In vivo MR imaging of intravascularly injected magnetically labeled mesenchymal stem cells in rat kidney and liver

PURPOSE: To evaluate in vivo magnetic resonance (MR) imaging with a conventional 1.5-T system for depiction and tracking of intravascularly injected superparamagnetic iron oxide (SPIO)-labeled mesenchymal stem cells (MSCs). MATERIALS AND METHODS: This study was conducted in accordance with French law governing animal research and met guidelines for animal care and use. Rat MSCs were labeled with SPIO and transfection agent. Relaxation rates at 1.5 T, cell viability, proliferation, differentiation capacity, and labeling stability were assessed in vitro as a function of SPIO concentration. MSCs were injected into renal arteries of healthy rats (labeled cells in four, unlabeled cells in two) and portal veins of rats treated with carbon tetrachloride to induce centrolobular liver necrosis (labeled cells and unlabeled cells in two each). Follow-up serial T2*-weighted gradient-echo MR imaging and R2* mapping were performed. MR imaging findings were compared histologically. RESULTS: SPIO labeling caused a strong R2* effect that increased linearly with iron dose; R2* increase for cells labeled for 48 hours with 50 microg of iron per milliliter was 50 sec(-1) per million cells per milliliter. R2* was proportional to iron load of cells. SPIO labeling did not affect cell viability (P > .27). Labeled cells were able to differentiate into adipocytes and osteocytes. Proliferation was substantially limited for MSCs labeled with 100 microg Fe/mL or greater. Label half-life was longer than 11 days. In normal kidneys, labeled MSCs caused signal intensity loss in renal cortex. After labeled MSC injection, diseased liver had diffuse granular appearance. Cells were detected for up to 7 days in kidney and 12 days in liver. Signal intensity loss and fading over time were confirmed with serial R2* mapping. At histologic analysis, signal intensity loss correlated with iron-loaded cells, primarily in renal glomeruli and hepatic sinusoids; immunohistochemical analysis results confirmed these cells were MSCs. CONCLUSION: MR imaging can aid in monitoring of intravascularly administered SPIO-labeled MSCs in vivo in kidney and liver.     

MR evaluation of the glomerular homing of magnetically labeled mesenchymal stem cells in a rat model of nephropathy

PURPOSE: To assess renal glomerular homing of intravenously injected superparamagnetic iron oxide (SPIO)-labeled mesenchymal stem cells (MSCs) at in vivo and ex vivo magnetic resonance (MR) imaging in an experimental rat model of mesangiolysis. MATERIALS AND METHODS: Animal procedures were performed in accordance with protocols approved by Institutional Animal Care and Use Committee. Fourteen rats were divided into two groups: one pathologic (n = 10), with persistent mesangiolysis following simultaneous injection of OX-7 monoclonal antibody and puromycin aminonucleoside in which 10(7) SPIO- and DiI-labeled MSCs were injected, and one control (n = 4). In vivo and ex vivo MR imaging examinations were performed with 4.7- and 9.4-T spectrometers, respectively, and T2*-weighted sequences. In vivo signal intensity variations were measured in the liver and kidney before and 6 days after MSC injection. Intrarenal signal intensity variations were correlated with histopathologic data by means of colocalization of DiI fluorescence, alpha-actin, and Prussian blue stain-positive cells. Histologic differences between the glomerular homing of MSCs in different kidney portions were correlated to the areas of MR signal intensity decrease with nonparametric statistical tests. RESULTS: On in vivo images, signal intensity measurements of pathologic kidneys following MSC injection did not show any signal intensity decrease (P = .7), whereas a 34% +/- 14 (mean +/- standard deviation) signal intensity decrease was observed in the liver (P < .01), where a substantial number of labeled cells were trapped. On ex vivo images, pathologic kidneys showed focal cortical (glomerular) areas of signal intensity loss, which was absent in controls. The areas of low signal intensity correlated well with alpha-actin and Prussian blue stain- and DiI-positive areas (P < .01), which indicates that MSCs specifically home to injured tissue. No MSCs were detected in the kidneys of control animals. CONCLUSION: Intravenously injected MSCs specifically home to focal areas of glomerular damage and can be detected at ex vivo MR imaging.     

