Jinglong Zhao

Targeted Tumor Computed Tomography Imaging Using Low‐Generation Dendrimer‐Stabilized Gold Nanoparticles

We report a facile approach to fabricating low-generation poly(amidoamine) (PAMAM) dendrimer-stabilized gold nanoparticles (Au DSNPs) functionalized with folic acid (FA) for in vitro and in vivo targeted computed tomography (CT) imaging of cancer cells. In this study, amine-terminated generation 2 PAMAM dendrimers were employed as stabilizers to form Au DSNPs without additional reducing agents. The formed Au DSNPs with an Au core size of 5.5 nm were covalently modified with the targeting ligand FA, followed by acetylation of the remaining dendrimer terminal amines to endow the particles with targeting specificity and improved biocompatibility. Our characterization data show that the formed FA-modified Au DSNPs are stable at different pH values (5-8) and temperatures (4-50 °C), as well as in different aqueous media. MTT assay data along with cell morphology observations reveal that the FA-modified Au DSNPs are noncytotoxic in the particle concentration range of 0-3000 nM. X-ray attenuation coefficient measurements show that the CT value of FA-modified Au DSNPs is much higher than that of Omnipaque (a clinically used CT contrast agent) at the same concentration of the radiodense elements (Au or iodine). Importantly, the FA-modified Au DSNPs are able to specifically target a model cancer cell line (KB cells, a human epithelial carcinoma cell line) over-expressing FA receptors and they enable targeted CT imaging of the cancer cells in vitro and the xenografted tumor model in vivo after intravenous administration of the particles. With the simple synthesis approach, easy modification, good cytocompatibility, and high X-ray attenuation coefficient, the FA-modified low-generation Au DSNPs could be used as promising contrast agents for targeted CT imaging of different tumors over-expressing FA receptors.

Lactobionic Acid-Modified Dendrimer-Entrapped Gold Nanoparticles for Targeted Computed Tomography Imaging of Human Hepatocellular Carcinoma

Development of novel nanomaterial-based contrast agents for targeted computed tomography (CT) imaging of tumors still remains a great challenge. Here we describe a novel approach to fabricating lactobionic acid (LA)-modified dendrimer-entrapped gold nanoparticles (LA-Au DENPs) for in vitro and in vivo targeted CT imaging of human hepatocellular carcinoma. In this study, amine-terminated poly(amidoamine) dendrimers of generation 5 pre-modified with fluorescein isothiocyanate and poly(ethylene glycol)-linked LA were employed as templates to form Au nanoparticles. The remaining dendrimer terminal amines were subjected to an acetylation reaction to form LA-Au DENPs. The prepared LA-Au DENPs were characterized via different methods. Our results reveal that the multifunctional Au DENPs with a Au core size of 2.7 nm have good stability under different pH (5-8) and temperature (4-50 °C) conditions and in different aqueous media, and are noncytotoxic to normal cells but cytotoxic to the targeted hepatocarcinoma cells in the given concentration range. In vitro flow cytometry data show that the LA-Au DENPs can be specifically uptaken by a model hepatocarcinoma cell line overexpressing asialoglycoprotein receptors through an active receptor-mediated targeting pathway. Importantly, the LA-Au DENPs can be used as a highly effective nanoprobe for specific CT imaging of hepatocarcinoma cells in vitro and the xenoplanted tumor model in vivo. The developed LA-Au DENPs with X-ray attenuation property greater than clinically employed iodine-based CT contrast agents hold a great promise to be used as a nanoprobe for targeted CT imaging of human hepatocellular carcinoma.

