| Part â… The expression of multidrug resistance in the tissues of human nasopharyngeal carcinomaObjectives:To assess the expression of multidrug resistance (MDR) in the tissues of human nasopharyngeal carcinoma (NPC) and analyze the relationship between their expression and therapeutic efficacy.Methods:One hundred and seventy two biopsy specimens of nasopharynx in the nasal endoscopy room at our hospital (Eye, Ear, Nose and Throat Hospital of Fudan University) were obtained during April of 2011 to April of 2012. Among them,75 patients were diagnosed with nasopharyngeal carcinoma. The patients, who were recieved the chemotherapy of cisplatin (DDP) and 5-fluorouracil (5-FU) were separated into two groups (complete remission and not complete remission) according to their therapeutic efficacy. There were a total of 8 patients in the group of not complete remission and were included as group B to analyze the expression of MDR. And 8 another patients in the complete remission group were selected as group A as control. The specimens were studied by real time PCR. The gene markers included dihydropyrimidine dehydrogenase (DPYD), thymidine phosphorylase (TYMP), thymidylate synthase (TS), orotate phosphoribosyl transferase (OPRT), dihydrofolate reductase (DHFR), methylenetetrahydrofolate reductase (MTHFR), excision repair cross complementing gene 1 (ERCC1), epidermal growth factor receptor, (EGFR), vascular endothelial growth factor-A (VEGF-A), multidrug resistance relation protein (MRP), lung-resistance related protein 1 (LRP1), breast cancer resistance protein (BCRP), topoisomeerase â…¡ (TOP2A). The difference of the expression in the gene markers were compared between group A and B. Then the relationship between their expression and therapeutic efficacy were analyzed.Results:The gene markers were expressed differently in all the 16 specimens. The expression of ERCC1 and TS in the group B were higher than the expression in the group B (P<0.05). Other parameters had no significant difference between group A and group B (P>0.05).Conclusions:There were many MDR gene markers expressed in the nasopharygeal carcinoma tissues. And their expression may play an important role in the therapeutic efficacy. The high expression of ERCC1 and TS may be related the poor response to treatment. expression of ERCC1 and TS in the group B were higher than the expression in the group B (P<0.05). Other parameters had no significant difference between group A and group B (P>0.05).Part â…¡ Establishment of nasopharyngeal carcinoma cisplatin-resistant cell line HNE1/DDP and its biological characteristicsObjectives:To establish a DDP-resistant cell line from human nasopharyngeal carcinoma cell line HNE1 and to investigate its biological characteristics.Methods:The human nasopharyngeal carcinoma cell line HNE1 was exposed to cisplatin with the approach of combination of pulse treatment selection and stepwise selection for I year to establish cisplatin resistance cell line (HNE1/DDP). And the drug sensitivity to the commonly used chemotherapy drugs (cisplatin, carboplatin, 5-FU and paclitaxel) in the NPC was detected in the HNE1/DPP and its parental cell HNE1. The morphology of the cells in the process of establishment was observed. And their ability of proliferation, migration and invasion were evaluated.Results:The HNE1/DDP cell line was developed after 1 year. It could stably grow and passage in the cell culture with lug/ml cisplatin. The resistance index of HNE1/DPP to cisplatin is 14.75. And it also exhibited cross-resistance to other chemotherapeutic agents (carboplatin,5-FU and paclitaxel). The morphology of HNE1 cell in the process of establishment changed. But the morphology of HNE1/DPP cell was almost same to the HNE1 except the doubling time was a little prolonged (41.85 h VS 37.44 h). The ability of migration and invasion had no difference between resistant cells (HNE1/DDP) and the parental cell (HNE1).Conclusions:We successfully established the cisplatin-resistance NPC cell lines HNE1/DDP. It could stably grow and passage in the cell culture with lug/ml cisplatin and showed showed various cross-resistance to carboplatin,5-FU and paclitaxel. The morphology and ability of migration and invasion of the HNE1/DPP did not change compared to HNE1. It was a typical and stable resistant NPC cells and can be used as a research model for downstream experiment.Part â…¢ The multidrug resistance expression in the nasopharygeal carcinoma cell line and its reversionObjectives:The objiect of third part is to detect the multidrug (MDR) expression of nasopharygeal carcinoma (NPC) cisplatin resistant cell line HNE1/DPP and its parental cell line HNE1 and to compare their difference. Then the role of Lidocaine in the reversion of the cisplatin in the HNE1/DPP was explored.Methods:The expression of MDR parameters were detected by real time PCR (RT-PCR), immunofluorescence (IF) and Western-Blot. The MDR parameters included dihydropyrimidine dehydrogenase (DPYD), thymidine phosphorylase (TYMP), thymidylate synthase (TS), orotate phosphoribosyl transferase (OPRT), dihydrofolate reductase (DHFR), methylenetetrahydrofolate reductase (MTHFR), excision repair cross complementing gene 1 (ERCC1), epidermal growth factor receptor, (EGFR), vascular endothelial growth factor-A (VEGF-A), multidrug resistance relation protein (MRP), lung-resistance related protein 1 (LRP1), breast cancer resistance protein (BCRP), topoisomeerase II (TOP2A). And the mitochondrial membrane potential (JC-1), reactive oxygen species (ROS) and flow cytometry (Annexin V-FICT/propidine iodide, PI) were used to explore the reversion role of the Lidocaine.Results:The results of RT-PCR, IF and Western-Blot showed the MDR protein were expressed in varying degrees in the HNE1/DPP and HNE1. The IC50 of HNE/DPP cell to the cisplatin decreased when the Lidocaine was added into the cell culture.Conclusions:The MDR resistance mechanisms of HNE1/DPP cell may be associated with kinds of molecular mechanisms. And Lidocaine has increased sensitivitity of HNE1/DPP to cisplatin. |
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