超声联合微泡开放大鼠血脑屏障提高阿糖胞苷脑组织药物浓度
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摘要
目的
探讨低功率超声联合微泡靶向、可逆的开放大鼠脑血脑屏障(BBB),增强脑组织阿糖胞苷(Ara-C)药物浓度的可行性。
方法
1、实验组45只成年雄性SD大鼠,根据不同辐照时间(10s、30s、60s)和超声功率(0.8W、1.9W、2.7W)组合,分为9组,利用MRI增强扫描测定靶点信号增强值,通过静注伊文氏蓝后脑组织切片以观察靶点蓝染情况,从而判断血脑屏障是否开放,然后通过组织切片在光镜下观察脑组织受损程度,寻找大鼠血脑屏障开放的最适参数。
2、常规剂量组25只SD大鼠,在超声联合微泡开放血脑屏障后的不同时间点(0h、2h、4h、6h、24h)经尾静脉注射常规剂量Ara-C(4mg/kg),通过MRI增强扫描测定靶点信号增强值,高效液相色谱法(HPLC)测定靶点和非靶点脑组织在不同时间点的Ara-C浓度。从而推测最适参数下超声联合微泡开放BBB后的BBB通透性的变化规律。
3、大剂量组5只SD大鼠按上述方法2中相同的步骤经尾静脉注射微泡,但不进行超声辐照,模拟全身大剂量化疗注射Ara-C(50mg/kg),利用MRI增强扫描来测定靶点信号强度增强值, HPLC测定脑组织内的Ara-C浓度。
4、分析实验组、对照组、大剂量组脑组织内Ara-C浓度的差异。测得的数值全部表示为均值±标准差(x±s),用SPSS13.0统计分析软件,各组数据之间的比较采用t检验,P<0.05为有统计学差异,P<0.01为有显著性差异。
结果
1.大鼠BBB开放的最适参数
1.1靶点MRI增强扫描结果:对各组大鼠脑组织行MRI增强扫描发现,0.8W+10s组、0.8W +30s组、0.8W+60s组、1.9W+10s组、2.7W+10s组的增强值与对照组相比无显著性差异;1.9W+30s组、1.9W+60s组、2.7W+30s组、2.7W+60s组的增强值明显高于对照组(P<0.01)。
1.2靶点EB染色结果:0.8W+10s组、0.8W +30s组、0.8W+60s组、1.9W+10s组、2.7W+10s组靶点均未见明显蓝染。1.9W+30s组、1.9W+60s组、2.7W+30s组、2.7W+60s组可见靶点蓝染,并且蓝染范围及程度随着幅照时长和超声功率的增加而增加。
1.3 HE染色组织学检查结果:0.8W+10s组、0.8W +30s组、0.8W+60s组、1.9W+10s组、2.7W+10s组光镜下没有看到明显的靶点组织损伤或红细胞渗出,与靶点外的组织无显著区别;1.9W+30s组可见微血管周围少量红细胞的渗出,以及血管内皮细胞的轻微肿胀,但神经细胞形态没有明显异常;1.9W+60s组、2.7W+30s组见红细胞明显渗出,微血管明显破坏,内皮细胞破损,细胞碎片可见,实质细胞也有不同程度受损,周围组织可见水肿带;2.7W+60s组血管破裂以及红细胞广泛的渗出,可见血管内血栓形成以及少量炎细胞,神经细胞明显肿胀,组织液化坏死。
2.最适参数下超声联合微泡开放BBB后其通透性变化规律超声功率1.9W辐照30s后0h、2h、4h靶点的MRI信号增强值和脑组织的Ara-C浓度明显高于非靶点处(P<0.01),2h达到峰值,而8h、24h组的测定值与对照组比较无显著性差异。
3. HPLC测得超声实验组的脑组织Ara-C浓度为7.69±1.65ug/g脑组织,常规剂量Ara-C组为1.75±0.76 ug/g脑组织,大剂量阿糖胞苷组为7.04±2.76ug/g脑组织。超声实验组与大剂量阿糖胞苷组无显著性差异(P>0.05),而超声实验组明显高于常规剂量Ara-C组,有显著性差异(P<0.01)。
结论
1、在适宜的辐照时间和功率参数条件下(30s+1.9W),低功率超声联合微泡能可逆性开放BBB,而脑组织损伤很小。
2、利用低功率超声联合微泡可逆开放BBB技术,能够显著增加脑组织Ara-C浓度,给予常规剂量即可达到大剂量Ara-C静脉给药相同效果,又避免了大剂量给药可能带来的全身副作用,效果明显优于单纯静脉给药,这种新型给药方式可能为防治中枢神经系统白血病提供一个新思路。
OBJECTIVE
To investigate the effects of reversible and targeted opening the blood–brain barrier (BBB) by MRI-guided focused ultrasound (FUS) Asssociated with Microbubbles and delivering cytrarabine(Ara-C) to the rat brain. Then to explore a effective administration route of curing central nervous system leukemia(CNS-L).
METHODS
1. According to different exposure times(10s,30s,60s) and different ultrasonic frequency(0.8W,1.9W,2.7W) respectively, 45 SD rats sonicated as experimental group(EG) by MRI-guided FUS were divided into 9 groups. then Evans blue extravasation , contrast-enhanced MRI scans and histologic examination were performed to evaluate the optimum condition for opening BBB with minimal damage.
2. 25 SD rats were injected with Normal doses of Ara-c(4mg/kg) through the tail vein at every time point(0h,2h,4h,6h,24h) after FUS with microbubbles. The target as experimental group and the non-target ascontrol group. The Ara-c concentration was analyzed by high– performance liquid chromatography (HPLC) and the Ara-c concentration and signal intensity enhancement of MRI was detected on every time point to determine the change tendency of permeability of BBB after sonication.
