Comparative Study Of Three Kinds Of Carbon Nanomaterials Modified By Hyaluronic Acid

Currently, cancer has become one of the leading causes of death, and seriously threated to human health and survival. Because of their optical anticancer activity, carbon nanomaterials have been widely used in biological medicine field, such as single-walled carbon nanotubes, graphene oxide and fullerenes. Wherein, with the perfect photothermal conversion property, the single-walled carbon nanotubes and graphene oxide can be used as photothermal agents, while the fullerenes act as a kind of photosensitizer depending on the photodynamic property. Although these carbon nanomaterials possess several advantages as follows, hollow structure, large surface area, stable physical and chemical properties, good biocompatibility and so on, as drug transporters, they still have a major flaw: poor water-solubility and lack of specific targeting to tumor cells, which leads drugs not to be delivered to the target site. The good news is that, a natural polysaccharide, hyaluronic acid can specifically bind to CD44 recepter, which is overpressed on a variety of tumor cell surface, in order to achieve active targeting of these materials to tumor cells. According to the above description, in this paper, we used hyaluronic acid to modify these carbon nanomaterials, to improve their defects as drug transporting vectors and strengthen their active targeting for tumor.Firstly, we modified single-walled carbon nanotubes, graphene oxide and fullerenes with hyaluronic acid by using ethylene diamine as a tether. The modification was on base of the amidation reaction between a carboxyl group and an amino group. After the synthesis of these targeting carbon nanomaterials, the following characterization was conducted: ultraviolet full wavelength scanning and infrared spectroscopy scanning showed that the modification of carbon nanomaterials by hyaluronic acid was successful. Transmission electron microscopy and energy spectrum analysis proved the above result intuitively. Ultraviolet spectrophotometry determined the modified degree of hyaluronic acid. Photothermal conversion in vitro test demonstrated excellent photothermal conversion property of HA-SWNT and HA-GO.Secondly, the uptake differences of these targeting carbon nanomaterials by tumor cells were compared qualitatively by fluorescence microscopy and laser scanning confocal microscopy and evaluated quantitatively by flow cytometry, the results showed that, the uptake of HA-GO by tumor cells was the fastest, then was HA-C60, the last was HA-SWNT. The reactive oxygen production of these targeting carbon nanomaterials under visible light irradiation was detected with active oxygen kit, the assay demonstrated that under visible light irradiation, among the three targeting carbon nanomaterials, active oxygen yield of HA-C60 was much higher than the others, which proved the good photodynamic therapy effect of HA-C60.The cytotoxic effect of these targeting carbon nanomaterials was measured in MCF-7 using SRB assay. Cytotoxicity experiments confirmed that, in the same concentration, among the three targeting carbon nanomaterials, the cytotoxicity of HA-SWNT was the lowest, then was HA-C60, the highest was HA-GO, which was consistent with the uptake degree of them; while after the laser irradiation, the cell inhibition degree of the above three targeting carbon nanomaterials was as follows: the highest was HA-SWNT, then was HA-GO, the last was HA-C60, which was related to the phototherapy property of them. In addition, the influence of these targeting carbon nanomaterials on tumor cell cycle and apoptosis was verified by flow cytometry. The result showed that these nanomaterials could block cell cycle in G1 phase, and could induce the appotosis of tumor cells.Finally, we used tumor-bearing mice as in vivo tumor model, continuously administrated several times during this period. We measured the weight of mice and the volume of tumors everyday. After the administration, tumor-bearing mice were sacrificed and the major organs and tumor were exteriorized for pathological reseach. Pharmacodynamics in vivo and anti-tumor activity experiments showed that the three targeting carbon nanomaterials and laser irradiation did not affect the physiological function of mice, and the inhibition degree for tumor of these targeting carbon nanomaterials was consistent with the cytotoxicity assay results. Then animal imager was used to observe the tumor targeting property of the targeting carbon nanomaterials.