發(fā)布時(shí)間：2019-02-24 09:13

【摘要】：目的:探索趨化因子受體CX3CR1對(duì)人肝細(xì)胞癌,以及通過(guò)調(diào)節(jié)巨噬細(xì)胞極化對(duì)腫瘤微環(huán)境的影響及其機(jī)制。方法:在體外實(shí)驗(yàn)中,用50ng/ml氟波脂(PMA)將人單核細(xì)胞THP-1誘導(dǎo)成巨噬細(xì)胞(M0),并用干擾RNA干擾M0細(xì)胞中的CX3CR1,然后通過(guò)RT-PCR的方法檢測(cè)CX3CR1的表達(dá)水平。處理后的巨噬細(xì)胞表型通過(guò)RT-PCR檢測(cè)M1和M2相關(guān)標(biāo)記來(lái)確定,然后分別收集巨噬細(xì)胞M0和干擾CX3CR1后巨噬細(xì)胞的條件培養(yǎng)基,并將條件培養(yǎng)基加入到Hep G2和7721細(xì)胞中,通過(guò)流式、MTT、劃痕、Transwell等試驗(yàn)方法觀察腫瘤細(xì)胞的增殖、凋亡、遷移、侵襲等生物學(xué)效應(yīng)的改變,同時(shí)用過(guò)表達(dá)質(zhì)粒和干擾RNA,過(guò)表達(dá)7721細(xì)胞中的CX3CR1、抑制HepG2細(xì)胞中的CX3CR1的表達(dá),通過(guò)MTT、流式、劃痕、Transwell等方法檢測(cè)腫瘤細(xì)胞增殖、遷移、侵襲等改變,最后通過(guò)Western blotting的方法檢測(cè)腫瘤細(xì)胞中PI3K-AKT和ERK-MAPK相關(guān)信號(hào)通路的變化,初步探究CX3CR1發(fā)揮調(diào)節(jié)作用的機(jī)制。結(jié)果:在用干擾RNA處理誘導(dǎo)成功的巨噬細(xì)胞后,巨噬細(xì)胞中的CX3CR1的水平受到了明顯的抑制,檢測(cè)巨噬細(xì)胞相關(guān)指標(biāo)發(fā)現(xiàn)M1型巨噬細(xì)胞相關(guān)標(biāo)記升高,M2型相關(guān)標(biāo)記下降,將收集的條件培養(yǎng)基加入Hep G2和7721細(xì)胞中觀察到,HepG2和7721細(xì)胞的增殖、遷移、侵襲能力受到明顯抑制,凋亡增加,過(guò)表達(dá)CX3CR1促進(jìn)了7721細(xì)胞的增殖、遷移和侵襲能力,干擾CX3CR1抑制了HepG2細(xì)胞的增殖、遷移和侵襲能了。結(jié)論:抑制了CX3CR1的表達(dá)后,可促進(jìn)巨噬細(xì)胞向M1型極化,并且抑制了HepG2和7721細(xì)胞的增殖、遷移、侵襲,促進(jìn)了腫瘤細(xì)胞的凋亡,過(guò)表達(dá)CX3CR1促進(jìn)了7721細(xì)胞的增殖、遷移和侵襲能力,干擾CX3CR1抑制了HepG2細(xì)胞的增殖、遷移和侵襲能了。
[Abstract]:Aim: to investigate the effects of chemokine receptor (CX3CR1) on human hepatocellular carcinoma (HCC) and the effect of macrophage polarization on tumor microenvironment. Methods: in vitro, human monocyte THP-1 was induced into macrophages (M0) by 50ng/ml fluroid (PMA), and CX3CR1, in M0 cells was interfered by RNA. Then the expression of CX3CR1 was detected by RT-PCR. The phenotypes of treated macrophages were determined by RT-PCR detection of M1 and M2 markers, then the macrophage M0 and conditioned medium interfering with CX3CR1 were collected, and the conditioned medium was added to Hep G2 and 7721 cells. The biological effects of proliferation, apoptosis, migration and invasion of tumor cells were observed by flow cytometry, MTT, scratch and Transwell. The expression plasmid and CX3CR1, in 7721 cells were also used to interfere with RNA, overexpression. The expression of CX3CR1 was inhibited in HepG2 cells. The changes of proliferation, migration and invasion of tumor cells were detected by MTT, flow, scratch and Transwell. Finally, the changes of PI3K-AKT and ERK-MAPK related signaling pathways in tumor cells were detected by Western blotting. To explore the mechanism of regulation of CX3CR1. Results: the level of CX3CR1 in macrophages induced by interfering RNA was significantly inhibited. The related markers of M 1 type macrophages were increased, while M 2 type related markers were decreased. It was observed that the proliferation, migration, invasion and apoptosis of HepG2 and 7721 cells were significantly inhibited and apoptosis increased. Overexpression of CX3CR1 promoted the proliferation, migration and invasion of 7721 cells. Interfering with CX3CR1 inhibits the proliferation, migration and invasion of HepG2 cells. Conclusion: inhibiting the expression of CX3CR1 can promote macrophage polarization to M1 type, inhibit the proliferation, migration and invasion of HepG2 and 7721 cells, promote the apoptosis of tumor cells, and over-express CX3CR1 can promote the proliferation of 7721 cells. The ability of migration and invasion inhibited the proliferation of HepG2 cells by interfering with CX3CR1.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R735.7

【參考文獻(xiàn)】

相關(guān)期刊論文 前4條

1 Hong-Yan Xie;Zhi-Min Shao;Da-Qiang Li;;Tumor microenvironment:driving forces and potential therapeutic targets for breast cancer metastasis[J];Chinese Journal of Cancer;2017年03期

2 Jinlu Dai;Yi Lu;Hernan Roca;Jill M.Keller;Jian Zhang;Laurie K.Mc Cauley;Evan T.Keller;;Immune mediators in the tumor microenvironment of prostate cancer[J];Chinese Journal of Cancer;2017年03期

3 Shweta Gera;Mark Ettel;Gabriel Acosta-Gonzalez;Ruliang Xu;;Clinical features,histology,and histogenesis of combined hepatocellular-cholangiocarcinoma[J];World Journal of Hepatology;2017年06期

4 Dimitrios N Samonakis;Elias A Kouroumalis;;Systemic treatment for hepatocellular carcinoma: Still unmet expectations[J];World Journal of Hepatology;2017年02期

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本文編號(hào)：2429399