R2 and R2* mapping for sensing cell-bound superparamagnetic nanoparticles: in vitro and murine in vivo testing

PURPOSE: To prospectively determine the cellular iron uptake by using R2 and R2* mapping with multiecho readout gradient-echo and spin-echo sequences. MATERIALS AND METHODS: All experiments were approved by the institutional animal care committee. Lung carcinoma cells were lipofected with superparamagnetic iron oxides (SPIOs). Agarose gel phantoms containing (a) 1 x 10(5) CCL-185 cells per milliliter of agarose gel with increasing SPIO load (0.01-5.00 mg of iron per milliliter in the medium), (b) different amounts (5.0 x 10(3) to 2.5 x 10(5) cells per milliliter of agarose gel) of identically loaded cells, and (c) free (non-cell-bound) SPIOs at the iron concentrations described for (b) were analyzed with 3.0-T R2 and R2* relaxometry. Iron uptake was analyzed with light microscopy, quantified with atomic emission spectroscopy (AES), and compared with MR data. For in vivo relaxometry, agarose gel pellets containing SPIO-labeled cells, free SPIO, unlabeled control cells, and pure agarose gel were injected into three nude mice each. Linear and nonlinear regression analyses were performed. RESULTS: Light microscopy and AES revealed efficient SPIO particle uptake (mean uptake: 0.22 pg of iron per cell +/- 0.1 [standard deviation] for unlabeled cells, 31.17 pg of iron per cell +/- 4.63 for cells incubated with 0.5 mg/mL iron). R2 and R2* values were linearly correlated with cellular iron load, number of iron-loaded cells, and content of freely dissolved iron (r(2) range, 0.92-0.99; P < .001). For cell-bound SPIO, R2* effects were significantly greater than R2 effects (P < .01); for free SPIO, R2 and R2* effects were similar. In vivo relaxometry enabled accurate prediction of the number of labeled cells. R2' (R2* - R2) mapping enabled differentiation between cell-bound and free iron in vitro and in vivo. CONCLUSION: Quantitative R2 and R2* mapping enables noninvasive estimations of cellular iron load and number of iron-labeled cells. Cell-bound SPIOs can be differentiated from free SPIOs with R2' imaging.     

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.     

MR Imaging Enables Measurement of Therapeutic Nanoparticle Uptake in Rat N1-S1 Liver Tumors after Nanoablation

PurposeTo test the hypothesis that magnetic resonance (MR) imaging can quantify intratumoral superparamagnetic iron oxide (SPIO) nanoparticle uptake after nanoablation.Materials and MethodsSPIO nanoparticles functionalized with doxorubicin were synthesized. N1-S1 hepatomas were successfully induced in 17 Sprague-Dawley rats distributed into three dosage groups. Baseline tumor R2* values (the reciprocal of T2*) were determined using 7-tesla (T) MR imaging. After intravenous injection of SPIO nanoparticles, reversible electroporation (1,300 V/cm, 8 pulses, 100-μs pulse duration) was applied. Imaging of rats was performed to determine tumor R2* values after the procedure, and change in R2* (ΔR2*) was calculated. Inductively coupled plasma mass spectrometry was used to determine intratumoral iron (Fe) concentration after the procedure, which served as a proxy for SPIO nanoparticle uptake. Mean tumor Fe concentration [Fe] and ΔR2* for each subject were assessed for correlation with linear regression, and mean [Fe] for each dosage group was compared with analysis of variance.ResultsΔR2* significantly correlated with tumor SPIO nanoparticle uptake after nanoablation (r = 0.50, P = .039). On average, each 0.1-ms⁻¹ increase in R2* corresponded to a 0.1394-mM increase in [Fe]. There was no significant difference in mean SPIO nanoparticle uptake among dosage groups (P = .57).ConclusionsIntratumoral SPIO nanoparticle uptake after nanoablation can be successfully quantified noninvasively with 7-T MR imaging. Imaging can be used as a method to estimate localized drug delivery after nanoablation.     

