【摘要】：目的研究PFDA、PFOS、PFOA對(duì)胃上皮細(xì)胞增殖的影響,并探究PFDA影響胃上皮細(xì)胞增殖的分子機(jī)制,這不僅可以為解析污染環(huán)境中胃癌的發(fā)生提供新的線索,更促使人類關(guān)注環(huán)境污染對(duì)人類健康造成的危害并采取相應(yīng)的治理措施。方法1.檢測(cè)PFDA、PFOS、PFOA對(duì)胃上皮細(xì)胞增殖情況的影響。以PFDA、PFOS、PFOA處理胃上皮細(xì)胞AGS,72h后各組取300個(gè)細(xì)胞進(jìn)行克隆形成實(shí)驗(yàn),單克隆生長(zhǎng)至肉眼可見(jiàn)時(shí),進(jìn)行結(jié)晶紫染色、計(jì)數(shù),檢測(cè)PFDA、PFOS、PFOA對(duì)胃上皮細(xì)胞增殖情況的影響。2.細(xì)胞凋亡、細(xì)胞周期、自噬、衰老以PFDA處理AGS細(xì)胞,72h后進(jìn)行FITC、PI染色,采用流式細(xì)胞術(shù)檢測(cè)PFDA對(duì)AGS細(xì)胞的細(xì)胞周期、細(xì)胞凋亡的影響。采用Western Blotting在蛋白質(zhì)水平檢測(cè)自噬相關(guān)蛋白P62、LC3的表達(dá)；β-半乳糖苷酶染色研究PFDA對(duì)AGS細(xì)胞衰老的影響,并探索衰老在PFDA促進(jìn)AGS細(xì)胞增殖中的分子機(jī)制。3.生物芯片分析PFDA促進(jìn)胃上皮細(xì)胞增殖可能的靶基因。以PFDA處理AGS細(xì)胞,72h后收取細(xì)胞,進(jìn)行表達(dá)譜生物芯片分析,并以qRT-PCR方法對(duì)顯著變化的靶基因在mRNA水平進(jìn)行驗(yàn)證。4. TCF4、PLA2G2A在PFDA促進(jìn)AGS細(xì)胞增殖的機(jī)制中的作用。以PFDA處理AGS細(xì)胞24h后,TCF4、PLA2G2A表達(dá)質(zhì)粒分別轉(zhuǎn)染,48h后提取RNA,進(jìn)行qRT-PCR,檢測(cè)TCF4、PLA2G2A mRNA的表達(dá),并進(jìn)行p-半乳糖苷酶染色檢測(cè)質(zhì)粒轉(zhuǎn)染后AGS細(xì)胞的衰老變化。結(jié)果1.克隆形成結(jié)果顯示,PFDA、PFOS、PFOA都能夠促進(jìn)AGS細(xì)胞的增殖,但促進(jìn)AGS細(xì)胞增殖的能力不同,大小順序依次為PFDAPFOSPFOA。2.流式細(xì)胞術(shù)結(jié)果顯示,PFDA對(duì)胃上皮細(xì)胞的細(xì)胞周期沒(méi)有影響,但能夠抑制細(xì)胞凋亡,但不明顯；同時(shí)自噬相關(guān)蛋白P62、LC3的表達(dá)上調(diào),表明自噬在一定程度上抑制了AGS細(xì)胞的增殖。3.基因芯片表達(dá)譜結(jié)果顯示,PFDA處理AGS細(xì)胞后,在mRNA水平PLA2G2A、TCF4的表達(dá)顯著降低。4. qRT-PCR結(jié)果表明,PFDA通過(guò)TCF4下調(diào)PLA2G2A的表達(dá)進(jìn)而抑制細(xì)胞的衰老,β-Galactosidase staining結(jié)果表明TCF4、PLA2G2A表達(dá)質(zhì)粒轉(zhuǎn)染后AGS衰老的細(xì)胞數(shù)量顯著增加。結(jié)論1. PFDA促進(jìn)胃上皮細(xì)胞AGS增殖。2. PFDA通過(guò)抑制胃上皮細(xì)胞的衰老進(jìn)而促進(jìn)細(xì)胞增殖。3. PFDA通過(guò)TCF4下調(diào)PLA2G2A的表達(dá)抑制細(xì)胞衰老,進(jìn)而發(fā)揮促進(jìn)細(xì)胞增殖的作用。
[Abstract]:Objective to study the effect of PFDA,PFOS,PFOA on the proliferation of gastric epithelial cells and explore the molecular mechanism of PFDA affecting the proliferation of gastric epithelial cells, which can not only provide a new clue for analyzing the occurrence of gastric cancer in polluted environment. It also urges human beings to pay attention to the harm to human health caused by environmental pollution and to take corresponding measures to deal with it. Method 1. The effect of PFDA,PFOS,PFOA on the proliferation of gastric epithelial cells was detected. After the gastric epithelial cells were treated with PFDA,PFOS,PFOA, 300 cells were taken from each group for clone formation test. When the clones were grown to be visible to the naked eye, crystal violet staining was carried out, counting, and detection of PFDA,PFOS, was carried out. Effect of PFOA on proliferation of gastric epithelial cells. Apoptosis, cell cycle, autophagy and