3. Five SD rats were injected with Microbubbles through the tail vein without sonication to model general chemotherapy by large doses of Ara-c(50mg/kg) as large doses group(LDG). The MRI signal intensity enhancement in the target locations was detected the specimens of the brain tissue were and the concentration of Ara-c in all the specimens was determined by HPLC.
4. To analyze and compare the concentration of Ara-c in the brain tissue specimens of 3 groups. Results are presented as mean±SD. Statistical differences in the experimental groups were analyzed with a t test, where P < .01 was considered statistically significant.
RESULTS
1. The optimum condition for opening BBB of rats.
1.1 Contrast-enhanced MRI: Significance difference of Ga retention was observed in 1.9W+30s group, 1.9W+60s group, 2.7W+30s group and 2.7W+60s group compared with the control group (P < 0.01).
1.2 Evans blue staning: Immediately performed after sonication. No visible blue stain was noted in the brain specimens exposed to 0.8W+10s group, 0.8W+30s group, 0.8W+60s group, 1.9W+10s group and 2.7W+10s group; While the EB stained positively in 1.9W+30s group, 1.9W+60s group, 2.7W+30s group, and 2.7W+60s group. EB extravasation was more broadly distributed and darker when sonication was performed for longer durations and higher power.
1.3. Hematoxylin(HE) and eosin histologic analysis at the targeted focal locations: Immediately after sonication, there was no damage and RBCs extravasation in the sonicated sites of 0.8W+10s group, 0.8W+30s group, 0.8W+60s group, 1.9W+10s group and 2.7W+10s group, and the findings of these sites differed little from those of the nonsonicated sites. In the sites of 1.9W+30s group, a few scattered erythrocytes were observed, and no obvious signs of parenchymal damage were detected. Sites of 1.9W+60s group and 2.7W+30s group performed extravasation of erythrocytes around microvessels, and signs of vasodilatation, stasis, and destruction were noted in the vessel walls. Parenchymal damage was mostly in the form of ischemic changes, and slight vacuolation of the neuropil was seen. In the sites of 2.7W+60s group, multiple areas of blood vessel destruction and extensive erythrocyte extravasations into brain parenchyma were noted, the brain tissue was edematous with fragmented axons, the vacuolation, local tissue necrosis, and neutrophil infiltration were observed.
2. The change tendency of permeability of BBB after sonication with Microbubbles:The MRI signal intensity enhancement and the concentration of Ara-c in brain tissue of specimens in sites of 0h, 2h and 4h was significant higher than that of nontargeted sites. It reached peak at 2h while it had no significant difference with control group at 8h and 24h(P<0.01).
3. Ara-c concentrations for all the groups: the Ara-c levels of EG, NDG and LDG were 7.69±1.65ug/g tissue, 1.75±0.76ug/g tissue and 7.04±2.76ug/g tissue respectively. No significant differences were observed between the EG and LDG(P>0.05). Ara-c concentration in the brain tissue of EG was significantly higher than that of NDG(P < 0.01).
CONCLUTION
1. MRI-guided focused ultrasound can open the BBB targeted and reversible with the minimal damage of brain tissue on the optimum condition (30s+1.9W).
2. MRI-guided FUS can open the BBB reversibly and deliver the intravenously administered Ara-c to the targeted brain locations; the application of this method brings about a substantial increase in the drug level in the targeted brain tissue and has similar effect to large drug doses delivery. In this way side-effect large drug doses delivery could be avoided. Accordingly, this effective administration route may hew out a new perspective of curing CNS-L.
探讨低功率超声联合微泡靶向、可逆的开放大鼠脑血脑屏障(BBB),增强脑组织阿糖胞苷(Ara-C)药物浓度的可行性。
方法
1、实验组45只成年雄性SD大鼠,根据不同辐照时间(10s、30s、60s)和超声功率(0.8W、1.9W、2.7W)组合,分为9组,利用MRI增强扫描测定靶点信号增强值,通过静注伊文氏蓝后脑组织切片以观察靶点蓝染情况,从而判断血脑屏障是否开放,然后通过组织切片在光镜下观察脑组织受损程度,寻找大鼠血脑屏障开放的最适参数。
2、常规剂量组25只SD大鼠,在超声联合微泡开放血脑屏障后的不同时间点(0h、2h、4h、6h、24h)经尾静脉注射常规剂量Ara-C(4mg/kg),通过MRI增强扫描测定靶点信号增强值,高效液相色谱法(HPLC)测定靶点和非靶点脑组织在不同时间点的Ara-C浓度。从而推测最适参数下超声联合微泡开放BBB后的BBB通透性的变化规律。
3、大剂量组5只SD大鼠按上述方法2中相同的步骤经尾静脉注射微泡,但不进行超声辐照,模拟全身大剂量化疗注射Ara-C(50mg/kg),利用MRI增强扫描来测定靶点信号强度增强值, HPLC测定脑组织内的Ara-C浓度。
4、分析实验组、对照组、大剂量组脑组织内Ara-C浓度的差异。测得的数值全部表示为均值±标准差(x±s),用SPSS13.0统计分析软件,各组数据之间的比较采用t检验,P<0.05为有统计学差异,P<0.01为有显著性差异。
结果
1.大鼠BBB开放的最适参数
1.1靶点MRI增强扫描结果:对各组大鼠脑组织行MRI增强扫描发现,0.8W+10s组、0.8W +30s组、0.8W+60s组、1.9W+10s组、2.7W+10s组的增强值与对照组相比无显著性差异;1.9W+30s组、1.9W+60s组、2.7W+30s组、2.7W+60s组的增强值明显高于对照组(P<0.01)。
