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        受翻譯調(diào)節(jié)的腫瘤蛋白在PRL-3促進(jìn)結(jié)腸癌轉(zhuǎn)移中的作用*

        2011-09-14 06:21:28褚忠華李守峰曾育杰關(guān)玉峰
        中國(guó)病理生理雜志 2011年10期
        關(guān)鍵詞:結(jié)腸癌克隆試劑盒

        褚忠華, 劉 璐, 來(lái) 偉, 李守峰, 曾育杰, 關(guān)玉峰

        (1中山大學(xué)孫逸仙紀(jì)念醫(yī)院胃腸外科,廣東廣州510120;2廣州市番禺中心醫(yī)院普通外科,廣東廣州511400)

        肝再生磷酸酶-3(phosphatase of regenerating liver-3,PRL-3)是近年來(lái)發(fā)現(xiàn)的一種具有促進(jìn)細(xì)胞增殖、遷移和侵襲的蛋白酪氨酸磷酸酶[1]。最初,Saha等[2]通過(guò)比較結(jié)直腸癌轉(zhuǎn)移灶與原發(fā)腫瘤之間的基因表達(dá)差異,發(fā)現(xiàn)PRL-3在結(jié)直腸癌肝轉(zhuǎn)移灶中高度表達(dá),同時(shí)存在基因拷貝數(shù)的增加,而在未轉(zhuǎn)移的結(jié)直腸癌和正常的結(jié)直腸上皮不表達(dá)。之后,陸續(xù)有研究發(fā)現(xiàn)PRL-3在卵巢癌、胃癌、乳腺癌和霍奇金淋巴瘤中表達(dá)升高[3-6]。目前,已有大量研究表明PRL-3在多種腫瘤細(xì)胞的轉(zhuǎn)移過(guò)程中均起到重要作用[3,4,6]。然而 PRL -3 促進(jìn)腫瘤轉(zhuǎn)移的具體分子機(jī)制仍然不清楚。受翻譯調(diào)節(jié)的腫瘤蛋白(translationally controlled tumor protein,TCTP),是一類廣泛存在于動(dòng)物、植物及酵母中,并在序列上高度保守且有很高同源性的蛋白家族[7]。最初認(rèn)為TCTP是一類生長(zhǎng)相關(guān)蛋白,但隨后的研究提示TCTP可能具有非常重要的生物學(xué)功能,包括調(diào)節(jié)細(xì)胞周期的進(jìn)程和惡性轉(zhuǎn)移,鈣結(jié)合蛋白和細(xì)胞外組胺釋放蛋白等[7]。更重要的是,新近的研究發(fā)現(xiàn),TCTP在前列腺癌和結(jié)腸癌細(xì)胞增殖、細(xì)胞凋亡、細(xì)胞遷移和侵襲及轉(zhuǎn)移均具有重要作用[8,9]。由于PRL-3與TCTP在腫瘤轉(zhuǎn)移的多個(gè)環(huán)節(jié)均具有重要調(diào)節(jié)作用,因此它們?cè)诎l(fā)揮促腫瘤轉(zhuǎn)移過(guò)程中可能存在一定的相關(guān)性。因此,本文主要探討PRL-3是否對(duì)TCTP的表達(dá)進(jìn)行調(diào)節(jié)及TCTP在PRL-3促結(jié)腸癌轉(zhuǎn)移中的作用。

