【摘要】：microRNA是最近研究發(fā)現(xiàn)的一類(lèi)長(zhǎng)度約為19-25個(gè)核苷酸的非編碼單鏈小RNA分子,其通過(guò)與靶基因的3’端非翻譯區(qū)互補(bǔ)配對(duì)結(jié)合的方法,從而調(diào)控內(nèi)源基因的表達(dá)。miRNA作為生物體生長(zhǎng)發(fā)育的重要調(diào)控基因,目前其主要研究方向是探明miRNAs與疾病的關(guān)系,尤其是miRNA與腫瘤的關(guān)系。鼻咽癌(Nasopharyngeal carcinoma, NPC)是廣東地區(qū)常見(jiàn)的惡性腫瘤之一,在頭頸部惡性腫瘤中占首位。鼻咽癌的發(fā)病人群以40-50歲的青壯年多見(jiàn),一旦發(fā)病對(duì)社會(huì)、經(jīng)濟(jì)和家庭造成較大影響。鼻咽癌原發(fā)部位隱蔽,不易觀察,且與鼻腔、鼻竇和顱內(nèi)相毗鄰,所以臨床癥狀出現(xiàn)較晚而且各異；鼻咽癌也是頭頸腫瘤中轉(zhuǎn)移率最高的。這些都導(dǎo)致了鼻咽癌常常容易被誤診或漏診,影響治療效果。因此,鼻咽癌的早期診斷和治療的研究一直是我國(guó)頭頸外科學(xué)研究的重點(diǎn)之一。目前鼻咽癌的治療還是以放射治療為主。對(duì)放療不敏感、放療后復(fù)發(fā)以及中晚期的患者,可采取化學(xué)藥物治療,但這只是輔助性治療或姑息性治療。鼻咽癌的手術(shù)治療也只在一些特殊情況下作為輔助方法采用。除了以上三種常規(guī)治療,近年來(lái)也有作者嘗試應(yīng)用光動(dòng)力療法和微波熱療治療鼻咽癌,但也只能作為輔助手段。重組DNA技術(shù)的發(fā)展使腫瘤的基因治療成為可能。根據(jù)腫瘤發(fā)生機(jī)理制定的基因治療方案已成為腫瘤治療的熱點(diǎn)。早期研究者主要運(yùn)用反義核酸技術(shù)干預(yù)腫瘤相關(guān)基因的表達(dá),取得了一定效果。近年來(lái),RNA干擾技術(shù)在哺乳動(dòng)物體內(nèi)的應(yīng)用,更為腫瘤的基因治療開(kāi)辟了一個(gè)新的視野。因此,我們意在探索性地研究miRNA在鼻咽癌發(fā)生發(fā)展中的調(diào)控作用。 我們通過(guò)Targetscan數(shù)據(jù)庫(kù),對(duì)差異表達(dá)miRNA的靶基因進(jìn)行預(yù)測(cè),并將預(yù)測(cè)結(jié)果輸入GenMAPP軟件進(jìn)行生物學(xué)通路分析。在通路分析中,大部分靶基因富集度高的通路都涉及到信號(hào)轉(zhuǎn)導(dǎo)及與癌癥發(fā)生相關(guān)的通路,例如EGFR-1信號(hào)通路,MAPK信號(hào)通路等等。其中，，小GTP酶介導(dǎo)的信號(hào)轉(zhuǎn)導(dǎo)通路最為顯著。同時(shí),Wnt信號(hào)通路的顯著性,則與我們實(shí)驗(yàn)室過(guò)去在鼻咽癌cDNA表達(dá)譜的研究結(jié)果相吻合。繼續(xù)深入研究這些差異]miRNA在生物學(xué)通路中的調(diào)控作用,以及尋找與鼻咽癌發(fā)生發(fā)展進(jìn)程相關(guān)的臨床診斷標(biāo)記物,則是我們未來(lái)努力的方向。 microRNA (miRNA)即微小RNA,是一種存在于植物和動(dòng)物基因組里的小分子RNA,約19～25nt(少數(shù)小于20nt),由一段具有發(fā)夾環(huán)結(jié)構(gòu)、長(zhǎng)度為70-80nt的單鏈RNA前體(Pre-miRNA)剪切后形成。它通過(guò)與其目標(biāo)mRNA分子的3’端非編碼區(qū)域(3'-untranslatedregion,3'UTR)完全或非完全互補(bǔ)匹配、參與基因轉(zhuǎn)錄后水平(蛋白表達(dá))的調(diào)控,在生物體內(nèi)各種生理和病理狀態(tài)都發(fā)揮著十分重要的作用。 miRNA最早由Lee等在1993年對(duì)線蟲(chóng)(Caenothabditis elegans)胚胎發(fā)育研究過(guò)程中發(fā)現(xiàn)的一種具有調(diào)控功能的非編碼RNA。首先,miRNA基因的初級(jí)轉(zhuǎn)錄產(chǎn)物(Pri-miRNA)在細(xì)胞核中被RNase (?) Drosha切割成為前體miRNA (pre-miRNA)。在最初的剪切后,Pre-miRNA在轉(zhuǎn)運(yùn)蛋白exPortin-5的作用下、由核內(nèi)轉(zhuǎn)到胞質(zhì)中,然后由另一種RNase (?) Dicer進(jìn)一步切割產(chǎn)生成熟的miRNA.這些成熟的miRNA與其他蛋白質(zhì)一起組成RISC (RNA-induced sileneing complex)復(fù)合體,從而引起靶mRNA的降解或者翻譯抑制。 