β‐Cell subcellular localization of glucose‐stimulated Mn uptake by X‐ray fluorescence microscopy: implications for pancreatic MRI

Manganese (Mn) is a calcium (Ca) analog that has long been used as a magnetic resonance imaging (MRI) contrast agent for investigating cardiac tissue functionality, for brain mapping and for neuronal tract tracing studies. Recently, we have extended its use to investigate pancreatic β‐cells and showed that, in the presence of MnCl₂, glucose‐activated pancreatic islets yield significant signal enhancement in T₁‐weigheted MR images. In this study, we exploited for the first time the unique capabilities of X‐ray fluorescence microscopy (XFM) to both visualize and quantify the metal in pancreatic β‐cells at cellular and subcellular levels. MIN‐6 insulinoma cells grown in standard tissue culture conditions had only a trace amount of Mn, 1.14 ± 0.03 × 10^?11 µg/µm², homogenously distributed across the cell. Exposure to 2 m m glucose and 50 µ m MnCl₂ for 20 min resulted in nonglucose‐dependent Mn uptake and the overall cell concentration increased to 8.99 ± 2.69 × 10^?11 µg/µm². When cells were activated by incubation in 16 m m glucose in the presence of 50 µ m MnCl₂, a significant increase in cytoplasmic Mn was measured, reaching 2.57 ± 1.34 × 10^?10 µg/µm². A further rise in intracellular concentration was measured following KCl‐induced depolarization, with concentrations totaling 1.25 ± 0.33 × 10^?9 and 4.02 ± 0.71 × 10^?10 µg/µm² in the cytoplasm and nuclei, respectively. In both activated conditions Mn was prevalent in the cytoplasm and localized primarily in a perinuclear region, possibly corresponding to the Golgi apparatus and involving the secretory pathway. These data are consistent with our previous MRI findings, confirming that Mn can be used as a functional imaging reporter of pancreatic β‐cell activation and also provide a basis for understanding how subcellular localization of Mn will impact MRI contrast. Copyright © 2011 John Wiley & Sons, Ltd.     

Relaxivity enhancement of aquated Tris(β‐diketonate)gadolinium(III) chelates by confinement within ultrashort single‐walled carbon nanotubes

Ultrashort single‐walled carbon nanotubes loaded with gadolinium ions (gadonanotubes) have been previously shown to exhibit extremely high T₁‐weighted relaxivities (>100 mm ^?1 s^?1). To further examine the effect of nanoconfinement on the relaxivity of gadolinium‐based contrast agents for magnetic resonance imaging, a series of ultrashort single‐walled carbon nanotube (US‐tube) materials internally loaded with gadolinium chelates have been prepared and studied. US‐tubes were loaded with Gd(acac)₃ · 2H₂O, Gd(hfac)₃ · 2H₂O, and Gd(thd)₃ (acac = acetylacetone, hfac = hexafluoroacetylacetone, thd = tetramethylheptanedione). The longitudinal relaxivities of the prepared materials determined at 25°C in a 1.5 T field were 103 mm ^?1 s^?1 for Gd(acac)₃ · 2H₂O@US‐tubes, 105 mm ^?1 s^?1 for Gd(hfac)₃ · 2H₂O@US‐tubes and 26 mm ^?1 s^?1 for Gd(thd)₃@US‐tubes. Compared with the relaxivities obtained for the unloaded chelates (<10 mm ^?1 s^?1) as well as accounting for the T₁ reduction observed for the empty US‐tubes, the boost in relaxivity for chelate‐loaded US‐tubes is attributed to confinement within the nanotube and depends on the number of coordinated water molecules. Copyright © 2014 John Wiley & Sons, Ltd.     

