癌细胞膜靶向相变纳米分子探针研制及光声/超声双模态显像与光热治疗乳腺癌

目的 制备一种癌细胞膜包被的相变纳米分子探针,观察其体外光声/超声双模态成像效果,并探讨其靶向同源乳腺癌细胞的能力及光热治疗(PTT)杀伤效果.材料与方法通过化学裂解和反复冻融提取乳腺癌4T1细胞膜(CCM),然后以双乳化法联合膜挤压法制备CCM包被的同时携载全氟戊烷(PFP)和Fe3O4的聚乳酸-羟基乙酸共聚物(PL...     

携载Ce6核壳纳米粒的双模态成像及光热治疗研究

【摘要】目的：构建介孔二氧化硅（MSN）-聚多巴胺（PDA）核壳纳米粒子（MSN-Ce6@PDA-Gd nanoparticles,MSN-Ce6@PDA-Gd NPs）,探讨其应用于体内外磁共振成像及荧光成像的潜能和光热治疗效用。方法：采用十六烷基三甲基氯化铵（CTAC）模板法、甲苯回流法制备氨基化介孔二氧化硅（MSN-NH2）纳米粒子,并通过静电作用包裹二氢卟吩e6（Ce6）,最后采用溶液氧化法进行PDA涂覆及钆离子（Gd3+）配位后,制备获得MSN-Ce6@PDA-Gd NPs。采用透射电子显微镜（TEM）、Zeta电位、紫外光谱等方法对MSN-Ce6@PDA-Gd NPs进行表征。采用红外热成像仪监测光热升温效果。采用细胞增殖及细胞毒性检测（MTT）法分析此纳米材料对小鼠胚胎成纤维细胞NIH-3T3和人乳腺癌细胞MDA-MB-231的细胞毒性及光热杀伤231细胞的效果。采用3.0T磁共振扫描仪观察此纳米诊疗剂的体内代谢途径及治疗前、后肿瘤大小,采用倒置荧光显微镜及荧光成像系统观察其体内外荧光成像效果。采用单因素方差分析进行数据的统计学分析。结果：MSN-Ce6@PDA-Gd NPs水合粒径为（142.10±0.29）nm,电位（-16.03±0.12）mV。Ce6的最高包封率为53.58%,负载量为17.65%。在不同浓度MSN-Ce6@PDA-Gd NPs溶液的作用下NIH-3T3和MDA-MB-231细胞存活率的差异均无统计学意义（F=2.317,P>0.05;F=2.344,P>0.05）。体外成像结果显示MDA-MB-231细胞内磁共振T1信号及荧光成像信号的增强具有浓度依赖性。体外光热治疗结果显示,随着MSN-Ce6@PDA-Gd NPs浓度的递增（0.0、12.5、25.0、50.0、100.0、150.0和200.0mg/L）,激光辐照组的MDA-MB-231细胞的存活率递减;无激光辐照组在纳米粒子浓度为0.0~200.0mg/L以及激光辐照组在纳米粒子浓度为0~50.0mg/L时均表现出较高的细胞存活率,当纳米粒子的浓度为100.0~200.0mg/L时,激光辐照组的细胞存活率显著降低。在体实验结果表明MSN-Ce6@PDA-Gd NPs可以实现对乳腺癌荷瘤裸鼠的的双模态成像以及对乳腺癌肿瘤有效的治疗作用。结论：制备了一种新型诊疗材料MSN-Ce6@PDA-Gd NPs,可成功实现对乳腺癌的体内外磁共振/荧光双模态显像和光热治疗。     

载IR-820相变型纳米粒双模态成像及其对类风湿关节炎成纤维样滑膜细胞光动力效应的体外研究

目的 探讨载IR-820及全氟正戊烷(PFP)的相变型多功能分子影像探针(IR-820/PFP@PLGA)在体外的超声/光声双模态显像特性及其对类风湿关节炎成纤维样滑膜细胞的光动力效应.方法 采用双乳化法制备IR-820/PFP@PLGA纳米粒,检测其基本表征、热致相变及光热效应,观察该纳米粒在体外的超声/光声双模态成...     

