STK33基因?qū)Ψ切〖?xì)胞肺癌相關(guān)信號(hào)通路Ras-MAPK的調(diào)控研究
發(fā)布時(shí)間：2018-09-19 09:21
【摘要】：目的：探討STK33在非小細(xì)胞肺癌(NSCLC)中對(duì)腫瘤相關(guān)信號(hào)通路Ras-MAPK的調(diào)控作用,從而為明確其在促進(jìn)NSCLC轉(zhuǎn)移及侵襲能力的發(fā)生機(jī)制方面提供一定的理論依據(jù),進(jìn)而為NSCLC的靶向治療尋找新靶點(diǎn)提供理論支持。方法：用已經(jīng)建立穩(wěn)定的高轉(zhuǎn)移性人大細(xì)胞肺癌細(xì)胞L9981和低轉(zhuǎn)移性人大細(xì)胞肺癌細(xì)胞NL9980,通過(guò)RT-PCR技術(shù)檢測(cè)L9981、NL9980中STK33的mRNA表達(dá)量。構(gòu)建STK33瞬時(shí)低表達(dá)質(zhì)粒并轉(zhuǎn)染細(xì)胞L9981,記為L(zhǎng)9981(-),構(gòu)建STK33瞬時(shí)高表達(dá)質(zhì)粒并轉(zhuǎn)染細(xì)胞NL9980,記為NL9980(+),采用RT-PCR鑒定轉(zhuǎn)染結(jié)果。轉(zhuǎn)染成功后,通過(guò)Western-blot技術(shù)檢測(cè)4組細(xì)胞中STK33和Ras-MAPK信號(hào)通路中重要蛋白PAK1、p38MAPK的蛋白表達(dá)量；通過(guò)免疫共沉淀,分析STK33和PAK1-p、p38MAPK-p之間的相互作用關(guān)系。結(jié)果：(1) STK33 在 NL9980中的mRNA表達(dá)量低于L9981(P0.001)；(2)成功構(gòu)建出STK33瞬時(shí)高表達(dá)質(zhì)粒和STK33瞬時(shí)低表達(dá)質(zhì)粒并轉(zhuǎn)染成功。(3)L9981(-)中STK33的蛋白表達(dá)量較L9981降低,NL9980(+)中STK33的蛋白表達(dá)量較NL9980增加；(4)當(dāng)STK33蛋白表達(dá)量變化時(shí),Ras-MAPK信號(hào)通路的重要蛋白PAK1,p38MAPK的非磷酸化蛋白表達(dá)水平變化不大,磷酸化蛋白在L9981(-)中的表達(dá)量低于L9981,在NL9980(+)中的表達(dá)量高于NL9980細(xì)胞。(5)用免疫共沉淀的方法檢測(cè)STK33與Ras-MAPK信號(hào)通路中磷酸化蛋白PAK1-p和p38MAPK-p的相互作用。實(shí)驗(yàn)結(jié)果證實(shí)STK33與PAK1-p無(wú)直接作用,但是與p38MAPK-p可能有直接作用。結(jié)論：STK33在不同轉(zhuǎn)移性肺癌細(xì)胞中的差異性表達(dá),提示它可能與肺癌的分期、分化程度有關(guān)。STK33表達(dá)量的變化能夠引起Ras-MAPK信號(hào)通路中磷酸化蛋白PAK1-p 和 p38MAPK-p表達(dá)量的變化,STK33高表達(dá)時(shí),磷酸化蛋白PAK1-p和p38MAPK-p表達(dá)量增加,并可能促進(jìn)上皮細(xì)胞—間充質(zhì)轉(zhuǎn)化(EMT),降低細(xì)胞凋亡,提示STK33可能能夠促進(jìn)NSCLC的轉(zhuǎn)移和侵襲能力；抑制STK33在肺癌細(xì)胞中的表達(dá),可能會(huì)使得NSCLC的轉(zhuǎn)移能力下降。STK33可能成為治療NSCLC的新靶點(diǎn)。
[Abstract]:Objective: to investigate the role of STK33 in the regulation of tumor related signal pathway (Ras-MAPK) in non-small cell lung cancer (NSCLC) (NSCLC), so as to provide a theoretical basis for the mechanism of NSCLC metastasis and invasion. Thus, it provides theoretical support for finding new targets for targeted therapy of NSCLC. Methods: the expression of STK33 mRNA in L9981NL9980 cells was detected by RT-PCR technique with the established stable high metastatic human lung cancer cell line L9981 and low metastatic human lung cancer cell line NL9980,. The transient low expression plasmid of STK33 was constructed and transfected into L9981 (-) cell line L9981 (-). The transient high expression plasmid of STK33 was constructed and the transfection cell NL9980, was recorded as NL9980 (),. The transfection results were identified by RT-PCR. After transfection, the expression of PAK1,p38MAPK, an important protein in STK33 and Ras-MAPK signaling pathway, was detected by Western-blot technique, and the interaction between STK33 and PAK1-p,p38MAPK-p was analyzed by co-immunoprecipitation. Results: (1) the mRNA expression of STK33 in NL9980 was lower than that in L9981 (P0.001); (2). The transient high expression plasmid of STK33 and the transient low expression plasmid of STK33 were successfully constructed and transfected successfully. (3) the expression of STK33 in L9981 (-) was lower than that in L9981 (-) and the expression of STK33 in L9980 (-) was higher than that in NL9980. (4) when the expression of STK33 protein changed, the expression level of non-phosphorylated protein of PAK1,p38MAPK, an important protein in Ras-MAPK signaling pathway, did not change significantly. The expression of phosphorylated protein in L9981 (-) was lower than that in L9981 (-) and was higher in NL9980 () than in NL9980 cells. (5) the interaction between STK33 and PAK1-p and p38MAPK-p in Ras-MAPK signaling pathway was detected by immunoprecipitation. The results show that STK33 has no direct effect on PAK1-p, but may have direct effect on p38MAPK-p. Conclusion the differential expression of TK33 in different metastatic lung cancer cells suggests that it may be related to the stage of lung cancer. The expression of phosphorylated protein PAK1-p and p38MAPK-p in Ras-MAPK signaling pathway increased when the expression of phosphorylated protein PAK1-p and p38MAPK-p increased when the expression level of STK33 was related to the degree of differentiation. It is suggested that STK33 can promote the metastasis and invasion of NSCLC and inhibit the expression of STK33 in lung cancer cells. STK 33 may be a new target for the treatment of NSCLC.