microRNAs具有重要的組織特異功能,在基因表達(dá)中發(fā)揮著總體調(diào)控作用,其中miR-421在腫瘤進(jìn)展調(diào)控,細(xì)胞增殖等方面起著重要的作用,目前對(duì)于鼻咽癌的調(diào)控作用尚缺少文獻(xiàn)報(bào)道。本課題將探討miR-421在鼻咽癌發(fā)生發(fā)展中的作用。本研究的目的是模擬天然miRNA構(gòu)建表達(dá)miRNA前體的質(zhì)粒,實(shí)現(xiàn)調(diào)控腫瘤相關(guān)基因miR-421的表達(dá),開(kāi)展對(duì)鼻咽癌的基因治療研究。 首先是調(diào)控目標(biāo)基因的miR-421表達(dá)質(zhì)粒的構(gòu)建和有效質(zhì)粒篩選。然后進(jìn)一步確定針對(duì)FOXO4基因的1niRNA的有效作用位點(diǎn),觀察構(gòu)建的質(zhì)粒對(duì)鼻咽癌中兩個(gè)靶基因的調(diào)控效應(yīng),為后續(xù)的體外實(shí)驗(yàn)奠定基礎(chǔ)。miRNA重組質(zhì)粒調(diào)控FOX04表達(dá)能有效抑制腫瘤生長(zhǎng),為進(jìn)一步的臨床研究打下了基礎(chǔ)。 第一部分 1、過(guò)表達(dá)miR-.421促進(jìn)鼻咽癌細(xì)胞增殖和抗凋亡的作用 目的：檢測(cè)過(guò)表達(dá)miR-421對(duì)于鼻咽癌細(xì)胞增殖和抗凋亡的能力。 方法：利用慢病毒感染技術(shù),將miR-421轉(zhuǎn)染到CNE2,觀察體外miR-421對(duì)CNE2增殖作用,并利用Annexin V法及TUNEL檢測(cè)細(xì)胞凋亡情況。 結(jié)果：軟瓊脂克隆形成實(shí)驗(yàn)表明,miR-421過(guò)表達(dá)細(xì)胞的增殖速度是對(duì)照組的3倍,表明miR-421上調(diào)可以顯著增強(qiáng)鼻咽癌細(xì)胞的生長(zhǎng)。此外,Annexin V法及TUNEL檢測(cè)表明,miR-421的過(guò)表達(dá)可增強(qiáng)鼻咽癌細(xì)胞對(duì)于化療藥物順鉑的凋亡抵抗能力。這些結(jié)果表明,體外實(shí)驗(yàn)中,miR-421在鼻咽癌中扮演著調(diào)控細(xì)胞基因的作用。 結(jié)論：miR-421上調(diào)可促進(jìn)入鼻咽癌細(xì)胞的增殖,提高細(xì)胞凋亡抵抗能力。 2、抑制miR-421減少鼻咽癌細(xì)胞增殖和誘導(dǎo)細(xì)胞凋亡的作用 目的：檢測(cè)抑制miR-421對(duì)于鼻咽癌細(xì)胞增殖和凋亡的影響 方法：利用慢病毒感染技術(shù),將抑制miR-421轉(zhuǎn)染CNE2,觀察體外miR-421對(duì)CNE2增殖作用,并利用Annexin V法及TUNEL檢測(cè)細(xì)胞凋亡情況。 結(jié)果：軟瓊脂克隆形成實(shí)驗(yàn)表明,抑制miR-421后細(xì)胞的增殖速度降低,對(duì)照組是抑制組的3倍,表明抑制miR-421可以顯著降低鼻咽癌細(xì)胞的生長(zhǎng)。此外,Annexin V法及TUNEL檢測(cè)表明,抑制miR-421可增強(qiáng)鼻咽癌細(xì)胞對(duì)于化療藥物順鉑的凋亡促進(jìn)能力。這些結(jié)果表明,體外實(shí)驗(yàn)中,miR-421在鼻咽癌中扮演著調(diào)控細(xì)胞基因的作用。 結(jié)論：抑制miR-421可降低人鼻咽癌細(xì)胞的增殖,促進(jìn)細(xì)胞凋亡。 第二部分 1、探討miR-421對(duì)FOXO4信號(hào)途徑的作用 目的：研究miR-421促進(jìn)鼻咽癌細(xì)胞增殖及抗凋亡的內(nèi)在分子機(jī)制是否通過(guò)FOXO4信號(hào)途徑調(diào)節(jié)。 方法：構(gòu)建針對(duì)耙基因FOXO4的熒光素酶報(bào)告基因載體,利用熒光照度計(jì)分別測(cè)定正常組、miR-421過(guò)表達(dá)組與miR-421抑制組的熒光值。并利用實(shí)時(shí)定量PCR及western blot檢測(cè)FOXO4靶基因p21, p27, Bim和FasL的mRNA及蛋白表達(dá)情況。 結(jié)果：1.在熒光素酶活性檢測(cè)中,FOXO4的熒光素酶活性在miR-421過(guò)表達(dá)的細(xì)胞中降低,而在miR-421抑制的細(xì)胞中表達(dá)增多。2.與此相一致的是,在正常組、miR-421過(guò)表達(dá)組與miR-421抑制組中我們發(fā)現(xiàn),FOXO4四個(gè)經(jīng)典靶點(diǎn)基因p21, p27, Bim和FasL的mRNA及蛋白表達(dá)在miR-421過(guò)表達(dá)組是下調(diào),而在miR-421抑制的細(xì)胞中,上述基因及蛋白上調(diào)。 