Paramagnetic Liposomes Research Articles

Persistent contrast enhancement by sterically stabilized paramagnetic liposomes in murine melanoma.

In the present research, we investigated the use of paramagnetic liposomes as contrast agents (CAs) for the detection of solid tumors. The liposomes were sterically stabilized by a polyethylene glycol (PEG) coating, and their size was constrained to approximately 100 nm. Dimyristoyl-sn-glycero-3-phosphoethanolamine-N-diethylene-triaminepentaacetate (DMPE-DTPA) was used as the gadolinium-carrying fatty acid chain. The relaxation properties were characterized through nuclear magnetic relaxation dispersion (NMRD) measurements, and analyzed with the use of theories and computer programs that are adequate for slowly rotating systems. Their relaxivity at 1.5 T was found to be acceptable for in vivo use. We then tested the liposomes against B16-F10 murine melanomas using standard T1-weighted schemes at 1.5 T, and concentrations corresponding to 0.03 mmol/kg of gadolinium (i.e., three to six times lower than the concentration of the small gadolinium complexes in clinical use). The blood half-life was found to be 120 +/- 20 min. The experiments show a good contrast enhancement in the tumor (33% +/- 22%) 2 hr after administration, a further increase (43 +/- 27%) 20 hr after administration, and a decrease (25% +/- 14%) 54 hr after administration. High persistence of the CA was also observed in the liver and intestine, as expected in a hepatobiliar excretion pathway.

Biodistribution of pH-responsive liposomes for MRI and a novel approach to improve the pH-responsiveness

The potential of pH-sensitive paramagnetic liposomes as a probe for monitoring acidic pH in tumours with magnetic resonance imaging has recently been demonstrated. If the blood retention time is prolonged, such liposomes can accumulate in tumour interstitium due to increased vascular permeability and interstitial retention. In the present study, biodistribution studies in healthy rats showed rapid clearance of the pH-sensitive system dipalmitoylphosphatidylethanolamine (DPPE)/dipalmitoylglycerosuccinate (DPSG) liposomal GdDTPA-BMA from the blood circulation with most of the Gd dose in the liver at 15 min post intravenous injection. Incorporation of 1.5 mol% polyethylene glycol (PEG) grafted DPPE (DPPE-PEG) in the above-mentioned formulation resulted in a significantly prolonged blood circulation time. However, the relaxometric pH-response of the DPPE/DPSG/DPPE-PEG system decreased as a function of mol% DPPE-PEG. Therefore, a compromise would be necessary between long blood residence time and a suitable pH-sensitivity of the liposomes. A possible approach to compensate for the reduced pH-sensitivity was investigated. Gadofosveset, a low-molecular weight Gd-chelate with high affinity for albumin, was encapsulated within DPPE/DPSG liposomes. This promising system showed in blood a markedly higher relaxometric response than the corresponding system with GdDTPA-BMA, due to release of gadofosveset at low pH and subsequent binding to albumin.

Novel pH-sensitive paramagnetic liposomes with improved MR properties.

The use of paramagnetic pH-sensitive liposomes was recently suggested as a new approach for monitoring pathologic changes in pH by MRI. Such liposomes must be stable in blood and selectively release the encapsulated paramagnetic agent when exposed to lower pH in the target tissue. In the present study, different liposomal systems were formulated and characterized by relaxometry, cryo-transmission electron microscopy (cryo-TEM), and MRI. The pH-sensitive system dipalmitoylphosphatidylethanolamine/palmitic acid (DPPE/PA) liposomal GdDTPA-BMA, which was previously shown to be unstable in blood, was modified to improve its stability. The incorporation of cholesterol into the DPPE/PA liposomes significantly increased their stability in blood, but the pH sensitivity was diminished. Polyethylene glycol (PEG)-modified DPPE/PA liposomes were pH-insensitive in buffer, and unstable in blood. However, exchanging PA with the double-chained amphiphile dipalmitoylglycerosuccinate (DPSG) yielded liposomes with improved properties. DPPE/DPSG liposomal GdDTPA-BMA was stable in blood at physiological pH, and displayed a marked pH sensitivity. The pH sensitivity was not diminished after preincubation in blood, contrary to what has been reported for analogues containing unsaturated lipids. The potential of this system for monitoring pH was demonstrated in an in vitro MRI phantom study.

