Synthesis, Characterization, In Vitro Phantom Imaging, and Cytotoxicity of A Novel Graphene-Based Multimodal Magnetic Resonance Imaging - X-Ray Computed Tomography Contrast Agent.

Graphene nanoplatelets (GNPs), synthesized using potassium permanganate-based oxidation and exfoliation followed by reduction with hydroiodic acid (rGNP-HI), have intercalated manganese ions within the graphene sheets, and upon functionalization with iodine, show excellent potential as biomodal contrast agents for magnetic resonance imaging (MRI) and computed tomography (CT). Structural characterization of rGNP-HI nanoparticles with low- and high-resolution transmission electron microscope (TEM) showed disc-shaped nanoparticles (average diameter, 200 nm, average thickness, 3 nm). Energy dispersive X-ray spectroscopy (EDX) analysis confirmed the presence of intercalated manganese. Raman spectroscopy and X-ray diffraction (XRD) analysis of rGNP-HI confirmed the reduction of oxidized GNPs (O-GNPs), absence of molecular and physically adsorbed iodine, and the functionalization of graphene with iodine as polyiodide complexes (I3- and I5-). Manganese and iodine content were quantified as 5.1 ± 0.5 and 10.54 ± 0.87 wt% by inductively-coupled plasma optical emission spectroscopy and ion-selective electrode measurements, respectively. In vitro cytotoxicity analysis, using absorbance (LDH assay) and fluorescence (calcein AM) based assays, performed on NIH3T3 mouse fibroblasts and A498 human kidney epithelial cells, showed CD50 values of rGNP-HI between 179-301 µg/ml, depending on the cell line and the cytotoxicity assay. CT and MRI phantom imaging of rGNP-HI showed high CT (approximately 3200% greater than HI at equimolar iodine concentration) and MRI (approximately 59% greater than equimolar Mn2+ solution) contrast. These results open avenues for further in vivo safety and efficacy studies towards the development of carbon nanostructure-based multimodal MRI-CT contrast agents.

[1]  Riichiro Saito,et al.  Raman spectroscopy of carbon nanotubes , 2005 .

[2]  Zhuang Liu,et al.  Nano-graphene oxide for cellular imaging and drug delivery , 2008, Nano research.

[3]  Sunny C. Patel,et al.  Fabrication and Characterization of Three-Dimensional Macroscopic All-Carbon Scaffolds. , 2013, Carbon.

[4]  A. Mikos,et al.  Tungsten disulfide nanotubes reinforced biodegradable polymers for bone tissue engineering. , 2013, Acta biomaterialia.

[5]  Xiaogang Qu,et al.  Long-circulating Gd(2)O(3):Yb(3+), Er(3+) up-conversion nanoprobes as high-performance contrast agents for multi-modality imaging. , 2013, Biomaterials.

[6]  Morteza Mahmoudi,et al.  Engineered nanoparticles for biomolecular imaging. , 2011, Nanoscale.

[7]  Mauro Ferrari,et al.  Geometrical confinement of gadolinium-based contrast agents in nanoporous particles enhances T1 contrast , 2010, Nature nanotechnology.

[8]  Jan Grimm,et al.  An X-ray computed tomography imaging agent based on long-circulating bismuth sulphide nanoparticles , 2006, Nature materials.

[9]  Hui‐Ming Cheng,et al.  The reduction of graphene oxide , 2012 .

[10]  A. Panich,et al.  Paramagnetic Impurities in Graphene Oxide , 2013 .

[11]  T. Button,et al.  Physicochemical characterization of a novel graphene-based magnetic resonance imaging contrast agent , 2013, International journal of nanomedicine.

[12]  K. Bolotin,et al.  Three-dimensional graphene foams promote osteogenic differentiation of human mesenchymal stem cells. , 2013, Nanoscale.

[13]  Z. Xia,et al.  X-ray diffraction patterns of graphite and turbostratic carbon , 2007 .

[14]  M. Fleischmann,et al.  X-ray diffraction from adsorbed iodine on graphite , 1980, Nature.

[15]  Xiaogang Qu,et al.  Hybrid mesoporous gadolinium oxide nanorods: a platform for multimodal imaging and enhanced insoluble anticancer drug delivery with low systemic toxicity , 2012 .

