Synthesis And Application Of Multifunctional CT/Fluorescent Nano-molecular Probes

In small animal imaging, X-ray computed tomography (CT) imaging and fluorescence molecular imaging each existed advantages and disadvantages, respectively. However, the combination of CT and fluorescence is an ideal bimodal molecular imaging model, which can combine the advantages of molecular functional imaging and tissue imaging perfectly and has a great significance. The CT/fluorescence dual functional agent is the premise and the key point in the combination of these two imaging techniques. At present, the CT/fluorescence probes were synthesized by precious metals or iodine with organic dyes, which had lots of problems in design, preparation and biological application of probes, of Some CT contrast agents had high fluorescence quenching, some probes’size, surface groups and biocompatibility had high toxicity in vivo, the CT angiography and the fluorescence intensity are weak, which were related to the CT contrast agent and the fluorescent compound. Compared to the reported CT contrast agents, Bi has the largest atomic number and X-ray attenuation coefficient, inexpensive raw materials and low toxicity, which imply bismuth (Bi) contrast agent might be the most promising contrast agents. Quantum dots (CdSe/ZnS) is an excellent fluorescent molecular probes, which had more advantages of surface functionalized easily and the high fluorescence yield in the biological applications of dual mode probes. Therefore, the probe had both dual modal CT/fluorescence imaging in vitro and in vivo, which has important clinical value in dual-mode imaging and life sciences. The main results are as following:(1) Probe bismuth sulfide modified with Pluronic F127(Bi2S3-PF127), which has high biocompatibility and dispersibility, is synthesized by using triblock copolymer Pluronic F127to modify hydrophobic Bi2S3nanoparticles that are prepared by a hot injection method. TEM results show that most of the probe has length of about14.85±1.70nm and breadth of about4.79±0.63nm. The stability of B12S3-PF127nanoparticles were evaluated by examining changes of mean particle size, polydispersity index (PDI) and Zeta potential during storage at4-37℃, the results showed that nanopartiles did not any significant changes, which indicated that probes had enough stability.After injected into the tail vein of a mouse, the probe has obvious CT contrast enhancement capability from X-ray CT imaging results. Meanwhile, the probe’s in vivo toxicity is also studied, it is found that hematoxylin and eosin stains of major organs have no change, biochemical analysis (alanine aminotransferase and aspartate aminotransferase) prove the probe has no adverse effect, the results of blood analysis (white blood cell count, red blood cell count, hemoglobin and platelet count) are also normal, the biological distribution of Bi by ICP-AES shows that most of nanoparticles are cleaned out after injection48h, and the circulation half-life of the probe is5.0h suggested that Bi2S3-PF127has a long circulation, indicating that Bi2S3-PF127probe has good biocompatibility and safety(2) Multifunction probe that has CT contrast enhancement and fluorescence imaging capabilities are prepared by one-pot in a reverse microemulsion system. In the system, Bi2S3nanoparticles are prepared firstly and then add with QDs, which are both wrapped in SiO2by hydrolysis of TEOS, lastly50nm diameter and monodispersed Bi2S3-QD@SiO2-PEG nanoparticles modified by PEG-silane are synthesized. Experimental results show that the nanoparticles of15mg/mL Bi give a CT number at700HU, which are higher than that of the commercial current iobitridol CT contrast agent. Furthermore, MTT experiments reveal that cellular survival rate reach80%with a concentration up to of2.70mg/mL Bi for24h. The nanoparticle not only has a longer circulation time (>4h) compared with the medical iodine contrast, but also has obvious CT contrast enhancement and fluorescence imaging capabilities in vitro and in vivo. Therefore, the multifunction probe has a great potential application in CT/fluorescence dual mode imaging.(3) PEG-phospholipids was used to synthesis nanoparticles of50nm diameter that contains hydrophobic bismuth sulfide nanoparticles and CdSe/ZnS quantum dots, and also studied the probe’s in vitro and in vivo toxicity, CT imaging and fluorescence imaging performance. Experimental results showed that the nanocomposite micelle has obvious CT contrast enhancement and fluorescence imaging capability in vitro and in vivo. It gave a CT number of700(HU) at15mg/mL, higher than that of the current iobitridol CT contrast agent. MTT experiments revealed that it had low cytotoxicity at concentration up to of3.14mg/mL of Bi. In vivo, the nanocomposite micelles had a longer circulation time (>4h) compared with iobitridol. We also performed an extensive study on its in vivo toxicity, including histopathological changes, biochemical analysis and blood analysis, which reveal the nanoparticles did not affect normal functioning of organs after injection.In summary, Bi2S3-PF127nanoparticles were synthysized based on the metal bismuth which had low toxicity, good biocompatibility and CT angiography. Bi2S3-QD@Si02-PEG nanoparticles were prepared combined with CdSe/ZnS that had CT/fluorescence imaging capability in vitro and in vivo. We improved the material and the design of probe, prepared Bi2S3-QD@DSPE nanoparticles uising PEG-phospholipids, not only had excellent CT/fluorescence imaging capability in vitro and in vivo, but also had good stability and high biocompatibility in vivo. The study provided the new methods for the dual-mode imaging design, the imaging in vitro and in vivo, evaluation in vivo, the tumor targeting diagnostics and imaging.