Seok Tae Lim

Coadministration of Angiopoietin-1 and Vascular Endothelial Growth Factor Enhances Collateral Vascularization

Using growth factors to induce vasculogenesis is a promising approach in the treatment of ischemic legs and myocardium. Because the vasculogenesis requires a cascade of growth factors, their receptors, and intracellular signals, such therapies may require the application of more than a single growth factor. We examined the effect of 2 endothelial cell–specific growth factors, angiopoietin-1 (Ang1) and vascular endothelial growth factor (VEGF), on primary cultured porcine coronary artery endothelial cells. VEGF, but not Ang1, increased DNA synthesis and cell number. Ang1 or VEGF induced migration and sprouting activity, increased plasmin and matrix metalloproteinase-2 secretion, and decreased tissue inhibitors of metalloproteinase type 2 secretion. A combination of the submaximal doses of Ang1 and VEGF enhanced these effects and was more potent than the maximal dose of either alone. In a rabbit ischemic hindlimb model, a combination of Ang1 and VEGF gene delivery produced an enhanced effect on resting and maximal blood flow and capillary formation that was greater than that of either factor alone. Angiographic analyses revealed that larger blood vessels were formed after gene delivery of Ang1 or Ang1 plus VEGF than after VEGF gene delivery. These results suggest that combined treatment of Ang1 and VEGF could be used to produce therapeutic vascularization.

Facile Nucleophilic Fluorination Reactions Using tert-Alcohols as a Reaction Medium: Significantly Enhanced Reactivity of Alkali Metal Fluorides and Improved Selectivity

Although protic solvents are generally not preferred for nucleophilic displacement reactions because of their partial positive charge and hydrogen-bonding capacity that solvate the nucleophile and reduce its reactivity, we recently reported a remarkably beneficial effect of using tertiary alcohols as a reaction media for nucleophilic fluorination with alkali metal fluorides, as well as fluorine-18 radiolabeling with [18F]fluoride ion for the preparation of PET radiopharmaceuticals. In this work, we investigate further the influence of the tert-alcohol reaction medium for nucleophilic fluorination with alkali metal fluorides by studying various interactions among tert-alcohols, the alkali metal fluoride (CsF), and the sulfonyloxy substrate. Factors such as hydrogen bonding between CsF and the tert-alcohol solvent, the formation of a tert-alcohol solvated fluoride, and hydrogen bonding between the sulfonate leaving group and the tert-alcohol appear to contribute to the dramatic increase in the rate of the nucleophilic fluorination reaction in the absence of any kind of catalyst. We found that fluorination of 1-(2-mesyloxyethyl)naphthalene (5) and N-5-bromopentanoyl-3,4-dimethoxyaniline (8) with Bu(4)N(+)F(-) in a tert-alcohol afforded the corresponding fluoro products in much higher yield than obtained by the conventional methods using dipolar aprotic solvents. The protic medium also suppresses formation of byproducts, such as alkenes, ethers, and cyclic adducts.

Mesoporous Silica Nanoparticle Pretargeting for PET Imaging Based on a Rapid Bioorthogonal Reaction in a Living Body

Last-minute labeling: Mesoporous silica nanoparticles (MSNs) were modified with a very short half-life fluorine-18-labeled azide radiotracer by a cycloaddition reaction after the MSNs had reached the tumor site in mice. The tumor could then be visualized successfully with positron emission tomography.

SPION-loaded chitosan-linoleic acid nanoparticles to target hepatocytes.

The aim of this study was to develop a novel polymeric magnetic nanoprobe as an MRI contrast agent to target hepatocytes, as well as to evaluate the targeting ability of the nanoprobe with MRI in vivo. Superparamagnetic iron oxide nanocrystals (SPIONs) were synthesized by a thermal decomposition and seed growth method. An 1-ethyl-3-(3-(dimethylamino)-propyl) carbodiimide (EDC)-mediated reaction coupled water-soluble chitosan (WSC) to linoleic acid (LA). Twelve-nanometer-sized SPIONs were incorporated into the core of self-assembled WSC-LA nanoparticles. The morphology and size distribution of the SPION-loaded WSC-LA nanoparticles (SCLNs) were determined by transmittance electron microscopy (TEM) and dynamic light scattering (DLS), respectively. The encapsulation of SPIONs in the WSC-LA nanoparticles reduced the cytotoxicity of bare iron particles and enhanced their dispersion ability in water. The clustering of SPIONs into WSC-LA nanoparticles showed ultrasensitive magnetic behavior. After in vivo intravascular SCLN injection, MRI revealed relative signal enhancement in the liver. The localization of SCLN in hepatocytes was confirmed by Prussian blue staining and TEM analysis. We have successfully developed an ultrasensitive SCLN that effectively targets hepatocytes. The SCLN can be used as a contrast agent to aid in the diagnosis of hepatic diseases.