Influence of molecular parameters and increasing magnetic field strength on relaxivity of gadolinium‐ and manganese‐based T1 contrast agents

Simulations were performed to understand the relative contributions of molecular parameters to longitudinal (r₁) and transverse (r₂) relaxivity as a function of applied field, and to obtain theoretical relaxivity maxima over a range of fields to appreciate what relaxivities can be achieved experimentally. The field‐dependent relaxivities of a panel of gadolinium and manganese complexes with different molecular parameters, water exchange rates, rotational correlation times, hydration state, etc. were measured to confirm that measured relaxivities were consistent with theory. The design tenets previously stressed for optimizing r₁ at low fields (very slow rotational motion; chelate immobilized by protein binding; optimized water exchange rate) do not apply at higher fields. At 1.5T and higher fields, an intermediate rotational correlation time is desired (0.5–4 ns), while water exchange rate is not as critical to achieving a high r₁. For targeted applications it is recommended to tether a multimer of metal chelates to a protein‐targeting group via a long flexible linker to decouple the slow motion of the protein from the water(s) bound to the metal ions. Per ion relaxivities of 80, 45, and 18 mM ^?1 s^?1 at 1.5, 3 and 9.4 T, respectively, are feasible for Gd³⁺ and Mn²⁺ complexes. Copyright © 2009 John Wiley & Sons, Ltd.     

Manganese cell labeling of murine hepatocytes using manganese(III)‐transferrin

Manganese(III)‐transferrin [Mn(III)–Tf] was investigated as a way to accomplish manganese‐labeling of murine hepatocytes for MRI contrast. It is postulated that Mn(III)–Tf can exploit the same transferrin‐receptor‐dependent and ‐independent metabolic pathways used by hepatocytes to transport the iron analog Fe(III)–Tf. More specifically, it was investigated whether manganese delivered by transferrin could give MRI contrast in hepatocytes. Comparison of the T₁ and T₂ relaxation times of Mn(III)–Tf and Fe(III)–Tf over the same concentration range showed that the r₁ relaxivities of the two metalloproteins are the same in vitro, with little contribution from paramagnetic enhancement. The degree of manganese cell labeling following incubation for 2–7 h in 31.5 µm Mn(III)–Tf was comparable to that of hepatocytes incubated in 500 µm Mn²⁺ for 1 h. The intrinsic manganese tissue relaxivity between Mn(III)–Tf‐labeled and Mn²⁺‐labeled cells was found to be the same, consistent with Mn(III) being released from transferrin and reduced to Mn²⁺. For both treatment regimens, manganese uptake by hepatocytes appeared to saturate in the first 1–2 h of the incubation period and may explain why the efficiency of hepatocyte cell labeling by the two methods appeared to be comparable in spite of the ～16‐fold difference in effective manganese concentration. Hepatocytes continuously released manganese, as detected by MRI, and this was the same for both Mn²⁺‐ and Mn(III)–Tf‐labeled cells. Manganese release may be the result of normal hepatocyte function, much in the same way that hepatocytes excrete manganese into the bile in vivo. This approach exploits a biological process—namely receptor binding, endocytosis and endosomal acidification—to initiate the release of an MRI contrast agent, potentially conferring more specificity to the labeling process. The ubiquitous expression of transferrin receptors by eukaryotic cells should make Mn(III)–Tf particularly useful for manganese labeling of a wide variety of cells both in culture and in vivo. Published in 2008 by John Wiley & Sons, Ltd.     

Mn‐loaded apoferritin: a highly sensitive MRI imaging probe for the detection and characterization of hepatocarcinoma lesions in a transgenic mouse model

Mn‐Apo is a highly sensitive MRI contrast agent consisting of ca. 1000 manganese atoms entrapped in the inner cavity of apoferritin. Part of the metallic payload is in the form of Mn²⁺ ions that endow the nano‐sized system with a very high relaxivity that can be exploited to detect hepatocellular carcinoma in mice. Cellular studies showed that Mn‐Apo is readily taken up by normal hepatocytes via the ferritin transporting route. Conversely, hepatoma cells (HTC) displayed a markedly reduced ability to entrap Mn‐Apo from the culture medium. The i.v. administration of Mn‐Apo into C57BL/6 J mice resulted in a marked liver tissue hyperintensity in T₁‐weighted MR image 20 min after injection. When injected into HBV‐tg transgenic mice that spontaneously develop hepatocellular carcinoma (HCC), Mn‐Apo allowed a clear delineation of healthy liver tissue and tumor lesions as hyperintense and hypointense T₁‐weighted MR images, respectively. Immunohistochemistry analysis correlated Mn‐Apo cellular uptake to SCARA5 receptor expression. When the MRI contrast induced by Mn‐Apo was compared with that induced by Gd–BOPTA (a commercial contrast agent known to enter mouse hepatocytes through organic anion transporters) it was found that only some of the lesions were detected by both agents while others could only be visualized by one of the two. These results suggest that Mn‐Apo may be useful to detect otherwise invisible lesions and that the extent of its uptake directly reports the expression/regulation of SCARA5 receptors. Mn‐Apo contrast‐enhanced MR images may therefore contribute to improving HCC lesion detection and characterization. Copyright © 2012 John Wiley & Sons, Ltd.     