Tuning the MR properties of blood-stable pH-responsive paramagnetic liposomes

Paramagnetic pH-responsive liposomes have recently been suggested as a promising approach for monitoring by magnetic resonance imaging (MRI) pH changes in tumours. In the present study, the effects of variations in bilayer composition on the relaxometric properties of diacylphosphatidylethanolamine (PE)/dipalmitoylglycerosuccinate (DPSG) liposomal GdDTPA-BMA were investigated both in buffer and blood. A factorial experimental design was used with the variables PE chain length and mol% DPSG. All the relaxometric profiles displayed a semi-sigmoidal shape with a minimum plateau at high pH ( r 1 min) and a maximum at low pH ( r 1 max,E). Relevant sigmoidal curve fit parameters were evaluated by partial least squares regression. Systematic variations in the relaxometric response ( r 1 max,E− r 1 min) were shown for the liposomal systems both in buffer and blood. The pH value at which the r 1 was 20% of r 1 max,E relative to r 1 min, i.e. pH 20, decreased significantly both in buffer and blood as a function of the mol% DPSG. This phenomenon could be understood by the increased surface charge density with increasing mol% DPSG and, hence, higher barrier against liposome aggregation with consequent leakage of contrast agent. Furthermore, the pH relaxometric profiles in blood were shifted laterally to higher, and likely more clinically relevant pH values than the corresponding profiles in buffer. The liposome formulations displayed minimal leakage of contrast agent after prolonged incubation in blood at physiological pH and retained their pH sensitivity after pre-incubation in blood.

Novel radical‐responsive MRI contrast agent based on paramagnetic liposomes

AbstractA novel, radical responsive MRI contrast agent based on a gadolinium chelate conjugated to a liposome through a disulfide linker was synthesized, with the aim of pursuing the in vivo mapping of radicals. The liposome was prepared by incorporating a thiol‐activated phospholipid, which was subsequently reacted with a gadolinium chelate containing a free thiol group. The long reorientational motion of the supramolecular adduct endows the paramagnetic agent with a relaxivity significantly higher than that of the free complex. The disulfide bond represents a radical‐sensitive moiety and a large decrease in contrast efficacy (T1 relaxivity) is shown upon its cleavage. A preliminary assessment of the system was made by means of in vitro gamma‐irradiation and thiol–disulfide bond exchange with dithiothreitol. Both methods showed a clear dose‐dependent decrease in T1‐relaxivity. Copyright © 2003 John Wiley & Sons, Ltd.

PH-sensitive paramagnetic liposomes for MRI: assessment of stability in blood

The pH-dependent stability of dipalmitoyl phosphatidyl ethanolamine/palmitic acid (DPPE/PA) liposomal GdDTPA-BMA was investigated in human blood and after exposure to selected blood components. Relaxometry, visual observations and cryo-transmission electron microscopy (cryo-TEM) were employed for the assessment of stability. The liposomes were stable in buffer at physiological pH and the T 1-relaxivity (r 1) of the system was significantly lowered compared to that of non-liposomal GdDTPA-BMA, which could be explained by an exchange limited relaxation process. Lowering the pH, however, gave a marked increase in r 1, due to liposome aggregation and subsequent leakage of GdDTPA-BMA. After a few minutes incubation in human blood the liposomes were destabilised and leaky at both high and low pH, and blood components likely to cause the instability were studied. Physiological level of Na + (150 mM) did not affect the relaxometric behavior of the liposomes at pH 7.4, but shifted the pH-r 1 profile laterally to higher pH-values compared to a level of 50 mM Na +. Increased screening of the surface charges and, concomitantly, a lowering of the energy-barrier against aggregation is a plausible explanation for this phenomenon. In contrast, both Ca 2+ and Mg 2+ (physiological level, both 2 mM) caused massive aggregation of the liposomes and leakage of contents, and were therefore much more detrimental to the stability of the liposomes than a physiological level of Na +. This could be due to the higher screening ability of divalent cations, but aggregation could also be induced through an inter-liposomal “bridging” effect. Physiological level of both Na + and Ca 2+ caused less leakage than for lower Na + concentration (50 mM Na + and 2 mM Ca 2+), probably due to competition for the negative surface charges. Albumin also destabilised the liposomes, and it was shown to be due to an interaction between albumin and PA in the liposomal membrane.

