In Vitro Differential Sensitivity of Head and Neck Squamous Cell Carcinoma to Cisplatin, Silver Nanoparticles, and Photodynamic Therapy

Abstract

Background: The clinical effect of photodynamic therapy (PDT) may be correlated with the degree of dysplasia of cancer tissues. The aim of this study was to compare the effects of cisplatin, silver nanoparticles (AgNps), and photodynamic therapy (PDT) using methylene blue (MB) photosensitizer on Head and Neck squamous cell carcinoma - cell line (HNSCC), Hep-2, through genes expression.

Methods: Hep-2 cells were divided into four groups: group I as control and without any treatment, group II and III were treated by cisplatin and AgNps, respectively, and group IV were incubated with MB for four minutes followed by PDT using laser irradiation at 650 nm for 8 minutes. The resulting toxicity was assessed in cell lines using MTT cytotoxicity assay. Further, apoptosis and the response to treatment was examined via RT-qPCR.

Results: MB-PDT inhibited the proliferation of Hep-2 cells. Following PDT, compared with AgNps cells and via MTT assay, a highly significant decrease was observed in cell proliferation in Cancer cells treated with AgNps and MB- PDT groups compared to cancer group cells and cancer cells treated with Cisplatin (p value< 0.001). Mechanistically, both the mRNA and protein expression levels of Bcl-2, Caspase-3, Cyclin-D, HIF-1, IL-8, MAPK-38, and ROS were found to be down regulated in Hep-2 cell line after MB-PDT.

Discussion: MB-PDT effectively killed Hep-2 cells in vitro, however, under the same conditions, the susceptibilities of the cell line to cisplatin, AgNps, and MB-PDT were different. Further studies are necessary to confirm whether this difference is present in clinical oral cancer lesions.

Keywords: Head and Neck squamous cell carcinoma (HNSCC); Methylene Blue (MB); Photosensitizer (PS); photodynamic therapy (PDT); silver nanoparticles (AgNps).

Fig. 1

Microscopic pictures of cultured cancer cells; A, Control cancer cells, B, Cells treated by Cisplatin, C, Cells treated by AgNps, and D, Cells treated by PDT.

Fig. 2

Cell proliferation mean levels among different studied groups. Data were expressed as mean±SD, p value< 0.05 was significant. (*) denotes significant difference versus cancer group, (#) denotes significant difference versus Hep-2 + Cisplatin group, ($) denotes significant difference versus Hep-2 +AgNps group.

Fig. 3

A, BCL-2 gene expression among different studied. B, Caspase -3 gene expression among different studied. Data were expressed as mean±SD, p value< 0.05 was significant. (*) denotes significant difference versus cancer group, (#) denotes significant difference versus Hep-2 + Cisplatin group, ($) denotes significant difference versus Hep-2 +AgNps group.

Fig. 4

Cyclin D1 gene expression: mean levels among different studied groups. Data were expressed as Mean ± SD, p value <0.05 was significant. (*) denotes significant difference versus cancer group., (#) denotes significant difference versus Hep-2 + Cisplatin group, ($) Denotes significant difference versus Hep-2 + AgNps group.

Fig. 5

HIF-1gene expression: mean levels among different studied groups. Data were expressed as mean±SD, p value< 0.05 was significant. (*) denotes significant difference versus cancer group, (#) denotes significant difference versus Hep-2 + Cisplatin group, ($) denotes significant difference versus Hep-2 +AgNps group.

Fig. 6

A, MAP Kinase protein level and B, IL-8 protein levels among different studied groups. Data were expressed as Mean±SD, p value< 0.05 was significant, (*) denotes significant difference versus cancer group, (#) denotes significant difference versus Hep-2 + Cisplatin group, ($) denotes significant difference versus Hep-2 +AgNps group.

Fig. 7

Ros level among different study groups. Data were expressed as Mean±SD, p value< 0.05 was significant. (*) denotes significant difference versus cancer group, (#) Denotes significant difference versus Hep-2 + Cisplatin group, ($) Denotes significant difference versus Hep-2 +AgNps group.