Evaluate the combination effect of modulated electro-hyperthermia (mEHT) and gold nanoparticles with different shapes in HepG2 hepatoma cell line

• Main Authors: Ya-Yun Chen, 陳雅筠
• Other Authors: Chuan-Lin Chen
• Format: Others
• Language: zh-TW
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/e67bhj

碩士 === 國立陽明大學 === 生物醫學影像暨放射科學系 === 106 === Abstract Purpose: Hyperthermia (HT) has been used as an adjuvant cancer therapy in the clinic. The Modulated electro-hyperthermia (mEHT) is a new loco-regional electromagnetic hyperthermia method for selectively targeting cancer cell hyperthermia treatment. Gold nanoparticles (AuNP) have been demonstrated as a promising nano-drug carrier and several studies indicated that AuNP can promote heat generation as exposed to the radiofrequency with the better result of cell killing effect. Till now, the AuNP was combined with mEHT for cancer cell hyperthermia therapy hasn’t been investigated and might possess synergistic effect for mEHT treatment. In this study, the different shapes (spherical, urchin-like or rod) AuNP were prepared and treated with HepG2 cell line under various HT conditions (including water bath and mEHT) to investigate the differences in the biological effect. Methods: The homemade spherical and urchin-like AuNPs were prepared according to different methods. The homogenous sizes and sharps AuNPs were obtained from the seed-mediated growth method with hydroquinone. Gold nanorod was purchased from Sigma-Aldrich. All nanoparticles were characterized by UV-VIS absorption spectrometry, dynamic light scattering (DLS), transmission electron microscope (TEM) and inductively coupled plasma mass spectrometry (ICP-MS). Oncothermia EHY-100 was used for investigating the heating effect of AuNP with different shapes and the killing effect of HepG2 hepatoma cells after the combination AuNP with mEHT. The cell survival was analyzed by MTT assay and Annexin V staining. Results: Gold nanoparticles were synthesized by Turkevich, seed-growth and hydroquinone reduction method. Among these methods, the seed-mediated growth approach with hydroquinone reduction method had the advantage in controlling particle sizes and sharps. The spherical and urchin-like gold nanoparticles were synthesized by adjusting the amount of hydroquinone. The diameter of ~50 nm spherical and urchin-like gold nanoparticles were obtained from 5x10^11~1x10^12 gold seeds reacted with 254 μmol HAuCl4, 22 nmol sodium citrate and 3 μmol or 30 μmol hydroquinone in 10 ml deionized water, respectively. The heating rate of AuNPs was around 0.4˚C/min by mEHT treatment and no significant differences were observed between deionized water and various sharps AuNPs solutions. After 24 h incubated with 50 nm AuNPs solution, the concentration of 1.56 to 100 ppm showed negligible cytotoxicity to HepG2 cells and no difference was obvious among various shapes of AuNP. HepG2 cells were exposed to HT treatment (water bath and mEHT). Compared with water bath (42˚C) treatment, the mEHT treatment showed significantly reduced survival ratio. While no significant synergistic effect on cell killing of HepG2 cell was noticed in the co-incubation with various AuNPs as mEHT treatment. However, compared to the control group (without AuNP incubation) or the co-incubation groups, the cell-killing effect of mEHT was significantly decreased in those groups which HepG2 cells pre-incubated with AuNPs for 24 h and then exposed to mEHT treatment. The results of annexin-V flow cytometry revealed that percentage of early apoptotic cells significantly reduced. Conclusion: This study has demonstrated that 50 nm of spherical and urchin-like gold nanoparticles could be synthesized by adjusting the reaction concentration of hydroquinone in the hydroquinone reduction method. Compare to water, there is no significant heat generation in different shapes of AuNPs after exposing to mEHT. Similar cell damage effects of mEHT in HepG2 cells were noticed between the control group (without AuNPs) and the co-incubation groups, while high cell survival and low apoptosis ratio was observed in the pre-incubation groups.