Superparamagnetic iron oxide nanoparticles as a dual imaging probe for targeting hepatocytes in vivo

Hepatocyte‐specific targeting agents are useful for evaluation of the hepatocytic function and the monitoring of disease progress. Superparamagnetic iron oxide nanoparticles (SPION) bearing terminal galactose groups exhibit a high affinity for the asialoglycoprotein receptor on the hepatocyte surface. In this study, we synthesized and characterized the dual probe SPION detectable by both nuclear and MR imaging modality for specifically targeting hepatocytes in vivo. SPION with 12‐nm diameter were functionalized with dopamine. Surface modification of the SPION was performed to target asialoglycoprotein receptor on hepatocytes, using lactobionic acid. Transmission electron microscope images demonstrated that SPION displayed highly uniform characteristics in terms of both particle size and shape. The X‐ray diffraction pattern of SPION revealed a nanocrystal structure of magnetite. To radiolabel the magnetite with 99mTc, diethylenetriaminepentaacetic acid was conjugated to unreacted functional groups of dopamine. 99mTc‐labeled lactobionic acid‐SPION showed high accumulation in liver, with 38.43 ± 6.45% injected dose per gram. In MR imaging, the reduction of the T2 signal in the liver by lactobionic acid‐SPION was approximately 50.8 ± 7.3%. Competition studies and transmission electron microscope images of liver tissues demonstrated that the lactobionic acid‐SPION were localized in hepatocytes. Therefore, the lactobionic acid‐SPION may be used as a hepatocyte‐targeted dual contrast agent for both nuclear and MR imaging. Magn Reson Med, 2009.

Oleyl-Chitosan Nanoparticles Based on a Dual Probe for Optical/MR Imaging in Vivo

Oleic acid-conjugated chitosan (oleyl-chitosan) is a powerful platform for encapsulating oleic acid-decorated iron oxide nanoparticles (ION), resulting in a good magnetic resonance imaging (MRI) probe. Oleyl-chitosan could self-assemble into core-shell structures in aqueous solution and provide the effective core compartment for loading ION. ION-loaded oleyl-chitosan nanoparticles showed good enhanced MRI sensitivity in a MR scanner. Cy5.5 dye was accessed to the oleyl-chitosan conjugate for near-infrared (NIR) in vivo optical imaging. After intravenous injection of ION-loaded Cy5.5-conjugated oleyl-chitosan (ION-Cy5.5-oleyl-chitosan) nanoparticles in tumor-bearing mice, both NIRF and MR imaging showed the detectable signal intensity and enhancement in tumor tissues via enhanced permeability and retention (EPR) effect. Tumor accumulation of the nanoparticles was confirmed through ex vivo fluorescence images and Prussian blue staining images in tumor tissues. It is concluded that ION-Cy5.5-oleyl-chitosan nanoparticle is highly an effective imaging probe for detecting tumor in vivo.

Synthesis of Iron Oxide Nanoparticles with Control over Shape Using Imidazolium-Based Ionic Liquids

Ionic liquid (IL)-coated iron oxide nanoparticles (IONs) were synthesized by the thermal decomposition of Fe(CO)(5) in an IL/DMF solvent system in the absence of stabilizing surfactants at 90 degrees C for 1 h. The IL, [omim][BF(4)], could regulate the shape and allow the preparation of anisotropic IONs, such as barlike and wirelike nanoparticles. The barlike IONs prepared using this method showed good superparamagnetic behavior in a magnetic field and a MR contrast enhancement effect in a MR scanner.

Superparamagnetic iron oxide nanoparticles-loaded chitosan-linoleic acid nanoparticles as an effective hepatocyte-targeted gene delivery system.