結(jié)論：通過(guò)構(gòu)建熒光素酶以及過(guò)表達(dá)和抑制miR-421實(shí)驗(yàn)結(jié)果表明,miR-421可抑制FOXO4信號(hào)通路。 2、miR-421與FOXO4調(diào)控關(guān)系探討 目的：探尋miR-421在基因調(diào)控中針對(duì)耙基因FOXO4調(diào)節(jié)作用。 方法：使用TargetScan軟件工具mir-421目標(biāo)預(yù)測(cè)分析表明,FOXO4是mir-421潛在目標(biāo),利用免疫印跡分析表明,在正常組、miR-421過(guò)表達(dá)組與miR-421抑制組中基因FOXO4的表達(dá)存在顯著的差異性,過(guò)表達(dá)組蛋白表達(dá)低于正常組,而抑制組蛋白表達(dá)高于正常組。 為了進(jìn)一步驗(yàn)證miR-421與FOXO4相互控制存在的關(guān)系,我們克隆了FOXO43'UTR片段,含有miR-421結(jié)合位點(diǎn)的psiCHECK-2熒光素酶載體,同時(shí)我們克隆了FOXO43'UTR突變載體,具體突變了二個(gè)堿基。 通過(guò)熒光素酶活性檢測(cè)表明,miR-421過(guò)表達(dá)組低于正常組,而miR-421抑制組高于正常組。 另外,為了進(jìn)一步分析miR-421與FOXO4在凋亡中的相互作用,在抑制miR-421的時(shí),同時(shí)抑制FOXO4作對(duì)照,實(shí)驗(yàn)說(shuō)明,抑制FOXO4基因后,細(xì)胞凋亡低于單純抑制miR-421組。 結(jié)果：1、miR-421過(guò)表達(dá)與miR-421抑制組中,基因FOXO4表達(dá)存在差異性；2、熒光素酶檢測(cè)表明,miR-421過(guò)表達(dá)與miR-421抑制組中熒光值存在差異性,而在突變體卻無(wú)此差異性；3、同時(shí)抑制miR-421時(shí),選擇下調(diào)基因FOXO4,結(jié)果表明,凋亡細(xì)胞低于單純抑制miR-421組。 結(jié)論：基因FOXO4是miR-421調(diào)控目的基因。 3. FOXO4是細(xì)胞生長(zhǎng)和抗凋亡的關(guān)鍵基因 目的：探尋FOXO4在細(xì)胞生長(zhǎng)和抗凋亡的調(diào)節(jié)作用。 方法：為了進(jìn)一步分析miR-421與FOXO4在凋亡中的作用,我們選擇將miR-421抑制時(shí),然后再單純抑制基因FOXO4來(lái)對(duì)照,檢測(cè)數(shù)據(jù)顯示,在同時(shí)抑制miR-421時(shí),單純抑制FOXO4后細(xì)胞凋亡數(shù)減少。 結(jié)果：同時(shí)抑制miR-421時(shí),選擇下調(diào)基因FOXO4,結(jié)果表明,凋亡細(xì)胞低于單純抑制miR-421組。 結(jié)論：FOXO4是細(xì)胞生長(zhǎng)和抗凋亡的關(guān)鍵基因。 4、在鼻咽癌組織中miR-421與FOXO4的臨床相關(guān)性 目的：研究臨床鼻咽癌患者的腫瘤標(biāo)本中miR-421與FOXO4的關(guān)系。 方法：7例新鮮收集的人鼻咽癌樣本組織,利用QPCR技術(shù)檢測(cè)miR-421與FOXO4的表達(dá)量,同時(shí)采用免疫印跡法檢測(cè)FOXO4表達(dá)水平,并研究?jī)烧咧g的關(guān)系。 結(jié)果：在7例新鮮收集的鼻咽癌組織樣本中miR-421表達(dá)量高者則其FOXO4表達(dá)量低,兩者呈負(fù)相關(guān)(r=-0.798,P=0.032)。 結(jié)論：miR-421表達(dá)量在鼻咽癌組織樣本中與FOXO4表達(dá)量呈負(fù)相關(guān)。 總之,miR-421上調(diào)可抑制FOXO4的表達(dá),從而導(dǎo)致鼻咽癌細(xì)胞增殖和凋亡抵抗；反之,實(shí)驗(yàn)中阻遏miR-421的表達(dá)則可抑制鼻咽癌細(xì)胞增殖并促進(jìn)其凋亡。
[Abstract]:MicroRNA is a non coding single strand RNA molecule of approximately 19-25 nucleotides, which is recently discovered, which is combined with the 3 'end non translation region of the target gene to regulate the expression of.MiRNA as an important regulatory gene for the growth and development of the organism. At present, the main research direction