The Comparison Of The Proferation And Migration Ability Of Tougue Squamous Cells Before And After The Exposure To Cisplatin

Oral and maxillofacial malignant tumor is one of the most common malignant tumor,and it has ranked sixth in the whole body malignant tumor.So although the treatment of head and neck malignant tumor in recent years has improved the quality life of patients in some sense, but it did not prolong the patient’s long-term survival.The patient’s death is mainly due to the tumor recurrence, cervical lymph node metastasis or distant organ metastasis, thus the formation of tumor and metastasis mechanisms become the research focus.Chemical drugs as an aid itreatment of middle-late cases before and after surgery, can effectively alleviate the growth of tumor, it can also be applied to patients with distant metastasis.However, some patients in still have tumor recurrence and metastasis after treatment, and prognosis is worse.At present, there is no research to compare migration ability and recurrence rate of tumor between the remaining cells after chemotherapy and cells without chemotherapy.To this end, the experimental objective is to study the Nanog and migration ability expression of tongue squamous cancer cells SCC- 15 before and after chemotherapy drugs.Try to explore with cisplatin tongue squamous cancer cells before and after the change of cell migration and proliferation ability.The reasonof choosing chemotherapy drugs and gene is as follows:1.Cisplatin(cisplatin, cisplatin, DDP) belongs to the cycle non-specific drug, as a first-line drug for the treatment of a variety of solid tumors.For multiple squamous cell carcinoma and urothelial carcinoma, such as head and neck, cervical, esophageal, and urinary tract tumors,it’s quite efficient.In 20 century 80 s, cisplatin begins to be applied in head and neck cancer chemotherapy, for head and neck squamous cell carcinomas have remarkable curative effect,and the effect of chemotherapy is better and less side effects than Docetaxe, it is widely used in tongue squamous carcinoma clinical medication.2.Nanog is widely known to participate in the regulation of embryonic stem cell self-renewal in tumor cells. The abnormal expression of Nanog and increasing ability of the cell proliferation in vitro and in vivo tumor growth is related to the ability to actively adjust the tumor cell activity and tumor metastasis.It is the key gene that maintain the stemness of tumor, and it help promote the cancer stem cell sample features of several kinds of cancer.In this paper, related research is divided into two parts:1.The Nanog expression between tongue squamous cancer cells of SCC-15 and 293 cell lines,and detection half inhibitory concentration IC50.Objective: to determine if there were any Nanog expression in SCC-15 cell lines.And compared to normal tissue cell lines, whether Nanog expression in human tongue squamous cancer cells is different.Cell immunofluorescence technique was applied to detect the Nanog expression between the two cell lines.Methods: firstly, immunofluorescence method is applied to detect if there are Nanog protein expression in SCC-15 cell lines and at the same time, comparing with 293 cell line of Nanog expression in comparison.If Nanog expression were observed in SCC-15 cell line, it can continue for subsequent experiments.CCK-8 kit was used to detect different half drug toxicity inhibition concentration(IC50) of cisplatin in SCC-15, to determine experimental conditions of each group.Results: 1.the tongue squamous cancer cells have Nanog expression in the system, and no expression in normal mature differentiated 293 cells, SCC- 15 can be used in the experiments related research.2.By cytotoxicity experiment,the drug toxicity effect inhibition rate of experimental group is calculated out successfully.The IC50 of the 24 h group is about 5 μg/ml, 48 h group is 4 μg/ml, 3.5 μg/ml in 72 h group, to determine the experimental conditions of the experimental group(see table 1.1).2.The second part of cisplatin and cell apoptosis, Nanog expression before and after contrast research migration abilityObjective: to apply various experimental technology to detect whether there are changes in apoptosis ability,Nanog expression,the ability to migrate.Methods: to compare the cisplatin effects before and after the cell apoptosis of SCC to 15,we apply the Annexin- FITC/PI double staining combined with flow cytometry instrument to test the cell apoptosis rate of experimental groups and the control group..q RT- PCR and Westernblot were applied to detect Nanog gene m RNA and protein expression level to quantitative analysis the expression of Nanog of each groups studied.Scratch test was applied to measure the cell migration ability of groups.Results: groups of apoptosis rate results are as follows:24 h group(13.34 + 4.14) %, 48 h group(24.75 + 1.48) %, 72 h group(25.92 + 1.78) %, the control group(6.61 + 0.64) %.Two two compared statistically analyzed that :24 h, 48 h and 72 h, respectively, compared with the control group,p value were 0.035, 0.000 and 0.000,which have statistical difference.Compared with control group, the apoptosis rate were higher in the experimental group respectively.This showed that Cisplatin promotes the apoptosis of cells.q RT- PCR results showed that: the m RNA level of Nanog is relatively higher in the sequence of 24 h, 48 h and 72 h.Expression quantity of experimental groups are(3.86±1.26,(5.11±1.35) and(7.92 ±0.80),compared with the control group(1.04 ± 0.36),they were higher(p = 0.040; p = 0.006; p = 0.000).That is:Nanog m RNA in experimental groups are relatively higher than the control group.In addition to Western results,Nanog protein of 24 h, 48 h, 72 h groups are :(0.057±0.013),( 0.131 ±0.031),(0.168 ±0.019),they are compared with control group(0.038 ± 0.005) separately(p = 0.541; p = 0.001; p = 0.000).Two groups of Nanog protein expression were higher than the control group.Scratch experiment shows: cells moving rate:24 h group(15.45±2.46) μm /h, 48 h group(35.74± 2.19) μm /h, 72 h group(45.70 ± 4.51)μm /h,respectively compared with the control group(11.42 ± 1.69)μm /h(p = 0.388;P = 0.000;P = 0.000).Longer visible through the cisplatin group, cell migration ability.Conclusion: after chemotherapy,tumor formation and migration ability of remaining cells are enhanced relatively after dealing with CDDP.