Study On Biotoxicity Of Graphene-titanium Dioxide Composites

Objective:To investigate the toxicity of graphene-titanium dioxide composites(GR-Ti O2), lysozyme(LYSO) was used as model protein and human liver cells(HL-7702) as cell model, Ti O2 as the control, the effect of GR-Ti O2 on the activity of LYSO and toxicity effect of HL-7702 cell was studied, the toxicity mechanism of GR-Ti O2 for HL-7702 cell was initially explored. To examine the anti-tumor activity of photocatalytic GR-Ti O2 in vivo, xenograft model of cervical cancer in nude mice was established, the effect of photocatalytic GR-Ti O2 on the growth of cervical cancer cells in vivo was investigated. Methods:Ti O2 as the control, the interaction of GR-Ti O2 with lysozyme(LYSO) was investigated by ultraviolet spectroscopy, fluorescence spectroscopy, synchronous fluorescence spectroscopy, three-dimensional fluorescence spectroscopy and circular dichroism, the effect of GR-Ti O2 on enzyme activity of LYSO was determined by enzyme activity experiment. HL-7702 Human liver cell line was employed to examine the cytotoxicity of GR-Ti O2 and compare with Ti O2 using MTT assay. AO/EB double staining method was used to explore the cytotoxicity mechanism of GR-Ti O2, confocal laser technology was applied to study the mode of GR-Ti O2 causing cytotoxicity. To investigate the inhibition of photocatalytic GR-Ti O2 and Ti O2 on the growth of cervical cancer cells in vivo, xenograft model of cervical cancer in nude mice was established by subcutaneously inoculating cervical cancer cells(Si Ha) in BALB/C nude mice. Before and after irradiating mice, tumor volume was measured and the effect of photocatalytic GR-Ti O2 on the growth of cervical cancer cells in vivo was observed. Results:The results of ultraviolet spectroscopy and fluorescence spectroscopy revealed that GR-Ti O2 interacted with LYSO. The endogenous fluorescence of LYSO was quenched by GR-Ti O2 and its mechanism was static quenching. The major driving forces were hydrogen bonds and van der Waals’ forces. These results of GR-Ti O2 were consistent with that of Ti O2. The results of synchronous fluorescence spectroscopy and circular dichroism further demonstrated that the conformation of LYSO was greater influenced by GR-Ti O2. The result of enzyme activity indicated that enzyme activity of LYSO was stronger inhibited by GR-Ti O2. When the concentration of Ti O2 and GR-Ti O2 was 200 mg·m L-1, the cell viability of HL-7702 cells decreased to 76.03% and 73.43% respectively. There was a significant difference between GR-Ti O2 and the control group. It showed that Ti O2 and GR-Ti O2 had obvious cytotoxicity on HL-7720 cells, causing change of cell morphology and inhibiting the cell growth. Under the same condition, the cytotoxicity effect of GR-Ti O2 and Ti O2 was more significant with their concentration increasing and the time interacted with cells prolonging. Confocal laser technology result initially indicated that GR-Ti O2 could enter into the cell through cell membrane. AO/EB double staining result demonstrated that GR-Ti O2 could induce apoptosis in HL-7720 cells. The induction of apoptosis was enhanced with their concentration increasing and the time interacted with cells prolonging. The anti-tumor experiment in vivo showed that the tumor volume of nude mice decreased from 64 mm3 to 51 mm3. Conclusion:Both GR-Ti O2 and Ti O2 could interact with LYSO, GR-Ti O2 stronger binded with LYSO. The conformation and enzyme activity of LYSO were greater influenced and stronger inhibited by GR-Ti O2. Under the same condition, GR-Ti O2 had stronger biological toxicity than that of Ti O2. The toxic effect of GR-Ti O2 and Ti O2 on HL-7720 cells was of dose and time dependent. GR-Ti O2 could induce apoptosis in HL-7720 cells through cell membrane and produce the cytotoxicity. The growth of cervical cancer cells in vivo was inhibited by photocatalytic GR-Ti O2.