The goal of this study was to develop a gene delivery imaging system that targets hepatocytes to help diagnose and treat various liver diseases. To this end, we prepared superparamagnetic iron oxide nanoparticles (SPIO)-loaded with water-soluble chitosan (WSC)-linoleic acid (LA) nanoparticles (SCLNs) that formed gene complexes capable of localizing specifically to hepatocytes. We confirmed that (99m)Tc-labeled SCLNs delivered into mice via intravenous injection accumulated mainly in the liver using nuclear and magnetic resonance imaging. SCLN/enhanced green fluorescence protein (pEGFP) complexes were also successfully formed and were characterized with a gel retardation assay. SCLN/pEGFP complexes were transfected into primary hepatocytes, where GFP expression was observed in the cytoplasm. In addition, the injection of the gene complexes into mice resulted in significantly increased expression of GFP in hepatocytes in vivo. Furthermore, gene silencing was effectively achieved by administration of gene complexes loaded with specific siRNAs. In conclusion, our results indicate that the SCLNs have the potential to be useful for hepatocyte-targeted imaging and effective gene delivery into hepatocytes.

Optical imaging to trace near infrared fluorescent zinc oxide nanoparticles following oral exposure

Background Understanding how nanomaterials are distributed in the body after exposure is important for assessing whether they are safe. In this study, we investigated the behavior and accumulation of nanoscaled and submicron-scaled zinc oxide (ZnO) particles in the body using optical imaging following oral exposure. Methods To trace these nanoparticles in the body, ZnO nanoparticles were conjugated with a monoreactive hydroxysuccinimide ester of Cy5.5 (Cy5.5-NHS), and the conjugation-stabilizing effect of Cy5.5 on the nanoparticles was evaluated in simulated gastric fluid (pH 1.2) for 7 hours. To compare the distribution of Cy5.5-NHS and Cy5.5-conjugated ZnO nanoparticles, Cy5.5-NHS 0.5 mg/kg and Cy5.5-conjugated ZnO nanoparticles 250 mg/kg were administered orally to healthy rats. We collected blood from the rats at predesignated time points for 7 hours after administration, and optical imaging studies were performed at 1, 2, 3, 5, and 7 hours after dosing. To investigate the extent of nanoparticle accumulation in the organs and tissues, the mice were sacrificed at 23 hours after administration, and the organs were removed and imaged. Results Cy5.5-conjugated ZnO nanoparticles were stable in simulated gastric fluid for 7 hours. The signal intensity of Cy5.5-NHS in blood was highest 3 hours after oral administration, and Cy5.5-conjugated ZnO nanoparticles showed the highest signal intensity in blood 5–7 hours after administration. In vivo optical images indicated that Cy5.5-NHS showed optical signals in the lung, liver, and gastrointestinal tract after oral administration, whereas Cy5.5-conjugated ZnO nanoparticles were seen only in the gastrointestinal tract. Seven hours following administration, biodistribution studies demonstrated that Cy5.5-NHS accumulated in the lung and liver, and Cy5.5-conjugated ZnO nanoparticles resulted in a strong signal in the kidney and liver. Different-sized ZnO nanoparticles showed dissimilar patterns of biodistribution in ex vivo optical images. Conclusion ZnO nanoparticles are absorbed into the tissues following oral exposure and their behavior can be monitored and evaluated using optical imaging.

F-18 Labeling Protocol of Peptides Based on Chemically Orthogonal Strain-Promoted Cycloaddition under Physiologically Friendly Reaction Conditions

We introduce the high-throughput synthesis of various (18)F-labeled peptide tracers by a straightforward (18)F-labeling protocol based on a chemo-orthogonal strain-promoted alkyne azide cycloaddition (SPAAC) using aza-dibenzocyclootyne-substituted peptides as precursors with (18)F-azide synthon to develop peptide based positron emission tomography (PET) molecular imaging probes. The SPAAC reaction and subsequent chemo-orthogonal purification reaction with azide resin proceeded quickly and selectively under physiologically friendly reaction conditions (i.e., toxic chemical reagents-free, aqueous medium, room temperature, and pH ≈7), and provided four (18)F-labeled tumor targetable bioactive peptides such as cyclic Arg-Gly-Asp (cRGD) peptide, bombesin (BBN), c-Met binding peptide (cMBP), and apoptosis targeting peptide (ApoPep) in high radiochemical yields as direct injectable solutions without any HPLC purification and/or formulation processes. In vitro binding assay and in vivo PET molecular imaging study using the (18)F-labeled cRGD peptide also demonstrated a successful application of our (18)F-labeling protocol.