Research On The Effect Of Protein Pin1 In The Apoptosis Of The Lung Cancer Cell A549 Induced By Paclitaxel

ObjectiveAS we have known that Pin1 is correlated with the differentiation and proliferation of tumor cells, it is highly expressed in tumor tissue and lowly expressed in normal tissue. Tumor cells will always have lower proliferation and higher apoptosis if the gene Pin1 is depressed. Protein Pin1 has effect on many critical signal factors such as p53 and cyclinD1. As is widely known, paclitaxel and other anti-microtubule agents facilitate the formation of the microtubule and inhibit it depolymerize, then the cell circle is inhibited in G2/M, the cell is more properly get apoptosis, gene bcl-2 depression and gene Bax up-regulation may be the possible mechanism. On the basis of conclusion of research before, gen Pin1 is a gene correlated with cell differentiation and apoptosis, we want to know the relationship between Pin1 and paclitaxel, our research is focus on the quantity change of protein Pin1 after we give paclitaxel and try to give the possible mechanism.Methods1.Effect of paclitaxel to the protein level of Pin1Cultivated the cell A549, and confirm the high expression of pin1 protein in cell A549 by immunocytochemistry. Then gave some paclitaxel to cell A549, first we observed the apoptosis of A549, second we get the total protein respectively after we added paclitaxel to cell A549oh,24h,48h,72h. At the same time we set up control group, then appraisal the protein level of Pin1 by western-blot. The research before have already shown that if we down-regulate the expression of protein Pin1, the tumor cell would get poorly proliferated and show more apoptosis, accordingly we draw a conclusion that there is a new apoptosis mechanism.2.Effect of paclitaxel to E2F1 expressionContinue to cultivate cell A549, and set a timepoint(when the protein pin1 level change significant by paclitaxel), set the test group and control group, took the total protein respectively, then western-blot to evaluate the change of the protein level of E2F1.We have known that E2F1 is in the up stream of Pin1,then we can deduce if there is a mechanism that the E2F1 was inhibited and pin1 was down-regulated.Results1.Effect of paclitaxel to the protein level of pin1Immunocytochemistry shown that protein pin1 was highly expressed in cell A549, contrast to the post MCF-7.After we gave paclitaxel, the apoptosis of the cell A549 was significant increased, the peak came by 24h, and continue to 48h.western-blot shown that the protein level of Pin1 decreased in 48h, and continue to72h, we conclude that paclitaxel can decrease the expression of protein Pin1 in A549, this is another mechanism of apoptosis.2. Effect of paclitaxel to E2F1 expressionWhen it come to 48h after we gave or not gave 1nmol/L paclitaxel, the protein level of E2F1 in test group was more remarkably decreased than the control group(p<0.05),it showed that paclitaxel affect the expression of protein E2F1,then affect the expression of protein Pin1,then the apoptosis increased. Conclusion1. Protein Pin1 was highly expressed in human lung adenocarcinoma cell A549, after interact with paclitaxel, more cell A549 get apoptosis, the expression of protein Pin1 also decreased as time delay, this showed that pin1 involved in the cell A549 apoptosis induced by paclitaxel.2. When it came to 48hafter the paclitaxel was delivered, protein E2F1 level was pin1remarkably decreased, then we knew that paclitaxel affect the expression of Pin1 by down regulated the expression of E2F1, thus increased the apoptosis of tumor cell. This was another apoptosis mechanism.