Evaluation Of The Effect Of Nisin Potentiated With Hyperthermia On Drug-Induced Cell Death Of Squamous Cell Carcinoma Cell Line

Abstract

Head and neck squamous cell carcinoma (HNSCC) is one of the leading cancers by incidence and is one of the main causes of death in the world. It is most commonly associated with some environmental and behavioral risk factors, including smoking, alcohol consumption, irradiation, few chemicals used in certain workplaces and some viruses, such as human papilloma virus. The prognosis for patients with head and neck squamous cell carcinoma is determined by early diagnosis and the extent of the tumor. Treatment of HNSCC requires balance between complete eradication of cancer and preservation of the surrounding normal tissue. Chemotherapeutic agents are cytotoxic agents used alone or in combination with other treatment modalities. There are several limitations for the chemotherapeutic agents including their bioavailability, metabolism and toxic side effects. So the need for treatment options that improve the clinical outcome and have less toxicity profile is mandatory. This study was conducted to evaluate the cytotoxic effect of the commonly used food preservative Nisin, an antimicrobial peptide, on squamous cell carcinoma cell line, by examining its antitumor potential. Also, hyperthermia was tested as an adjuvant therapy for Nisin to determine whether it could improve the apoptotic effect of Nisin on the cancer cells. This apoptotic effect was evaluated by marking the level of the apoptotic gene (CHAC1) in treated cancer cells. In this study, the apoptotic effect of Nisin was tested by evaluating its cytotoxic effect on laryngeal Hep2 cell line using three concentrations. Hyperthermia was added to the cells before and after treatment, to determine whether it had a potentiating cytotoxic effect on the cancer cells. RT-PCR was the technique used to detect CHAC1 gene expression in treated and untreated Hep2 cell line owing to the sensitivity of this test. Results of this study showed that the addition of Nisin to Hep2 cell line was accompanied by an increase in the values of cell death when compared to the control untreated Hep2 cells. In both durations 24 and 48 hours, the cytotoxic effect of Nisin was in a concentration dependent manner. That is, upon increasing the dose of Nisin applied to the cancer cells from 20µg/ml to 80µg/ml, a significant decrease in the viable cells was noticed. Regarding the use of hyperthermia in our present study, results showed that on application of hyperthermia alone on the cell line, there was a cytotoxic effect of hyperthermia when compared with the control cells in both durations. Also, a difference was achieved on comparing the apoptotic CHAC1 gene expression in control cells and cells treated with hyperthermia in both durations. These results support the fact that application of hyperthermia, to a specific level, is apoptotic to cancer cells. Also on comparing the effect of the three concentrations of Nisin used in this study on CHAC1 expression in Hep-2 cell line used separately with their effect upon application of hyperthermia for 24 and 48 hours, an obvious difference was noticed indicating the possible enhancing effect of hyperthermia on the apoptotic effect of Nisin and the potential use of the agent with hyperthermia as an adjuvant therapy in cancer treatment. A difference was also noticed in the values of gene expression when comparing the values achieved by application of Nisin with hyperthermia on the cancer cells in 24 hours and those achieved for the same groups in 48 hours. Again, this finding may indicate an important role of the duration of application of Nisin on its apoptotic effect on cancer cells. It was concluded that Nisin could be considered a potent anticancer agent inducing apoptosis in low doses that are proven to be safe for human intake. The combination of Nisin with hyperthermia showed a synergistic effect when compared to the effect of Nisin applied alone and the effect of application of hyperthermia separately on the cancer cells.