senescence of AGS cells were treated with PFDA. FITC,PI staining was performed after 72 hours. The effects of PFDA on cell cycle and apoptosis of AGS cells were detected by flow cytometry. Western Blotting was used to detect the expression of autophagy-associated protein P62 and LC3 at the protein level, 尾-galactosidase staining was used to study the effect of PFDA on the senescence of AGS cells, and the molecular mechanism of senescence in the promotion of AGS cell proliferation by PFDA was explored. Biochip was used to analyze the possible target genes of PFDA promoting the proliferation of gastric epithelial cells. AGS cells were treated with PFDA and collected 72 hours later. The expression profiles of AGS cells were analyzed by biochip analysis. The significantly changed target genes were verified by qRT-PCR at mRNA level. 4. The role of TCF4,PLA2G2A in the mechanism of PFDA promoting the proliferation of AGS cells. AGS cells were treated with PFDA for 24 hours and then transfected with TCF4,PLA2G2A expression plasmid. 48 hours later, RNA, was extracted and qRT-PCR, was used to detect the expression of TCF4,PLA2G2A mRNA, and the senescence changes of AGS cells were detected by pgalactosidase staining. Outcome 1. The results of clone formation showed that PFDA,PFOS,PFOA could promote the proliferation of AGS cells, but the ability to promote the proliferation of AGS cells was different, and the order of size was PFDAPFOSPFOA.2.. The results of flow cytometry showed that PFDA had no effect on the cell cycle of gastric epithelial cells, but could inhibit the apoptosis of gastric epithelial cells, but it was not obvious. At the same time, the expression of autophagy-associated protein P62 and LC3 was up-regulated, indicating that autophagy inhibited the proliferation of AGS cells to a certain extent. The gene chip expression profile showed that the expression of PLA2G2A,TCF4 in AGS cells treated with PFDA decreased significantly at the mRNA level. 4. The results of qRT-PCR showed that PFDA inhibited the senescence of AGS cells by down-regulating the expression of PLA2G2A through TCF4. The results of 尾-Galactosidase staining showed that the number of senescent cells of AGS transfected with TCF4,PLA2G2A expression plasmid increased significantly. Conclusion 1. PFDA promotes the proliferation of AGS in gastric epithelial cells. PFDA promotes cell proliferation by inhibiting the aging of gastric epithelial cells. 3. PFDA inhibits cell senescence by down-regulating the expression of PLA2G2A through TCF4, thus promoting cell proliferation.
【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：R735.2

