-apoptotic proteins (caspase-3and Bax) expression.