Mn‐alginate gels as a novel system for controlled release of Mn2+ in manganese‐enhanced MRI

The aim of the present study was to test alginate gels of different compositions as a system for controlled release of manganese ions (Mn²⁺) for application in manganese‐enhanced MRI (MEMRI), in order to circumvent the challenge of achieving optimal MRI resolution without resorting to high, potentially cytotoxic doses of Mn²⁺. Elemental analysis and stability studies of Mn‐alginate revealed marked differences in ion binding capacity, rendering Mn/Ba‐alginate gels with high guluronic acid content most stable. The findings were corroborated by corresponding differences in the release rate of Mn²⁺ from alginate beads in vitro using T₁‐weighted MRI. Furthermore, intravitreal (ivit) injection of Mn‐alginate beads yielded significant enhancement of the rat retina and retinal ganglion cell (RGC) axons 24 h post‐injection. Subsequent compartmental modelling and simulation of ivit Mn²⁺ transport and concentration revealed that application of slow release contrast agents can achieve a significant reduction of ivit Mn²⁺ concentration compared with bolus injection. This is followed by a concomitant increase in the availability of ivit Mn²⁺ for uptake by RGC, corresponding to significantly increased time constants. Our results provide proof‐of‐concept for the applicability of Mn‐alginate gels as a system for controlled release of Mn²⁺ for optimized MEMRI application. Copyright © 2012 John Wiley & Sons, Ltd.     

Concentration‐independent MRI of pH with a dendrimer‐based pH‐responsive nanoprobe

The measurement of extracellular pH (pH_e) has significant clinical value for pathological diagnoses and for monitoring the effects of pH‐altering therapies. One of the major problems of measuring pH_e with a relaxation‐based MRI contrast agent is that the longitudinal relaxivity depends on both pH and the concentration of the agent, requiring the use of a second pH‐unresponsive agent to measure the concentration. Here we tested the feasibility of measuring pH with a relaxation‐based dendritic MRI contrast agent in a concentration‐independent manner at clinically relevant field strengths. The transverse and longitudinal relaxation times in solutions of the contrast agent (GdDOTA‐4AmP)₄₄‐G5, a G5–PAMAM dendrimer‐based MRI contrast agent in water, were measured at 3 T and 7 T magnetic field strengths as a function of pH. At 3 T, longitudinal relaxivity (r₁) increased from 7.91 to 9.65 mM^?1 s^?1 (on a per Gd³⁺ basis) on changing pH from 8.84 to 6.35. At 7 T, r₁ relaxivity showed pH response, albeit at lower mean values; transverse relaxivity (r₂) remained independent of pH and magnetic field strengths. The longitudinal relaxivity of (GdDOTA‐4AmP)₄₄‐G5 exhibited a strong and reversible pH dependence. The ratio of relaxation rates R₂/R₁ also showed a linear relationship in a pH‐responsive manner, and this pH response was independent of the absolute concentration of (GdDOTA‐4AmP)₄₄‐G5 agent. Importantly, the nanoprobe (GdDOTA‐4AmP)₄₄‐G5 shows pH response in the range commonly found in the microenvironment of solid tumors. Copyright © 2015 John Wiley & Sons, Ltd.     