Solvent 1H NMRD study of biotinylated paramagnetic liposomes containing Gd-bis-SDA-DTPA or Gd-DMPE-DTPA

The relaxometric properties of two biotinylated paramagnetic liposomes with different lipophilic complexes have been investigated by water proton nuclear magnetic resonance dispersion. The proton relaxivity was found to have a peak at the proton Larmor frequencies generally used in MRI, and to be largely affected by the residence lifetime of the water molecule in the coordination site of the metal chelate. The measurements also indicate that a local motion in the nanosecond time scale, i.e. much faster than the rotational time of the whole liposome, is effective. Possible explanations for this behavior are discussed, that may provide guidelines for the design of second-generation paramagnetic liposomes as contrast agents.

PH-sensitive paramagnetic liposomes as MRI contrast agents: in vitro feasibility studies

A novel type of pH-sensitive paramagnetic contrast agent is introduced; a low molecular weight gadolinium (Gd) chelate (GdDTPA-BMA) encapsulated within pH-sensitive liposomes. The in vitro relaxometric properties of the liposomal Gd chelate were shown to be a function of the pH in the liposomal dispersion and the membrane composition. Only a minor pH-dependency of the T 1 relaxivity (r 1) was observed for liposomal GdDTPA-BMA composed of the unsaturated lipids dioleoyl phosphatidyl ethanolamine (DOPE) and oleic acid (OA). On the other hand, the r 1 of GdDTPA-BMA encapsulated within saturated dipalmitoyl phosphatidyl ethanolamine/palmitic acid (DPPE/PA) liposomes demonstrated a strong pH-dependency. At physiological pH and above, the r 1 of this system was significantly lowered compared to that of non-liposomal Gd chelate, which was explained by an exchange limited relaxation process. Lowering the pH below physiological value, however, gave a sharp and 6–7 fold increase in r 1, due to liposome destabilisation and subsequent leakage of entrapped GdDTPA-BMA. The pH-sensitivity of the DPPE/PA liposome system was confirmed in an in vitro magnetic resonance imaging (MRI) phantom study.

Thermosensitive paramagnetic liposomes for temperature control during MR imaging-guided hyperthermia: In vitro feasibility studies

Magnetic resonance (MR) imaging-based temperature monitoring has gained interest for use in general hyperthermia treatment of tumors. Such therapy requires an accurate control of the temperature, which should range from 41 degrees to 45 degrees C. A novel type of thermosensitive MR agent is proposed: liposome-encapsulated gadolinium chelates whose temperature response is linked to the phase-transition properties of the liposome carrier. In vitro relaxometry and MR imaging were used to evaluate the thermosensitivity of the contrast properties of liposomal gadolinium diethylenetriaminepentaacetic acid bis(methylamide) (Gd-DTPA-BMA). T1 relaxivity (rl) measurements of liposomal Gd-DTPA-BMA were undertaken at 0.47 T and at temperatures of 20 degrees-48 degrees C. MR imaging was performed at 2.0 T with a gel phantom containing inserts of liposomes. Diffusion-weighted and T1-weighted gradient-recalled echo images were acquired as the phantom was heated from 22 degrees to about 65 degrees C. At ambient temperature, the r1 of liposomal Gd-DTPA-BMA was exchange limited due to slow water exchange between the liposome interior and exterior. A sharp, marked increase in r1 occurred as the temperature reached and exceeded the gel-to-liquid crystalline phase-transition temperature (Tm) of the liposomes (42 degrees C). The relaxation enhancement was mainly attributable to the marked increase in transmembrane water permeability, yielding fast exchange conditions. There was good correlation between the relaxometric and imaging results; the signal intensity on T1-weighted gradient-recalled echo images increased markedly as the temperature approached Tm. The temperature sensitivity of the diffusion-weighted technique differed from that of the liposome-based T1-weighted approach, with an apparent water diffusion coefficient increasing linearly with temperature. Since the transition from low to high signal intensity occurred in the temperature range of 38 degrees - 42 degrees C, the investigated paramagnetic liposomes have a potential role as "off-on" switches for temperature control during hyperthermia treatment.

