Effects Of Arsenic Trioxide On The Apoptosis And Proliferation Of Human Lung Cancer Cell At Hypoxia

【Background and Objective】Lung cancer is one of the most serious malignancies with a rapidly growing morbidity and mortality. It has posed great threaten on human being's health in recent 100 years. According to some reports, there are about one million newly diagnosed lung cancer patients in the world every year. In China, the incidence of lung cancer has grown about 11.9% year by year from 1973 to 1990. It is estimated that the total death caused by lung cancer will reach 90 million every year by 2025 in China. Report by the WHO, lung cancer and AIDS will be the most serious health problems in the 21st century .The treatment of lung cancer has been from surgery or radiotherapy access to surgery based multimodality therapy. However, common chemotherapy drugs such as paclitaxel, docetaxel, oxaliplatin, gemcitabine, etc. can not distinguish tumor cells from normal cells. So they have relatively strong toxicity. Targeting treatment, which is recently emerged, has small toxicity, but these drugs are very expensive and have relatively low efficiency. So it is significant to look for a new anticancer drug which has low toxicity and high efficiency.Arsenic trioxide (As₂O₃) comes from traditional Chinese drug. Arsenic apparently affects numerous intracellular signal transduction pathways and causes many alterations in cellular functions, resulting in the induction of apoptosis, the inhibition of growth and angiogenesis, and the promotion of differentiation. Recently, it has been successfully employed in the treatment of acute promyelocytic leukemia (APL) in China where several reports have shown that low doses of As₂O₃ can with minimal toxicity induce complete remission in patients with relapsed APL. Moreover, the potency of As₂O₃ to induce cell death in vitro has been shown in a variety of cancer cells including non-APL leukemia and lymphoma cells as well as solid tumors like tumors originating from prostate, kidney, liver and bladder.Hypoxic microenvironment is frequently found in solid tumors as a result of inefficient vascular supply and high oxygen consumption of rapidly proliferating malignant cells. Tumor hypoxia is associated with malignant progression, lower sensitivity to chemotherapy and radiotherapy, increased metastatic potential, and poor prognosis [2,3]. So it is therefore of great interest to evaluate the efficiency of As₂O₃ treatment of malignant tumors at both normoxia and hypoxia. According to some researches, some tumor cells are responsive to arsenic trioxide at both normoxia and hypoxia [4,5,6,7]. It is significant to discuss the efficiency of As₂O₃ treatment of lung cancer at hypoxia.We apply three different concentrations of As₂O₃ to the human lung cancer cell A549 at both normoxia and hypoxia to find the effects of arsenic trioxide on the apoptosis and proliferation of human lung cancer cell at hypoxia.【Experimental content】1. Effects of arsenic trioxide on the apoptosis and proliferation of human lung cancer cell at normoxiaMaterial and method: We used the human lung cancer A549 cell line in this study. The A549 cell line was grown in F12K. The media were supplemented with 10% FCS, penicillin (100 units/mL), and streptomycin (100μg/mL). The cells were cultured at 37°C in a 95% air/5% CO2 humidified incubator. Three kinds of As₂O₃ with different concentrations were used(1umol/L,2 umol/L and 4umol/L)to treat the cells which were in a logarithmic growth phase and was observed at 12h ,24h and 48h. MTT assay and FCM were used to detect apoptosis in A549 cells.Result:1. MTT assay : There were significant differences (P﹤0.05)of the growth inhibition ratio between As₂O₃ and control groups. There were also significant differences (P﹤0.05)of the growth inhibition ratio in different concentration As₂O₃ groups and different time groups. As₂O₃ could inhibit growth of human lung cancer cell A549 at normoxia. 2. Apoptosis Detection: There were significant differences (P﹤0.05)of the apoptotic ratio between As₂O₃ and control groups. There were also significant differences (P﹤0.05)of the apoptotic ratio in different concentration As₂O₃ groups and different time groups. As₂O₃ induce apoptosis of human lung cancer cell A549 at normoxia.. It was obvious that apoptosis became more marked along with the prolongation of reaction and the increasing of the concentration of the chemical agents as well. 2. Effects of arsenic trioxide on the apoptosis and proliferation of human lung cancer cell at hypoxiaMaterial and method: We used the human lung cancer A549 cell line in this study. The A549 cell line was grown in F12K. The media were supplemented with 10% FCS, penicillin (100 units/mL), and streptomycin (100μg/mL). The cells were cultured at 37°C in a 95% air/5% CO₂ humidified incubator. Hypoxic conditions, 5% O₂, were created by Genbag Microaer (BioMerieux Technology, France). Three kinds of As₂O₃ with different concentrations were used(1umol/L,2 umol/L and 4umol/L)to treat the cells which were in a logarithmic growth phase and was observed at 24h ,48h and 72h. MTT assay and FCM were used to detect apoptosis in A549 cells.Result:1. MTT assay : There were significant differences (P﹤0.05)of the growth inhibition ratio between As₂O₃ and control groups. There were also significant differences of the growth inhibition ratio in different concentration As₂O₃ groups and different time groups. As₂O₃ could inhibit growth of human lung cancer cell A549 at hypoxia. But there were not great significant differences of the growth inhibition ratio(P>0.05) between hypoxia and normoxia groups. 2. Apoptosis Detection: There were significant differences (P﹤0.05)of the apoptotic ratio between As₂O₃ and control groups. There were also significant differences (P﹤0.05)of the apoptotic ratio in different concentration As₂O₃ groups and different time groups. As₂O₃ induce apoptosis of human lung cancer cell A549 at hypoxia. It was obvious that apoptosis became more marked along with the prolongation of reaction and the increasing of the concentration of the chemical agents as well. But there were not great significant differences of the apoptotic ratio(P>0.05)between hypoxia and normoxia groups. 【Conclusion】1.As₂O₃ could inhibit the proliferation and induce apoptosis of human lung cancer cell A549. It was obvious that the efficiency of As2O3 became more marked along with the prolongation of reaction and the increasing of the concentration of the chemical agents as well.2.Human lung cancer cell A549 was responsive to arsenic trioxide at both normoxia and hypoxia. There were not great significant differences of the efficiency of As2O3 between hypoxia and normoxia groups.3.At hypoxia and normoxia after 12 hours of treatment with 1μmol/L As2O3, the cultures could display high sensitive to As₂O₃.4.As2O3 was a potential candidate drug in treatment of solid tumors with hypoxic areas like lung cancer.