胶质细胞源性神经营养因子对胰腺癌细胞侵袭的影响

Influence of glial cell derived neurotrophic factor on invasion of pancreatic carcinoma

目的：观察胶质细胞源性神经营养因子（GDNF）在胰腺癌细胞侵袭中的作用及可能的机制。方法四甲基偶氮唑盐（MTT）法分别检测促分裂原活化蛋白激酶通道（MAPK）阻断剂PD98059和磷脂酰肌醇3激酶通道（PI3K）阻断剂LY294002在不同浓度、不同作用时间条件下对胰腺癌细胞MIA Paca-2存活率的影响，利用Trans-well装置在boyden小室的下室中分别加入不同的细胞因子（GDNF、GDNF抗体、PD98059、LY294002），通过计数各组跨膜细胞的数量并进行统计学分析，检测GDNF对胰腺癌细胞的趋化作用和PD98059、LY294002的阻断效果。结果 MTT实验中，培养基中分别加入不同浓度的PD98059或LY294002后，在不同作用时间下胰腺癌细胞形态未发生明显改变，各组的细胞存活率总体水平大都维持在90％～100％，且随药物作用时间或药物浓度变化均无明显规律性变化。Trans-well实验中，下室中加入GDNF后增加了Boyden小室跨膜细胞数量，加入阻断剂PD98059后跨膜细胞数量明显减少，差异有统计学意义（P＝0.014），加入阻断剂LY294002后跨膜细胞数量有减少趋势，差异无统计学意义（P＝0.221），同时加入两种阻断剂后跨膜细胞数量减少，差异有统计学意义（P＜0.01）。结论 GDNF可增强胰腺癌细胞MIA Paca-2的侵袭转移能力，发挥这一作用的途径可能与MAPK通道和PI3 K通道有关。

Objective To observe the effect and the possible mechanism of glial cell derived neuro-trophic factor （GDNF）on invasive pancreatic carcinoma. Methods Methyl thiazolyl tetrazolium （MTT） as-say were used to detect the effect of PD98059-blocker of mitogen-activated protein kinase （MAPK）pathway and LY294002-blocker of phosphatidyl inositol 3-kinase （PI3K）pathway on the survival rate of pancreatic cancer cell MIA Paca-2 under different drug dosage and different reaction time. Different cytokines （GDNF、anti-GDNF antibody、PD98059、LY294002）were added in the lower chamber of the boyden chamber. Trans-membrane cell numbers of each group were analyzed statistically. Chemotaxis of GDNF on pancreatic cancer cell and the blocking effect of PD98059 and LY294002 by Trans-well device were also analysed.Results In the MTT assay,pancreatic cancer cell morphology did not change significantly when treated with different concen-trations of PD98059 and LY294002 in different reaction time. No significant difference was identified in the survival rate of each group. The overall survival rate maintained mostly from 90%to 1 00%,There was no sig-nificant change with different action time or drug concentration. In the Trans-well assay,when treated with GD-NF,the number of trans-membrane cells in boyden chamber increases while reduces when treated with PD98059,with significance （P=0.01 4）. LY29400 also reduce the number,but without significant （P=0.221 ）. The difference is obvious when added with both blockers simutaneously （P〈0.01 ）. Conclusion GDNF can enhance the ability of invasive and metastatic ability of MIA Paca-2. The mechanism may relate with MAPK path and PI3 K pathway.