3.0 T血氧水平依赖功能MRI技术对注入碘对比剂后肾脏损害评价的实验研究

目的 探讨3.0 T血氧水平依赖功能MRI(fMRI-BOLD)技术在碘对比剂大鼠肾脏损害评价中的应用价值.方法 29只SD大鼠分别于注入碘对比剂前、注入碘对比剂碘普胺后20 min、24 h、48 h、72 h不同时间点进行常规MR序列及BOLD序列扫描;注入碘对比剂前作为对照组.BOLD序列扫描图像在工作站处理后,测量大鼠肾脏皮质、外髓及内髓的T2*值,计算表观自旋-自旋弛豫率(R2*值)(R2*值=1/T2 *值).对照组大鼠双肾不同部位R2*值的比较采用方差分析,对照组和实验组不同时间、不同部位右肾和左肾R2*值的比较采用t检验.结果 对照组大鼠左、右肾脏外髓R2*值[分别为(31.76±2.73)/s和(32.77±3.07)/s]均高于相应皮质[分别为(30.20±3.48)/s和(28.84±3.11)/s]及内髓[分别为(29.54±2.42)/s和(28.37±2.80)/s],F值分别为3.357和14.961,P值均＜0.05.左肾和右肾内髓、外髓和皮质的R2*值差异均无统计学意义(P值均＞0.05).注入碘对比剂后不同时间点,以外髓R2*值变化明显.在注射对比剂20 min时,肾脏外髓R2*值最高[右肾和左肾分别为(44.58±3.13)/s和(43.57±3.84)/s],24 h后R2*值逐渐降低[右肾和左肾分别为(42.89±3.40)/s和(42.07±4.82)/s].内髓及皮质R2*值仅表现为轻微改变.结论 R2*值能反映出大鼠肾脏皮、髓质氧分压的变化,判断髓质缺血缺氧程度.BOLD成像技术能反映肾脏皮髓质氧代谢情况,评估肾脏损害程度,是一种简单、可行及重复性较好的方法.     

A molecular MRI probe to detect treatment of cardiac apoptosis in vivo

Cell death by apoptosis is critical in myocardial diseases, and noninvasive detection of early, reversible apoptosis might be useful clinically. Exogenous Annexin‐V (ANX) protein binds membrane phosphatidylserine, which is externalized in early apoptosis. A molecular MRI probe was constructed with superparamagnetic iron oxide (SPIO) conjugated to recombinant human ANX (ANX‐SPIO). Apoptosis was induced with doxorubicin, a cardiotoxic cancer drug, in culture in neonatal rat ventricular myocytes, cardiac fibroblasts, and mesenchymal stem cells, and in vivo in the mouse heart. ANX‐SPIO was validated using T2*‐weighted 3T MRI. ANX‐SPIO produced T2* signal loss, reflecting iron content, that correlated highly with independent apoptosis markers; bound with high affinity to apoptotic myocytes by competition assay (Ki 69 nM); detected apoptosis in culture much earlier than did TUNEL stain; and revealed fibroblast resistance to apoptosis. With apoptosis in vivo, ANX‐SPIO produced diffuse myocardial T2* signal loss that correlated with increased iron stain and caspase activity. Treatment with an alpha‐1‐adrenergic agonist in vivo reversed apoptosis and eliminated the ANX‐SPIO MRI signal. It is concluded that cardiac MRI of ANX‐SPIO detects early, nonischemic cardiac apoptosis in culture and in vivo, and can identify reversibly injured cardiac cells in diseased hearts, when treatment is still possible. Magn Reson Med, 2011. © 2011 Wiley‐Liss, Inc.     