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 怡悅;芫荽促進(jìn)毛囊上皮細(xì)胞增殖的作用[J];國(guó)外醫(yī)學(xué)(中醫(yī)中藥分冊(cè));1999年04期

2 彭惠,李平華,羅莉霞,駱云鵬,湯為學(xué);堿性成纖維細(xì)胞生長(zhǎng)因子促進(jìn)兔晶體上皮細(xì)胞增殖的實(shí)驗(yàn)研究[J];中國(guó)病理生理雜志;2000年10期

3 王娟,李筱榮,張曉紅,孫慧敏,袁佳琴;表皮生長(zhǎng)因子對(duì)晶體上皮細(xì)胞增殖影響的研究[J];中華眼科雜志;1999年04期

4 彭惠,李平華,羅莉霞,駱云鵬,湯為學(xué);堿性成纖維細(xì)胞生長(zhǎng)因子對(duì)兔晶體上皮細(xì)胞增殖作用的免疫組織化學(xué)研究[J];中國(guó)病理生理雜志;2000年10期

5 高虹,王吉耀,沈錫中,劉建軍;幽門螺桿菌感染對(duì)胃黏膜上皮細(xì)胞增殖的影響[J];中華消化雜志;2000年06期

6 羅莉霞;藥物抑制晶體上皮細(xì)胞增殖的研究進(jìn)展[J];重慶醫(yī)科大學(xué)學(xué)報(bào);2002年03期

7 陳祥貴,王瑞淑,鄧茂先,冉新澤,粟永萍;外源核苷酸對(duì)腸上皮細(xì)胞增殖和遷移的作用[J];四川工業(yè)學(xué)院學(xué)報(bào);2003年S2期

8 劉曉明,孫秀坤,齊欣,林熙然;20種中藥灌胃對(duì)小鼠上皮細(xì)胞增殖和表皮細(xì)胞分化及血漿內(nèi)皮素-1的影響[J];中華皮膚科雜志;2001年04期

9 胡金瑋,黃維國(guó),姜鴻彥,邱建華,劉順利,王錦玲;豚鼠發(fā)育過(guò)程前庭上皮細(xì)胞增殖和凋亡的關(guān)系[J];中華耳鼻咽喉科雜志;2001年03期

10 楊藝,鄧長(zhǎng)生,彭俊忠;幽門螺桿菌對(duì)體外培養(yǎng)的胃上皮細(xì)胞增殖與凋亡的影響[J];中華微生物學(xué)和免疫學(xué)雜志;2002年01期

相關(guān)會(huì)議論文 前10條

1 霍亞楠;姚玉峰;裘文亞;;氧自由基在表皮生長(zhǎng)因子刺激的角膜上皮細(xì)胞增殖中的作用[A];2006年浙江省眼科學(xué)術(shù)會(huì)議論文集[C];2006年

2 侯素春;劉曉明;林熙然;郭永江;;六種中藥方劑灌胃對(duì)小鼠上皮細(xì)胞增殖和表皮細(xì)胞分化及血漿內(nèi)皮素-1的影響[A];2002中國(guó)中西醫(yī)結(jié)合皮膚性病學(xué)術(shù)會(huì)議論文匯編[C];2002年

3 劉佳;儲(chǔ)臘萍;王笑云;;血管緊張素Ⅱ及氟伐他汀對(duì)腎小管上皮細(xì)胞增殖的影響[A];中華醫(yī)學(xué)會(huì)腎臟病學(xué)分會(huì)2006年學(xué)術(shù)年會(huì)論文集[C];2006年

4 熊錫山;梅長(zhǎng)林;付莉莉;李林;段文虎;蔣華良;;新型酪氨酸激酶抑制劑Zhao260054抑制囊腫襯里上皮細(xì)胞增殖機(jī)制初探[A];中華醫(yī)學(xué)會(huì)腎臟病學(xué)分會(huì)2006年學(xué)術(shù)年會(huì)論文集[C];2006年

