發(fā)布時(shí)間：2018-05-01 15:41

  本文選題：survivin + 膀胱癌T24　； 參考：《桂林醫(yī)學(xué)院》2014年碩士論文

【摘要】：目的：survivin是近年來發(fā)現(xiàn)的凋亡抑制蛋白（IAP）家族中的新成員，是目前發(fā)現(xiàn)的最強(qiáng)的凋亡抑制因子，并且survivin廣泛高表達(dá)于人類大多數(shù)惡性腫瘤組織中，在正常組織很少表達(dá)或不表達(dá)。有研究表明，該因子通過抑制腫瘤細(xì)胞凋亡，在腫瘤的形成和發(fā)展中起著重要的作用。本研究通過使用不同濃度化療藥物吡柔比星(Pirarubicin，THP)作用人膀胱癌T24細(xì)胞一定時(shí)間后，檢測T24細(xì)胞的細(xì)胞增殖情況，并檢測Survivin mRNA和Survivin蛋白在不同濃度化療藥物作用下細(xì)胞中的表達(dá)水平，探討Survivin在THP誘導(dǎo)人膀胱癌T24細(xì)胞凋亡過程中的作用機(jī)制和原理，為吡柔比星在臨床上膀胱癌的灌注治療中提供一定的理論依據(jù)，并為膀胱癌的發(fā)生發(fā)展以及分子靶向治療提供一定的理論基礎(chǔ)。 方法：1.配置成不同終濃度的THP(0μg/ml、0.1μg/ml、1μg/ml、10μg/ml)，分別作用于人膀胱癌T24細(xì)胞一定時(shí)間，在設(shè)置空白對照組以及各藥物實(shí)驗(yàn)組的前提下，應(yīng)用MTT法檢測不同濃度THP對人膀胱癌T24細(xì)胞分別作用24、48、72小時(shí)后增殖抑制率的影響。2.應(yīng)用RT-PCR方法檢測在不同濃度(0、0.1、1、10μg/ml)THP處理膀胱癌T24細(xì)胞24小時(shí)后，T24細(xì)胞中Survivin mRNA的表達(dá)水平。3.應(yīng)用Western-blot蛋白印跡技術(shù)檢測人膀胱癌T24細(xì)胞在不同濃度(0、0.1、1、10μg/ml)THP處理24小時(shí)后，T24細(xì)胞中Survivin蛋白的表達(dá)水平變化。應(yīng)用SPSS.13.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)學(xué)分析，數(shù)據(jù)以“均值±標(biāo)準(zhǔn)差”（X±s）表示。兩樣本均數(shù)比較采用t檢驗(yàn)，多樣本均數(shù)比較采用完全隨機(jī)的單因素（One-ANOVA）方差分析。P0.05表示具有差異，有統(tǒng)計(jì)學(xué)意義，P0.01表示差異顯著。 結(jié)果：1.經(jīng)MTT法檢測空白組、24h，48h，72h四組細(xì)胞在藥物作用后各組的OD值，根據(jù)測得的OD值，比較各組細(xì)胞生長情況。與對照組比較，各組濃度的THP在24h，48h，72h均可以抑制膀胱癌T24細(xì)胞的增殖，差異具有統(tǒng)計(jì)學(xué)意義（P0.05），計(jì)算加入藥物組24h，48h，72h增殖抑制率，發(fā)現(xiàn)其抑制率呈現(xiàn)一定的濃度及時(shí)間依賴性（P0.05）。2.不同濃度(0、0.1、1、10μg/ml)THP處理T24細(xì)胞24小時(shí)后，均使Survivin mRNA的表達(dá)明顯下調(diào)，并表現(xiàn)為一定的濃度依賴性（P0.01）。3.應(yīng)用western-blot檢測不同濃度THP作用膀胱癌T24細(xì)胞24小時(shí)后，，各組不同濃度間Survivin蛋白質(zhì)的表達(dá)顯著下調(diào)，并且具有一定的濃度依賴性，隨著藥物濃度的升高，Survivin蛋白下調(diào)的更加明顯，表現(xiàn)出一定范圍的濃度依賴性（P0.05）。 結(jié)論：吡柔比星在體外的細(xì)胞實(shí)驗(yàn)中，對膀胱癌T24細(xì)胞有明顯的抑制作用，并且具有一定的濃度和時(shí)間依賴性。即隨著THP濃度的升高和作用時(shí)間的延長，吡柔比星對T24細(xì)胞的抑制作用更加明顯。在THP對膀胱癌T24細(xì)胞的體外抑制實(shí)驗(yàn)中，survivin mRNA和survivin蛋白質(zhì)隨著化療藥物THP對T24細(xì)胞作用的增強(qiáng)，即THP濃度的升高和作用時(shí)間的延長，survivin mRNA和survivin蛋白質(zhì)的表達(dá)下降的更加明顯，表現(xiàn)出一定的濃度和時(shí)間的依賴性。綜上所述，THP很可能是通過下調(diào)survivin凋亡抑制因子的表達(dá)這條通路來促使T24細(xì)胞的凋亡，在上述原理下來達(dá)到抑制膀胱癌T24細(xì)胞的生長的。
[Abstract]:Objective: survivin is a new member of the apoptosis suppressor protein (IAP) family found in recent years. It is the strongest inhibitor of apoptosis, and survivin is widely expressed in most of human malignant tumor tissues and is rarely expressed or expressed in normal tissues. The formation and development of tumor play an important role in the development of human bladder cancer T24 cells with different concentrations of Pirarubicin, THP, to detect the cell proliferation of T24 cells and to detect the expression of Survivin mRNA and Survivin protein in cells under the action of different concentrations of chemotherapeutic drugs. To explore the mechanism and principle of Survivin in the process of THP induced apoptosis of human bladder cancer T24 cells, providing a theoretical basis for the treatment of pirubicin in the treatment of bladder cancer in clinic, and providing a theoretical basis for the development of bladder cancer and molecular targeting therapy.