        材料和方法

        1 材料

        結(jié)腸癌細(xì)胞株LoVo購(gòu)自中山大學(xué)醫(yī)學(xué)院細(xì)胞庫(kù),大腸桿菌DH-5α購(gòu)自上海生工公司,真核載體pAcGFP -C3購(gòu)自 Clontech,T4連接酶、Xho I、EcoR I內(nèi)切酶、Trizol和RT-PCR試劑盒購(gòu)自TaKaRa,Lipofectamine 2000和G418購(gòu)自Invitrogen。兔抗PRL-3多克隆抗體和兔抗TCTP多克隆抗體購(gòu)自Santa Cruz,辣根過(guò)氧化物酶標(biāo)記的羊抗兔Ⅱ抗購(gòu)自Santa Cruz。siRNA干擾序列由上海吉馬公司合成,CCK-8試劑盒購(gòu)自碧云天公司,Transwell雙層細(xì)胞培養(yǎng)板(直徑 6.5 mm,孔徑 8.0 μm)購(gòu)自 Corning,matrigel購(gòu)自BD,4%多聚甲醛和結(jié)晶紫購(gòu)自廣州威佳公司。

        2 方法

        2.1 質(zhì)粒的構(gòu)建 根據(jù)GenBank中人PRL-3 mRNA的序列設(shè)計(jì)擴(kuò)增引物序列,上游引物5'-CCGCTCGAGATGGCTCGGATGAACC -3',下游引物5'- CGGAATTCCTACATAACGCAGCACCG - 3'。RT-PCR擴(kuò)增目的基因,并將產(chǎn)物在37℃下Xho I、EcoR I內(nèi)切酶作用2 h,1%瓊脂糖凝膠電泳并切膠回收。同時(shí)用Xho I、EcoR I內(nèi)切酶酶切pAcGFPC3空載體并回收酶切后大片段。將回收的PRL-3酶切產(chǎn)物和pAcGFP1-C3(Xho I、EcoR I雙酶切)載體與室溫下過(guò)夜連接,構(gòu)建pAcGFP-C3-PRL-3真核載體。

        2.2 LoVo細(xì)胞的轉(zhuǎn)染和篩選 對(duì)數(shù)生長(zhǎng)期的LoVo細(xì)胞以 2×108cells/L鋪于24孔板中,按照 Lipofectamine 2000說(shuō)明書進(jìn)行轉(zhuǎn)染,實(shí)驗(yàn)組轉(zhuǎn)染pAcGFP-C3-PRL-3載體,對(duì)照組轉(zhuǎn)染空質(zhì)粒pAcGFPC3。6 h后更換培養(yǎng)基,24 h后開始進(jìn)行G418篩選,待形成陽(yáng)性單細(xì)胞克隆群落后,在熒光顯微鏡下觀察并用尖吸管吸取陽(yáng)性單克隆培養(yǎng)。

        2.3 熒光定量PCR Trizol提取轉(zhuǎn)細(xì)胞總RNA,使用RT-PCR試劑盒逆錄得到cDNA,隨后使用2×SYBR Premix Ex TaqTM試劑盒擴(kuò)增PRL-3或TCTP基因,定量分析PRL-3或TCTP表達(dá)水平。PRL-3引物如下:上游引物5'-CCGCTCGAGATGGCTCGGATGAACC-3',下游引物5'- CGGAATTCCTACATAACGCAGCACCG -3'。TCTP引物如下:上游引物 5'- TGAAGAACAGAGACCAGAAAG -3',下游引物 5'-CACGGTAGTCCAATAGAGCAAC-3'。GAPDH引物如下:上游引物5'-GCACCGTCAAGGCTGAGAAC -3',下游引物 5'-TGGTGAAGACGCCAGTGGA-3'。反應(yīng)條件:95℃ 30 s預(yù)變性后,95℃ 5 s,60℃ 20 s,40個(gè)循環(huán),最后95℃ 0 s,65℃15 s,95 ℃ 0 s。

        2.4 Western blotting檢測(cè) RIPA提取細(xì)胞總蛋白,每組細(xì)胞以30 μg等量蛋白進(jìn)行12%SDS-PAGE電泳。電泳完畢后轉(zhuǎn)膜至PVDF膜,然后使用含有5%脫脂奶TBST進(jìn)行封閉。加入兔抗PRL-3多克隆抗體或兔抗TCTP多克隆抗體后4℃孵育過(guò)夜,內(nèi)參照用兔抗GAPDH多克隆抗體4℃孵育過(guò)夜,辣根過(guò)氧化物酶標(biāo)記的羊抗兔Ⅱ抗常溫孵育1 h。ECL kit試劑盒顯色并曝光。