Improving T1 and T2 magnetic resonance imaging contrast agents through the conjugation of an esteramide dendrimer to high‐water‐coordination Gd(III) hydroxypyridinone complexes

Commercial gadolinium magnetic resonance imaging (MRI) contrast agents are limited by low relaxivity (r₁) and coordination to only a single water molecule (q = 1). Consequently, gram quantities of these agents must be injected to obtain sufficient diagnostic contrast. In this study, MRI contrast agents for T₁ and T₂ relaxivity were synthesized using hydroxypyridinone and terephthalamide chelators with mesityl and 1,4,7‐triazacyclononane capping moieties. When covalently conjugated to a highly biocompatible esteramide dendrimer, T₂ relaxation rates up to 52 m m ^?1 s^?1 and T₁ relaxation rates up to 31 m m ^?1 s^?1 per gadolinium were observed under clinically relevant conditions. These values are believed to be brought about by using a dendritic macromolecule to decrease the molecular tumbling time of the small molecule complexes. These agents also show high aqueous solubility and low toxicity in vitro. In this study we report six new compounds: three discrete complexes and three dendrimer conjugates. Copyright © 2012 John Wiley & Sons, Ltd.     

Disruptive chemical doping in a ferritin‐based iron oxide nanoparticle to decrease r2 and enhance detection with T1‐weighted MRI

Inorganic doping was used to create flexible, paramagnetic nanoparticle contrast agents for in vivo molecular magnetic resonance imaging (MRI) with low transverse relaxivity (r₂). Most nanoparticle contrast agents formed from superparamagnetic metal oxides are developed with high r₂. While sensitive, they can have limited in vivo detection due to a number of constraints with T₂ or T₂*‐weighted imaging. T₁‐weighted imaging is often preferred for molecular MRI, but most T₁‐shortening agents are small chelates with low metal payload or are nanoparticles that also shorten T₂ and limit the range of concentrations detectable with T₁‐weighting. Here we used tungsten and iron deposition to form doped iron oxide crystals inside the apoferritin cavity to form a WFe nanoparticle with a disordered crystal and un‐coupled atomic magnetic moments. The atomic magnetic moments were thus localized, resulting in a principally paramagnetic nanoparticle. The WFe nanoparticles had no coercivity or saturation magnetization at 5 K and sweeping up to ±20 000 Oe, while native ferritin had a coercivity of 3000 Oe and saturation at ±20 000 Oe. This tungsten–iron crystal paramagnetism resulted in an increased WFe particle longitudinal relaxivity (r₁) of 4870 mm ^?1 s^?1 and a reduced transverse relaxivity (r₂) of 9076 mm ^?1 s^?1 compared with native ferritin. The accumulation of the particles was detected with T₁‐weighted MRI in concentrations from 20 to 400 nm in vivo, both injected in the rat brain and targeted to the rat kidney glomerulus. The WFe apoferritin nanoparticles were not cytotoxic up to 700 nm particle concentrations, making them potentially important for targeted molecular MRI. Copyright © 2014 John Wiley & Sons, Ltd.     

Nano assembly and encapsulation; a versatile platform for slowing the rotation of polyanionic Gd3+‐based MRI contrast agents

Encapsulating discrete Gd³⁺ chelates in nano‐assembled capsules (NACs) is a simple and effective method of preparing an MRI contrast agent capable of delivering a large payload of high relaxivity imaging agent. The preparation of contrast agent containing NACs had previously focussed on preparations incorporating GdDOTP^5‐ into the internal aggregate. In this report we demonstrate that other Gd³⁺ chelates bearing overall charges as low as 2‐ can also be used to prepare NACs. This discovery opens up the possibility of using Gd³⁺ chelates that have inner‐sphere water molecules that could further increase the relaxivity enhancement associated with the long τ_R that arises from encapsulation. However, encapsulation of the q = 1 chelate GdDTPA^2‐ did not give rise to a significant increase in relaxivity relative to encapsulation of the outer‐sphere chelate GdTTHA^3‐. This leads us to the conclusion that in the NAC interior proton transport is not mediated by movement of whole water molecules and the enhanced relaxivity of Gd³⁺ chelate encapsulated within NACs arises primarily from second sphere effects. Copyright © 2015 John Wiley & Sons, Ltd.     