聚乳酸羟基乙酸CXCR4-miRNA纳米微粒抑制人肾癌细胞生长的研究

目的探讨聚乳酸羟基乙酸CXCR4-miRNA纳米复合微粒在体外对人肾癌细胞增殖的抑制作用。方法运用二次超声乳化和溶剂挥发法制备聚乳酸羟基乙酸CXCR4-miRNA纳米粒;通过MTT法观察纳米微粒对肾癌细胞A498增殖的抑制作用;并运用流式细胞术检测纳米微粒作用后A498细胞的凋亡情况。结果制备的聚乳酸羟基乙酸CXCR4-miRNA纳米粒外观呈圆型,平均粒径为280 nm,平均载药量为(0.515±0.023)%,平均包封率为50.2%。通过MTT实验证明纳米微粒可以较好的抑制人肾癌细胞(A498)的增殖,随着聚乳酸羟基乙酸CXCR4-miRNA纳米微球浓度的增加,药物作用时间越长,细胞增殖受到抑制的效果越明显。流式细胞术检测可见凋亡峰出现,细胞周期阻滞在S+G2/M期。结论聚乳酸羟基乙酸CXCR4-miRNA纳米粒具有抑制肾癌细胞A498增殖的作用,其效果与药物浓度及作用时间长短有关。     

卡铂-乳酸/羟基乙酸共聚物微球的制备和体外性质

目的制备卡铂-乳酸/羟基乙酸共聚物(PLGA)微球,比较不同方法所得微球的形态、载药量和体外释药特点。方法采用相分离法和溶剂挥发法制备卡铂-PLGA微球,显微镜下测定微球的粒径和粒径分布,电子扫描显微镜观察微球表面形态。用电感耦合等离子体发射光谱法(ICP-AES)测定微球含药量,计算包封率,考察微球体外释药行为。结果两种方法所得微球球形较好,相分离法制得的卡铂-PLGA微球,平均粒径为22~31μm,含药量为42~61μg·mg-1、包封率21%~31%;体外释放试验中药物于24h完全溶出。溶剂挥发法所得微球平均粒径为38~54μm,含药量为7.2μg·mg-1、包封率约为20%;体外药物突释率约为39%,缓释期药物释放符合Higuchi模型,PLGA75/25、η=0.19和PLGA50/50、η=0.18的微球药物释放速度常数分别为2.40h-1/2和0.85h-1/2;体外14d累计释药分别达到71%和54%。结论相分离法制备卡铂-PLGA微球含药量高,但体外释药快,没有缓释作用;溶剂挥发法所得微球药物突释率较低,体外能控制药物缓慢释放。     

磁性纳米氧化铁粒子在肿瘤影像及治疗中的应用及进展

磁性纳米氧化铁粒子是指直径在纳米范围的氧化铁粒子,具有化学性质稳定、血液滞留时间长、低毒性、顺磁性、生物可降解性等特点,可通过静电作用或化学作用耦联多肽、单克隆抗体、化疗药物、基因片段等靶肿瘤功能分子,因而在肿瘤影像、治疗、研究中有着广泛的应用.该文对近年来磁性纳米氧化铁粒子在肿瘤影像及治疗中的应用及进展进行了评述.     

