本文選題：趨化因子CXCL1 + 腫瘤微環(huán)境��； 參考：《廣西醫(yī)科大學(xué)》2016年碩士論文

【摘要】：目的：目前研究認(rèn)為腫瘤的許多生物特性與腫瘤微環(huán)境有著密切的關(guān)系。腫瘤微環(huán)境中的許多細(xì)胞因子可以促進(jìn)腫瘤發(fā)展,也因其組織間質(zhì)壓力高、供血不足、營養(yǎng)相對缺乏,易引起腫瘤細(xì)胞發(fā)生內(nèi)質(zhì)網(wǎng)應(yīng)激,激活相關(guān)通路,促進(jìn)細(xì)胞因子表達(dá)以及腫瘤發(fā)展。許多報(bào)道認(rèn)為趨化因子CXCL1與腫瘤發(fā)生、發(fā)展、遷移、侵襲具有相關(guān)性,如在一些黑色素瘤、肝癌、胃癌、卵巢癌等均有高表達(dá),且高表達(dá)的腫瘤其惡性程度也增加。然而趨化因子CXCL1為什么在腫瘤微環(huán)境中高表達(dá),又是如何影響肝癌發(fā)生、發(fā)展、遷移、侵襲報(bào)道很少。因此,本研究從肝癌微環(huán)境的角度研究CXCL1對腫瘤作用。為進(jìn)一步研究CXCL1提供基礎(chǔ)及肝癌治療提供新的思路。方法：首先用不同濃度的衣霉素(TM)刺激HepG2細(xì)胞,引起細(xì)胞發(fā)生內(nèi)質(zhì)網(wǎng)應(yīng)激(endoplasmic reticulum stress ER Stress),提取總RNA后反轉(zhuǎn)錄,再利用Q-PCR技術(shù)檢測ELR+家族趨化因子的mRNA表達(dá)水平,同時(shí)利用ELISA方法檢測培養(yǎng)基中CXCL1的水平。用衣霉素刺激HepG2細(xì)胞12h,引起細(xì)胞發(fā)生內(nèi)質(zhì)網(wǎng)應(yīng)激后換成新鮮培養(yǎng)基,培養(yǎng)24h,收集培養(yǎng)基用于刺激HepG2細(xì)胞及誘導(dǎo)成熟的THP-1細(xì)胞,提取總RNA后反轉(zhuǎn)錄,再利用Q-PCR技術(shù)檢測其內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)基因ATF-6、PERK、IRE1的mRNA表達(dá)水平。合成SiRNA,瞬時(shí)轉(zhuǎn)染到HepG2細(xì)胞中,敲低其CXCL1表達(dá),再用衣霉素刺激HepG2細(xì)胞12h后,換成新鮮培養(yǎng)基,培養(yǎng)24h,收集培養(yǎng)基用于刺激HepG2細(xì)胞及誘導(dǎo)成熟的THP-1細(xì)胞,提取mRNA,反轉(zhuǎn)錄后利用Q-PCR技術(shù)檢測其內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)基因ATF-6、PERK、IRE1的mRNA表達(dá)水平。最后通過基因重組技術(shù)構(gòu)建CXCL1載體,確認(rèn)其在真核細(xì)胞表達(dá)后,瞬時(shí)轉(zhuǎn)染到HepG2細(xì)胞內(nèi),使CXCL1過表達(dá),再提取mRNA,反轉(zhuǎn)錄后利用Q-PCR技術(shù)檢測內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)基因ATF-6、PERK、IRE1的mRNA水平。以上所得數(shù)據(jù)均用SPSS 16.0軟件分析,采用單因素方差分析對結(jié)果進(jìn)行比較,P0.05認(rèn)為具有統(tǒng)計(jì)學(xué)意義。結(jié)果：HepG2在不同濃度的衣霉素刺激下,CXCL1、CXCL2、CXCL3的mRNA表達(dá)水平與對照組相比均升高(P0.05),CXCL8的mRNA水平降低,而CXCL5、CXCL6、CXCL7、CXCLR1、CXCLR2的mRNA未檢測出；培養(yǎng)基中的CXCL1水平升高。用TM處理過的培養(yǎng)基刺激HepG2與對照組相比內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)基因ATF-6、PERK、IRE1的mRNA表達(dá)水平均升高(P0.05)；當(dāng)CXCL1被敲低時(shí),內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)基因ATF-6、PERK、IRE1的mRNA表達(dá)水平均降低(P0.05)；CXCL1載體構(gòu)建成功并在真核細(xì)胞中表達(dá),當(dāng)CXCL1在HepG2細(xì)胞過表達(dá)時(shí),HepG2細(xì)胞的內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)基因ATF-6、PERK、IRE1的mRNA水平均升高(P0.05)。結(jié)論：在肝癌微環(huán)境中,當(dāng)細(xì)胞發(fā)生內(nèi)質(zhì)網(wǎng)應(yīng)激時(shí),可促進(jìn)CXCL1、CXCI2、 CXCL3的表達(dá),CXCL8降低,而CXCL1通過自分泌或者旁分泌又引起細(xì)胞發(fā)生內(nèi)質(zhì)網(wǎng),兩者相互促進(jìn)。
[Abstract]:Objective: many biological characteristics of tumor are closely related to tumor microenvironment. Many cytokines in tumor microenvironment can promote the development of tumor. Because of the high interstitial pressure, insufficient blood supply and relative lack of nutrition, tumor cells are prone to endoplasmic reticulum stress and activation of related pathways. Promote cytokine expression and tumor development. Many reports suggest that chemokine CXCL1 is associated with tumorigenesis, development, migration and invasion, such as high expression of CXCL1 in some melanoma, liver cancer, gastric cancer, ovarian cancer and so on. However, there are few reports on why the chemokine CXCL1 is overexpressed in tumor microenvironment and how it affects the occurrence, development, migration and invasion of HCC. Therefore, the effect of CXCL1 on tumor was