1.2靶点EB染色结果:0.8W+10s组、0.8W +30s组、0.8W+60s组、1.9W+10s组、2.7W+10s组靶点均未见明显蓝染。1.9W+30s组、1.9W+60s组、2.7W+30s组、2.7W+60s组可见靶点蓝染,并且蓝染范围及程度随着幅照时长和超声功率的增加而增加。
1.3 HE染色组织学检查结果:0.8W+10s组、0.8W +30s组、0.8W+60s组、1.9W+10s组、2.7W+10s组光镜下没有看到明显的靶点组织损伤或红细胞渗出,与靶点外的组织无显著区别;1.9W+30s组可见微血管周围少量红细胞的渗出,以及血管内皮细胞的轻微肿胀,但神经细胞形态没有明显异常;1.9W+60s组、2.7W+30s组见红细胞明显渗出,微血管明显破坏,内皮细胞破损,细胞碎片可见,实质细胞也有不同程度受损,周围组织可见水肿带;2.7W+60s组血管破裂以及红细胞广泛的渗出,可见血管内血栓形成以及少量炎细胞,神经细胞明显肿胀,组织液化坏死。
2.最适参数下超声联合微泡开放BBB后其通透性变化规律超声功率1.9W辐照30s后0h、2h、4h靶点的MRI信号增强值和脑组织的Ara-C浓度明显高于非靶点处(P<0.01),2h达到峰值,而8h、24h组的测定值与对照组比较无显著性差异。
3. HPLC测得超声实验组的脑组织Ara-C浓度为7.69±1.65ug/g脑组织,常规剂量Ara-C组为1.75±0.76 ug/g脑组织,大剂量阿糖胞苷组为7.04±2.76ug/g脑组织。超声实验组与大剂量阿糖胞苷组无显著性差异(P>0.05),而超声实验组明显高于常规剂量Ara-C组,有显著性差异(P<0.01)。
结论
1、在适宜的辐照时间和功率参数条件下(30s+1.9W),低功率超声联合微泡能可逆性开放BBB,而脑组织损伤很小。
2、利用低功率超声联合微泡可逆开放BBB技术,能够显著增加脑组织Ara-C浓度,给予常规剂量即可达到大剂量Ara-C静脉给药相同效果,又避免了大剂量给药可能带来的全身副作用,效果明显优于单纯静脉给药,这种新型给药方式可能为防治中枢神经系统白血病提供一个新思路。
OBJECTIVE
To investigate the effects of reversible and targeted opening the blood–brain barrier (BBB) by MRI-guided focused ultrasound (FUS) Asssociated with Microbubbles and delivering cytrarabine(Ara-C) to the rat brain. Then to explore a effective administration route of curing central nervous system leukemia(CNS-L).
METHODS
1. According to different exposure times(10s,30s,60s) and different ultrasonic frequency(0.8W,1.9W,2.7W) respectively, 45 SD rats sonicated as experimental group(EG) by MRI-guided FUS were divided into 9 groups. then Evans blue extravasation , contrast-enhanced MRI scans and histologic examination were performed to evaluate the optimum condition for opening BBB with minimal damage.
2. 25 SD rats were injected with Normal doses of Ara-c(4mg/kg) through the tail vein at every time point(0h,2h,4h,6h,24h) after FUS with microbubbles. The target as experimental group and the non-target ascontrol group. The Ara-c concentration was analyzed by high– performance liquid chromatography (HPLC) and the Ara-c concentration and signal intensity enhancement of MRI was detected on every time point to determine the change tendency of permeability of BBB after sonication.
3. Five SD rats were injected with Microbubbles through the tail vein without sonication to model general chemotherapy by large doses of Ara-c(50mg/kg) as large doses group(LDG). The MRI signal intensity enhancement in the target locations was detected the specimens of the brain tissue were and the concentration of Ara-c in all the specimens was determined by HPLC.
4. To analyze and compare the concentration of Ara-c in the brain tissue specimens of 3 groups. Results are presented as mean±SD. Statistical differences in the experimental groups were analyzed with a t test, where P < .01 was considered statistically significant.
RESULTS
1. The optimum condition for opening BBB of rats.
1.1 Contrast-enhanced MRI: Significance difference of Ga retention was observed in 1.9W+30s group, 1.9W+60s group, 2.7W+30s group and 2.7W+60s group compared with the control group (P < 0.01).
1.2 Evans blue staning: Immediately performed after sonication. No visible blue stain was noted in the brain specimens exposed to 0.8W+10s group, 0.8W+30s group, 0.8W+60s group, 1.9W+10s group and 2.7W+10s group; While the EB stained positively in 1.9W+30s group, 1.9W+60s group, 2.7W+30s group, and 2.7W+60s group. EB extravasation was more broadly distributed and darker when sonication was performed for longer durations and higher power.
1.3. Hematoxylin(HE) and eosin histologic analysis at the targeted focal locations: Immediately after sonication, there was no damage and RBCs extravasation in the sonicated sites of 0.8W+10s group, 0.8W+30s group, 0.8W+60s group, 1.9W+10s group and 2.7W+10s group, and the findings of these sites differed little from those of the nonsonicated sites. In the sites of 1.9W+30s group, a few scattered erythrocytes were observed, and no obvious signs of parenchymal damage were detected. Sites of 1.9W+60s group and 2.7W+30s group performed extravasation of erythrocytes around microvessels, and signs of vasodilatation, stasis, and destruction were noted in the vessel walls. Parenchymal damage was mostly in the form of ischemic changes, and slight vacuolation of the neuropil was seen. In the sites of 2.7W+60s group, multiple areas of blood vessel destruction and extensive erythrocyte extravasations into brain parenchyma were noted, the brain tissue was edematous with fragmented axons, the vacuolation, local tissue necrosis, and neutrophil infiltration were observed.