Size-controlled synthesis of dendrimer -stabilized silver nanoparticles for X-ray computed tomography imaging applications

We report a facile size-controlled synthesis of dendrimer-stabilized silver nanoparticles (Ag DSNPs) for X-ray computed tomography (CT) imaging applications. Amine-terminated generation 5 poly(amidoamine) dendrimers were used as templates to complex Ag(I) ions for subsequent reductive formation of dendrimer-entrapped Ag nanoparticles. Following a one-step acetylation reaction to transform dendrimer terminal amine to acetyl groups, Ag DSNPs can be formed. The formed Ag DSNPs were characterized using 1H NMR, UV-Vis spectrometry, transmission electron microscopy, and ζ-potential measurements. We show that through the variation of the dendrimer/Ag salt molar ratio, the size of Ag DSNPs can be controlled at the range of 8.8–23.2 nm. The formed Ag DSNPs are stable not only in water, PBS buffer, and fetal bovine serum, but also at different pH conditions (pH 5–8) and temperatures (20–50 °C). X-Ray absorption coefficient measurements show that the attenuation of Ag DSNPs is size-dependent, and the Ag DSNPs with a diameter of 16.1 nm display an X-ray attenuation intensity close to that of a clinically used iodine-based contrast agent (Omnipaque) at the same molar concentration of the active element (Ag versus iodine). This suggests that Ag DSNPs with an appropriate size have a great potential to be used as a CT imaging contrast agent, although the atomic number of Ag is lower than that of iodine. Furthermore, CT scanning showed prolonged enhancement at the point of mice injected subcutaneously with Ag DSNPs, rendering them as a promising contrast agent in CT imaging applications.

Facile hydrothermal synthesis of low generation dendrimer-stabilized gold nanoparticles for in vivo computed tomography imaging applications

We report a facile approach to synthesizing low generation poly(amidoamine) (PAMAM) dendrimer-stabilized gold nanoparticles (Au DSNPs) for in vivo computed tomography (CT) imaging applications. In this study, amine-terminated generation 2 PAMAM dendrimers were employed as stabilizers to form gold nanoparticles via a simple hydrothermal method. The formed aminated Au DSNPs were then acetylated to neutralize the dendrimer terminal amines, rendering the particles with improved biocompatibility. The final formed acetylated Au DSNPs were characterized via different techniques. We show that the formed Au DSNPs with an Au core size of 5.6 nm are relatively uniform and stable at different pH (5–8) and temperature (4–50 °C) conditions. X-ray attenuation coefficient measurements show that the Au DSNPs display approximately the same X-ray attenuation property as that of Omnipaque, a clinically used iodinated contrast agent. Importantly, the acetylated Au DSNPs showed much better performance in CT imaging of the major organs of rats (heart, liver, kidney, spleen, and bladder) in vivo than Omnipaque, likely due to their nanometer size and thus prolonged blood circulation time. The formed Au DSNPs may be used as a promising contrast agent for CT imaging of different biological systems.

Tunable synthesis and acetylation of dendrimer-entrapped or dendrimer-stabilized gold–silver alloy nanoparticles

In this study, amine-terminated generation 5 poly(amidoamine) dendrimers were used as templates or stabilizers to synthesize dendrimer-entrapped or dendrimer-stabilized Au-Ag alloy nanoparticles (NPs) with different gold atom/silver atom/dendrimer molar ratios with the assistance of sodium borohydride reduction chemistry. Following a one-step acetylation reaction to transform the dendrimer terminal amines to acetyl groups, a series of dendrimer-entrapped or dendrimer-stabilized Au-Ag alloy NPs with terminal acetyl groups were formed. The formed Au-Ag alloy NPs before and after acetylation reaction were characterized using different techniques. We showed that the optical property and the size of the bimetallic NPs were greatly affected by the metal composition. At the constant total metal atom/dendrimer molar ratio, the size of the alloy NPs decreased with the gold content. The formed Au-Ag alloy NPs were stable at different pH (pH 5-8) and temperature (4-50°C) conditions. X-ray absorption coefficient measurements showed that the attenuation of the binary NPs was dependent on both the gold content and the surface modification. With the increase of gold content in the binary NPs, their X-ray attenuation intensity was significantly enhanced. At a given metal composition, the X-ray attenuation intensity of the binary NPs was enhanced after acetylation. Cytotoxicity assays showed that after acetylation, the cytocompatibility of Au-Ag alloy NPs was significantly improved. With the controllable particle size and optical property, metal composition-dependent X-ray attenuation characteristics, and improved cytocompatibility after acetylation, these dendrimer-entrapped or dendrimer-stabilized Au-Ag alloy NPs should have a promising potential for CT imaging and other biomedical applications.

