The Effect Of Lipid Raft Damaged On Invasion And Metastasis Ability Of Human Nasopharyngeal Carcinoma Line CNE1-LMP1

Objective:In this paper, we analysis the LMP1 gene affecting CNE1 cells proliferation,metastasis and invasion. And we investigate the inpact of lipid raft damaged on lipid raft-related protein expression, especially metastasis and invasion of CNE1-LMP1 in vitro. To sum up, the datas provide some new clues of nasopharyngeal transfer mechanisms for us.Methods:1. Human NPC cells CNE1, CNE1-LMP1 was selected as the research objects,the latter one was already stable transfected into LMP1 gene, and attested by Western Blot. Mean while we indentified some biological characters of CNE1-LMP1 by CCK8?Transwell.2. The reasonable time and concentration of lipid raft disrupting agents(M?CD)must be determined to establish lipid rafts damaged model. We observed the changes of cytoactive by CCK8-kit. Then we tested the cholesterol content of CNE1-LMP1 before and after deal with M?CD at several different time and concentration.3. Furthermore, Western-Blot methods was uesd to evaluate the changes of protein content of caveolin-1 and LMP1 in CNE1-GL cells, when RT-PCR was applied to evaluate the level of m RNA, after disrupted LRs integrity. We just want to know the influence of lipid raft broken on cell LR-associated proteins.4. Transwell experiments were taken to discover the changes of invision and transfer ability of CNE1-LMP1 after pretreated by M?CD.Results:1. CNE1-LMP1 lines indeed expressed high level of LMP1. Several capabilities of CNE1 were enhanced by this special gene, including accelerating proliferation, promoting metastasis and invasion, and there was a statistical discrepancy(P<0.05).2. Lipid raft disrupting agent(M?CD) would not bring remarkable damages to CNE1-GL cell(P>0.05) within 1 hour. But the CNE1-GL cytoactive decreased with raised concentration when prolonged the time to 2h, the data was prominently difference(P<0.01).3. The membrane level of cholesterol would prominently decreased along with raised concentration of M?CD(P<0.01), when controled the drug treatment within1 hour.4. The protein and m RNA level of Caveolin-1 and LMP1 had a prominent decreased(P<0.01), when 10 m M M?CD pre-treated on CNE1-LMP1 cells for actual 1 hour, and there is a statistical discrepancy.5. Transwell invasion and migration assays showed the influence of M?CD work on CNE1-LMP1 cells, the drug group had lower capability of invasion and metastasis.Conclusions:LMP1 gene can accelerates NPC cells self-proliferation, spread and attack.M?CD is a reagent that can bring cholesterol away from cell membrane,and that is the way to destroy the integrity of caveolae. The reasonable time and concentration of M?CD can be acquired to get the model successfully. The experiment datas demonstrate that the protein and m RNA level of caveolin-1 anf LMP1 turns to descend along with decline of the amount of cholesterol in cells. Finally, the ability of invade and transfer of CNE1-LMP1 weakened accompany with the reduced of cholesterol.