2. The change tendency of permeability of BBB after sonication with Microbubbles:The MRI signal intensity enhancement and the concentration of Ara-c in brain tissue of specimens in sites of 0h, 2h and 4h was significant higher than that of nontargeted sites. It reached peak at 2h while it had no significant difference with control group at 8h and 24h(P<0.01).
3. Ara-c concentrations for all the groups: the Ara-c levels of EG, NDG and LDG were 7.69±1.65ug/g tissue, 1.75±0.76ug/g tissue and 7.04±2.76ug/g tissue respectively. No significant differences were observed between the EG and LDG(P>0.05). Ara-c concentration in the brain tissue of EG was significantly higher than that of NDG(P < 0.01).
CONCLUTION
1. MRI-guided focused ultrasound can open the BBB targeted and reversible with the minimal damage of brain tissue on the optimum condition (30s+1.9W).
2. MRI-guided FUS can open the BBB reversibly and deliver the intravenously administered Ara-c to the targeted brain locations; the application of this method brings about a substantial increase in the drug level in the targeted brain tissue and has similar effect to large drug doses delivery. In this way side-effect large drug doses delivery could be avoided. Accordingly, this effective administration route may hew out a new perspective of curing CNS-L.
引文
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[9] Barnard JW, Fry WJ, Fry FJ, et al. Effects of high intensity ultrasound on the central nervous system of the cat[J]. J Comp Neurol. 1955,103:459–84
[10] Ballantine Jr HT, Bell E, Manlapaz J. Progress and problems in the neurological applications of focused ultrasound[J]. J Neurosurg. 1960,17:858–76
[11] Patrick JT, Nolting MN, Goss SA, et al. Ultrasound and the blood–brain barrier[J], Adv Exp Med Biol. 1990,267:369–381
[12] Vykhodtseva N, Hynynen K, Damianou C. Histologic effects of high intensity pulsed ultrasound exposure with subharmonic emission in rabbit brain in vivo[J]. Ultrasound Med Biol,1995,21:969-79
[13] Mychaskiw G,Badr AE,Tibbs R,et al.Optison(FS069)disrupts the blood—brain barrier in rats[J], Anesth Analg.2000,91:798-803
[14] Hynynen K, McDannold N, Vykhodtseva N, et al.Noninvasive MR imaging-guidedfocal opening of the blood-brain barrier in rabbits[J]. Radiology. 2001,220:640–46
[15] Vykhodtseva NI, McDannold N, Agabian S,et al. Histology findings after ultrasound exposure of the brain with ultrasound contrast agent—role of ischemia and apoptosis[A].Chapelon JY, Lafon C(eds): 3rd International Symposium on Therapeutic Ultrasound[C]. Lyon, France: INSERM,2003,80–85
[16] Hynynen K, McDannold N, Martin H, et al. The threshold for brain damage in rabbits induced by bursts of ultrasound in the pressence of an ultrasound contrast agent (option), Ultrasound in Med.&Biol, 2003,29(3): 473-481.
[17] Xiangtao Yin, Kullervo Hynynen. A numerical study of transcranial focused ultrasound bean propagation at low frequency. Phys. Med. Biol. 2005,50:1821-1836
[18] Hynynen K, McDannold N, Martin H, et al. The threshold for brain damage in rabbits induced by bursts of ultrasound in the pressence of an ultrasound contrast agent (option), Ultrasound in Med.&Biol, 2003,29(3): 473-481.
[19] Illum L . Nasal drug delivery--possibilities,problems and Solutions(J). J Control Release,2003,87(1-3):187—198
[20] Sheikov N, McDannold N, Vykhodtseva N, et al. Cellular mechanisms of the blood-brain barrier opening induced by ultrasoundin presence of microbubbles. Ulresound in Med.&Biol, 2004,30(7):979-989.
[21] Laske DW,Youle RJ,Oldfield EH.Tumor regression with regional distribution of the targeted toxin TF-CRM107 in patients with malignant brain tumors.Nat Med,1997,3(12):1362-1368.
[22] Weaver M, Laske DW. Transferrin receptor ligand-targeted toxin conjugate (Tf-CRM107) for therapy of malignant gliomas.J Neurooncol,2003,65(1):3-13.
[23] Haar,GT Therapeutic applications of ultrasound [J]Prog Biophys MolBiol,2007,93(1-3):111-129.
[24] Rouviere O,Lyonnet D,Raudrant A,et a1.MRI appearance of prostate following transrectal HIFU ablation of localized cancer[J].Eur Urol,2001,4O(3):265—274
[25]胡俊,丁仕义,黎海涛.血脑屏障开放的检测方法[J].重庆医学.2003,32(12): 1744-1746
[26] Carrera RM,Pacheco AM,Mastroti RA.Qualitative evaluation of the blood-brain barrier after the use of hypertonic saline solution in young rats[J].Eur Surg Res,2001,33:311-9
[27] Kinoshita M, McDannold N,Jolesz FA.et al.Targeted delivery of antibodies through the blood–brain barrier by MRI-guided focused ultrasound[J]. Biochemical and Biophysical Research Communications . 2006,340:1085–90
[28] Hynynen K, McDannold N, Vykhodtseva N, et al.Noninvasive MR imaging-guided focal opening of the blood-brain barrier in rabbits[J]. Radiology. 2001,220:640–46
[29] Chen WS,Matula TJ,Brayman AA,et al.A comparison of the fragmentation thresholds and inertial cavitation doses of different ultrasound contrast agents[J].J Acoust Soc Am.2003,113:643-51
[30] Yang FY,Fu WM,Yang RS, et al. Quantitative Evaluation of Focused Ultrasound with a Contrast Agent on Blood-Brain Barrier Disruption[J]. Ultrasound in Medicine and Biology.2007,33:1421-7
[1] Murphy SB,Bowman WP,Abromowitch M,et al.Result of treatment of advanced-stage Burkitt'S lymophoma and B cell(Sig+) acute lymphocytic leukemia with high-dose fractionated cyclophosphamide and coordinated high-dose methotrexate and cytarabine.J Clin Oncol,1986,4(12):1 732～1 739.