新型MR对比剂长循环超顺磁氧化铁脂质体纳米微粒的体内外实验研究

目的 评价自制新型MR对比剂:长循环超顺磁性氧化铁(superparamagnetic iron oxide,SPIO)脂质体纳米微粒的药代动力学.方法 以加入SPIO为对照组,加入长循环SPIO脂质体纳米微粒为实验组.(1)巨噬细胞的体外吞噬实验:将巨噬细胞株RAW 264.7经复苏、培养,细胞达到80%～90%融合后,以每孔2.5×105个细胞接种于培养板,对照组和实验组细胞与不同浓度药物孵化后,分别测定细胞中Fe浓度和Fe染色情况,并用析因设计的方差分析对结果进行统计学处理.(2)小鼠体内药物分布实验:C57BL/6J雄性小鼠6只,每组2只,设为空白对照组、对照组、实验组,经尾静脉分别注射生理盐水、SPIO以及长循环SPIO脂质体后牺牲动物,经病理检查观察2组药物在体内的分布情况.(3)荷肺癌裸鼠MR成像实验:建立20只BALB/c裸鼠移植型肺癌模型,分为对照组和实验组,每组10只,分别经尾静脉注入SPIO和长循环SPIO脂质体纳米微粒后行MR扫描,测量增强前后2组实验动物各时间点肝脏和肿瘤ROI的信号强度,绘制信噪比(SNR)时间动态变化曲线,采用协方差分析比较2组动物肝脏及肿瘤增强12 h 后SNR差异有无统计学意义.(4)细胞毒性实验(噻唑蓝,MTT法):检查2种药物对人肝细胞株HL-7702细胞的毒性作用,用析因设计的方差分析对结果进行统计学处理.结果 (1)巨噬细胞的体外实验:析因分析结果表明,对照组细胞内Fe含量明显高于实验组,差异有统计学意义(F=296 839.9,P=0.000).普鲁士蓝染色显示对照组巨噬细胞染色较深,实验组巨噬细胞染色较浅.(2)小鼠体内药物分布实验:对照组肝、脾、肺、肾组织内蓝染颗粒多于实验组.(3)荷肺癌裸鼠MR成像实验:平扫对照组肝脏SNR为31.47±0.56,实验组为30.89±1.41;增强扫描12 h后对照组为17.00±0.96,实验组为22.29±0.73.平扫对照组肿瘤SNR为58.41±0.61,实验组为58.44±1.08;增强扫描12 h后对照组为58.50±0.63,实验组为52.47±1.18.经协方差分析表明增强12 h后对照组肝脏SNR显著低于实验组(F=167.022,P=0.000);而实验组肿瘤 SNR显著低于对照组(F=266.106,P=0.000).(4)细胞毒性实验:随着培养液中Fe浓度的增加,HL-7702细胞生存率均有下降趋势,但析因分析结果表明,实验组药物的细胞毒性较对照组低(F=2256.204,P=0.000).结论 长循环SPIO脂质体纳米微粒在体内外均有较好的抗巨噬系统吞噬功能,在体内实现了长循环,对非网状内皮系统的肿瘤也具有良好的T2负性被动靶向强化效果;同时还降低了SPIO对细胞的毒性作用.

Abstract：

Objective To evaluate pharmacodynamics of prepared long-circulating superparamagnetic iron oxide (SPIO) liposomes. Methods Control and experimental groups were established after adding SPIO or long-circulating SPIO liposomes as agents. (1)Macrophages experiment in vitro: the RAW 264.7 macrophage cell strains were recovered, cultured and seeded in the culture plate at a density of 2.5×105 cells/well until they reached 80%-90% confluence. The intracellular Fe uptake of control and experimental group cells were quantified by Ferrozine assay after incubation with different concentrations of drugs. Factorial design analysis of variance was used as statistics method. Prussian blue staining method was used to detect staining of experimental cells.(2)Drug biodistribution in mice: C57BL/6J(n=6) were classified into blank control group (n=2), control group(n=2) and experimental group(n=2).Saline, SPIO and long circulating SPIO were injected via the tail vein in the blank control group, control group and experimental group respectively. Then distribution of drugs in the body was observed by pathological examination.(3) MR imaging of tumor-bearing nude mice: 20 BALB/c nude mice bearing lung cancer models were established and classified into control group and experimental group. After administration of drugs, all animals underwent MR scanning. Signal intensities of livers and tumors were measured, SNR-time dynamic curves were drew. Covariance analysis was used to compare post-enhanced SNR at the 12th hour. (4)Cytotoxicity studies (MTT): cytotoxicity of both drugs on human liver cell line HL-7702 was studied, and statistically analyzed using factorial design analysis of variance. Results (1) Macrophages experiment in vitro: The nanoparticle uptake by macrophage cells evaluated by ferrozine assay showed the uptake of blank SPIO was higher than long-circulating SPIO liposomes. Compared with the blank control group, there was strong blue staining in the macrophages with Prussian staining in the control group and little blue staining in the experimental group. (2) Drug biodistribution in mice: for blue stained cells composed of iron particles, the amounts in the liver, spleen, lung, kidney of the control group were more than those in the experimental group. (3) MR imaging of tumor-bearing nude mice: the non-enhanced SNR of livers and tumors in the control group and experimental group were 31.47 ± 0.56, 30.89 ± 1.41, 58.41 ± 0.61, 58.44 ± 1.08, respectively. After injecting of contrast agents, SNR of livers and tumors in the control group and experimental group were 17.00 ± 0.96, 22.29 ± 0.73, 58.50 ± 0.63, 52.47 ± 1.18, respectively. The covariance analysis showed that SNR of the livers in the control group after 12 hours was significantly lower than the experimental group (F=167.022, P=0.000); while the SNR of the tumors in the experimental group was significantly lower than the control group (F=266.106, P=0.000).(4) Cytotoxicity of nanoparticles by MTT method: the viability of HL-7702 cells tend to decrease with the increase of Fe concentration. Cytotoxicity in the long-circulating SPIO liposomes was lower than the SPIO(F=2256.204,P=0.000). Conclusions Long-circulating SPIO liposomes we prepared reveal suitable sizes, even distribution, and good anti-macrophage ability in vitro and in vivo. They have long circulation characteristic and T2 negative enhancement effect in the transplanted lung cancers, while they still maintain low cytotoxicity.     