Enhanced MRI relaxivity of Gd3+‐based contrast agents geometrically confined within porous nanoconstructs

Gadolinium chelates, which are currently approved for clinical MRI use, provide relaxivities well below their theoretical limit, and they also lack tissue specificity. Recently, the geometrical confinement of Gd³⁺‐based contrast agents (CAs) within porous structures has been proposed as a novel, alternative strategy to improve relaxivity without chemical modification of the CA. Here, we have characterized and optimized the performance of MRI nanoconstructs obtained by loading [Gd(DTPA)(H₂O)]^2? (Magnevist®) into the pores of injectable mesoporous silicon particles. Nanoconstructs with three different pore sizes were studied, and at 60 MHz, they exhibited longitudinal relaxivities of ~24 m m ^?1 s^?1 for 5–10 nm pores and ~10 m m ^?1 s^?1 for 30 – 40 nm pores. No enhancement in relaxivity was observed for larger pores sizes. Using an outer‐sphere compound, [GdTTHA]^3?, and mathematical modeling, it was demonstrated that the relaxivity enhancement is due to the increase in rotational correlation times (CA adsorbed on the pore walls) and diffusion correlation times (reduced mobility of the water molecules), as the pore sizes decreases. It was also observed that extensive CA adsorption on the outer surface of the silicon particles negates the advantages offered by nanoscale confinement. Upon incubation with HeLa cells, the nanoconstructs did not demonstrate significant cytotoxicity for up to 3 days post incubation, at different particle/cell ratios. In addition, the nanoconstructs showed complete degradation after 24 h of continuous agitation in phosphate‐buffered saline. These data support and confirm the hypothesis that the geometrical confinement of Gd³⁺‐chelate compounds into porous structures offers MRI nanoconstructs with enhanced relaxivity (up to 6 times for [Gd(DTPA)(H₂O)]^2?, and 4 times for [GdTTHA]^3?) and, potentially, improved stability, reduced toxicity and tissue specificity. Copyright © 2012 John Wiley & Sons, Ltd.     

Bifunctional Gd(III) and Tb(III) chelates based on a pyridine–bis(iminodiacetate) platform,suitable optical probes and contrast agents for magnetic resonance imaging

To study the physicochemical properties of lanthanide complexes derived from a bifunctional chelating agent based on a PMN‐tetraacetic acid moiety {PMN‐tetraacetic acid (1): [2,6‐pyridinediylbis(methylene nitrilo)‐tetraacetic acid]}, 4‐carboxylic acid substituted pyridine derivative (2) was synthesized. This ligand forms heptadentate (N₃O₄) Ln(III) complexes (Ln = Gd, Eu, Tb), with two water molecules completing the inner coordination sphere of the metal. The parameters that govern the relaxivity of the Gd(III) complex and the luminescence of Eu(III) and Tb(III) complexes were obtained by ¹⁷O and ¹H NMR studies and time‐resolved fluorescence experiments, respectively. The gadolinium and terbium complexes show interesting properties either for MRI or FOR optical imaging; that is, for the Gd complex, a high proton relaxivity (r₁ = 6.4 s^?1 mm ^?1 at 20 MHz) with short water residence time (τ_M = 38.5 ns); for the Tb complex, a luminescence lifetime of 1.22 ms at room temperature and a luminescence quantum yield of 10%. The kinetic stability of these complexes toward blood protein, cation or bioactive oxyanion was also examined. The Gd(2)(H₂O)₂ complex does not interact with human serum albumin, but undergoes a transmetalation reaction with Zn(II) in a phosphate buffer solution (pH 7.4), rather similar to that of Gd–DTPA–BMA(H₂O). On the other hand, as observed for Eu and Tb complexes, these chelates do not form ternary complexes with bidentate anions such as l ‐lactate, citrate or carbonate. Finally, a phosphatidylserine‐specific hexapeptide (TLVSSL) was grafted on Gd or Tb chelates, and the Gd–peptide conjugate was used in vitro for targeting apoptotic cells. Copyright © 2014 John Wiley & Sons, Ltd.     