is to explore miRNAs The relationship with the disease, especially the relationship between miRNA and tumor. Nasopharyngeal carcinoma (NPC) is one of the most common malignant tumors in Guangdong area. It is the first in the head and neck malignant tumor. The incidence of nasopharyngeal carcinoma is more common in 40-50 years old, once the disease has a great influence on the society, economy and family. Nasopharyngeal carcinoma is the primary cancer. It is hidden, not easy to observe, and adjacent to the nasal cavity, sinus and intracranial phase, so the clinical symptoms are late and different, and nasopharyngeal carcinoma is also the highest metastatic rate in the head and neck tumor. All these cause nasopharyngeal carcinoma to be easily misdiagnosed or missed and affect the treatment effect. Therefore, the early diagnosis and treatment of nasopharyngeal carcinoma have been the research of our country. One of the key points in the scientific research of the head and neck. At present, the treatment of nasopharyngeal carcinoma is mainly radiation therapy. Chemotherapy is not sensitive to radiotherapy. Chemotherapy can be taken in patients with relapse and middle and late stage, but it is only adjuvant therapy or palliative treatment. The surgical treatment of nasopharyngeal carcinoma is also used only as an auxiliary method in some special cases. In addition to the three conventional treatments, the author has also tried to use photodynamic therapy and microwave thermotherapy in recent years to treat nasopharyngeal carcinoma, but it can only be used as an auxiliary means. The development of recombinant DNA technology makes it possible for tumor gene therapy. The gene therapy scheme based on the mechanism of tumor has become a hot spot in cancer treatment. The researchers mainly use antisense nucleic acid technology to interfere with the expression of tumor related genes, which has achieved some effect. In recent years, the application of RNA interference technology in mammals has opened up a new field of vision for the gene therapy of tumor. Therefore, we intend to explore the regulatory role of miRNA in the development of nasopharyngeal carcinoma.