[2] kantarjian HM,O'Brien S,Smith TL,et al.Result of treatment with hyper-CVAD, a dose-intensive regimen,in adult acute lymphocytic leukemia.J Clin Oncol,2000,18(3):547～561
[3] kantarjian H,Thomas D,O'Brien,et al.Long-term follow-up results of hyperfractionated cyclophosphamide,vincristine,doxorubincin,and dexamethasone (Hyper-CVAD), A does-intensive regimen,in adult acute lymphocytic leukemia. Cancer,2004,101(12):2 788～2 801.
[4] Yang JL , Cheng EH , Capizzi RL , et al . Effect s of uracil arabinoside on metabolism and cytotoxicity of cytosine arabinoside in L5178γmurine leukemia. J Clin Invest , 1985 , 75 : 141-146.
[5] Peters WG, Colly LP , Willemze R. High-dose cytosine arabinoside : pharmacological and clinical aspect s. Blut , 1988 , 56 :1211.
[6] Pui CH,Sandlund JT,Pei D,et al.Improved outcome for children with acute lymphoblastic leukemia;results of total Therapy Study XⅢB at St Jude Children’s Research Hospital.Blood,2004,104;2690-2696.
[7] Pui CH,Cheng C,Leung W,et al.Extended follow-up of long-term survivors of childhood acute lymphoblastic leukemia.N Engl J Med,2003,349;640-649.
[8] Pardridge WM. Blood–brain barrier genomics and the use of endogenous transporters to cause drug penetration into the brain[J]. Curr Opin Drug Discov Devel. 2003,6:683–91.
[9] Doolittle ND, Miner ME, Hall WA, et al. Safety and efficacy of a multicenter study using intraarterial chemotherapy in conjunction with osmotic opening of the blood–brain barrier for the treatment of patients with malignant brain tumors[J]. Cancer. 2000,88:637–47.
[10] Laske DW, Youle RJ, Oldfield EH. Tumor regression with regional distribution of the targeted toxin TF-CRM107 in patients with malignant brain tumors[J]. NatMed.1997, 3:1362–1368.
[11] Choi JJ, Pernot M, Small SA, et al. Noninvasive, transcranial and localized opening of the blood-brain barrier using focused ultrasound in mice[J]. Ultrasound Med Biol. 2007, 33:95–104.
[12] Kinoshita M, McDannold N, Jolesz FA, et al. Targeted delivery of antibodies through the blood-brain barrier by MRI-guided focused ultrasound[J]. Biochem Biophys Res Commun. 2006,340:1085–90.
[13] Unger EC,Porter T,Culp W.et a1.Therapeutic applications of lipid-coated mierobubbles. Adv Drug Deliv Rev, 2004,56(9):1291-314.
[14] Unger EC,Hersh E,Vannan M,et a1.Local drug and gene delivery through microbubble [J].Prog Cardiovasc Dis,2001,4445—4454.Crowder KC, Hughes M S, Hughes M S,et a1.Sonic activation of molecularly-targeted nanopartides accelerates transmemhrane lipid delivery to cancer cells through contact mediated mechanisms:implications for enhanced local drug dlivery. Ultrasound Med Biol,2005,31(12):1693—1700.
[15] Porder JB,Porder KN,Meltzer RS Ultrasound bioeffects[J],Echocardiography,1987,4:89—99.
[16] Hynynen K, McDannold N, Vykhodtseva N, et al. Noninvasive MR imaging-guided focal opening of the blood-brain barrier in rabbits[J]. Radiology. 2001,220:640–6
[17] McDannold N, Vykhodtseva N, Raymond S, et al. MRI-guided targeted blood-brain barrier disruption with focused ultrasound :histological findings in rabbits[J]. Ultrasound Med Biol. 2005,31:1527–37
[18] Hynynen K, McDannold N, Vykhodtseva N, et al. Focal disruption of the blood-brain barrier due to 260-kHz ultrasound burst: a method for molecular imaging and targeted drug delivery[J]. J Neurosurg 2006,105:445–54
[1]. Gramiak R, Shah PM.Echocardiography of the aortic root.Invest Radiol, 1968, 3(5): 356-366
[2]. Miller M W, Miller D L, Brayman A A, et a1.A review of in vitro bioeffects. Ultrasounic cavitation from a mechanistic perspective[J]. Ultrasound Med Biol, 1996, 22(9):1131-1154.
[3].冉海涛,任红,王志刚,等.超声波空化效应对体外培养细胞细胞膜作用的实验研究[J].中华超声影像学杂志,2003,12(8):499-501.
[4]. WANG X, LIANG H D, DONG B, et a1. Gene transfer with microbubble ultrasound and plasmid DNA into skeletal nluscle of mice: comparison between commercially available microbubble contrast agents[J]. Radiology, 2005, 237(1):224-229.
[5]. KOCH S, POHL P, COBET U, et a1. Ultrasound enhancement of liposome: mediated cell transfection is caused by cavitation effects[J]. Ultrasound Med Biol, 2000, 26(5): 897-903.
[6]. Dayton PA, Ferrara KW. Targeted imaging using ultrasound[J]. Magn Reson Imagingt 2002, 16(4):362-377.
[7]. Youk JH, Lee JM,Kim CS. Therapeutic response evaluation of malignant hepatic masses treated by interventional procedures with contrast-enhanced agent detection imaging[J]. Ultrasound Med, 2003, 22(9):911-920
[8].徐亚丽,高云华.超声介导造影微泡在验证靶向方面的研究进展.中华超声影像学杂志,2005,14(4):310-312.