分析1.5T与3.0T MR定量评价体外铁浓度模型比较研究

目的:探讨MRI对体外不同铁浓度模型定量测量的可靠性,并比较1.5T和3.0T MR对体外铁浓度模型定量测定的准确性.方法:制作2组不同铁浓度体外模型,一组铁浓度范围为0～5.0 mg/mL.另一组铁浓度范围为0～1.0 mg/mL.两组铁浓度模型分别在1.5T及3.0T MR上进行扫描.扫描序列包括短轴面FSE/T1WI、FRFSE/T2WI、T2 map和T2*map.评价测量指标包括T1WI和T2WI信噪比、T2、R2、T2*和R2*值.评价指标的一致性采用组内相关系数分析.铁浓度与T1WI信噪比、T2WI信噪比、T2、R2、T2*和R2*值的关系应用Pearson相关分析.结果:两位研究者对体外铁浓度模型MR图像测量指标T1WI信噪比、T2WI信噪比、T2、R2、T2*和R2*值一致性评价的组内相关系数均大于0.900(P＜0.001).T2WI信噪比、T2和T2*值与铁浓度在1.5T和3.0T MR上均呈非线性相关.在1.5T MR上,当铁浓度≤1.5 mg/mL时,T1WISNR、R2与铁浓度呈显著性正相关(P＜0.05).R2*则在铁浓度≤2.5 mg/mL时,与铁浓度呈显著正性线性相关(P＜0.001).而在3.0T MR上,当铁浓度≤0.90 mg/mL时,R2和R2*值与铁浓度呈正相关(P＜0.05).结论:MRI定量测定体外不同铁浓度模型的可重复性好;在1.5T或3.0T MR各测量参数中,R2*值可准确定量反映体外铁浓度,且1.5T MR检测铁浓度范围宽于3.0T MR.     

Magnetic resonance detection of kidney iron deposition in sickle cell disease: a marker of chronic hemolysis

PURPOSE: To study the pattern, etiology, and significance of renal iron accumulation in chronically transfused sickle cell disease (SCD) and thalassemia major (TM) patients using magnetic resonance imaging (MRI). MATERIALS AND METHODS: Magnetic resonance imaging (MRI) was performed in 75 SCD patients, 73 TM patients, and 16 healthy controls. Multiecho gradient echo protocols were used to measure T2* reciprocals (R2*) in the kidney, liver, and heart. Kidney R2* was compared to tissue iron estimates, serum iron markers, and surrogates of intravascular hemolysis by univariate regression. RESULTS: Mean R2* in SCD patients was 55.3+/-45.3 Hz, compared with 22.1+/-11 Hz in TM patients and 21.3+/-5.8 Hz in control subjects (P<0.001). Kidney R2* decreased with advancing age (R2=0.09, P<0.02). Kidney R2* correlated strongly with increased serum lactate dehydrogenase levels found in SCD (R2=0.55, P<0.001), but was independent of hepatic iron concentration and cardiac R2*. Kidney R2* did not correlate with blood pressure, creatinine, cardiac index, or right and left ejection fraction. CONCLUSION: Intravascular hemolysis, not chronic transfusion, causes renal hemosiderosis in SCD. Prospective trials are necessary to determine whether kidney R2* is a biomarker for hemolysis-mediated vascular complications in SCD.     

Superparamagnetic iron oxide-enhanced MRI of atherosclerotic plaques in Watanabe hereditable hyperlipidemic rabbits