Application of a superparamagnetic iron oxide (Resovis®) for MR imaging of human cerebral blood volume

The authors describe the feasibility of dynamic MRI using a novel superparamagnetic iron oxide contrast agent. Resovist® was injected as a bolus at doses of 4, 8, and 16 μmol Fe/kg bodyweight in three consented patients participating in a Phase 2 clinical multicenter trial for hepatic MRI. Dynamic images of the brain were obtained with a conventional FLASH sequence. Results were analyzed by evaluation of dynamic images, cerebral blood volume maps, and normalized signal intensity time curves. Resovist® enabled rapid injections and a dose-dependent strong reduction in gray and white matter signal intensity. The small injection volume and good tolerability may enable Resovist® to serve as a perfusion agent. Dedicated clinical trials are warranted to assess the potentials of Resovist® for perfusion MRI and fMRI.     

Targeting chemo-proton therapy on C6 cell line using superparamagnetic iron oxide nanoparticles conjugated with folate and paclitaxel

Purpose: This report presents novel nanoparticle-based drug delivery system (NPDDS) aiming to targeting chemo-proton therapy (TCPT) to improve the therapeutic efficacy on brain cancer treatments.

Materials and methods: A NPDDS, superparamagnetic iron oxide nanoparticles conjugated with folate and paclitaxel, was synthesized and applied to C6 brain cancer cell line that was prepared for TCPT. The characterization of NPDDS was analyzed by transmission electron microscope (TEM) and Fourier transform infrared (FTIR) spectroscopy. The uptake of NPDDS into the cytoplasm of C6 cells was observed by confocal laser scanning microscopy (CLSM). The therapeutic efficacy of proton beam was quantitatively evaluated by flow cytometry and clonogenic assay at various radiation dose.

Results: NPDDS was synthesized in the uniform size distribution with a mean diameter of 5.44?±?0.70?nm, and it showed no significant cytotoxicity at the concentration lower than 200?ng/mL. Radiosensitization enhancement factors of PTX, D-SPIONs and FA-PTX-D-SPIONs were 1.35, 1.16 and 1.52, respectively.

Conclusions: It was demonstrated that TCPT improved the therapeutic efficacy of the proton beam therapy when the synthesized novel NPDDS was administrated. The improvement in therapeutic efficacy was achieved by the synergetic effect of drug delivery increased by FA, radiosensitivity increased by PTX and absorption of proton energy increased by SPIONs.     