[16]  Weihong Tan,et al.  Synthesis and Characterization of Fluorescent, Radio‐Opaque, and Paramagnetic Silica Nanoparticles for Multimodal Bioimaging Applications , 2005 .

[17]  Zhen Yao,et al.  Catalyst-free synthesis of iodine-doped graphene via a facile thermal annealing process and its use for electrocatalytic oxygen reduction in an alkaline medium. , 2012, Chemical communications.

[18]  G. Tracton,et al.  Image registration: an essential part of radiation therapy treatment planning. , 1998, International journal of radiation oncology, biology, physics.

[19]  A. Mikos,et al.  Two-dimensional nanostructure-reinforced biodegradable polymeric nanocomposites for bone tissue engineering. , 2013, Biomacromolecules.

[20]  A. Mikos,et al.  Bismuth@US-tubes as a Potential Contrast Agent for X-ray Imaging Applications. , 2013, Journal of materials chemistry. B.

[21]  Scott C. Brown,et al.  Multimodal nanoparticulate bioimaging contrast agents. , 2010, Methods in molecular biology.

[22]  M. Dresselhaus,et al.  Stacking nature of graphene layers in carbon nanotubes and nanofibres , 1997 .

[23]  M van Herk,et al.  Definition of the prostate in CT and MRI: a multi-observer study. , 1999, International journal of radiation oncology, biology, physics.

[24]  Vladimir P Torchilin,et al.  Multifunctional nanocarriers. , 2006, Advanced drug delivery reviews.

[25]  A. Alexandrov,et al.  Advanced multimodal CT/MRI approaches to hyperacute stroke diagnosis, treatment, and monitoring , 2012, Annals of the New York Academy of Sciences.

[26]  Zhuang Liu,et al.  Graphene-based magnetic plasmonic nanocomposite for dual bioimaging and photothermal therapy. , 2013, Biomaterials.

[27]  Jagdish Singh,et al.  Iodinated Contrast Media and Their Adverse Reactions* , 2008, Journal of Nuclear Medicine Technology.

[28]  M. Mahmoudi,et al.  Graphene: promises, facts, opportunities, and challenges in nanomedicine. , 2013, Chemical reviews.

[29]  Zhuang Liu,et al.  Graphene based gene transfection. , 2011, Nanoscale.

[30]  Xin Lu,et al.  Fast and Facile Preparation of Graphene Oxide and Reduced Graphene Oxide Nanoplatelets , 2009 .

[31]  Dar-Bin Shieh,et al.  In vitro and in vivo studies of FePt nanoparticles for dual modal CT/MRI molecular imaging. , 2010, Journal of the American Chemical Society.

[32]  Andre K. Geim,et al.  Raman spectrum of graphene and graphene layers. , 2006, Physical review letters.

[33]  Tae Sup Lee,et al.  Amphiphilic polymer-coated hybrid nanoparticles as CT/MRI dual contrast agents , 2011, Nanotechnology.

[34]  P. Avti,et al.  Luminescent single-walled carbon nanotube-sensitized europium nanoprobes for cellular imaging , 2012, International journal of nanomedicine.

[35]  A. Panich,et al.  Magnetic resonance evidence of manganese-graphene complexes in reduced graphene oxide , 2012 .

[36]  J. Mintorovitch,et al.  Comparison of Magnetic Properties of MRI Contrast Media Solutions at Different Magnetic Field Strengths , 2005, Investigative radiology.

[37]  Xin Cai,et al.  Graphene-based contrast agents for photoacoustic and thermoacoustic tomography☆ , 2013, Photoacoustics.

[38]  David A Jaffray,et al.  Multimodal Contrast Agent for Combined Computed Tomography and Magnetic Resonance Imaging Applications , 2006, Investigative radiology.

[39]  H. Lien,et al.  Enhanced dehalogenation of halogenated methanes by bimetallic Cu/Al. , 2002, Chemosphere.

[40]  Meyya Meyyappan,et al.  Nanotechnology: Opportunities and Challenges , 2003 .

[41]  Jonas Björk,et al.  Gadolinium contrast media are more nephrotoxic than iodine media. The importance of osmolality in direct renal artery injections , 2006, European Radiology.