RATIONALE AND OBJECTIVES: Inflammatory atherosclerotic plaques are characterized by increased endothelial permeability and multiple macrophages. Blood-pool MRI contrast agents like superparamagnetic iron oxide (SPIO) have an affinity for the monocyte-macrophage system and thus, may label inflammatory plaques. The objective was to demonstrate SPIO uptake in plaques of atherosclerotic rabbits by MRI and histology. METHODS: Aortas of anesthetized Watanabe hereditable hyperlipidemic rabbits were studied with a moderately T2*-weighted gradient-echo sequence at 1.5 T. Four groups of five animals each were studied: (1) without ultrasmall SPIO (carboxydextran coating; particle size, 25 nm; estimated plasma half-life, 6 hours) or with imaging after intravenous injection of SPIO at a dose (micromol Fe/kg) and postcontrast time delay (hours) of 50/8 (2), 50/24 (3), or 200/48 (4). In vivo MRI was compared with corresponding ex vivo histological iron stains. RESULTS: Animals receiving 200 micromol Fe/kg demonstrated areas of focal signal loss clearly confined to the aortic wall on a mean of 24 +/- 9 (31% +/- 11%) of 76 +/- 5 images compared with 0 +/- 0 of 76 +/- 5 images in controls (P = 0.009). The number of images with this finding in groups 2 and 3 was not significantly different compared with controls. By microscopy, SPIO-iron was seen in the endothelial cells and subendothelial intimal macrophages of atherosclerosis-prone aortic wall segments. Atherosclerotic lesions demonstrating iron uptake also showed a high macrophage content. CONCLUSIONS: SPIO accumulates in aortic plaques of atherosclerotic rabbits, producing a characteristic MRI finding. As SPIO accumulates in plaques with increased endothelial permeability and a high macrophage content, two established features of plaque inflammation, it may have a potential for noninvasive assessment of inflammatory atherosclerotic plaques.     

Atherosclerotic imaging using 4 types of superparamagnetic iron oxides: New possibilities for mannan-coated particles

Purpose

We used magnetic resonance imaging (MRI) and histologic techniques to compare the uptake by the rabbit atherosclerotic wall of 4 types of superparamagnetic iron oxide (SPIO) particles, i.e. SPIO, mannan-coated SPIO (M-SPIO), ultrasmall SPIO (USPIO), and mannan-coated USPIO (M-USPIO).

Materials and methods

All experimental protocols were approved by our institutional animal experimentation committee. We intravenously injected 12 Watanabe heritable hyperlipidemic rabbits with one of the 4 types of SPIO (0.8 mmol Fe/kg). Two other rabbits served as the control. The rabbits underwent in vivo contrast-enhanced magnetic resonance angiography (MRA) before- and 5 days after these injections; excised aortae were subjected to in vitro MRI. In the in vivo and in vitro studies we assessed the signal intensity of the vessels at identical regions of interest (ROI) and calculated the signal-to-noise ratio (SNR). For histologic assessment we evaluated the iron-positive regions in Prussian blue-stained specimens.

Results

There were significant differences in iron-positive regions where M-USPIO > USPIO, M-SPIO > SPIO, USPIO > SPIO (p < 0.05) but not between M-USPIO and M-SPIO. The difference between the pre- and post-injection SNR was significantly greater in rabbits treated with M-USPIO than USPIO and in rabbits injected with M-SPIO than SPIO (p < 0.05). On in vitro MRI scans SNR tended to be lower in M-USPIO- and M-SPIO- than USPIO- and SPIO-treated rabbits (p < 0.1).

Conclusion

Histologic and imaging analysis showed that mannan-coated SPIO and USPIO particles were taken up more readily by the atherosclerotic rabbit wall than uncoated SPIO and USPIO.     

Evaluation of superparamagnetic iron oxide for MR imaging of liver injury: proton relaxation mechanisms and optimal MR imaging parameters.

PURPOSE: To investigate the proton relaxation mechanisms and the optimal MR imaging parameters in superparamagnetic iron oxide (SPIO)-enhanced MR imaging of liver injury. METHODS: A liver injury model was created in the rat using carbon tetrachloride. The T1 and T2 relaxation effects of SPIO in normal and injured liver were estimated by ex vivo relaxometry. In vivo laser confocal microscopy of the liver was performed to simulate the distribution and clustering of SPIO particles in the hepatic macrophages. SPIO-enhanced MR imaging (1.5T) of normal and diseased rats was performed with variable parameters. The liver specimens were prepared for histopathological examination. RESULTS: Histopathological and laser confocal microscopic findings showed diffuse macrophage distribution but decreased intracellular clustering of SPIO in injured liver. Ex vivo relaxometry showed sustained T1 and T2 relaxation effects of SPIO in liver injury. On MR images obtained with moderate echo time (spin echo [SE] 2000/40 and gradient echo [GRE] 130/9.0/60 degrees), injured liver showed significantly lower decrease in signal-to-noise ratio (SNR) than the normal liver, whereas little difference in SNR was found between the normal and injured liver on heavily T2-(SE 2000/80) and T1-weighted (SE 300/11 and GRE 130/2.0/90 degrees) MR images. CONCLUSION: Pulse sequences with a moderately long echo time (TE) may be more appropriate than heavily T1- or T2-weighted images for distinguishing normal and injured liver in SPIO-enhanced MR imaging because of the maintained T1 and T2 relaxation effect but decreased T2* relaxation effect of SPIO in injured liver.     