Methods: 1. THP (0 g/ml, 0.1 u g/ml, 1 g/ml, 10 mu g/ml) were configured for a certain time of human bladder cancer T24 cells respectively. Under the premise of setting blank control group and each drug experiment group, the effect of THP on the proliferation inhibition rate of T24 cells in human bladder cancer after 24,48,72 hours was detected by MTT method. 2. RT-PCR method was used to detect the expression level of Survivin mRNA in T24 cells after 24 hours of T24 cells treated with different concentrations of 0,0.1,1,10 g/ml THP to detect the expression level of Survivin mRNA in T24 cells using Western-blot blot technique to detect the expression level of human bladder cancer T24 cells at different concentrations (0,0.1,1,10 muon) for 24 hours SPSS.13.0 statistical software was used to carry out statistical analysis with "mean mean standard deviation" (X + s). The average number of two samples was compared with t test. The diversity of the average numbers compared with the random single factor (One-ANOVA) variance analysis.P0.05 indicated that there were differences, and there were statistical significance, P0.01 indicated that the difference was significant.
Results: 1. after the MTT method was used to detect the blank group, 24h, 48h, 72h four groups of cells in each group after the action of drug action, according to the measured OD value, compare the growth of each group. The THP concentration in each group of 24h, 48h, 72h can inhibit the proliferation of bladder cancer T24 cells, the difference has statistical significance (P0.05), the addition of drug group 24h, 4. 8h, 72h proliferation inhibition rate, it was found that the inhibition rate showed a certain concentration and time dependence (P0.05).2. different concentrations (0,0.1,1,10 mu g/ml) THP treatment T24 cells 24 hours after the Survivin mRNA expression obviously down, and showed a certain concentration dependence (P0.01).3. application detection of different concentrations of bladder cancer cells After 24 hours, the expression of Survivin protein decreased significantly between different concentrations, and had a certain concentration dependence. With the increase of drug concentration, the down regulation of Survivin protein was more obvious, showing a certain range of concentration dependence (P0.05).
Conclusion: in vitro, pirirubicin has a significant inhibitory effect on bladder cancer T24 cells and has a certain concentration and time dependence. That is, the inhibition of pirarubicin to T24 cells is more obvious with the increase of the concentration of THP and the prolongation of the action time. In vitro inhibition test of THP to T24 cells of bladder cancer, Su Rvivin mRNA and survivin protein, with the enhancement of the effect of chemotherapeutic drug THP to T24 cells, that is, the increase of THP concentration and the prolongation of the action time, the expression of Survivin mRNA and survivin protein decreases more clearly, showing a certain concentration and time dependence. In summary, THP is likely to reduce the inhibitory cause of survivin apoptosis. The pathway of expression of T24 promotes the apoptosis of T24 cells and inhibits the growth of bladder cancer cells.

【學(xué)位授予單位】：桂林醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R737.14

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