[9]. HUANG X, MOLEMA G, KING S, et a1. Tumor infarction in mice by antibody directed targeting of tissue factor to tumor vascu-lature[J]. Science, 1997,275(5299): 547-550.
[10]. UNGER EC, MAT SUNAGA TO, MCCREERY T, et a1. Therapeulie applications of microbubbles[J]. Eur J Radiol, 2002, 42(1): 160-168.
[11]. EMOTO M, TACHIBANA K, IWASAKI H, et a1. Antitumor effect of TNP470, an angiogenesis inhibitor, combined with ultrasound irradiation for human uterine sarcoma xenografts evaluated using contrast color Doppler ultrasound[J]. Cancer Sci, 2007, 98(6):929. 35.
[12]. Unger EC, Hersh E, Vannan M, et al.Local drug and gene delivery through microbubbles.Prog Cardiovase Diseases, 2001, 44(1):45-54.
[13]. HOWARD C M, FORSBERG F, MINIMO C, et a1. Ultrasound guided site specific gene delivery system using adenoviral vectors and commercial ultrasound contrast agents[J]. J Cell Physiol, 2006, 209(2): 4l3421.
[14].吴巍,宁新宝,姜藻,等.低频超声辐射Levovist试剂致家兔肝脏微血管栓塞的研究.东南大学学报(自然科学版).2003,33(3):300-302.
[15]. Frenkel PA, Chen S, Thai T , et a1. DNA-loaded albumin microbubbles enhance ultrasound-mediated transfection in vitro. Ultr asound Med Biol, 2002, 28(6):817-822.
[16]. Chen S, Shohet RV, Bekeredjian R, et al. Optimization of ultrasound parameters for cardiac gene delivery of adenoviral or plasmid deoxyribonucleic acid by ultrasound-targeted microbubble destruction.Am Coll Cardiol, 2003,42(2):301-308.
[17]. ter Haar GR, Ultrasonic contrast agents: safety considerations reviewed. Eur J Radiol.2002,41(3):217-221
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[7] Rubin LL, Staddon JM. The cell biology of the blood-brain barrier[J]. Annu Rev Neurosci. 1999,22:11–28。
[8] Vykhodtseva N,McDannold N, Hynynen K. Progress and problems in the application of focused ultrasound for blood–brain barrier disruption[J]. Ultrasonics. 2008,48:279-96
[9] Abbott NJ, Romero IA. Transporting therapeutics across the blood-brain barrier. Mol Med Today,1996, 2(3):106-113。
[10] Kroll RA, Neuwelt EA. Outwitting the blood-brain barrier for therapeutic purposes: osmotic opening and other means. Neurosurgery 1998,42(5):1083-1099.
[11] Laske DW,Youle RJ,Oldfield EH.Tumor regression with regional distribution of the targeted toxin TF-CRM107 in patients with malignant brain tumors.Nat Med,1997,3(12):1362-1368。
[12] Hynynen K, McDannold N, Vykhodtseva N, et al. Non-invasive opening of BBB byfocused ultrasound[J]. Acta Neurochir Suppl. 2003,86:555–8
[13]程远,于锐,宋或,等.低频超声联合微泡经颅开放血脑屏障初步研究.中国医学影像技术2006年第22卷第l期,42-44
[14] Yang JL , Cheng EH , Capizzi RL , et al . Effect s of uracil arabinoside on metabolism and cytotoxicity of cytosine arabinoside in L5178γmurine leukemia. J Clin Invest , 1985 , 75 : 141-146.
[15] Peters WG, Colly LP , Willemze R. High-dose cytosine arabinoside : pharmacological and clinical aspect s. Blut , 1988 , 56 :1211
[16]大剂量阿糖胞苷治疗时血浆和脑脊液中药物浓度测定的研究,谢晓恬、李本尚等,Shanghai Med J , 2005 ,Vol 28 ,No 3。
[17] Pardridge, WM. Drug and gene delivery to the brain: the vascular route[J]. Neuron. 2002, 36:555–8
[1] Ehrlich, P. Das Sauerstoffbedurfnis des Organismus[M]. Berlin: Eine farbenanalytische Studie. Hirschwald, 1885
[2] Pardridge WM. The blood-brain barrier: bottleneck in brain drug development[J]. NeuroRx. 2005,2:3–14
[3] Vykhodtseva N,McDannold N, Hynynen K. Progress and problems in the application of focused ultrasound for blood–brain barrier disruption[J]. Ultrasonics. 2008,48:279-96
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[6] Rubin LL, Staddon JM. The cell biology of the blood-brain barrier.Annu Rev Neurosci 1999;22:11–28
[7] Harkins CP, Mackenize F,Tofts P, et al. Patterns of blood-brain barrier breakdown in inflammatory demyelination.Brain, 1991,114:801
[8] Lynn JG, Putnam TJ. Histology of cerebral lesions produced by focused ultrasound[J]. Am J Pathol. 1944,20:637–52.