磁标记大鼠骨髓间充质干细胞活体内移植后向肝癌细胞趋向性迁移的研究

目的 探讨磁标记大鼠骨髓间充质干细胞(BMSCs)活体内移植后对大鼠肝细胞癌的趋向性迁移及其机制.方法 培养大鼠BMSCs,超顺磁性氧化铁粒子标记.制备大鼠肝癌模型24只,数字表法随机分为3组:实验组(n=12)经脾植入磁标记的BMSCs;对照组A(n=6)移植未标记的BMSCs;对照组B(n=6)不作任何处理.分别于移植前及移植后1、3、7和14 d行MR扫描,选用T_2*WI序列进行移植细胞的示踪并测量肿瘤组织与正常肝组织的信号强度的比值(SI/SI*),结果行单因素方差分析;取肿瘤组织、瘤旁正常肝组织行普鲁士蓝染色,分析BMSCs在体内的分布并与MR对照.结果 BMSCs的磁标记率为90%以上.移植后实验组T_2*WI显示肿瘤信号强度值明显减低,移植前及移植后1、3、7和14 d的SI/SI*值分别为3.18±0.21、1.98±0.20、2.38±0.28、2.70±0.25及3.16±0.24,差异有统计学意义(F=56.65,P<0.05);与移植前相比,1、3、7 d肿瘤信号强度的减低有统计学意义(t值分别为1.20、0.79、0.48,P值均<0.05).对照组移植前后各SI/SI*值差异无统计学意义(P>0.05).免疫组织化学显示实验组肿瘤边缘及内部有大量监染的普鲁士蓝阳性细胞分布,标记细胞在肿瘤内的分布与MR信号改变基本一致.对照组肿瘤组织普鲁士蓝染色均为阴性结果.结论 BMSCs在活体内对肝癌细胞有明显的趋向迁移特性,有望成为基因治疗肝细胞癌的载体.     

MRI in focal liver disease: A comparison of small and ultra-small superparamagnetic iron oxide as hepatic contrast agents

The purpose of this study was to compare small and ultrasmall superparamagnetic iron oxide particles (SPIO and USPIO, respectively) as MR contrast agents for the evaluation of focal hepatic disease. In two different patient groups (SPIO [n = 53], USPIO [n = 27]), with focal liver disease (metastases, hepatocellular carcinoma [HCC], hepatocellular adenoma [HCA], and focal nodular hyperplasia [FNH]), spin-echo T1- and T2-weighted images (T1WI, T2WI) were obtained at 1.0T, before and after intravenous contrast administration. The percentage signal-to-noise ratio (SNR) change and lesion-to-liver contrast (LLC) were measured and statistically compared. The liver decreased in signal intensity (SI) after SPIO administration (?28%) and increased after USPIO administration (+16%) on T1WI. On T2WI, the liver decreased in SI on postcontrast images with both agents (?78% SPIO, ?73% USPIO). This difference was not statistically significantly different (P ? .07). Both SPIO and USPIO provided >500% improvement in LLC on T2WI. On T1WI, LLC was increased in metastases (120%) and HCC (325%) with SPIO. Post-USPIO, LLC was increased on T1WI only in metastases (>500%). Both SPIO and USPIO show excellent hepatic uptake, presumed secondary to reticuloendothelial activity, based on the degree of %SI change seen in the liver after administration of contrast on T2WI. However, USPIO preparations exhibit blood pool activity that may aid in further characterization of focal liver lesions, as is evidenced by their greater T1 effect in the liver and in some focal liver lesions.     