        2.5 siRNA干擾 設(shè)計(jì)并合成針對(duì)TCTP的siRNA序列和陰性對(duì)照siRNA序列,TCTP-siRNA上游引物 5'-GGUAACAUUGAUGACUCGCdTdT -3',下游引物5'-GCGAGUCAUCAAUGUUACCdTdT -3'。Lo-Vo-PRL-3細(xì)胞接種于6孔板(2×105cells/well)。24 h后,按照Lipofectamine 2000說(shuō)明書,將 siRNA(轉(zhuǎn)染濃度為100 nmol/L)轉(zhuǎn)染至LoVo-PRL-3細(xì)胞。在轉(zhuǎn)染后的24 h、48 h和72 h,收集細(xì)胞進(jìn)行real-time PCR,Western blotting及細(xì)胞遷移和侵襲實(shí)驗(yàn)。

        2.6 細(xì)胞增殖實(shí)驗(yàn) 將LoVo-control與 LoVo-PRL-3細(xì)胞分別接種于 96孔板(5×103cells/well)。24 h后,將 TCTP-siRNA與 control-siRNA(轉(zhuǎn)染濃度為100 nmol/L)分別轉(zhuǎn)染至LoVo-PRL-3細(xì)胞。在轉(zhuǎn)染后的24 h、48 h和72 h,將CCK8分別加至LoVo-control細(xì)胞、LoVo-PRL-3細(xì)胞、TCTP-siRNA干擾后的細(xì)胞和control-siRNA處理后的細(xì)胞,并于37℃培養(yǎng)1 h,酶標(biāo)儀檢測(cè)450 nm吸光度值。

        2.7 細(xì)胞遷移和侵襲實(shí)驗(yàn) siRNA轉(zhuǎn)染LoVo-PRL-3細(xì)胞48 h后,消化并收集轉(zhuǎn)染后細(xì)胞,接種于Transwell雙層細(xì)胞培養(yǎng)板上室(1×105cells/well)并加入0.1 mL無(wú)血清RPMI-1640培養(yǎng)基,下室加入0.6 mL含有10%胎牛血清的RPMI-1640培養(yǎng)基。培養(yǎng)20 h后,取出上室,用棉簽擦去上室上表面的殘留細(xì)胞,將上室置于4%多聚甲醛中固定上室下表面的細(xì)胞約20 min,將膜自然晾干,結(jié)晶紫染色細(xì)胞約10 min。進(jìn)行侵襲實(shí)驗(yàn)時(shí)在上室內(nèi)鋪一層30 μL基質(zhì)膠(1∶6稀釋),其余步驟同遷移實(shí)驗(yàn)。顯微鏡下觀察每個(gè)孔上、下、左、右、中隨機(jī)選取5個(gè)視野計(jì)數(shù)細(xì)胞(×200),結(jié)果表示穿過(guò)聚碳酸酯膜的細(xì)胞數(shù)目。

        3 統(tǒng)計(jì)學(xué)處理

        結(jié) 果

        1 轉(zhuǎn)染PRL-3顯著上調(diào)LoVo細(xì)胞TCTP mRNA和蛋白的表達(dá)

        檢測(cè)發(fā)現(xiàn)轉(zhuǎn)染PRL-3后LoVo-PRL-3細(xì)胞的PRL-3表達(dá)顯著增高,見(jiàn)圖1。與對(duì)照細(xì)胞LoVo-control相比,LoVo-PRL-3細(xì)胞的 TCTP mRNA及蛋白表達(dá)均上調(diào),TCTP mRNA在LoVo-PRL-3細(xì)胞中上調(diào)1.8倍,TCTP蛋白在LoVo-control細(xì)胞中的相對(duì)表達(dá)量為0.78±0.14,而在LoVo-PRL-3細(xì)胞中的相對(duì)表達(dá)量為1.63±0.11,見(jiàn)圖2。

