The Preliminary Study Of The Effect And Mechanism Of Metastasis And Drug Resistance Involved With XCT In Nasopharyngeal Carcinoma

Background & Objective:Nasopharyngeal Carcinoma(NPC)is one of the common malignant tumors that seriously damage human health in south China. Because the primary anatomical site of tumor growth is located in a cryptic area, with slightly early symptoms, the NPC patients tend to present a more advanced stage of disease in clinic with higher metastatic potential. Therefore, as much as 60-85% of NPC patients already have clinically detectable aggressive metastasis in the regional lymph nodes.The main treatment of nasopharyngeal carcinoma is radiotherapy and it is often combined with chemotherapy to improve prognosis. But in the course of therapy, nasopharyngeal carcinoma often gives rise to drug resistance. However,the function and mechanism of NPC's invasion and dug risistance are still unclear. xCT is the functional subunit of the cystine/glutamate transporter Xc-system and it plays a critical role in the maintenance balance of intracellular glutathione and redox.In recent years, more and more research shows that xCT is closely related to proliferation, drug resistance and metastasis of many tumor cells,but the relationship with NPC has not reported yet.So our study is focused on the role of xCT in the metastasis and drug resistance of NPC to provide new evidences for the mechanism of NPC's metastasis and drug resistance and to enhance effects of chemotherapy.Methods:(1) Inununochemistry was used to detect the expression of xCT in normal nasopharyn membran,nasopharyngeal carcinoma with lymph nodes metastasis and nasopharyngeal carcinoma without lymph nodes metastasis;(2) Fluorescence quantitative PCR and western blot were used to detect the xCT expression of 5-8F and 6-10B; (3)To treat 5-8F with SASP,an inhibitor of xCT, then MTS assay and plate colony formation assay were used to detect the effect of xCT on cell proliferation; (4)To construct xCT eukaryotic expression vector transient transfecting 6-10B cell line.To treat 5-8F with SASP and xCT antisense oligodeo-xynucleotide (ASO). Scratching assay and transwell method were employed to detect the cell metastasis and invasion abilities.Cell adhesion assay was used to detect cell-extracellular matrix adhesion.MTS assay was used to detect the sensitivities of all the cell lines to Cisplatin(cDDP), Pingyangmycin(PYM) and 5-fluorouracil(5-Fu).The expressions of MMP1 and cyclinD1 of all the cell lines were surveyed by western blot.Results:(1)The positive expression of xCT in normal nasopharyn membran was 10%, which was obviously lower than that of nasopharyngeal carcinoma without lymph node metastasis(35%).The positive expression rate of xCT in nasopharyngeal carcinoma with lymph node metastasis was 85.7%,which was obviously higher than that of nasopharyngeal carcinoma without lymph node metastasis; (2)The results of fluorescence quantitative PCR and Western blot showed that the expression of xCT in 5-8F was higher than 6-1 OB in mRNA and protein levers; (3) The growth inhibition rate of 5-8F cells increased with the SASP concentration increased gradually.When the 5-8F cells were treated with 0.5mM SASP, the colony formation ratio was 1.85 folds over control group(P<0.05); (4)Exogenous overexpression of xCT enhanced metastasis/invasion ability and cell-extracellular matrix adhesion of 6-1 OB cells.Upregulation of xCT resulted the resistance of 6-10B to cDDP was 3.98 folds over control group respectively (P<0.05). (5)Silencing and inhibition of xCT decreased the metastasis/invasion abilities and cell-extracellular matrix adhesion of 5-8F.The sensitivity of 5-8F/xCT ASO and 5-8F/SASP cells to cDDP was 1.91 and 1.99 folds over corresponding control groups (P<0.05). (6)The expressions of MMP1 and cyclinD1 proteins in 5-8F were higher than 6-10B,and they were positively correlated with the expression of xCT.Conclusion:(1)xCT is related to the metastasis/invasion and proliferation abilities of nasopharyngeal carcinoma.(2)xCT of nasopharyngeal carcinoma can induce resistance to cisplatin. (3)xCT may effect proliferation and matastasis/invasion of NPC cells by regulating the expression of cyclinDl and MMP1.