ICAM-1 expression in autoimmune encephalitis visualized using magnetic resonance imaging

The expression of leukocyte adhesion molecules in the intact brains of mice with experimental autoimmune encephalitis (EAE) was visualized by Magnetic Resonance Imaging (MRI) through the use of a new, target-specific MR contrast agent. Antibody-conjugated paramagnetic liposomes (ACPLs) were designed to achieve in vivo targeting of molecules expressed on vascular endothelium, while providing sufficient signal enhancement at these sites for detection by MRI. ACPLs targeted to intercellular adhesion molecule-1 (ICAM-1), an endothelial leukocyte receptor upregulated on cerebral microvasculature during EAE, were administered to diseased mice. Fluorescence microscopy confirmed that fluorescently-tagged ACPLs were localized to central nervous system (CNS) microvasculature in a pattern consistent with ICAM-1 upregulation described immunohistochemically. High resolution MRI of mouse brains ex vivo demonstrated that ACPL binding conferred significant enhancement of signal intensity (SI) as compared to control images. These results suggest that ACPLs can be used as MRI contrast agents to visualize specific molecules expressed on vascular endothelium during disease.

Paramagnetic liposomes as MRI contrast agents: influence of liposomal physicochemical properties on the in vitro relaxivity.

The in vitro contrast efficacy of liposome encapsulated gadolinium-[10-(2-hydroxypropyl)-1,4,7,10-tetraazacyclododecane-1, 4,7-triacetic acid] (GdHPDO3A) has been assessed by relaxometry. The internal concentrations were 150 and 250 mM Gd. Two types of liposome compositions were investigated: a phospholipid blend consisting of both hydrogenated phosphatidylcholine (HPC) and phosphatidylserine (HPS) with a gel-to-liquid crystalline phase transition temperature (Tm) of 50 degrees C, and a mixture of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG) with a Tm of 41 degrees C. The investigated liposome size range was 70-400 nm. The T1 and T2 relaxivities (r1 and r2) of liposome encapsulated GdHPDO3A were significantly reduced at 37 degrees C and 0.47 T, compared to those of non-liposomal metal chelate, due to an exchange limitation of the dipolar relaxation process. The highest relaxivity values were obtained for the DPPC/DPPG liposomes, and were attributed to a higher liposome water permeability and to a more efficient water exchange across the membrane. A reduction in liposome size increased the r1, confirming the exchange limited dipolar relaxation. The increased r1 with increasing temperature demonstrated the prerequisite of rapid water exchange between the interior and exterior of the liposome for efficient dipolar relaxation enhancement. Susceptibility effects were present in the liposome systems as the r2/r1 ratio increased with increasing liposome size and internal Gd concentration. In summary, the current work has shown the influence of key physicochemical properties, such as liposome size, membrane composition and permeability, on the in vitro relaxivity of liposome encapsulated GdHPDO3A.

Paramagnetic liposomes as magnetic resonance imaging contrast agents. Assessment of contrast efficacy in various liver models.

Liposomal gadolinium (Gd)-HP-DO3A has been evaluated as a contrast agent for liver magnetic resonance imaging. The influence of various liposomal physicochemical properties on the liver uptake and contrast efficacy was investigated in various ex vivo and in vivo liver models. Liposomes of different size and membrane properties were prepared. The liposome size ranged from 74 to 304 nm. Two types of phospholipid compositions were studied; a mixture of hydrogenated phosphatidylcholine (HPC) and hydrogenated phosphatidylserine (HPS) with a phase transition temperature (Tm) of 51 degrees C and, a blend composed of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG) displaying a Tm of 41 degrees C. Ex vivo tissue relaxometry and in vivo liver imaging were used to study the influence of liposome composition on the liver uptake and contrast efficacy of intravenously injected liposomes. The influence of liposome size and composition on the kinetics of liver uptake and imaging effect was assessed ex vivo in the perfused rat liver. The HPC/HPS preparations showed generally a higher and faster liver uptake than the DPPC/DPPG preparations due to a higher stability in blood/perfusate (high Tm) and to the HPS component. The liposome size modulated the extent and kinetics of liver uptake; the larger the size, the faster and more extensive was the liver uptake. Both types of liposome preparations were shown to be efficient liver susceptibility agents both ex vivo and in vivo due to their uptake by the Kupffer cells of liver. The lack of full correlation between the extent of liver uptake and degree of contrast enhancement might be attributed to different regimes of susceptibility-based relaxation. The present study has demonstrated the influence of key liposomal physicochemical properties on the liver uptake and contrast efficacy of liposome-encapsulated Gd chelates, exemplified by Gd-HP-DO3A.