5 陳祥貴;王瑞淑;鄧茂先;冉新澤;粟永萍;;外源核苷酸對(duì)腸上皮細(xì)胞增殖和遷移的作用[A];達(dá)能營(yíng)養(yǎng)中心青年科學(xué)工作者論壇優(yōu)秀論文集2002年第3期[C];2002年

6 朱海萍;寧宏;;Skp2在晶體上皮細(xì)胞增殖中的作用及與p27 kip1關(guān)系的實(shí)驗(yàn)研究[A];中華醫(yī)學(xué)會(huì)第十二屆全國(guó)眼科學(xué)術(shù)大會(huì)論文匯編[C];2007年

7 劉曉明;孫秀坤;齊欣;林熙然;;20種中藥灌胃對(duì)小鼠上皮細(xì)胞增殖和表皮細(xì)胞分化及血漿內(nèi)皮素-1的影響[A];2001年中國(guó)中西醫(yī)結(jié)合皮膚性病學(xué)術(shù)會(huì)議論文匯編[C];2001年

8 朱莉;靳蓉;王鴻娟;李惠;詹泉;劉文莉;全小麗;;兒童CagA~+幽門螺桿菌感染根治前后胃粘膜上皮細(xì)胞增殖、凋亡和P53蛋白水平表達(dá)的對(duì)比研究[A];2006年消化系病學(xué)術(shù)年會(huì)暨胰腺疾病專題講座論文匯編[C];2006年

9 劉平;石廣森;;三氧化二砷對(duì)兔晶體上皮細(xì)胞增殖影響的研究[A];中華醫(yī)學(xué)會(huì)第十二屆全國(guó)眼科學(xué)術(shù)大會(huì)論文匯編[C];2007年

10 崔龍;孫紅梅;許紅;吳海霞;白麗敏;;中藥對(duì)EAE大鼠脊髓室膜上皮細(xì)胞增殖及髓鞘再生的影響[A];中國(guó)神經(jīng)科學(xué)學(xué)會(huì)第六屆學(xué)術(shù)會(huì)議暨學(xué)會(huì)成立十周年慶祝大會(huì)論文摘要匯編[C];2005年

相關(guān)重要報(bào)紙文章 前1條

1 白玉;四環(huán)生物“博弈”納米新藥[N];醫(yī)藥經(jīng)濟(jì)報(bào);2003年

相關(guān)碩士學(xué)位論文 前8條

1 彭艷萍;全氟癸酸（PFDA）促進(jìn)胃上皮細(xì)胞增殖的機(jī)制研究[D];山東大學(xué);2016年

2 楊靜;8-氯腺苷對(duì)人晶體上皮細(xì)胞增殖影響的體外研究[D];華中科技大學(xué);2006年

3 彭惠;堿性成纖維細(xì)胞生長(zhǎng)因子對(duì)兔晶體上皮細(xì)胞增殖的影響及與藥物相互作用的實(shí)驗(yàn)研究[D];重慶醫(yī)科大學(xué);2001年

4 趙杰;雷帕霉素對(duì)體外培養(yǎng)的兔晶體上皮細(xì)胞增殖的影響[D];山東大學(xué);2005年

5 張玉霞;堿性成纖維細(xì)胞生長(zhǎng)因子對(duì)犬角膜上皮細(xì)胞增殖影響的研究[D];華中農(nóng)業(yè)大學(xué);2007年

6 楊慧慧;PPA對(duì)人呼吸道上皮細(xì)胞增殖、TLR4、Act1、A20表達(dá)的影響[D];中南大學(xué);2011年

7 王丹;蘚化飲對(duì)體外培養(yǎng)的口腔上皮細(xì)胞增殖活性的影響及PCNA在口腔扁平苔蘚組織中的表達(dá)及意義[D];河北醫(yī)科大學(xué);2008年

8 高宇端;連接蛋白43在晶體上皮細(xì)胞增殖過(guò)程中表達(dá)變化的研究[D];中國(guó)醫(yī)科大學(xué);2005年

，

本文編號(hào)：2465373