Facile formation of folic acid-modified dendrimer-stabilized gold–silver alloy nanoparticles for potential cellular computed tomography imaging applications

We report a facile approach to fabricating dendrimer-stabilized gold-silver alloy nanoparticles (Au-Ag alloy DSNPs) for targeted in vitro computed tomography (CT) imaging of cancer cells. In this study, folic acid (FA)-modified amine-terminated generation 5 poly(amidoamine) dendrimers (G5·NH2-FA) were used as stabilizers to prepare Au-Ag alloy DSNPs by simultaneously reducing both gold and silver salts, followed by acetylation of the dendrimer terminal amines. The formed Au-Ag alloy DSNPs were characterized via different techniques. We show that the formed Au-Ag alloy DSNPs are spherical in shape with a relatively narrow size distribution, have good water solubility and colloidal stability, and display higher X-ray attenuation intensity than the iodine-based contrast agent of Omnipaque at the same molar concentration of the active element (i.e., Au plus Ag, or iodine). Cytotoxicity assay results show that the Au-Ag alloy DSNPs are cytocompatible in a given concentration range. Importantly, the formed Au-Ag alloy DSNPs are able to be specifically taken up by cancer cells overexpressing FA receptors and enable targeted CT imaging of the cancer cells. Given the unique structural characteristics of dendrimers and the facile synthesis of DSNPs, the developed Au-Ag alloy DSNPs may be used for various biomedical applications in sensing, diagnosis, and therapeutics.

Facile synthesis of acetylated dendrimer-entrapped gold nanoparticles with enhanced gold loading for CT imaging applications

We describe a facile approach to synthesizing acetylated dendrimer-entrapped gold nanoparticles (Au DENPs) with enhanced Au loading in the dendrimer interior. In this study, amine-terminated generation 5 poly(amidoamine) (PAMAM) dendrimers (G5.NH2) were used as templates to form Au DENPs via a stepwise Au salt complexation/reduction approach, followed by acetylation of the dendrimer terminal amines. The formed Au DENPs before and after acetylation were characterized with different techniques. We show that the stepwise complexation/reduction of HAuCl4 is able to significantly improve the loading amount of Au within the dendrimer interior. UV-Vis spectroscopy reveals that the intensity of the surface plasmon resonance (SPR) band increases with the Au loading, confirming the stepwise loading synthesis of Au DENPs. TEM images show that the synthesized Au DENPs have a quite uniform size distribution with sizes tunable in the range of 2-4 nm depending on the Au loading. The formed acetylated Au DENPs with enhanced Au loading are very stable under different pH and temperature conditions. Importantly, computed tomography (CT) imaging experiments reveal that the formed acetylated Au DENPs have higher attenuation intensity than a clinically used iodinated contrast agent, Omnipaque, at the same molar concentration of active elements (Au or iodine), and enable significantly enhanced CT imaging of rat heart in vivo. The acetylated Au DENPs with enhanced Au loading formed via the facile stepwise approach may be used as contrast agents for highly sensitive CT imaging applications.

Magnetic resonance imaging of glioma with novel APTS-coated superparamagnetic iron oxide nanoparticles

We report in vitro and in vivo magnetic resonance (MR) imaging of C6 glioma cells with a novel acetylated 3-aminopropyltrimethoxysilane (APTS)-coated iron oxide nanoparticles (Fe3O4 NPs). In the present study, APTS-coated Fe3O4 NPs were formed via a one-step hydrothermal approach and then chemically modified with acetic anhydride to generate surface charge-neutralized NPs. Prussian blue staining and transmission electron microscopy (TEM) data showed that acetylated APTS-coated Fe3O4 NPs can be taken up by cells. Combined morphological observation, cell viability, and flow cytometric analysis of the cell cycle indicated that the acetylated APTS-coated Fe3O4 NPs did not significantly affect cell morphology, viability, or cell cycle, indicating their good biocompatibility. Finally, the acetylated APTS-coated Fe3O4 nanoparticles were used in magnetic resonance imaging of C6 glioma. Our results showed that the developed acetylated APTS-coated Fe3O4 NPs can be used as an effective labeling agent to detect C6 glioma cells in vitro and in vivo for MR imaging. The results from the present study indicate that the developed acetylated APTS-coated Fe3O4 NPs have a potential application in MR imaging.

