發(fā)布時間：2018-07-17 16:21

【摘要】：P-糖蛋白(P-glycoprotein, P-gp)是一種ATP依賴性的跨膜轉運蛋白，可以將多種結構和藥效不同的化合物排出細胞，是產生多藥耐藥的主要原因之一。目前P-糖蛋白在多要耐藥方面的研究主要集中在腫瘤及難治性癲癇的發(fā)生，其中腫瘤的發(fā)生是原癌基因的激活、抑癌基因的抑制、凋亡基因及修復基因的改變等多因素綜合作用的結果，而P-糖蛋白的升高是癲癇發(fā)展為難治性癲癇的重要生物學基礎，，因此，P-糖蛋白在難治性癲癇中的作用機制研究尤為重要。目前關于P-糖蛋白基因調控及相應的信號傳導通路成為P-糖蛋白的研究熱點，但是P-糖蛋白處于不斷的合成與降解的動態(tài)平衡，因此，從P-糖蛋白降解的角度揭示P-糖蛋白在多藥耐藥中的作用機制是一個新思路。 目的 研究神經細胞內P-糖蛋白的降解途徑并探討P-糖蛋白在神經細胞內降解對難治性癲癇治療的臨床意義。 方法 1.腺病毒作為載體，將ABCB1基因轉染到U251細胞，建立高表達P-糖蛋白的細胞模型。 2.在已建立的高表達P-糖蛋白的細胞模型上，不同濃度的特異性蛋白酶體抑制劑MG132、溶酶體抑制劑ChlQ處理細胞24h，同時設定時間依賴組。Western blot檢測P-糖蛋白水平的變化情況，明確神經細胞內P-糖蛋白的降解途徑。 結果 1.蛋白酶體抑制劑MG132處理組，P-糖蛋白在蛋白酶體抑制劑處理后表達明顯升高，且P-糖蛋白表達隨著MG132濃度的升高而升高。P-糖蛋白表達與蛋白酶體抑制劑處理時間無明顯相關性，但是蛋白酶體抑制劑處理可以明顯增加P-糖蛋白的表達。 2.溶酶體抑制劑ChlQ處理組，P-糖蛋白的表達與對照組相比無明顯異常，且不隨溶酶體抑制劑濃度及處理時間變化而變化。 結論 1.神經細胞內P-糖蛋白的降解與溶酶體途徑無關。 2.神經細胞內P-糖蛋白的降解通過泛素-蛋白酶體途徑。 3.抑制泛素-蛋白酶體功能可以增加神經細胞內泛素化P-糖蛋白的含量，引起神經細胞內P-糖蛋白積聚。
[Abstract]:P-glycoprotein (P-gp) is an ATP-dependent transmembrane transporter, which can expel many compounds with different structures and effects, and is one of the main reasons for multidrug resistance. At present, the research on multidrug resistance of P- glycoprotein mainly focuses on the occurrence of tumor and refractory epilepsy, in which tumor is activated by proto-oncogene and suppressor gene is inhibited. The change of apoptosis gene and repair gene is the result of many factors, and the increase of P-glycoprotein is an important biological basis for the development of epilepsy into intractable epilepsy, so it is very important to study the mechanism of P- glycoprotein in refractory epilepsy. At present, the regulation of P- glycoprotein gene and the corresponding signal transduction pathway have become the research hotspot of P- glycoprotein, but P- glycoprotein is in the dynamic balance of synthesis and degradation, so, It is a new idea to reveal the action mechanism of P- glycoprotein in multidrug resistance from the point of view of P- glycoprotein degradation. Objective to study the degradation pathway of P-glycoprotein in nerve cells and the clinical significance of P-glycoprotein degradation in the treatment of intractable epilepsy. Method 1. Adenovirus was used as a vector to transfect ABCB1 gene into U251 cells to establish a cell model of high expression of P- glycoprotein. 2. In the established cell model with high expression of P-glycoprotein, the cells were treated with different concentrations of specific proteasome inhibitor MG132 and lysosomal inhibitor ChlQ for 24 h. To identify the degradation pathway of P-glycoprotein in nerve cells. Result 1. The expression of P- glycoprotein in proteasome inhibitor MG132 group was significantly increased after treatment with proteasome inhibitor, and the expression of P- glycoprotein increased with the increase of MG132 concentration. There was no significant correlation between the expression of P- glycoprotein and the treatment time of proteasome inhibitor. However, proteasome inhibitor treatment significantly increased the expression of P-glycoprotein. 2. 2. The expression of P- glycoprotein in the lysosomal inhibitor ChlQ group was not abnormal compared with the control group, and did not change with the concentration of lysosomal inhibitor and the treatment time. Conclusion 1. The degradation of P-glycoprotein in nerve cells was not related to lysosomal pathway. 2. Degradation of P-glycoprotein in nerve cells through the ubiquitin-proteasome pathway. 3. Inhibiting the function of ubiquitin-proteasome can increase the content of ubiquitin-glycoprotein and induce the accumulation of P- glycoprotein in nerve cells.
【學位授予單位】：泰山醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R742.1

【參考文獻】

相關期刊論文 前1條

1 李建;姜德春;張國君;蔡立新;樸月善;王育琴;;難治性癲癇患者腦組織內致癇灶與其周圍組織P糖蛋白表達量的比較[J];中國藥物依賴性雜志;2008年03期

本文編號：2130225