超声微泡增效骨髓干细胞移植效率的实验研究

目的在动物水平研究超声联合微泡增效骨髓干细胞(BMSCs)心肌移植的作用,为临床上提高心肌梗死后BMSCs移植效率提供一种新的思路。方法12只中国家猪分别体外分离纯化BMSCs,超顺磁性氧化铁颗粒标记;介入法建立急性心肌梗死模型,成功后14d经冠状动脉注入自体BMSCs;实验组先注入微泡后再注入BMSCs,同时行超声辐射。术后行磁共振检查;心肌做病理切光镜、透射电镜观察。结果①两组在心肌梗死区和梗死周边区磁共振上均见SPIO标记的BMSCs形成的低密度区;②实验组较对照组普鲁士蓝染色阳性细胞明显增多(P<0.01);③两组心肌超微结构无差异,但B1组见血管内皮间隙增宽。结论超声辐射微泡可增加自体BMSCs心脏的移植效率。     

超声联合一氧化氮微泡对大鼠心肌组织的生物学效应

目的:本研究旨在探讨超声联合一氧化氮(NO)微泡的作用对大鼠心肌组织局部基质衍生因子-1(SDF-1)的影响,及其对心肌组织作用的安全性?方法:将32只SPF级SD雄性大鼠随机分为4组,每组8只?第1?2?3组分别经尾静脉输入1 ml的NO微泡?普通脂质微泡以及PBS,并采用频率为1 MHz?强度为1 W/cm2的超声辐照大鼠心前区,辐照时间为10 min;第4组为空白对照组?4 h后每组处死4只大鼠,留取心肌组织进行HE染色,观察局部的炎症反应情况;同时留取血清标本用于检测肌酸激酶(CK),乳酸脱氢酶(LDH)和肌钙蛋白I(TnI)的含量?1周后处死剩余大鼠,取心肌组织进行RT-PCR和Western blot分别检测SDF-1的基因及蛋白相对表达量?结果:辐照4 h后留取心肌组织行HE染色显示各组均可见少量炎性细胞浸润,组间没有明显差异;血清学指标检测显示,第1组与第2组血清CK和LDH高于第3组和第4组,且差异有统计学意义(P < 0.05);第1组血清TnI低于第2组,但高于第3组和第4组,且差异有统计学意义(P < 0.05);辐照1周后留取的心肌组织行RT-PCR和Western blot分析结果均显示第1组SDF-1相对表达量最多,各组间进行两两比较,差异均有统计学意义(P < 0.05)?结论:超声联合NO微泡辐照大鼠心肌组织,与内含全氟显气体的普通脂质微泡一样,对心肌组织无明显破坏;但能使大鼠心肌组织SDF-1表达增加,其程度高于应用超声联合普通脂质微泡,因此,NO微泡应用于干细胞归巢将优于目前普通脂质微泡?     

超声微泡造影剂在聚焦超声开放血脑屏障中的应用

超声微泡造影剂在聚焦超声无创性开放血脑屏障的过程中具有非常重要的作用,本文就其发展历程、机制、安全性、时效和量效关系进行慨述和探讨。     

Apoptosis Induced by Microbubble-Assisted Acoustic Cavitation in K562 Cells: The Predominant Role of the Cyclosporin A-Dependent Mitochondrial Permeability Transition Pore

Acoustic cavitation of microbubbles has been described as inducing tumor cell apoptosis that is partly associated with mitochondrial dysfunction; however, the exact mechanisms have not been fully characterized. Here, low-intensity pulsed ultrasound (1 MHz, 0.3-MPa peak negative pressure, 10% duty cycle and 1-kHz pulse repetition frequency) was applied to K562 chronic myelogenous leukemia cells for 1 min with 10% (v/v) SonoVue microbubbles. After ultrasound exposure, the apoptotic index was determined by flow cytometry with annexin V–fluorescein isothiocyanate/propidium iodide. In addition, mitochondrial membrane potential (ΔΨ_m) was determined with the JC-1 assay. Translocation of apoptosis-associated protein cytochrome c was evaluated by Western blotting. We found that microbubble-assisted acoustic cavitation can increase the cellular apoptotic index, mitochondrial depolarization and cytochrome c release in K562 cells, compared with ultrasound treatment alone. Furthermore, mitochondrial dysfunction and apoptosis were significantly inhibited by cyclosporin A, a classic inhibitor of the mitochondrial permeability transition pore; however, the inhibitor of Bax protein, Bax-inhibiting peptide, could not suppress these effects. Our results suggest that mitochondrial permeability transition pore opening is involved in mitochondrial dysfunction after exposure to microbubble-assisted acoustic cavitation. Moreover, the release of cytochrome c from the mitochondria is dependent on cyclosporin A–sensitive mitochondrial permeability transition pore opening, but not formation of the Bax-voltage dependent anion channel complex or Bax oligomeric pores. These data provide more insight into the mechanisms underlying mitochondrial dysfunction induced by acoustic cavitation and can be used as a basis for therapy.     

