| Breast cancer is one of the most common female malignant tumors, and the incidence is trending to rapidly ascend. The onset age of the patient gradually gets younger.It is seriously threatening female health.About one third of breast cancer patients whose estrogen receptor (ER) expression are negative.Compared with breast cancer patients with ER-positive,their diseases are high malignant and prognosis is bad. Although comprehensive treatments with primary surgery integrated with chemotherapy and/or radiotherapy have improved the survival rate of the patients, the recurrence rate and the metastasis rate is still high. Arsenic trioxide, commomly known as Pishuang in China, was used for the treatment of acute promylocytic leukemia in the 70s of 20th centrury. Since then, it has become the investigative hotspot of international medical circles because of its remarkable curative effects and few side effects. More and more researches show arsenic trioxide can inhibit the growth and/or induce the apoptosis of multiple solid tumors besides haemal tumor. But there is relative little research on the use of arsenic trioxide in breast cancer now. We will observe the influence of arsenic trioxide on proliferation, morphology, apoptosis and apoptosis-related genes of estrogen receptor negative human breast cancer cell line MDA-MB-231 and explore the possible mechanism in order to provide experimental basis for possible clinical applications in the future and open up a new therapeutic approach for these patients.ObjectiveTo observe the infact of arsenic trioxide in different concentrations on human breast cancer MDA-MB-231 cell proliferation and apoptosis and discuss the mechanism of apoptosis induced.MethodsMDA-MB-231 cells was incubated with arsenic trioxide in different concentrations for different time points.Then the inhibition of cell growth was estimated by MTT assay; The cell morphologic changes were observed under inverted microscope; The cell apoptosis was detected by terminal deoxynucletidyl transferase (TdT) mediated biotin-dUTP nick end labeling (TUNEL);The expression of apoptosis-associated protein survivin and caspase-3 were analyzed by immunohistochemistry.Results1. Arsenic trioxide in different concentrations could inhibit MDA-MB-231 cell growth in time- and dose- dependent manner in vitro(P<0.001). The 24-hour 50% inhibitory concentration (IC50) of arsenic trioxide is 43.13μM, the 48 h IC50 is 5.64μM and 72h IC50 is 1.96μM.2. MDA-MB-231 cell morphology variation with concentration were observed under the inverted microscope. Cells in the control group (no treated with arsenic trioxide), ovoid or spindle in shape, intercellular tight junction, had transparent and homogeneous cytoplasm and were growing with good adherence. With the increase of treatment concentration, gradually, adherence cells decreased and floating cells increased in the culture flask, cells became round and small, the cytoplasmic granule increased and the intercellular space became large.3. Detecting the cell apoptosis by TUNEL.The result showed under optical microscope:There were positive staining cells both in treatment groups and control (no treated with As2O3).In the control group there were few positive cells. The mean gray scale value of TUNEL positive area was different among every group (P<0.05) and became high with the increase of treatment concentration.4. Immunohistochemical staining result indicated the expression of survivin protein in MDA-MB-231 cell decreased and the caspase-3 increased as the concentration of arsenic trioxide(P<0.05).The correlation analysis indicated that the expression of survivin protein has a negative correlation with caspase-3 (P<0.001).Conclusion1. Arsenic trioxide can inhibit MDA-MB-231 cell proliferation in a dose- and time- dependent manner in vitro.2. Arsenic trioxide can induce the MDA-MB-231 cell apoptosis in vitro, and the apoptosis effect has a positive correlation with the treatment concentration.3. Arsenic trioxide inhibiting survivin protein expression and activating caspase-3 may be one of mechanisms of its inducing MDA-MB-231 cell apoptosis in vitro.
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