Synthesis and evaluation of a polydisulfide with Gd–DOTA monoamide side chains as a biodegradable macromolecular contrast agent for MR blood pool imaging

Macromolecular Gd(III)‐based contrast agents are effective for contrast‐enhanced blood pool and cancer MRI in preclinical studies. However, their clinical applications are impeded by potential safety concerns associated with slow excretion and prolonged retention of these agents in the body. To minimize the safety concerns of macromolecular Gd contrast agents, we have developed biodegradable macromolecular Gd contrast agents based on polydisulfide Gd(III) complexes. In this study, we designed and synthesized a new generation of the polydisulfide Gd(III) complexes containing a macrocyclic Gd(III) chelate, Gd–DOTA monoamide, to improve the in vivo kinetic inertness of the Gd(III) chelates. (N₆‐Lysyl)lysine‐(Gd–DOTA) monoamide and 3‐(2‐carboxyethyldisulfanyl)propanoic acid copolymers (GODC) were synthesized by copolymerization of (N₆‐lysyl)lysine DOTA monoamide and dithiobis(succinimidylpropionate), followed by complexation with Gd(OAc)₃. The GODC had an apparent molecular weight of 26.4 kDa and T₁ relaxivity of 8.25 m m ^?1 s^?1 per Gd at 1.5 T. The polymer chains of GODC were readily cleaved by l ‐cysteine and the chelates had high kinetic stability against transmetallation in the presence of an endogenous metal ion Zn²⁺. In vivo MRI study showed that GODC produced strong and prolonged contrast enhancement in the vasculature and tumor periphery of mice with breast tumor xenografts. GODC is a promising biodegradable macromolecular MRI contrast agent with high kinetic stability for MR blood pool imaging. Copyright © 2013 John Wiley & Sons, Ltd.     

The Gd3+ complex of 1,4,7,10‐tetraazacyclododecane‐1,4,7,10‐tetraacetic acid mono(p‐isothiocyanatoanilide) conjugated to inulin: a potential stable macromolecular contrast agent for MRI

Reaction of DOTA–NCSA [1,4,7,10‐tetraazacyclododecane‐1,4,7,10‐tetraacetic acid mono(p‐isothiocyanatoanilide)] with O‐(aminopropyl)inulin (degree of polymerization 25) provided a chelate that formed a kinetically extremely stable Gd³⁺ complex. No transmetalation was observed with Zn²⁺. The conjugate has a relaxivity of 21.7 s^?1 m m ^?1 at 20 MHz and 37 °C, and each molecule of the inulin carries on average 35 Gd³⁺ ions. The parameters governing the relaxivity of this material and of a low‐molecular‐weight model compound prepared by conjugation of DOTA–NCSA and propylamine were evaluated by investigation of their water ¹H longitudinal relaxation rate enhancements at different magnetic fields (NMRD) and by studying variable temperature ¹⁷O NMR data. The high relaxivity of the inulin conjugate can be ascribed to the efficient slowing down of the molecular tumbling by this carrier. The rotational correlation time at 37 °C of this material is 1460 ps, whereas that of the model compound is 84 ps. Furthermore, both complexes do not interact significantly with human serum albumin, as shown by their NMRD profiles, and do not undergo transmetallation by zinc ions. The inulin conjugate thus has potential for application as a contrast agent for MRI, particularly as a blood pool agent. Copyright © 2011 John Wiley & Sons, Ltd.     

Determination of liver‐specific r2* of a highly monodisperse USPIO by 59Fe iron core‐labeling in mice at 3 T MRI

Accurate determination of tissue concentration of ultrasmall superparamagnetic iron oxide nanoparticles (USPIO) using T₂* MR relaxometry is still challenging. We present a reliable quantification method for local USPIO amount with the estimation of the liver specific relaxivity r₂* using monodisperse ⁵⁹Fe‐core‐labeled USPIO (⁵⁹FeUSPIO). Dynamic and relaxometric in vivo characteristics of unlabeled monodisperse USPIO were determined in MRI at 3 T. The in vivo MR studies were performed for liver tissue with ⁵⁹FeUSPIO using iron dosages of 9 (n = 3), 18 (n = 2) and 27 (n = 3) µmol Fe kg^?1 body weight. The R₂* of the liver before and after USPIO injection (?R₂*) was measured and correlated with ⁵⁹Fe activity measurements of excised organs by a whole body radioactivity counter (HAMCO) to define the dependency of ?R₂* and ⁵⁹FeUSPIO liver concentration and calculate the r₂* of ⁵⁹FeUSPIO for the liver. Ultrastructural analysis of liver uptake was performed by histology and transmission electron microscopy. ?R₂* of the liver revealed a dosage‐dependent accumulation of ⁵⁹FeUSPIO with a percentage uptake of 70–88% of the injection dose. Hepatic ?R₂* showed a dose‐dependent linear correlation to ⁵⁹FeUSPIO activity measurements (r = 0.92) and an r₂* in the liver of 481 ± 74.9 mm ^?1 s^?1 in comparison to an in vitro r₂* of 60.5 ± 3.3 mm ^?1 s^?1. Our results indicate that core‐labeled ⁵⁹FeUSPIO can be used to quantify the local amount of USPIO and to estimate the liver‐specific relaxivity r₂*. Copyright © 2014 John Wiley & Sons, Ltd.     

