本文關(guān)鍵詞： 外周T細胞淋巴瘤 GATA3 JAK-STAT通路 相關(guān)性　出處：《北京協(xié)和醫(yī)學(xué)院》2016年博士論文　論文類型：學(xué)位論文

【摘要】：背景和目的外周T細胞淋巴瘤(PTCL)是一組少見、異質(zhì)性很強的非霍奇金淋巴瘤,侵襲性強、預(yù)后較差,暫無規(guī)范有效的治療方案。尋找新的分子標(biāo)記物并探索其在成瘤過程中的具體機制,對于新型靶向藥物的研發(fā)具有重要的意義。近年來國外基于高通量基因組層面的研究結(jié)果顯示轉(zhuǎn)錄因子GATA3對于PTCL的發(fā)生發(fā)展可能起到促進作用,而廣泛參與體內(nèi)生理過程的JAK-STAT信號通路對于多種腫瘤的形成和侵襲密切相關(guān)。過敏性疾病的研究顯示二者之間具有一定調(diào)控的作用。本研究擬通過在體外條件下探究GATA3和JAK-STAT通路之間相互的關(guān)系,為進一步對新型靶向藥物的研究提供新的思路。方法在體外條件下通過慢病毒轉(zhuǎn)染法建立GATA3過表達的Hut78 T細胞淋巴瘤細胞系,并結(jié)合之前已有的GATA3敲低細胞系,在RNA和蛋白層面檢測GATA3水平變化條件下JAK-STAT通路的成員STAT1、STAT6的變化情況。再反過來通過使用JAK-STAT通路特異性抑制劑及siRNA干擾的方法分別降低細胞的STAT1、 STAT6水平,檢測GATA3水平的變化。結(jié)果對T淋巴瘤細胞系Hut78應(yīng)用慢病毒轉(zhuǎn)染技術(shù)建立GATA3過表達的細胞系Hut78-pLV-GATA3,并結(jié)合已獲得的GATA3敲低的細胞系Hut78-shGATA3進行STAT1和STAT6水平檢測。與對照組相比,隨著GATA3水平降低,在RNA和蛋白層面STAT1水平均升高,STAT6水平均降低；而隨著GATA3水平升高,在RNA和蛋白層面STAT1水平均降低,RNA層面STAT6水平升高,但蛋白水平有所下降。使用JAK2抑制劑AG490處理Hut78細胞系,在RNA和蛋白層面均有GATA3和STAT6的升高和STAT1水平的降低；使用STAT1抑制劑Fludarabine處理使STAT1水平降低后,STAT6的RNA水平升高,蛋白水平變化輕度下降,GATA3的RNA水平也有下降,蛋白水平無明顯變化。使用siRNA對Hut78細胞系的STAT6進行干擾后,隨著STAT6水平的下降,STAT1的RNA水平明顯升高,蛋白水平變化不明顯；GATA3 RNA和蛋白水平均明顯下降。結(jié)論在PTCL中,GATA3水平的變化不能完全解釋STAT1和STAT6水平的變化,提示GATA3直接位于JAK-STAT通路上游的可能性不大。而STAT水平變化后,GATA3水平在RNA和蛋白水平均有所改變,提示GATA3位于STAT下游的可能性較大。
[Abstract]:Background and objective Peripheral T-cell lymphoma (PTCLs) is a rare heterogeneous non-Hodgkin 's lymphoma with strong invasion and poor prognosis. Find new molecular markers and explore their specific mechanisms in tumorigenesis. In recent years, foreign research based on high-throughput genome level results show that transcription factor GATA3 may play a role in the development of PTCL. The JAK-STAT signaling pathway, which is widely involved in physiological processes in vivo, is closely related to the formation and invasion of many kinds of tumors. To explore the relationship between GATA3 and JAK-STAT pathway in vitro. Methods GATA3 over-expressed Hut78 T cell lymphoma cell lines were established by lentivirus transfection in vitro. Combined with previous GATA3 knockout cell lines, the STAT1 of JAK-STAT pathway was detected at the RNA and protein levels. The changes of STAT6. In turn, the levels of STAT1 and STAT6 were decreased by using JAK-STAT pathway specific inhibitor and siRNA interference, respectively. Results Lentivirus transfection technique was used to establish GATA3 overexpression cell line Hut78-pLV-GATA3 in T lymphoma cell line Hut78. The levels of STAT1 and STAT6 were detected by combining the obtained GATA3 knockout cell line Hut78-shGATA3. Compared with the control group, the level of GATA3 decreased. The levels of STAT1 increased and STAT6 decreased in both RNA and protein levels. However, with the increase of GATA3 level, the level of STAT1 in both RNA and protein level decreased and STAT6 level increased. JAK2 inhibitor AG490 was used to treat Hut78 cell line. The levels of GATA3 and STAT6 increased and the level of STAT1 decreased at the level of RNA and protein. After treatment with STAT1 inhibitor Fludarabine, the RNA level of STAT6 increased and the protein level decreased slightly after STAT1 level decreased. The RNA level of GATA3 also decreased, but the protein level did not change significantly. After siRNA was used to interfere with the STAT6 of Hut78 cell line, the level of STAT6 decreased with the decrease of STAT6 level. The RNA level of STAT1 increased significantly, but the protein level did not change significantly. Conclusion the changes of GATA3 level in PTCL can not fully explain the changes of STAT1 and STAT6 levels. The results suggest that it is not possible for GATA3 to be located directly upstream of JAK-STAT pathway, but the level of GATA3 changes in RNA and protein after the change of STAT level. It suggests that GATA3 is more likely to be located downstream of STAT.
【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：R733.1

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