studied from the point of view of microenvironment of liver cancer. It provides a new idea for the further study of CXCL1 and the treatment of liver cancer. Methods: HepG2 cells were stimulated with different concentrations of chlortetracycline (TM) to induce endoplasmic reticulum stress (ER stress), and reverse transcription was performed after total RNAs were extracted. The expression of chemokines in the ELR family was detected by Q-PCR. At the same time, the level of CXCL1 in culture medium was detected by Elisa. HepG2 cells were stimulated by itriamycin for 12 hours, and then changed into fresh medium after endoplasmic reticulum stress. The culture medium was used to stimulate HepG2 cells and induce mature THP-1 cells for 24 hours. The total RNA was extracted and reverse transcripted. Q-PCR technique was used to detect the mRNA expression level of ER stress-related gene ATF-6 PERKN IRE1. SiRNAs were synthesized and transfected into HepG2 cells, and the expression of CXCL1 in HepG2 cells was reduced. After 12 h of stimulation with itamycin, HepG2 cells were transferred to fresh culture medium for 24 h. The culture medium was used to stimulate HepG2 cells and induce mature THP-1 cells. After mRNAs were extracted, Q-PCR technique was used to detect the mRNA expression level of ER stress-related gene ATF-6 and PERKN IRE1. Finally, CXCL1 vector was constructed by gene recombination technique. After eukaryotic expression, CXCL1 was transiently transfected into HepG2 cells to overexpression CXCL1, then mRNAs were extracted. Q-PCR technique was used to detect the mRNA level of ER stress-related gene ATF-6PERKN IRE1. All the above data were analyzed by SPSS 16.0 software, and the results were compared by single factor variance analysis (P0.05). Results the mRNA expression level of CXCL1, CXCL2, CXCL2, CXCL8, CXCL8, CXCL8, CXCL8, and CXCL1 in CXCL1, CXCL1, CXCL7, CXCL6, CXCL6, CXCL7, CXCL1, and CXCL1 in culture medium of HepG2 were significantly higher than those in control group (P0.05), while the mRNA expression of CXCL1, CXCL8, CXCL8, CXCL8, CXCL8 and CXCL1 in culture medium were not detected. Compared with the control group, the mRNA expression of ER stress-related gene ATF-6PERKnIRE1 was increased in HepG2 treated with TM (P0.05), and the mRNA expression level of ER stress-related gene ATF-6 / PERKN IRE1 was decreased when CXCL1 was knocked down (P0.05). CXCL1 vector was successfully constructed and expressed in eukaryotic cells. When CXCL1 was overexpressed in HepG2 cells, the mRNA level of ER stress-related gene ATF-6 / PERK-1 IRE1 was increased in HepG2 cells (P0.05). Conclusion: in the microenvironment of liver cancer, the expression of CXCL1, CXCI2 and CXCL3 can be decreased when the endoplasmic reticulum stress occurs, while CXCL1 can induce the endoplasmic reticulum formation through autocrine or paracrine.
【學(xué)位授予單位】：廣西醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2016
【分類號】：R735.7

【相似文獻(xiàn)】

相關(guān)期刊論文 前2條

1 黎艷聰;舒嘯;黃秀華;;慢性阻塞性肺疾病患者急性加重期治療前后誘導(dǎo)痰趨化因子受體2及其配體CXCL1表達(dá)的研究[J];醫(yī)學(xué)綜述;2013年14期

2 ;[J];;年期

相關(guān)碩士學(xué)位論文 前1條

1 陳偉;CXCL1對肝癌內(nèi)質(zhì)網(wǎng)應(yīng)激作用的相關(guān)性研究[D];廣西醫(yī)科大學(xué);2016年

，

本文編號：2110634