        Figure 1.The expression of PRL-3 in LoVo-PRL-3 cells and LoVo-control cells.±s.n=3.**P<0.01 vs control.A:PRL - 3 mRNA was determined by realtime PCR.The results show that expression level of PRL-3 mRNA in LoVo-PRL-3 cells was 430-fold higher than that in LoVo-control cells.B:Cell lysates(30 μg)from stable cell lines were used to confirm PRL-3 expression by immunoblotting with anti-PRL-3 antibody.The expression level of PRL-3 protein in LoVo-PRL-3 cells was significantly increased but not detected in LoVo-control cells.圖1 LoVo-PRL-3和LoVo-control細(xì)胞PRL-3表達(dá)水平比較

        Figure 2.The expression of TCTP in LoVo-PRL-3 cells and LoVo-control cells.±s.n=3.*P<0.05 vs control.A:TCTP mRNA was determined by real- time PCR.The results show that expression level of TCTP mRNA in LoVo-PRL-3 cells was 1.83-fold higher than that in LoVo - control cells.B:Western blotting analysis of TCTP protein level.Cell lysates(30 μg)from stable cell lines were used to confirm TCTP expression by immunoblotting with anti-TCTP antibody.The expression level of TCTP protein in LoVo-PRL-3 cells was significantly increased compared with LoVo-control cells.圖2 LoVo-PRL-3和LoVo control細(xì)胞TCTP表達(dá)水平比較

        2 TCTP-siRNA有效抑制LoVo-PRL-3細(xì)胞TCTP mRNA和蛋白的表達(dá)

        Real-time PCR結(jié)果顯示,轉(zhuǎn)染TCTP-siRNA 24 h、48 h和72 h后能顯著抑制TCTP mRNA在Lo-Vo-PRL-3細(xì)胞中的表達(dá)(P<0.01),見(jiàn)圖3A。Western blotting結(jié)果顯示,在轉(zhuǎn)染TCTP-siRNA 48h和72h后 LoVo-PRL-3細(xì)胞 TCTP蛋白的表達(dá)受到顯著抑制(P<0.01),見(jiàn)圖3B。

        Figure 3.Gene silencing of TCTP by siRNA transfection.±s.n=3.**P<0.01 vs LoVo-PRL-3 or control-siRNA.A:The level of TCTP expression in LoVo-PRL-3,control-siRNA and TCTP-siRNA cells were examined 24,48 and 72 h after siRNA transfection using quantitative real-time PCR.B:Western blotting also showed that 48 and 72 h after transfection with siRNA,LoVo-PRL-3 cells transfected with TCTP-siRNA dramatically down-regulated the expression of TCTP protein.圖3 siRNA干擾抑制TCTP表達(dá)

        3 siRNA干擾TCTP表達(dá)抑制PRL-3引起的細(xì)胞增殖

        如圖4所示,與對(duì)照細(xì)胞LoVo-control相比,轉(zhuǎn)染PRL-3后細(xì)胞的增殖能力顯著增強(qiáng)(P<0.05),siRNA干擾TCTP表達(dá)后又能顯著抑制PRL-3引起的細(xì)胞增殖(P<0.05)。

        4 siRNA干擾TCTP表達(dá)抑制PRL-3引起的細(xì)胞遷移和侵襲

        如圖5所示,與對(duì)照細(xì)胞LoVo-control相比,轉(zhuǎn)染PRL-3后細(xì)胞的遷移和侵襲能力顯著增強(qiáng)(P<0.05),siRNA干擾TCTP表達(dá)后又能顯著抑制PRL-3引起的細(xì)胞遷移和侵襲(P<0.01)。