Relaxation effects of clustered particles.

Relations between spatial distribution of superparamagnetic iron oxide (SPIO) particles and the image contrast caused by SPIO were investigated. Actual clustering pattern of particles was measured in the liver and spleen of animals using intravital laser confocal microscopy. SPIO-doped phantoms with and without Sephadex beads were made to simulate these patterns, and relaxation parameters were measured using a 1.5-T clinical scanner. Finally, these results were compared to clinical image data using SPIO particulate agent. Intravital microscopy indicated that the clustering of latex beads was more predominant in hepatic Kupffer cells than in splenic macrophages (P < 0.001). Phantoms without Sephadex beads showed an approximately linear increase of 1/T1 (R1), 1/T2 (R2) and 1/T2* (R2*) values with increasing SPIO concentration. However, with Sephadex beads, R1 and R2 showed little change with increasing SPIO concentration, while R2* showed the same linear increase with SPIO. Also, the R2* values were higher with Sephadex beads. These results were consistent with the clinical imaging data, where signal reduction was significantly smaller in the spleen (-0.4% +/- 27.4%) than in the liver (50.4% +/- 16.8%, P < 0.00001) on T2*-weighted images, but the reduction in the spleen (47.2% +/- 16.1%) was equivalent to the liver (38.8% +/- 26.0%) on T2-weighted images.     

Magnetic resonance imaging enhanced by superparamagnetic iron oxide particles: Usefulness for distinguishing between focused ultrasound‐induced blood–brain barrier disruption and brain hemorrhage

Purpose

To investigate the usefulness of a fully flow‐compensated heavy T2*‐weighted imaging enhanced by superparamagnetic iron oxide (SPIO) particles for distinguishing between focused ultrasound‐induced disruption of blood–brain barrier (BBB) and brain hemorrhage.

Materials and Methods

Focused ultrasound (frequency: 1.5 MHz) was used to induce disruption of the BBB in 39 rats. Two T2*‐weighted images were obtained before and after SPIO administration. Preenhanced T2*‐weighted images were used to detect hemorrhage. Detection of BBB disruption was performed on SPIO‐enhanced images. Thirty‐four rats were sacrificed after magnetic resonance (MR) scanning for histological confirmation of brain lesions. Theremaining five animals were followed up for 35 days. Prussian blue staining was performed on histological sections to detect SPIO particles in the brain.

Results

After SPIO injection the areas of BBB disruption in rat brain were significantly enlarged. The area of mismatch between the T2*‐weighted images indicated a safe region where BBB opening occurred without hemorrhagic complications. In the longitudinal study, removal of SPIO occurred at a faster rate in hemorrhagic areas, albeit being closer to that occurring in the liver. The presence of SPIO was confirmed by Prussian blue staining in brain parenchyma and capillary endothelial cells in areas of BBB disruption.

Conclusion

T2*‐weighted images—either with and without SPIO enhancement—may differentiate focused ultrasound‐induced BBB disruption from brain hemorrhage. J. Magn. Reson. Imaging 2009;29:31–38. © 2008 Wiley‐Liss, Inc.     

Low‐intensity pulsed ultrasound increases cellular uptake of superparamagnetic iron oxide nanomaterial: Results from human osteosarcoma cell line U2OS

Purpose:

To determine whether low‐intensity pulsed ultrasound (LIPUS) is able to facilitate the uptake of a superparamagnetic iron oxide (SPIO) nanomaterial by cells that do not express high endocytosis capacity.