Paramagnetic polymerized liposomes as new recirculating MR contrast agents.

We describe a well-tolerated blood pool contrast agent with extended recirculatory half-life based on paramagnetic polymerized liposomes (PPLs). PPLs were constructed from a new type of polymerizable lipid molecule that has a derivative of gadopentetate dimeglumine as the hydrophilic head group and diacetylene groups in the hydrophobic acyl chains, which cross-link when irradiated with ultraviolet (UV) light. Biodistribution, blood pool half-life, and MR image enhancement were determined for PPLs composed of 10% of the gadopentetate dimeglumine lipid and 90% of ditricosadiynoyl tricosadiynayl phosphatidylcholine (DAPC) at a dose of 0.015 mmol Gd+3/kg in rats. In T1-weighed MR images (TR/TE = 400/18 msec), an average signal enhancement of 34% in the kidneys and 20% in the liver was observed, which persisted for at least 90 minutes after administration of the PPLs. Biodistribution studies using radiolabeled PPLs confirmed that 80% of the injected dose remained in the blood pool after 2 hours. The half-life of elimination from the blood pool was 19 hours. The preparation was well tolerated in rats and produced similar MR contrast enhancement of the blood pool as produced by other liposome contrast agents. However, the half-life of PPL elimination from the blood pool was prolonged relative to other liposome systems.

Generation of contrast-carrying liposomes of defined size with a new continuous high pressure extrusion method

A novel continuous high pressure extrusion method was evaluated for the generation of radiopaque and paramagnetic liposomes. The magnetic resonance contrast agent gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA) as well as the X-ray contrast agent iopromide were used as water-soluble model substances for liposomal encapsulation. The continuous process, which is introduced here, allows the fast and efficient extrusion of large batches of liposomal preparations with maximum flow rates of 500 ml/min. Applying high pressures up to 10.5 MPa, MLV prepared by the film method were sequentially extruded through polycarbonate membranes of decreasing pore size. Encapsulation efficiency was found to be dependent on lipid composition and concentration, amount of contrast agent in the preparation as well as choice of final pore size for extrusion. Application of freeze-thaw cycles markedly improved the entrapment of iopromide, whereas for Gd-DTPA freeze-thaw surprisingly turned out to have only minor effects. Entrapment values rose with increasing lipid concentration and fell sharply with increasing solute concentration. Mean liposome diameters could be varied using polycarbonate membranes of differing pore sizes. Smaller final pore sizes led to vesicle populations with smaller mean diameters and lower encapsulation efficiencies. Excellent maximum encapsulation efficiencies of more than 50% for iopromide and over 60% for Gd-DTPA were obtained for vesicles with mean diameters of around 100 nm, as determined by photon correlation spectroscopy (PCS) and confirmed by negative-staining electron microscopy. Employing medium contrast (100 mg/g iodine and 180μmol/g Gd) and lipid concentrations (150 mg/g), entrapment values as high as 40% for the X-ray and 50% for the paramagnetic contrast agent could still be achieved. Best results were obtained using a lipid mixture of soy phosphatidylcholine (SPC), cholesterol (Chol) and soy phosphatidylglycerol (SPG) in a molar ratio of 6:3:1 for iopromide and SPC, Chol 7:3 for Gd-DTPA. Liposomal preparations remained stable upon storage at 2–8°C for 6 months. The new continuous high pressure extrusion method proved to be suitable for the generation of large volumes of stable, contrast-carrying liposomes with outstanding encapsulation efficiencies.

Relaxometry of paramagnetic liposomes.

Relaxometry of paramagnetic liposomes.

Paramagnetic Liposomes as Magnetic Resonance Contrast Agents

Paramagnetic Liposomes as Magnetic Resonance Contrast Agents

ChemInform Abstract: Novel Nitroxides for Spin-Labelling, -Trapping, and Magnetic Resonance Imaging Applications