        討 論

        結(jié)直腸上皮組織的惡性轉(zhuǎn)變及其進(jìn)一步的轉(zhuǎn)移是由多種分子共同參與完成的[10]。因此發(fā)現(xiàn)并研究腫瘤轉(zhuǎn)移過(guò)程中的重要相關(guān)分子及其信號(hào)通路對(duì)于預(yù)防和治療腫瘤轉(zhuǎn)移具有重要意義。已有大量研究表明PRL-3在腫瘤細(xì)胞轉(zhuǎn)移過(guò)程中起到重要作用[11]。目前認(rèn)為,PRL-3促進(jìn)腫瘤轉(zhuǎn)移主要通過(guò)促進(jìn)腫瘤細(xì)胞增殖、促進(jìn)腫瘤細(xì)胞遷移和侵襲及促進(jìn)血管形成等方面起作用。已有研究發(fā)現(xiàn)PRL-3高表達(dá)與卵巢癌的進(jìn)展相關(guān),并在體外通過(guò)RNA干擾方法抑制卵巢癌細(xì)胞的PRL-3表達(dá)能顯著抑制細(xì)胞增殖[4]。Zeng等[11]發(fā)現(xiàn) PRL -3 轉(zhuǎn)染至無(wú)轉(zhuǎn)移潛能的中國(guó)倉(cāng)鼠卵巢細(xì)胞(Chinese hamster ovary cell,CHO cell)后,CHO細(xì)胞的遷移和侵襲能力明顯增強(qiáng),并在裸鼠模型中形成轉(zhuǎn)移灶。此外,通過(guò)RNA干擾方法抑制具有轉(zhuǎn)移能力的結(jié)腸癌細(xì)胞DLD-1的內(nèi)源性PRL-3表達(dá)后能明顯抑制其在小鼠體內(nèi)的轉(zhuǎn)移成瘤能力[12]。同時(shí),已有研究表明,PRL -3表達(dá)增高的轉(zhuǎn)移瘤細(xì)胞可以在血管內(nèi)萌芽生長(zhǎng)[13]。在侵潤(rùn)性乳腺癌的脈管系統(tǒng)中發(fā)現(xiàn)PRL-3的表達(dá)上調(diào)[14]。綜上所述,PRL-3促進(jìn)腫瘤轉(zhuǎn)移是多種分子參與的多方面作用的結(jié)果。因此,發(fā)現(xiàn)并研究PRL-3促腫瘤轉(zhuǎn)移過(guò)程中的關(guān)鍵性分子對(duì)于預(yù)防和治療腫瘤轉(zhuǎn)移具有重要意義。

        Figure 4.Knockdown of TCTP inhibited the PRL-3-promoted proliferation of LoVo cells.±s.n=3.*P<0.05 vs LoVo-control or control-siRNA.The LoVo-control and LoVo-PRL-3 cells(5×103cells/well)were cultured in 96-well plates.After 24h,LoVo-PRL-3 cells were transfected with 100 nmol/L TCTP-siRNA or control-siRNA.Then,24,48 and 72h after transfection,cell proliferation was measured.Results show that elevated PRL-3 expression in LoVo cells promoted cell growth compared with LoVo-control cells,while knockdown of up-regulated TCTP expression in LoVo-PRL-3 cells led to suppression of cell growth.圖4 siRNA干擾TCTP表達(dá)抑制PRL-3引起的細(xì)胞增殖