Dynamic manganese‐enhanced magnetic resonance imaging can detect chronic cryoinjury‐induced infarction in pig hearts in vivo

The purpose was to investigate whether MnCl₂ can serve as an MRI contrast agent to detect chronic cryoinjury infarction in pigs in vivo and whether MnCl₂ causes significant hemodynamic disturbances. Hearts were subjected to a topical 2 min cryothermia to establish myocardial infarction (MI). Thereafter GdDTPA‐enhanced MRI was performed at 0, 1, 2 and 3 weeks using a 3 T scanner. Four weeks post‐cryoinjury the pigs underwent in vivo Mn‐enhanced magnetic resonance imaging (MEMRI). MnCl₂ (70 μmol/kg, 14 min) was infused i.v. intermittently (n = 4) or continuously (n = 5) and T₁‐weighted images were acquired every 2 min simultaneously recording heart rate and arterial blood pressure. Either infusion scheme led to an immediate increment in MR signal intensity (SI) within the left ventricular (LV) blood pool and LV normal and cryoinjured myocardium, which reached a maximum at the end of infusion. No significant difference was observed between the normal and cryoinjured myocardium. After infusion termination, SI decreased faster within the LV blood pool and the MI, as compared with the normal myocardium in either group, resulting in significant contrast between the MI and normal tissue (intermittent: 18 ± 7 vs 49 ± 13%, p = 0.002; continuous: 19 ± 8 vs 36 ± 9%, p = 0.004). Infarction sizes were similar in Mn²⁺‐ and GdDTPA‐enhanced images at 4 and 3 weeks post injury, respectively. Thus, in vivo MEMRI differentiated infarcted from normal myocardium in pig hearts subjected to 4‐week cryoinjury. Compared with intermittent infusion, continuous infusion minimized hemodynamic fluctuations. Copyright © 2011 National Research Council Canada.     

Optimized synthesis of 100 nm diameter magnetoliposomes with high content of maghemite particles and high MRI effect

Magnetoliposomes are liposomes surrounding an iron oxide core, which are used as contrast enhancing agents in magnetic resonance imaging (MRI). One method for producing magnetoliposomes consists of hydration of a lipid film with citrate‐coated iron oxide particles followed by extrusion. Two parameters are of major importance for in vivo applications of magnetoliposomes, namely their size, which must be small, optimally around 100 nm diameter, in order to ensure their prolonged circulation in the bloodstream, and their iron content, which must be maximal for generating high MRI effect. We studied the formation of magnetoliposomes by passive encapsulation of maghemite (γ‐Fe₂O₃) particle suspensions of varying concentrations, with the objective of producing magnetoliposomes of small size and high iron content. The iron to lipid ratio was used to determine the iron content of the magnetoliposomes after the successive purification steps and cryo‐TEM was used to characterize their size, their homogeneity and the efficiency of purification. The size of citrate‐coated maghemite clusters was found to be of critical importance for obtaining magnetoliposomes smaller than 200 nm. We were able to reproducibly synthesize magnetoliposomes of 100 nm diameter with high iron content –up to 77 particles per liposome (5.6 moles iron per mole lipid) – and high r₂ MRI relaxivity – up to 320 m m^?1 . s^?1. The magnetoliposomes present improved characteristics compared with previous reports. Future research will focus on using these magnetoliposomes as drug delivery systems for in vivo diagnostics or therapeutics applications. Copyright © 2011 John Wiley & Sons, Ltd.     