[9] Barnard JW, Fry WJ, Fry FJ, et al. Effects of high intensity ultrasound on the central nervous system of the cat[J]. J Comp Neurol. 1955,103:459–84
[10] Ballantine Jr HT, Bell E, Manlapaz J. Progress and problems in the neurological applications of focused ultrasound[J]. J Neurosurg. 1960,17:858–76
[11] Patrick JT, Nolting MN, Goss SA, et al. Ultrasound and the blood–brain barrier[J], Adv Exp Med Biol. 1990,267:369–381
[12] Vykhodtseva N, Hynynen K, Damianou C. Histologic effects of high intensity pulsed ultrasound exposure with subharmonic emission in rabbit brain in vivo[J]. Ultrasound Med Biol,1995,21:969-79
[13] Mychaskiw G,Badr AE,Tibbs R,et al.Optison(FS069)disrupts the blood—brain barrier in rats[J], Anesth Analg.2000,91:798-803
[14] Hynynen K, McDannold N, Vykhodtseva N, et al.Noninvasive MR imaging-guidedfocal opening of the blood-brain barrier in rabbits[J]. Radiology. 2001,220:640–46
[15] Vykhodtseva NI, McDannold N, Agabian S,et al. Histology findings after ultrasound exposure of the brain with ultrasound contrast agent—role of ischemia and apoptosis[A].Chapelon JY, Lafon C(eds): 3rd International Symposium on Therapeutic Ultrasound[C]. Lyon, France: INSERM,2003,80–85
[16] Hynynen K, McDannold N, Martin H, et al. The threshold for brain damage in rabbits induced by bursts of ultrasound in the pressence of an ultrasound contrast agent (option), Ultrasound in Med.&Biol, 2003,29(3): 473-481.
[17] Xiangtao Yin, Kullervo Hynynen. A numerical study of transcranial focused ultrasound bean propagation at low frequency. Phys. Med. Biol. 2005,50:1821-1836
[18] Hynynen K, McDannold N, Martin H, et al. The threshold for brain damage in rabbits induced by bursts of ultrasound in the pressence of an ultrasound contrast agent (option), Ultrasound in Med.&Biol, 2003,29(3): 473-481.
[19] Illum L . Nasal drug delivery--possibilities,problems and Solutions(J). J Control Release,2003,87(1-3):187—198
[20] Sheikov N, McDannold N, Vykhodtseva N, et al. Cellular mechanisms of the blood-brain barrier opening induced by ultrasoundin presence of microbubbles. Ulresound in Med.&Biol, 2004,30(7):979-989.
[21] Laske DW,Youle RJ,Oldfield EH.Tumor regression with regional distribution of the targeted toxin TF-CRM107 in patients with malignant brain tumors.Nat Med,1997,3(12):1362-1368.
[22] Weaver M, Laske DW. Transferrin receptor ligand-targeted toxin conjugate (Tf-CRM107) for therapy of malignant gliomas.J Neurooncol,2003,65(1):3-13.
[23] Haar,GT Therapeutic applications of ultrasound [J]Prog Biophys MolBiol,2007,93(1-3):111-129.
[24] Rouviere O,Lyonnet D,Raudrant A,et a1.MRI appearance of prostate following transrectal HIFU ablation of localized cancer[J].Eur Urol,2001,4O(3):265—274
[25]胡俊,丁仕义,黎海涛.血脑屏障开放的检测方法[J].重庆医学.2003,32(12): 1744-1746
[26] Carrera RM,Pacheco AM,Mastroti RA.Qualitative evaluation of the blood-brain barrier after the use of hypertonic saline solution in young rats[J].Eur Surg Res,2001,33:311-9
[27] Kinoshita M, McDannold N,Jolesz FA.et al.Targeted delivery of antibodies through the blood–brain barrier by MRI-guided focused ultrasound[J]. Biochemical and Biophysical Research Communications . 2006,340:1085–90
[28] Hynynen K, McDannold N, Vykhodtseva N, et al.Noninvasive MR imaging-guided focal opening of the blood-brain barrier in rabbits[J]. Radiology. 2001,220:640–46
[29] Chen WS,Matula TJ,Brayman AA,et al.A comparison of the fragmentation thresholds and inertial cavitation doses of different ultrasound contrast agents[J].J Acoust Soc Am.2003,113:643-51
[30] Yang FY,Fu WM,Yang RS, et al. Quantitative Evaluation of Focused Ultrasound with a Contrast Agent on Blood-Brain Barrier Disruption[J]. Ultrasound in Medicine and Biology.2007,33:1421-7
[1] Murphy SB,Bowman WP,Abromowitch M,et al.Result of treatment of advanced-stage Burkitt'S lymophoma and B cell(Sig+) acute lymphocytic leukemia with high-dose fractionated cyclophosphamide and coordinated high-dose methotrexate and cytarabine.J Clin Oncol,1986,4(12):1 732～1 739.
[2] kantarjian HM,O'Brien S,Smith TL,et al.Result of treatment with hyper-CVAD, a dose-intensive regimen,in adult acute lymphocytic leukemia.J Clin Oncol,2000,18(3):547～561
[3] kantarjian H,Thomas D,O'Brien,et al.Long-term follow-up results of hyperfractionated cyclophosphamide,vincristine,doxorubincin,and dexamethasone (Hyper-CVAD), A does-intensive regimen,in adult acute lymphocytic leukemia. Cancer,2004,101(12):2 788～2 801.
[4] Yang JL , Cheng EH , Capizzi RL , et al . Effect s of uracil arabinoside on metabolism and cytotoxicity of cytosine arabinoside in L5178γmurine leukemia. J Clin Invest , 1985 , 75 : 141-146.
[5] Peters WG, Colly LP , Willemze R. High-dose cytosine arabinoside : pharmacological and clinical aspect s. Blut , 1988 , 56 :1211.
[6] Pui CH,Sandlund JT,Pei D,et al.Improved outcome for children with acute lymphoblastic leukemia;results of total Therapy Study XⅢB at St Jude Children’s Research Hospital.Blood,2004,104;2690-2696.
[7] Pui CH,Cheng C,Leung W,et al.Extended follow-up of long-term survivors of childhood acute lymphoblastic leukemia.N Engl J Med,2003,349;640-649.
[8] Pardridge WM. Blood–brain barrier genomics and the use of endogenous transporters to cause drug penetration into the brain[J]. Curr Opin Drug Discov Devel. 2003,6:683–91.
[9] Doolittle ND, Miner ME, Hall WA, et al. Safety and efficacy of a multicenter study using intraarterial chemotherapy in conjunction with osmotic opening of the blood–brain barrier for the treatment of patients with malignant brain tumors[J]. Cancer. 2000,88:637–47.
