Growth Inhibition Effect and Validation of Stable Expressed Reference Genes for Near-Infrared Photothermal Therapy Mediated by Gold Nanoflower Particles on Human Tongue Carcinoma Tca-8113 Cells.

Abstract

Compared with surgery, radiotherapy, chemotherapy and other conventional cancer treatments, near-infrared (NIR) photothermal therapy mediated by gold nanoparticles had advantages of non-invasive, safe, non-drug resistance and small side effects. Especially for tongue cancer and other head and neck tumors, NIR gold nanoparticle-mediated photothermal therapy is more conducive to preserve the anatomic and functional integrity of stomatognathic system, improving the life quality of patients. However, the toxic organic phase substances are commonly used in conventional gold nanoparticles preparation process in order to increase the stability of the material which limited its application in the biomedical field. In this study, a new type of gold nanoflower (GNF) particles were synthesized by seeding approach in aqueous phase, with better biocompatibility and free of organic substances. Then, the photo-thermal conversion curve under the irradiation of NIR laser ray were investigated. The absorption peak of prepared GNF particles was 719 nm and they had high efficient photothermal conversion in near-infrared region. Furthermore, we investigated the inhibition effect of NIR photothermal therapy with different laser power mediated by the GNF particles on human tongue carcinoma cells by detecting the relative growth rate, cell cycle, and apoptosis and necrosis. The results showed that NIR photothermal therapy mediated by prepared GNF particles could inhibit proliferation of tongue carcinoma Tca-8113 cells obviously, and the relative growth rate of Tca-8113 cells decreased along with the increasing of GNF concentration and laser power. At last, we validated the expression stabilities of 12 commonly used reference genes (18S, ACTB, ALAS1, GAPDH, TBP, HPRT1, RPL29, HMBS, PPIA, PUM1, GUSB, and B2M) after the above treatment through real-time quantitative polymerase chain reaction (RT-qPCR). Three commonly used software, geNorm, NormFinder, and BestKeeper are used to compare stability between reference genes. The final ranking of the candidate reference genes expression stability was TBP > PPIA > RPL29 > B2M > 18S rRNA > GAPDH > GUSB > ACTB > HPRT1 > ALAS1 > HMBS > PUM1. The combination of TBP + PPIA is best for normalizing the relative quantitation. The results suggested that GNF particles had high photothermal conversion in NIR region and was ideal mediated material for tumor NIR photothermal therapy. NIR photothermal therapy mediated by GNFs could inhibit the growth rate of tongue carcinoma Tca-8113 cells obviously. The recommend reference genes can be used for improving the accuracy of gene expression studies in the molecular mechanism of near-infrared (NIR) photothermal therapy mediated by the gold nanoflower (GNF) particles.