Positive MRI contrast enhancement in THP‐1 cells with Gd2O3 nanoparticles

There is a demand for more efficient and tissue‐specific MRI contrast agents and recent developments involve the design of substances useful as molecular markers and magnetic tracers. In this study, nanoparticles of gadolinium oxide (Gd₂O₃) have been investigated for cell labeling and capacity to generate a positive contrast. THP‐1, a monocytic cell line that is phagocytic, was used and results were compared with relaxivity of particles in cell culture medium (RPMI 1640). The results showed that Gd₂O₃‐labeled cells have shorter T₁ and T₂ relaxation times compared with untreated cells. A prominent difference in signal intensity was observed, indicating that Gd₂O₃ nanoparticles can be used as a positive contrast agent for cell labeling. The r₁ for cell samples was 4.1 and 3.6 s^?1 mm ^?1 for cell culture medium. The r₂ was 17.4 and 12.9 s^?1 mm ^?1, respectively. For r₁, there was no significant difference in relaxivity between particles in cells compared to particles in cell culture medium, (p_r1 = 0.36), but r₂ was significantly different for the two different series (p_r2 = 0.02). Viability results indicate that THP‐1 cells endure treatment with Gd₂O₃ nanoparticles for an extended period of time and it is therefore concluded that results in this study are based on viable cells. Copyright © 2008 John Wiley & Sons, Ltd.     

Evaluation of rHA labeled with Gd–DTPA for blood pool imaging and targeted contrast delivery

A new contrast agent was developed by linking Gd–DTPA chelate to recombinant human albumin in the laboratory. The molar relaxivity of the new agent was tested in aqueous solution at B₀ 1.5 T and temperature 20°C. The soluble compound had a higher molar longitudinal relaxivity and molar transverse relaxivity in water (r₁ = 7.2 s^?1 mM ^?1, r₂ = 18.4 s^?1 mM ^?1) than those measured for Gd–DTPA solution (r₁ = 3.5 s^?1 mM ^?1, r₂ = 5.5 s^?1 mM ^?1). The performance of the compound as a blood pool agent was investigated with soluble and microparticulate forms of the compound and comparisons were made with Gd–DTPA and the polymeric blood‐pool agent, Gadomer. T₁‐weighted imaging experiments show that the soluble compound acts as a highly effective blood pool agent with hyperintensity in the vasculature persisting beyond 2 h post administration, compared with free Gd–DTPA, which was cleared from the blood pool after approximately 10 min. The clearance kinetics of the new agents were examined, due to the incomplete elimination within 14 days post injection; both rHA labeled compounds are probably not suitable for development as routine blood pool contrast media. However, with free sites on the Gd‐loaded rHA molecule, there are possibilities for binding the agent to antibodies in the laboratory, which was demonstrated, and thus there exist potential applications for in vivo molecular imaging with this agent. Copyright © 2010 John Wiley & Sons, Ltd.     

Targeted RGD nanoparticles for highly sensitive in vivo integrin receptor imaging

A new magnetic resonance imaging (MRI) contrast bearing RGD peptide is reported. In this study, ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles with various sizes were prepared. Particles sizes between 6 and 13 nm were tuned by varying the stirring rate. Remarkable negative contrast was observed because USPIO nanoparticles (13.1 ± 2.1 nm) generated high transversal relaxivity r₂ (188 ± 3 m m ^?1 s^?1) and saturation magnetization (94 emu g^?1 Fe). The USPIO nanoparticles were coated with PDA [2‐(pyridyldithio)‐ethylamine; PDA nanoparticles] containing functional polymer, which can be readily synthesized by Michael addition. The PDA nanoparticles were conjugated with RGD peptide (RGD nanoparticles) for targeting the specific site. The target specificity and high relaxivity allowed RGD nanoparticles to differentiate the expression level of integrin receptor on several cell lines and tumors (MCF‐7, A‐549, HT‐29 and HT‐1080) by in vitro and in vivo MR imaging. Importantly, a remarkable negative contrast (?51.3 ± 6.7%) was observed for in vivo MR imaging of MCF‐7 tumor. This result implies that the RGD nanoparticles that greatly enhance the MR imaging are highly sensitive for early stage tumor detection. Copyright © 2012 John Wiley & Sons, Ltd.