本文選題：前列腺癌 + RelB��； 參考：《蘇州大學(xué)》2014年碩士論文

【摘要】：目的本研究試圖闡明NF-κB家族成員中的RelB在前列腺癌細(xì)胞中的作用以及探討潛在的生物學(xué)機(jī)制。 方法用Western blot法檢測(cè)三種前列腺癌細(xì)胞株中NF-κB家族成員的蛋白表達(dá)情況。建立穩(wěn)定轉(zhuǎn)染RelB-siRNA的DU145細(xì)胞株。Western blot法檢測(cè)RelB沉默后細(xì)胞的NF-κB家族其它成員的蛋白表達(dá)情況。用xCELLigence系統(tǒng)實(shí)時(shí)動(dòng)態(tài)觀察RelB沉默后細(xì)胞生長(zhǎng)，，以及細(xì)胞遷移和細(xì)胞侵襲的變化。用流式細(xì)胞術(shù)檢測(cè)RelB沉默后細(xì)胞的細(xì)胞周期、細(xì)胞增殖以及細(xì)胞凋亡的變化。用qRT-PCR技術(shù)檢測(cè)基因的mRNA水平的表達(dá)。用細(xì)胞劃痕實(shí)驗(yàn)觀察RelB沉默后細(xì)胞遷移能力的變化。用明膠酶譜MMP活性檢測(cè)方法檢測(cè)RelB沉默后細(xì)胞的MMP2、MMP9活性變化。 結(jié)果RelA、p50、RelB和p52蛋白在雄激素非依賴型前列腺癌細(xì)胞株DU145和PC-3中呈持續(xù)性高表達(dá)，RelB在DU145細(xì)胞中的表達(dá)最為顯著。在DU145細(xì)胞株中分別穩(wěn)定轉(zhuǎn)染pSilencer3.1-siRelB質(zhì)粒和pSilencer3.1質(zhì)粒，成功獲得實(shí)驗(yàn)組RelB低表達(dá)DU145-siRelB細(xì)胞和對(duì)照組DU145-control細(xì)胞。RelB的沉默對(duì)細(xì)胞中NF-κB家族其它成員無(wú)明顯影響。DU145-siRelB細(xì)胞的生長(zhǎng)速度明顯慢于DU145-control細(xì)胞，并且遷移、侵襲能力均較DU145-control細(xì)胞顯著減弱。細(xì)胞劃痕實(shí)驗(yàn)也表明了DU145-siRelB細(xì)胞的遷移能力明顯較DU145-control細(xì)胞弱。DU145-siRelB細(xì)胞與DU145-control細(xì)胞在細(xì)胞增殖和細(xì)胞周期上無(wú)明顯差異。DU145-siRelB細(xì)胞的凋亡細(xì)胞百分比比DU145-control細(xì)胞明顯增多。DU145-siRelB細(xì)胞中的bcl-2基因mRNA表達(dá)水平較DU145-control細(xì)胞明顯下調(diào)，其余相關(guān)基因Bcl-xl、ICAP1、Bax、A20、Bim、Mcl-1mRNA表達(dá)水平無(wú)明顯變化。DU145-siRelB細(xì)胞中的ITGB1基因的mRNA和蛋白質(zhì)水平的表達(dá)均較DU145-control細(xì)胞顯著下調(diào)。DU145-siRelB細(xì)胞中的MMP2、MMP9蛋白質(zhì)水平的表達(dá)均較DU145-control細(xì)胞顯著下調(diào)，同時(shí)明膠酶譜MMP活性檢測(cè)方法檢測(cè)顯示DU145-siRelB細(xì)胞MMP2和MMP9的表達(dá)顯著低于DU145-control細(xì)胞。 結(jié)論在雄激素非依賴型前列腺癌細(xì)胞株中，RelB在DU145細(xì)胞中表達(dá)最為顯著。DU145-siRelB細(xì)胞的生長(zhǎng)明顯慢于DU145-control細(xì)胞，并且遷移、侵襲能力均較DU145-control細(xì)胞顯著減弱。RelB的缺失顯著抑制了前列腺癌細(xì)胞株DU145細(xì)胞的生長(zhǎng)，與細(xì)胞凋亡增多相關(guān)，bcl-2基因的下調(diào)可能是潛在的原因之一。RelB缺失顯著抑制了腫瘤細(xì)胞的遷移和侵襲能力，ITGB1基因表達(dá)的下調(diào)以及MMP2和MMP9活力的減弱是潛在的原因之一。
[Abstract]:Objective to elucidate the role of RelB from NF- 魏 B family in prostate cancer cells and to explore the potential biological mechanisms. Methods Western blot assay was used to detect the protein expression of NF- 魏 B family members in three prostate cancer cell lines. A stable DU145 cell line transfected with RelB-siRNA was established. Western blot assay was used to detect the protein expression of other members of NF- 魏 B family after the silencing of RelB. XCELLigence system was used to observe the cell growth, cell migration and cell invasion after RelB silencing. Cell cycle, cell proliferation and apoptosis were detected by flow cytometry. The mRNA level of the gene was detected by qRT-PCR. The cell migration ability was observed by cell scratch assay after RelB silencing. The changes of MMP2MMP9 activity were detected by gelatinase assay. Results the expression of RelAp50A RelB and p52 protein in androgen independent prostate cancer cell lines DU145 and PC-3 was significantly higher than that in DU145 cells. After stable transfection of pSilencer3.1-siRelB and pSilencer3.1 plasmids in DU145 cell line, the silencing of DU145-siRelB cells in experimental group and DU145-siRelB cells in control group had no significant effect on the growth rate of DU145-siRelB cells compared with that of DU145-control cells, and the growth rate of DU145-siRelB cells was significantly slower than that of DU145-control cells. Moreover, migration and invasion ability of DU 145-control cells were significantly decreased compared with those of DU145-control cells. Cell scratch assay also showed that the migration ability of DU145-siRelB cells was significantly weaker than that of DU145-control cells. There was no significant difference in cell proliferation and cell cycle between DU145-siRelB cells and DU145-siRelB cells. The percentage of apoptotic cells in DU145-siRelB cells was significantly higher than that in DU145-control cells. The expression of bcl-2 mRNA in DU145-control cells was significantly lower than that in DU145-control cells. The mRNA and protein levels of ITGB1 gene in DU145-siRelB cells were significantly down-regulated than those in DU145-RelB cells, and the expression of MMP2mMP9 in DU145-RelB cells was significantly lower than that in DU145-control cells, and the expression of MMP2mMP9 in DU145-RelB cells was significantly lower than that in DU145-control cells. The expression of MMP2 and MMP9 in DU145-siRelB cells was significantly lower than that in DU145-control cells. Conclusion the expression of MMP2 and MMP9 in androgen independent prostate cancer cells is the most significant in DU145 cells. Significantly slower than DU145-control cells, The migration and invasion ability of DU145-control cells were significantly lower than those of DU145-control cells. The loss of RelB significantly inhibited the growth of prostate cancer cell line DU145. The down-regulation of bcl-2 gene associated with increased apoptosis may be one of the potential reasons. The deletion of RelB significantly inhibits the migration and invasion of tumor cells. The down-regulation of ITGB1 gene expression and the decrease of MMP2 and MMP9 activity are one of the underlying reasons.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R737.25

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