Comparison study on the effect of gold nanoparticles shape in the forms of star,hallow, cage,rods, and Si-Au and Fe-Au core-shell on photothermal cancer treatment

Gold nanoparticles (GNPs) indicate potential in the development of cancer treatments as vehicles for thermal damage of cancer cells because of their photothermal heating capability. Herein, we aim to investigate the effect of GNPs geometry as photothermal transducers on cellular uptake and photothermal therapy (PTT) efficacy.For this aim, seven different shapes of anisotropic GNPs: stars, hollow, rods, cages, spheres, Fe-Au, and Si-Au core shells were synthesized and investigate the effect of shape on GNPs optical properties. The physic-chemical characterization of prepared GNPs was investigated by UV–vis, DLS-Zeta, and TEM analysis. The effect of GNPs geometry on cellular uptake was investigated by ICP-MS and flow cytometry method. The PTT potential of these GNPs was compared on MCF7 cells in vitro using MTT assay, cell cycle, and Annexin-V apoptosis assay.While all these GNPs could absorb and convert near-infrared light into heat, gold nanostars exhibited the lowest cytotoxicity, highest cellular uptake and highest heat generation compared to other structures. Following photothermal treatment, due to substantial heat production in MCF7 cells, the apoptosis induction rate was greatly increased for all anisotropic gold nanostructures (stars, hollow, rods, and cages) especially gold nanostars.Combined, we can conclude that GNPs geometry affects cellular uptake and heat generation amount as well as cell destruction by apoptosis pathway. The gold nanostar is promising candidates for photothermal destruction.     

对比剂铁羧葡胺在正常兔MR脑灌注的初步实验研究

目的 探讨铁羧葡胺(Resovist)应用于脑MR灌注加权成像(PWI)的可行性、给药方法以及最佳剂量.方法 健康新西兰大白兔30只,随机数字法平均分为A、B、C、D、E组.其中A、B、C、D 4组分别给予4、8、16、32 μmol Fe/kg;E组设为对照,给予0.2 mmol/kg的钆喷替酸葡甲胺(Gd-DTPA).所有动物均行MR PWI,获得相应信号强度一时间曲线图,并分别计算脑灰质、白质的最大信号下降百分比(SRRmax)、局部脑血容量(rCBV)和灰质与白质rCBV之比(QRCBV)、SRRmax之比(QRR max).所得数据,根据资料性质,行配对t检验和单因素方差分析.结果 Resovist能快速团注,4组均获得满意的信号强度一时间曲线图;4种剂量的Resovist对实验兔脑灰质和白质均有良好的分辨率.A、B、C、D、E 5组脑灰质和白质的rCBV分别为(50.48±3.84)、(25.57±2.10),(94.69±2.60)、(45.33±3.14),(141.13±6.26)、(67.67±4.65),(243.75±5.90)、(162.06±5.14),(84.60±3.60)、(41.36±2.18)ml/100 g;灰质和白质的SRRmax分别为:(13.70±1.50)%、(7.38±0.41)%,(31.01±4.06)%、(16.49±2.35)%,(43.81±3.42)%、(21.64±4.14)%,(64.49±5.35)%、(43.61±5.78)%,(27.78±2.98)%、(14.42±2.25)%;各组脑灰质与白质检测数据比较差异均有统计学意义(P值均<0.01).A、B、C、D、E 5组QrCBV分别为1.98±0.07、2.09±0.11、2.09±0.07、1.50±0.01、2.05±0.03;QSRRmax分别为:1.85±0.11、1.88±0.06、2.06±0.25、1.49±0.09、1.94±0.12;5组间差异均有统计学意义(QrCBV的F值为85.076,QSRR max的F值为13.915,P均<0.01).A、B、C 3组QrCBV值和QSRR max值差异无统计学意义(P>0.05);而D组QrCBV值和QSRR max值显著低于A组(P<0.01).结论 Resovist应用于MR脑灌注是可行的,适宜剂量4～16 μmol Fe/kg.     

SPIO粒子增强MRI检测腋窝转移性淋巴结的实验研究

目的:探讨超顺磁性氧化铁粒子(SPIO)增强MRI检测腋窝转移性淋巴结的价值.方法:选取健康成年新西兰白兔14只,采用随机数字法将其分为反应性增生淋巴结组(8只)及肿瘤转移淋巴结组(6只).MRI平扫后,3只反应性增生淋巴结皮下间隙注射SPIO(10 μmol Fe/单侧乳腺)后第1、3、6、12、24和48 h点扫描...