We describe the synthesis of the isotopically labelled nitrone spin traps 7,9, and 10, and spin labelled cytochalasin B derivatives 15 23. We describe a new general route to proxy1 nitroxide spin labelled fatty acids, for example, 28 and 29, and the 15Nnonadeutero 1 1-proxylpalmitic acid P-OH. New types of paramagnetic liposomes derived from bis nitroxide quaternary salts 31 and 32 have been prepared. Molecules designed as molecular amplifiers such as hexaamine 35a are described. Paramagnetic centers such as a nitroxide epoxide or Gd complex 37 may be attached giving unique spin relaxers such as 35b and 35c to which an isothiocyanate reactive group may be added giving 36b and 36c. INTRODUCTION Stable nitroxide free radicals continue to play a central role in a wide variety of investigations, especially involving spin labelling and spin trapping applications. More recently, novel applications have arisen in which nitroxides are being developed as contrast-enhancing agents for magnetic resonance imaging (MRI) and as paramagnetic agents for the relatively new field of imaging by electron spin resonance (ESR) spectroscopy. In this paper we summarize several recent developments in our laboratory in the nitroxide field. I. Development of *Hand '5N-isotopically substituted nitrone spin traps organic free radicals. In this method an appropriate nitrone or nitroso compound reacts with the short lived radical intermediate producing a nitroxide with a lifetime considerably greater than that of the parent radical. Recent advances in spin trapping include the preparation of a family of pyrrolidine-I-oxide spin traps with enhanced lipophilicity and a decreased tendency toward oxidation3 and the preparation of nitrones which react preferentially with hydroxyl radical over superoxide.4 Spin trapping12 has proven to be a powerful tool for the identification of biologically generated transient One difficulty encountered in the spin trapping of certain free radical intermediates, for example superoxide, is the rather slow rate (-10 M-hec-1) with which nitrones react with the intermediate. Thus potentially toxic concentrations of the nitrone are required for efficient trapping in biological systems. Our aim in this section is to develop nitrone-based spin traps with improved sensitivity owing to isotopic substitution. Our approach is based on the pioneering work of Beth et al.5 and Janzen et a1.6 These groups showed that a significant increase in ESR sensitivity could be achieved by preparing nitroxides that contain 2H and 15N in place of IH and 14N, respectively. Herein we report the synthesis of a family of such isotopically substituted nitrones and a study of their ability to spin trap superoxide and hydroxyl radicals generated from a model superoxide-generating system. 00GcX; C I I RyJ RQ; /O' 4, X, Y = / O H 1, X,Y=15N D 7, R=C&;N=% 8, R = CH3; N 14N 9, R = CD3; N = 15N 10, R = CH3; N = 14N

Novel nitroxides for spin-labelling, -trapping, and magnetic resonance imaging applications

Abstract

Hepatic metastases: liposomal Gd-DTPA-enhanced MR imaging.

Liposomal gadolinium diethylenetriaminepentaacetic acid (DTPA) encapsulated within 70- and 400-nm vesicles was tested as a contrast agent for magnetic resonance (MR) imaging of the liver in rats with hepatic metastases. Liposomal Gd-DTPA caused significant improvement in contrast between liver and tumor (P less than .005) on T1-weighted MR images. Smaller 70-nm liposomal Gd-DTPA vesicles caused greater contrast enhancement, reflecting the larger surface-area-to-volume ratio of the smaller vesicles. Liposomal Gd-DTPA-enhanced images permitted significant improvement in metastasis detection by five blinded radiologists (P less than .005). By comparison, free Gd-DTPA without liposomes caused a statistically significant reduction in contrast between tumor and liver and reduced lesion detection (P less than .01). Liposomal Gd-DTPA also resulted in sustained vascular enhancement for 1 hour after administration. The results suggest that paramagnetic liposomes may become a useful MR imaging contrast agent.

Gadolinium-DTPA liposomes as a potential MRI contrast agent. Work in progress.

To evaluate the use of liposomes containing Gadolinium-DTPA (Gd-DTPA) as potential intravascular contrast agents, we synthesized and tested Gd-DTPA liposomes. A freeze-thaw extrusion process was used to synthesize neutral unilamellar vesicles. Using this technique, we prepared 0.4 micron vesicles with encapsulation efficiency as high as 39% for Gd-DTPA. In vitro dialysis showed that essentially 100% of the Gd-DTPA was retained with the liposomes after 72 hours of dialysis. MR imaging of in vitro samples showed concentration-dependent increase in signal intensity with Gd-DTPA liposomes. Imaging of rats after intravenous injection of Gd-DTPA liposomes showed sustained intravascular contrast enhancement of vascular structures and liver greater than free Gd-DTPA. There was no evidence of acute toxicity in rats during the imaging experiments or on follow-up of two months. Paramagnetic liposomes may be useful to enhance the vasculature, liver, and spleen.