Effect of Photodynamic Therapy on Cyclin D1 and P53 mRNA Levels in Head and Neck Squamous Cell Carcinoma Cell Line

Abstract

Background: Photodynamic Therapy (PDT) is considered as one of the alternative treatments for Head and Neck Squamous Cell Carcinoma (HNSCC). P53 mRNA is a tumor suppressor gene whose mutation increases the likelihood of uncontrolled cancer-like divisions. The mutation of CCND1 oncogenic increases the production of cyclin D1 as a tumorigenic protein. Objectives: This study aimed to determine the effect of PDT using toluidine blue as a photosensitizer on the CCND1 and P53 mRNA levels in the HNSCC cell line. Methods: A human HNSCC cell line from NCBI.C196 designation HN5 was used. Cells were divided into four groups: Group I (HNSCC cell line under the influence of toluidine blue and laser irradiation), group II (HNSCC cell line under the influence of toluidine blue), group III (HNSCC cell line under laser irradiation), group IV (control group, HNSCC cell line). A 660 nm THOR laser using toluidine blue as a photosensitizer was applied. The RNA extraction was performed in three steps, including cell degradation, purification, and precipitation by alcohol. The cDNA was prepared using Takara Kit. The Amplicon kit and Real-Time PCR analysis were used for the assessment of mRNA relative levels. Results: The P53 mRNA relative levels were 1.50 ± 0.33 in group I (P = 0.65), 1.49 ± 0.23 (P =0.5) in group II, and 1.40 ± 1.05 (P = 0.63) in group III. Compared to the control group, the mean increases in CCND1 mRNA were 18.01 ± 3.37 (p=0.04) in group I, 17.69 ± 3.3 (P = 0.03) in group II, and 9.01 ± 6.17 (P = 0.20) in group III. Conclusions: The comparison of the fold change index for P53 and CCND1 mRNA by the one-way ANOVA test showed that despite the increased expression of P53 and CCND1 mRNA in treatment groups compared to the control group, there was no statistically significant difference between the increases in P53 mRNA (P = 0.99) and CCND1 mRNA (P = 0.22) index between the groups. The results of this study could be a starting point for a better understanding of the mechanism of genes in PDT of the HNSCC cell line.