Materials and Methods:

The human osteosarcoma cell line U2OS and a silica‐coated SPIO functionalized peripherally with amines groups (overall diameter 8 nm) were used in this study. Adherent U2OS cells were labeled with SPIO by incubating with culture media containing the SPIO at 4.5 μg[Fe]/mL. LIPUS with the same parameters as those used in clinical application to accelerate bone fracture healing (1.5 MHz, duty cycle 1:4, spatial‐average temporal‐average intensity 30 mW/cm²) was applied to the cells at the beginning of the labeling process for 0, 0.5, 1, or 3 hours. The total incubation time with SPIO was 12 hours. SPIO labeling efficiency was evaluated with Prussian blue staining and a blueness measurement method, and magnetic resonance imaging (MRI) of cell pellets via measuring areas of SPIO‐induced signal void.

Results:

Both Prussian blue staining and in vitro MRI demonstrated that LIPUS application increased the SPIO nanomaterial labeling efficiency for U2OS cells in an exposure‐duration‐dependent manner.

Conclusion:

This study is a “proof of concept” that LIPUS can facilitate the cellular take‐up of SPIO nanomaterial. J. Magn. Reson. Imaging 2010;31:1508–1513. © 2010 Wiley‐Liss, Inc.     

Superparamagnetic iron oxide-mediated hepatic signal intensity change in patients with and without cirrhosis: pulse sequence effects and Kupffer cell function

PURPOSE: To analyze superparamagnetic iron oxide (SPIO)-mediated hepatic signal intensity change in cirrhotic and noncirrhotic liver and to investigate the relationship between pulse sequence effects in SPIO-enhanced magnetic resonance (MR) imaging for hepatic cirrhosis. MATERIALS AND METHODS: Twelve patients with and 12 patients without cirrhosis underwent T2-weighted fast spin-echo, T2*-weighted gradient-echo (GRE), and T1-weighted GRE MR imaging before and twice (early and late phase) after SPIO administration. To assess the effect of SPIO, postcontrast relative signal-to-noise ratio (SNR) changes were statistically analyzed with repeated measurements analysis of variance for each pulse sequence. RESULTS: No interaction was shown between groups and data time points for any pulse sequence. There was no significant difference in mean hepatic relative SNR change on T2-weighted fast spin-echo images between the cirrhotic group and noncirrhotic group (-38.6% and -40.7%, early phase; -42.2% and -49.6%, late phase, respectively). For GRE images, statistically significant differences in mean hepatic relative SNR change were found between the cirrhotic group and noncirrhotic group (-14.2% and -44.5%, early phase; -28.5% and -56.4%, late phase on T2*-weighted GRE images (P <.001); 31.8% and 12.9%, early phase; 23.8% and 2.2%, late phase on T1-weighted GRE images (P <.05), respectively. CONCLUSION: Decreased overall phagocytic activity in cirrhotic liver is more likely due to Kupffer cell dysfunction than to Kupffer cell depletion, since magnetic susceptibility effects on T2*-weighted GRE images depend on intracellular SPIO cluster size.     

Comparison of various techniques of iron oxide-enhanced breath-hold MR imaging of hepatocellular carcinoma

The purpose of our study is to compare qualitatively and quantitatively the abilities of various superparamagnetic iron oxide (SPIO)-enhanced breath-hold magnetic resonance imaging (MRI) techniques to detect hepatocellular carcinoma (HCC). Eight patients with HCCs were imaged. The images were obtained with conventional T2-weighted spin-echo imaging (CSE), half-Fourier single-shot turbo spin-echo (HASTE), single-shot gradient-echo type echo planar imaging (GE-EPI), and single-shot spin-echo type echo planar imaging (SE-EPI) before and after SPIO administration. The liver signal-to-noise ratios (SNRs) in CSE and each EPI sequence were significantly decreased after SPIO administration. GE-EPI had the highest decrease ratio (DR) of liver SNR, followed by SE-EPI (TE=98), SE-EPI (TE=28), CSE, and HASTE in this order. The relative contrasts with GE-EPI and SE-EPI (TE=98) were significantly higher than that with CSE after SPIO administration. On receiver operating characteristic (ROC) analysis, diagnostic accuracy did not differ significantly among the pulse sequences after SPIO administration. GE-EPI and SE-EPI (longer TE) were useful for SPIO-enhanced breath-hold MRI performed to detect HCC.