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：R734.2
本文編號(hào)：2249712
[Abstract]:Objective: to investigate the role of STK33 in the regulation of tumor related signal pathway (Ras-MAPK) in non-small cell lung cancer (NSCLC) (NSCLC), so as to provide a theoretical basis for the mechanism of NSCLC metastasis and invasion. Thus, it provides theoretical support for finding new targets for targeted therapy of NSCLC. Methods: the expression of STK33 mRNA in L9981NL9980 cells was detected by RT-PCR technique with the established stable high metastatic human lung cancer cell line L9981 and low metastatic human lung cancer cell line NL9980,. The transient low expression plasmid of STK33 was constructed and transfected into L9981 (-) cell line L9981 (-). The transient high expression plasmid of STK33 was constructed and the transfection cell NL9980, was recorded as NL9980 (),. The transfection results were identified by RT-PCR. After transfection, the expression of PAK1,p38MAPK, an important protein in STK33 and Ras-MAPK signaling pathway, was detected by Western-blot technique, and the interaction between STK33 and PAK1-p,p38MAPK-p was analyzed by co-immunoprecipitation. Results: (1) the mRNA expression of STK33 in NL9980 was lower than that in L9981 (P0.001); (2). The transient high expression plasmid of STK33 and the transient low expression plasmid of STK33 were successfully constructed and transfected successfully. (3) the expression of STK33 in L9981 (-) was lower than that in L9981 (-) and the expression of STK33 in L9980 (-) was higher than that in NL9980. (4) when the expression of STK33 protein changed, the expression level of non-phosphorylated protein of PAK1,p38MAPK, an important protein in Ras-MAPK signaling pathway, did not change significantly. The expression of phosphorylated protein in L9981 (-) was lower than that in L9981 (-) and was higher in NL9980 () than in NL9980 cells. (5) the interaction between STK33 and PAK1-p and p38MAPK-p in Ras-MAPK signaling pathway was detected by immunoprecipitation. The results show that STK33 has no direct effect on PAK1-p, but may have direct effect on p38MAPK-p. Conclusion the differential expression of TK33 in different metastatic lung cancer cells suggests that it may be related to the stage of lung cancer. The expression of phosphorylated protein PAK1-p and p38MAPK-p in Ras-MAPK signaling pathway increased when the expression of phosphorylated protein PAK1-p and p38MAPK-p increased when the expression level of STK33 was related to the degree of differentiation. It is suggested that STK33 can promote the metastasis and invasion of NSCLC and inhibit the expression of STK33 in lung cancer cells. STK 33 may be a new target for the treatment of NSCLC.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：R734.2
【參考文獻(xiàn)】
相關(guān)期刊論文 前2條
1 范小英;左建宏;羅招陽(yáng);郭東銘;文容;;波形蛋白與腫瘤關(guān)系的研究進(jìn)展[J];現(xiàn)代生物醫(yī)學(xué)進(jìn)展;2014年13期
2 陳萬(wàn)青;張思維;曾紅梅;鄭榮壽;鄒小農(nóng);趙平;吳良有;李光琳;赫捷;;中國(guó)2010年惡性腫瘤發(fā)病與死亡[J];中國(guó)腫瘤;2014年01期
相關(guān)碩士學(xué)位論文 前1條
1 魏學(xué)強(qiáng);STK33在肺癌細(xì)胞轉(zhuǎn)移中作用的研究[D];昆明醫(yī)科大學(xué);2013年
，本文編號(hào)：2249712
本文鏈接：http://sikaile.net/yixuelunwen/zlx/2249712.html
最近更新
教材專(zhuān)著