The antivascular action of physiotherapy ultrasound on a murine tumor: role of a microbubble contrast agent

This study investigated whether a microbubble-containing ultrasound contrast agent had a role in the antivascular action of physiotherapy ultrasound on tumor neovasculature. Ultrasound images (B-mode and contrast-enhanced power Doppler [0.02 mL Definity]) were made of 22 murine melanomas (K1735²²). The tumor was insonated (I_SATA = 1.7 W cm^–2, 1 MHz, continuous output) for 3 min and the power Doppler observations of the pre- and postinsonation tumor vascularities were analyzed. Significant reductions (p = 0.005 for analyses of color-weighted fractional area) in vascularity occurred when a contrast-enhanced power Doppler study occurred before insonation. Vascularity was unchanged in tumors without a pretherapy Doppler study. Histologic studies revealed tissue structural changes that correlated with the ultrasound findings. The underlying etiology of the interaction between the physiotherapy ultrasound beam, the microbubble-containing contrast agent and the tumor neovasculature is unknown. It was concluded that contrast agents play an important role in the antivascular effects induced by physiotherapy ultrasound. (E-mail: sehgalc@uphs.upenn.edu)     

低频超声辐射微泡造影剂对人乳腺癌细胞的生物学效应研究

目的观察微泡造影剂不同浓度、不同声学参数对体外培养人乳腺癌细胞的生物学效应。方法人乳腺癌细胞株MDA-MB-231,受不同剂量,频率为300kHz的超声波辐照破坏微泡声学造影剂,采用噻唑蓝(MTT)法检测细胞存活率。结果在微泡浓度为5%时,0.25W/cm2×10s与0.5W/cm2×5s组细胞存活率未见明显改变,而0.5W/cm2×10s、0.5W/cm2×20s、0.5W/cm2×40s、1.0W/cm2×10s、2.0W/cm2×10s各组与对照组比较,细胞存活率明显下降。辐照强度为0.5W/cm2、照射时间10s时,随着微泡浓度增加,超声对细胞的杀伤效应越强,与对照组相比,5%微泡浓度组P<0.05,其余各组P<0.01。结论超声破坏微泡声学造影剂对人乳腺癌细胞的杀伤程度与超声辐照时间、声强及微泡浓度有关。     

小鼠颊癌颈部淋巴结超声造影的实验研究

目的 探讨局部注射造影剂对移植颊癌原发灶及颈部转移淋巴结增强显像效果.方法 建立小鼠颊癌颈部转移淋巴结动物模型后,经原发灶局部注入自制表面活性剂类超声微泡,以谐波显像方式观察原发灶及颈部淋巴结增强显像效果,同时进行回声强度的比较.结果 造影前,转移淋巴结检出率为61%（17/28）,炎性淋巴结检出率为50%（6/12）;造影后转移淋巴结检出率为82%（23/28）,炎性淋巴结检出率为75%（9/12）.与造影前比较,造影后原发灶和淋巴结的回声强度增大,差异有统计学意义（P〈0.01）结论自制表面活性剂类超声微泡能够经原发灶注入,进入淋巴道,实现显著的淋巴结超声显像增强效果.     

Creating Brain Lesions with Low-Intensity Focused Ultrasound with Microbubbles: A Rat Study at Half a Megahertz

Low-intensity focused ultrasound was applied with microbubbles (Definity, Lantheus Medical Imaging, North Billerica, MA, USA; 0.02 mL/kg) to produce brain lesions in 50 rats at 558 kHz. Burst sonications (burst length: 10 ms; pulse repetition frequency: 1 Hz; total exposure: 5 min; acoustic power: 0.47–1.3 W) generated ischemic or hemorrhagic lesions at the focal volume revealed by both magnetic resonance imaging and histology. Shorter burst time (2 ms) or shorter sonication time (1 min) reduced the probability of lesion production. Longer pulses (200 ms, 500 ms and continuous wave) caused significant near-field damage. Using microbubbles with focused ultrasound significantly reduced acoustic power levels and, therefore, avoided skull heating issues and potentially can extend the treatable volume of transcranial focused ultrasound to brain tissues close to the skull.     

Ultrasound with Microbubble Contrast Agent and Urokinase for Thrombosis

This study was aimed at assessing the effects of urokinase (UK) in combination with ultrasound and microbubbles in in vitro and in vivo thrombolytic therapy for the treatment of deep vein thrombosis (DVT). Thrombi with formation times of 1, 3, 7, 14 and 21 d were used for thrombolysis. Forty-five adult mongrel dogs were used to evaluate thrombosis in vivo. Both in vitro and in vivo analyses revealed that UK?+?microbubbles had the best effect among the combinations. Thrombolysis <7 d was more effective at a thrombolysis rate of about 50%, but the thrombolytic effect of thrombi >7 d was poor at thrombolysis rates <30%. Ultrasound?+?UK significantly increased the thrombolysis rate of thrombi <7 d. These results suggest that the combination of ultrasound with microbubble contrast agents and UK may have a synergistic effect on thrombolysis.     

诊断超声联合微泡靶向开放单侧大鼠血脑屏障的初步研究

目的初步探讨诊断超声靶向击破微泡开放大鼠单侧血脑屏障通透性的可行性、有效性及安性。方法25只健康SD大鼠,诊断超声经颞骨照射单侧大脑半球,对侧大脑半球作对照。对伊文思蓝外渗范围进行分级和萃取定量。透射电镜观察硝酸镧分布评价血脑屏障超微结构变化,并对双侧脑实质的病理损伤进行评价和分级。结果照射侧大脑半球可见以海马为中心的片状均匀伊文思蓝渗出,而对侧脑实质未见渗出;透射电镜照射侧大脑半球硝酸镧渗出血管并分布于神经纤维间隙,而对照侧未见硝酸镧外漏;超微结构显示少量神经元轻度肿胀。病理组织学观察照射侧大脑半球可见少量红细胞外漏,未见明显组织细胞损伤。结论诊断超声靶向击破微泡能安全有效靶向开放单侧大鼠血脑屏障且呈海马为中心分布,这对于颅内以海马为中心分布疾病的靶向性药物治疗具有潜在的临床价值。