A comparison of 3D-CTA and 4D-CE-MRA for the dynamic monitoring of angiogenesis in a rabbit VX2 tumor.

PURPOSE To compare three-dimensional computed tomography angiography (3D-CTA) and four-dimensional contrast-enhanced magnetic resonance angiography (4D-CE-MRA) for the in vivo monitoring of tumor angiogenesis. MATERIALS AND METHODS VX2 tumors were implanted into the right thigh muscle of 30 New Zealand white rabbits. The animals were randomly assigned to 5 groups, which, respectively, were scanned by 3D-CTA and 4D-CE-MRA on day 4, 7, 10, 13, or 16 after tumor implantation. After scanning, tumors were resected and processed for conventional histology and CD-31 immunohistochemistry. Tumor volume measurements derived from CT and MR imaging were compared with histopathological data. The minimum tumor diameter and the number of new tumor blood vessels detectable by 3D-CTA and 4D-CE-MRA were also compared. RESULTS There were no significant differences in the tumor volume measurements derived from CT, MR, and histological analysis. The minimum diameter of tumor vessels detectable by 3D-CTA (0.68 ± 0.07 mm) was significantly less than that by 4D-CE-MRA (0.85 ± 0.12 mm) (P=0.005). The number of tumor vessels detected by each imaging method was not significantly different until day 13 after implantation, when 3D-CTA detected a greater number (P<0.001). The morphologic process of tumor angiogenesis was demonstrated dynamically by 3D-CTA and 4D-CE-MRA in vivo. CONCLUSIONS Tumor angiogenesis can be dynamically monitored in vivo by 3D-CTA and 4D-CE-MRA. Of the two methods, 3D-CTA has better spatial resolution, but 4D-CE-MRA allows temporal resolution of tumor angiogenesis.

Combination of vascular endothelial growth factor antisense oligonucleotide therapy and radiotherapy increases the curative effects against maxillofacial VX2 tumors in rabbits.

PURPOSE To study the effects of combination of vascular endothelial growth factor (VEGF) antisense oligonucleotide therapy and radiotherapy on maxillofacial VX2 tumors in rabbits. METHODS We used 24 New Zealand white rabbits as a model to induce maxillofacial VX2 tumor. The rabbits were randomly divided into the following 4 groups: radiotherapy group (group A), treated with 16 Gy of radiotherapy; VEGF antisense oligonucleotide treatment group (group B), treated with an injection of 150 μg of VEGF antisense oligonucleotide into the local tumor; VEGF antisense oligonucleotide combined with radiotherapy group (group C), treated with an injection of 150 μg of VEGF antisense oligonucleotide into the local tumor immediately after 16 Gy of radiotherapy; and control group (group D), treated with an injection of 300 μl 5% aqueous glucose solution into the local tumor. On days 3 and 14 after treatment, dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) was performed to calculate maximal enhancement ratio (MER), slope of enhancement (SLE), and tumor volume change. Rabbits were killed on day 14 to obtain samples for pathological examination and immunohistochemical staining for VEGF. RESULTS In group C, tumor volume was significantly reduced on day 14 after treatment, and the difference was statistically different as compared to that before treatment, on day 3 after treatment and other groups (P < 0.01). Values of both MER and SLE after treatment were significantly lower than the values before treatment (P < 0.05). Pathological specimen revealed tumor cell edema, bleeding, necrosis, vascular wall thickening and occlusion, and decreased VEGF expression. The immunohistochemical score (IHS) of group C was significantly different from groups A and D respectively (P < 0.05). CONCLUSION Injecting the tumor with VEGF antisense oligonucleotide immediately after radiotherapy can enhance the curative effect on rabbit maxillofacial VX2 tumor, and DCE-MRI can serve as a reliable technique for in vivo monitoring.