[10] Laske DW, Youle RJ, Oldfield EH. Tumor regression with regional distribution of the targeted toxin TF-CRM107 in patients with malignant brain tumors[J]. NatMed.1997, 3:1362–1368.
[11] Choi JJ, Pernot M, Small SA, et al. Noninvasive, transcranial and localized opening of the blood-brain barrier using focused ultrasound in mice[J]. Ultrasound Med Biol. 2007, 33:95–104.
[12] Kinoshita M, McDannold N, Jolesz FA, et al. Targeted delivery of antibodies through the blood-brain barrier by MRI-guided focused ultrasound[J]. Biochem Biophys Res Commun. 2006,340:1085–90.
[13] Unger EC,Porter T,Culp W.et a1.Therapeutic applications of lipid-coated mierobubbles. Adv Drug Deliv Rev, 2004,56(9):1291-314.
[14] Unger EC,Hersh E,Vannan M,et a1.Local drug and gene delivery through microbubble [J].Prog Cardiovasc Dis,2001,4445—4454.Crowder KC, Hughes M S, Hughes M S,et a1.Sonic activation of molecularly-targeted nanopartides accelerates transmemhrane lipid delivery to cancer cells through contact mediated mechanisms:implications for enhanced local drug dlivery. Ultrasound Med Biol,2005,31(12):1693—1700.
[15] Porder JB,Porder KN,Meltzer RS Ultrasound bioeffects[J],Echocardiography,1987,4:89—99.
[16] Hynynen K, McDannold N, Vykhodtseva N, et al. Noninvasive MR imaging-guided focal opening of the blood-brain barrier in rabbits[J]. Radiology. 2001,220:640–6
[17] McDannold N, Vykhodtseva N, Raymond S, et al. MRI-guided targeted blood-brain barrier disruption with focused ultrasound :histological findings in rabbits[J]. Ultrasound Med Biol. 2005,31:1527–37
[18] Hynynen K, McDannold N, Vykhodtseva N, et al. Focal disruption of the blood-brain barrier due to 260-kHz ultrasound burst: a method for molecular imaging and targeted drug delivery[J]. J Neurosurg 2006,105:445–54
[1]. Gramiak R, Shah PM.Echocardiography of the aortic root.Invest Radiol, 1968, 3(5): 356-366
[2]. Miller M W, Miller D L, Brayman A A, et a1.A review of in vitro bioeffects. Ultrasounic cavitation from a mechanistic perspective[J]. Ultrasound Med Biol, 1996, 22(9):1131-1154.
[3].冉海涛,任红,王志刚,等.超声波空化效应对体外培养细胞细胞膜作用的实验研究[J].中华超声影像学杂志,2003,12(8):499-501.
[4]. WANG X, LIANG H D, DONG B, et a1. Gene transfer with microbubble ultrasound and plasmid DNA into skeletal nluscle of mice: comparison between commercially available microbubble contrast agents[J]. Radiology, 2005, 237(1):224-229.
[5]. KOCH S, POHL P, COBET U, et a1. Ultrasound enhancement of liposome: mediated cell transfection is caused by cavitation effects[J]. Ultrasound Med Biol, 2000, 26(5): 897-903.
[6]. Dayton PA, Ferrara KW. Targeted imaging using ultrasound[J]. Magn Reson Imagingt 2002, 16(4):362-377.
[7]. Youk JH, Lee JM,Kim CS. Therapeutic response evaluation of malignant hepatic masses treated by interventional procedures with contrast-enhanced agent detection imaging[J]. Ultrasound Med, 2003, 22(9):911-920
[8].徐亚丽,高云华.超声介导造影微泡在验证靶向方面的研究进展.中华超声影像学杂志,2005,14(4):310-312.
[9]. HUANG X, MOLEMA G, KING S, et a1. Tumor infarction in mice by antibody directed targeting of tissue factor to tumor vascu-lature[J]. Science, 1997,275(5299): 547-550.
[10]. UNGER EC, MAT SUNAGA TO, MCCREERY T, et a1. Therapeulie applications of microbubbles[J]. Eur J Radiol, 2002, 42(1): 160-168.
[11]. EMOTO M, TACHIBANA K, IWASAKI H, et a1. Antitumor effect of TNP470, an angiogenesis inhibitor, combined with ultrasound irradiation for human uterine sarcoma xenografts evaluated using contrast color Doppler ultrasound[J]. Cancer Sci, 2007, 98(6):929. 35.
[12]. Unger EC, Hersh E, Vannan M, et al.Local drug and gene delivery through microbubbles.Prog Cardiovase Diseases, 2001, 44(1):45-54.
[13]. HOWARD C M, FORSBERG F, MINIMO C, et a1. Ultrasound guided site specific gene delivery system using adenoviral vectors and commercial ultrasound contrast agents[J]. J Cell Physiol, 2006, 209(2): 4l3421.
[14].吴巍,宁新宝,姜藻,等.低频超声辐射Levovist试剂致家兔肝脏微血管栓塞的研究.东南大学学报(自然科学版).2003,33(3):300-302.
[15]. Frenkel PA, Chen S, Thai T , et a1. DNA-loaded albumin microbubbles enhance ultrasound-mediated transfection in vitro. Ultr asound Med Biol, 2002, 28(6):817-822.
[16]. Chen S, Shohet RV, Bekeredjian R, et al. Optimization of ultrasound parameters for cardiac gene delivery of adenoviral or plasmid deoxyribonucleic acid by ultrasound-targeted microbubble destruction.Am Coll Cardiol, 2003,42(2):301-308.
[17]. ter Haar GR, Ultrasonic contrast agents: safety considerations reviewed. Eur J Radiol.2002,41(3):217-221