Influence Of Acoustic Contrast Agent With Diagnostic Ultrasound On Rabbits' Testicular Germ Cells

Objective:To study the influence of acoustic contrast agent with diagnostic ultrasound on rabbits' testicular germ cells by injecting Sonovue with different dosage or different ways.Methods:Thirty three Nea Zealand white rabbits(12-16 weeks after birth)were randomly divided into 3 groups.In groupⅠ,there were three rabbits without sonicate and contrast agent as the contrast group.In groupⅡ,eighteen rabbits were bolus injected Sonovue from veins of the ear limbus,and physiological saline with dosage of 1ml were injected fast soon after Sonovue.The rabbits were divided into 6 group according to different dosage and different time to draw the materials from rabbiets,including inject single Sonovue with dosage of 0.1ml/kg(A1,A2),single Sonovue with dosage of 1ml/kg (C1,C2) and inject thrice Sonovue with dosage of 0.1ml/kg interval 15min(B1,B2). In groupⅢ,twelve rabbits were bolus injected Sonovue from veins of the ear limbus, and physiological saline with dosage of 1ml were injected fast soon after Sonovue,then breakout the agent bubbles when the Sonovue suffused the whole testis.The rabbits were divided into 4 group according to different dosage and different time to draw the materials from rabbiets,including inject Sonovue with dosage of 0.1ml/kg(D1,D2) and with dosage of 1ml/kg(E1,E2).The contrast parameter between two groups with different dosage(0.1ml/kg and 1ml/kg) of Sonovue was compared and analyzed.All testes were removed immediately or the time 24 hours after test,and HE Stain was done for microscopic observation;apoptosis was detected by in situ terminal deoxynucleotidyl transferase mediated dUTP nick end labeling(TUNEL);ultra-structural changes were observed by transmission electron microscope.Interclass's difference was compared with one-factor analysis of variance.RESULT:CEUS time-enhancement curve of contralateral testes showed no statistical significance between the different dosage(0.1 ml/kg and 1 ml/kg) groups in base intensity(BI),Arrival time(AT),Time to Peak(TTP),etc(P＞0.05),and there were distinct difference in peak intensity(PI),alpha(A=PI-BI),ascending velocity(v),etc (P＜0.05).The is no distinct changes were observed in tests in group A1～C2,and focal and mild pathological changes were observed in testes in group D1～E2,including the occasional presence of vacuoles in small or large clusters,the structural disorder in seminiferous epithelium,exfoliation of germ cells,insufficient spermatozoa,interstitial arterial hyperemia,as well as the presence of multinucleated cells.Johnsen score showed no statistical significance between the control group and group A1～E2 (P＞0.05).Apoptosis detected by terminal deoxynucleotidyl transferase-mediated deoxyuride triphosphate nick-end labeling(TUNEL) showed the total number of appotosis cells in group D1～E2 were significantly more than the control group as well as group A1～C2,while no statistical significance exsisted between group A1～C2 and the control group(P＞0.05).Apoptosis occurred predominantly in spermatocyte and spermatid. The focal ultra-structural damage were observed by transmission electron microscope in group C1 and C2,such as intercellular space widening,mitochondria swelling,and no distinct damage were observed in the group A1～B2.The distinct ultrastructural damage were observer by transmission electron microscope in group D1～E2,such as intercellular space widening,mitochondria swelling,cellular edema,especially spotty necrosisetc was observe in D1 and E1.CONCLUSION:There was some relationship between the dosage of contrast agent and perfusion imaging of testis flood flow.Intravenous injected with comparatively large dosage of contrast agent with low mechanical index,testis would come to light pathological change and focal ultra-structure damage,and the change and damage would vanish 24hours later.But when the agent bubble was broken,the light distinct pathological change and ultra-structure damage was observed in testis and would not vanish 24hours later.There is some significance in using contrast agent during the clinical work.