[42]  Ji Liang,et al.  Crystallization behavior of the amorphous carbon nanotubes prepared by the CVD method , 2001 .

[43]  Omid Akhavan,et al.  Size-dependent genotoxicity of graphene nanoplatelets in human stem cells. , 2012, Biomaterials.

[44]  Bhavna S. Paratala,et al.  Physicochemical Characterization, and Relaxometry Studies of Micro-Graphite Oxide, Graphene Nanoplatelets, and Nanoribbons , 2012, PloS one.

[45]  Kai Yang,et al.  Multimodal Imaging Guided Photothermal Therapy using Functionalized Graphene Nanosheets Anchored with Magnetic Nanoparticles , 2012, Advanced materials.

[46]  P. Couvreur,et al.  Nanoparticles in cancer therapy and diagnosis. , 2002, Advanced drug delivery reviews.

[47]  Balaji Sitharaman,et al.  Gadofullerenes and Gadonanotubes: A New Paradigm for High-Performance Magnetic Resonance Imaging Contrast Agent Probes , 2007 .

[48]  Krause.  Delivery of diagnostic agents in computed tomography. , 1999, Advanced drug delivery reviews.

[49]  L. Wilson,et al.  Preparation of I2@SWNTs: synthesis and spectroscopic characterization of I2-loaded SWNTs. , 2006, The journal of physical chemistry. B.

[50]  Vivek B Shenoy,et al.  Structural evolution during the reduction of chemically derived graphene oxide. , 2010, Nature chemistry.

[51]  Sachio Kuribayashi,et al.  Virtual monochromatic spectral imaging with fast kilovoltage switching: improved image quality as compared with that obtained with conventional 120-kVp CT. , 2011, Radiology.

[52]  M. Umeno,et al.  Iodine doping in solid precursor-based CVD growth graphene film , 2011 .

[53]  Xiaogang Qu,et al.  PEGylated hybrid ytterbia nanoparticles as high-performance diagnostic probes for in vivo magnetic resonance and X-ray computed tomography imaging with low systemic toxicity. , 2013, Nanoscale.

[54]  松本 一宏,et al.  Virtual monochromatic spectral imaging with fast kilovoltage switching : improved image quality as compared with that obtained with conventional 120-kVp CT , 2011 .

[55]  Chenjie Xu,et al.  Size and Concentration Effect of Gold Nanoparticles on X-ray Attenuation As Measured on Computed Tomography. , 2008, Chemistry of materials : a publication of the American Chemical Society.

[56]  M. Ferrari.  Cancer nanotechnology: opportunities and challenges , 2005, Nature Reviews Cancer.

[57]  Francis Vocanson,et al.  Gadolinium chelate coated gold nanoparticles as contrast agents for both X-ray computed tomography and magnetic resonance imaging. , 2008, Journal of the American Chemical Society.

[58]  Hui-Ming Cheng,et al.  Direct reduction of graphene oxide films into highly conductive and flexible graphene films by hydrohalic acids , 2010 .

[59]  G. Lalwani,et al.  MULTIFUNCTIONAL FULLERENE- AND METALLOFULLERENE-BASED NANOBIOMATERIALS , 2013 .

[60]  S. Stankovich,et al.  Synthesis of graphene-based nanosheets via chemical reduction of exfoliated graphite oxide , 2007 .

[61]  B. Sitharaman,et al.  The magnetic, relaxometric, and optical properties of gadolinium-catalyzed single walled carbon nanotubes. , 2013, Journal of applied physics.

[62]  S. Kanakia,et al.  The effects of graphene nanostructures on mesenchymal stem cells. , 2014, Biomaterials.

[63]  Shelton D Caruthers,et al.  Revisiting an old friend: manganese-based MRI contrast agents. , 2011, Wiley interdisciplinary reviews. Nanomedicine and nanobiotechnology.

[64]  B. Sitharaman,et al.  Gadonanotubes as new high-performance MRI contrast agents , 2006, International journal of nanomedicine.

[65]  H. Krug,et al.  Oops they did it again! Carbon nanotubes hoax scientists in viability assays. , 2006, Nano letters.