        TCTP是一類廣泛存在于動(dòng)植物細(xì)胞中在序列上高度保守且同源性很高的蛋白家族。在TCTP與腫瘤相關(guān)功能研究中,Tuynder等[15]發(fā)現(xiàn),TCTP在逆轉(zhuǎn)的白血病、乳腺癌、結(jié)腸癌、肺癌和黑色素瘤細(xì)胞中均顯著下調(diào),通過(guò)反義cDNA或siRNA干擾抑制腫瘤細(xì)胞TCTP表達(dá),會(huì)導(dǎo)致腫瘤細(xì)胞惡性表型的抑制。更重要的是,最近有研究發(fā)現(xiàn),TCTP在前列腺癌和結(jié)腸癌細(xì)胞轉(zhuǎn)移的不同階段:細(xì)胞增殖、細(xì)胞凋亡、細(xì)胞遷移和侵襲及轉(zhuǎn)移均具有重要作用[8,9]。此外,已有研究表明,編碼TCTP蛋白的基因TPT1受轉(zhuǎn)錄因子CREB的調(diào)控[16]。轉(zhuǎn)錄因子CREB的活化需要其第133個(gè)絲氨酸殘基的磷酸化,細(xì)胞外信號(hào)調(diào)節(jié)激酶1/2(ERK1/2)就是引起它第133個(gè)絲氨酸殘基磷酸化的激酶之一[17]。而PRL-3可以使整合素β1磷酸化水平下調(diào),從而引起ERK1/2磷酸化水平的增加從而發(fā)揮促進(jìn)腫瘤轉(zhuǎn)移作用[18]。因此,我們猜想PRL-3可能通過(guò)上調(diào)TCTP來(lái)促進(jìn)腫瘤細(xì)胞轉(zhuǎn)移。

        Figure 5.Down-regulation of TCTP by RNAi attenuated the PRL-3-promoted migration and invasion activity of LoVo cells.±s.n=3.**P <0.01 vs LoVo-control or control-siRNA.A:LoVo-PRL-3 cells were treated with TCTP-siRNA specific for TCTP or control-siRNA.Forty-eight hours after transfection,an aliquot of 1×105cells was placed in upper chamber with 0.1 mL serum-free medium,whereas the lower chamber was loaded with 0.6 mL of medium containing 10%fetal bovine serum.After 20 h of incubation,the cells migrating or invading to the underside of filter were stained and observed under a microscope(×200).B,C:Statistical plots of the number of cells invading the Transwell membranes by migration assay and matrigel invasion assay.Results showed that PRL - 3 promoted the migration and invasion of LoVo cells,while knockdown of up-regulated TCTP expression in LoVo-PRL-3 cells led to suppression of migration and invasion.圖5 siRNA干擾TCTP表達(dá)抑制PRL-3引起的細(xì)胞遷移和侵襲

        為了證實(shí)TCTP蛋白在PRL-3促進(jìn)腫瘤轉(zhuǎn)移過(guò)程中的重要作用,我們首先將PRL-3基因轉(zhuǎn)染到內(nèi)源性低表達(dá)PRL-3的結(jié)腸癌細(xì)胞株LoVo中,并檢測(cè)LoVo-PRL-3細(xì)胞及未轉(zhuǎn)染PRL-3的對(duì)照細(xì)胞LoVo-control的TCTP表達(dá)。結(jié)果發(fā)現(xiàn)TCTP mRNA和蛋白在轉(zhuǎn)染PRL-3后分別上調(diào)1.8倍和2.1倍。之后,為了證實(shí)TCTP的上調(diào)在PRL-3促腫瘤轉(zhuǎn)移中的作用,我們通過(guò)siRNA干擾抑制LoVo-PRL-3細(xì)胞中上調(diào)的TCTP表達(dá)。結(jié)果發(fā)現(xiàn),抑制TCTP表達(dá)能顯著降低PRL-3引起的細(xì)胞增殖、遷移和侵襲能力。因此我們認(rèn)為,TCTP是PRL-3促結(jié)腸癌轉(zhuǎn)移過(guò)程中的重要分子,PRL-3通過(guò)上調(diào)TCTP表達(dá)促進(jìn)結(jié)腸癌細(xì)胞增殖、遷移和侵襲;用siRNA靶向抑制TCTP表達(dá)可能是預(yù)防和治療結(jié)腸癌轉(zhuǎn)移的有效手段。

        [1]Kim KA,Song JS,Jee J,et al.Structure of human PRL -3,the phosphatase associated with cancer metastasis[J].FEBS Lett,2004,565(1 -3):181 -187.

        [2]Saha S,Bardelli A,Buckhaults P,et al.A phosphatase associated with metastasis of colorectal cancer[J].Science,2001,294(5545):1343 -1346.

        [3]Miskad UA,Semba S,Kato H,et al.Expression of PRL -3 phosphatase in human gastric carcinomas:close correlation with invasion and metastasis[J].Pathobiology,2004,71(4):176-184.

        [4]Polato F,Codegoni A,F(xiàn)ruscio R,et al.PRL -3 phosphatase is implicated in ovarian cancer growth[J].Clin Can cer Res,2005,11(19 Pt 1):6835 -6839.

        [5]Schwering I,Brauninger A,Distler V,et al.Profiling of Hodgkin's lymphoma cell line L1236 and germinal center B cells:identification of Hodgkin's lymphoma-specific genes[J].Mol Med,2003,9(3 -4):85 -95.

        [6]Wang L,Peng L,Dong B,et al.Overexpression of phosphatase of regenerating liver-3 in breast cancer:association with a poor clinical outcome[J].Ann Oncol,2006,17(10):1517-1522.

        [7]Bommer UA,Thiele BJ.The translationally controlled tumour protein(TCTP)[J].Int J Biochem Cell Biol,2004,36(3):379 -385.

        [8]Gnanasekar M,Thirugnanam S,Zheng G,et al.Gene silencing of translationally controlled tumor protein(TCTP)by siRNA inhibits cell growth and induces apoptosis of human prostate cancer cells[J].Int J Oncol,2009,34(5):1241-1246.

        [9]Ma Q,Geng Y,Xu W,et al.The role of translationally controlled tumor protein in tumor growth and metastasis of colon adenocarcinoma cells[J].J Proteome Res,2010,9(1):40-49.

        [10]崔 翼,汪建平,彭俊生,等.結(jié)腸癌的蛋白質(zhì)組學(xué)研究[J].中國(guó)病理生理雜志,2008,24(5):1013-1017.

        [11]Zeng Q,Dong JM,Guo K,et al.PRL -3 and PRL -1 promote cell migration,invasion,and metastasis[J].Cancer Res,2003,63(11):2716 -2722.

        [12]Kato H,Semba S,Miskad UA,et al.High expression of PRL-3 promotes can cer cell motility and liver metastasis in human colorectal cancer:a predictive molecular marker of metachronous liver and lung metastases[J].Clin Cancer Res,2004,10(21):7318 -7328.

        [13]Guo K,Li J,Tang JP,et al.Catalytic domain of PRL - 3 plays an essential role in tumor metastasis:formation of PRL -3 tumors inside the blood vessels[J].Cancer Biol Ther,2004,3(10):945 -951.

        [14]Parker BS,Argani P,Cook BP,et al.Alterations in vascular gene expression in invasive breast carcinoma[J].Cancer Res,2004,64(21):7857 -7866.

        [15]Tuynder M,F(xiàn)iucci G,Prieur S,et al.Translationally controlled tumor protein is a target of tumor reversion[J].Proc Natl Acad Sci U S A,2004,101(43):15364 -15369.

        [16]Andree H,Thiele H,F(xiàn)ahling M,et al.Expression of the human TPT1 gene coding for translationally controlled tumor protein(TCTP)is regulated by CREB transcription factors[J].Gene,2006,380(2):95 -103.

        [17]Shaywitz AJ,Greenberg ME.CREB:a stimulus-induced transcription factor activated by a diverse array of extracellular signals[J].Annu Rev Biochem,1999,68:821 -861.

        [18]Peng L,Xing X,Li W,et al.PRL -3 promotes the motility,invasion,and metastasis of LoVo colon cancer cells through PRL-3-integrin beta1-ERK1/2 and-MMP2 signaling[J].Mol Cancer,2009,8:110.

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