We used the Targetscan database to predict the target gene for differentially expressed miRNA, and put the predicted results into the GenMAPP software for biological pathway analysis. In the pathway analysis, most of the high target gene enrichment pathways involved signal transduction and the pathway related to cancer, such as the EGFR-1 signaling pathway, the MAPK signaling pathway. And so on. Among them, small GTP enzyme mediated signal transduction pathway is most significant. At the same time, the significance of the Wnt signaling pathway is consistent with our laboratory's previous study of cDNA expression profiles in nasopharyngeal carcinoma. Further study of these differences in the regulatory role of these differences in the biological pathways and to search for the progression of nasopharyngeal carcinoma. Clinical diagnostic markers are the direction of our future efforts.
MicroRNA (miRNA), a small RNA, is a small molecule RNA in the genome of plants and animals, about 19 to 25nt (a few less than 20nt), formed by a single strand RNA precursor (Pre-miRNA) with a clip ring structure and a length of 70-80nt. It passes through the 3 'end non coding region of its target mRNA molecule (3'-untranslatedregion, 3'UTR). Complete or incomplete complementarity matches the regulation of the post transcriptional level (protein expression), which plays a very important role in various physiological and pathological conditions in the organism.
MiRNA first found by Lee and so on in the study of the Caenothabditis elegans embryo development in 1993, a non coded RNA., first of which the primary transcriptional product of the miRNA gene (Pri-miRNA) was cut into the precursor miRNA (pre-miRNA) by RNase (?) Drosha in the nucleus. After the initial shear, the Pre-miRNA was transported. Under the action of protein exPortin-5, it is transferred from the nucleus to the cytoplasm, and then another RNase (?) Dicer is further cut to produce mature miRNA., the mature miRNA, together with other proteins, to form a RISC (RNA-induced sileneing complex) complex, which causes the target mRNA degradation or translation inhibition.
MicroRNAs plays an important role in gene expression, which plays an important role in the regulation of gene expression, and miR-421 plays an important role in the regulation of tumor progression, cell proliferation, and so on. The role of miR-421 in the development of nasopharyngeal carcinoma is not reported. This study will discuss the role of this study in nasopharyngeal carcinoma. The aim is to simulate the construction of plasmid expressing miRNA precursor by natural miRNA, and to control the expression of tumor associated gene miR-421, and to carry out gene therapy for nasopharyngeal carcinoma.
The first is to regulate the construction of the miR-421 expression plasmid and the effective plasmid screening of the target gene, and then to further determine the effective action loci of the 1niRNA for the FOXO4 gene, and observe the regulatory effect of the constructed plasmid on the two target genes in nasopharyngeal carcinoma, which can effectively inhibit the expression of FOX04 expression by the recombinant plasmid based on the.MiRNA recombinant plasmid in the following in vitro experiment. Tumor growth has laid the foundation for further clinical research.
Part one
1, overexpression of miR-.421 promotes proliferation and anti apoptosis of nasopharyngeal carcinoma cells.
Objective: to detect the ability of over expression of miR-421 in nasopharyngeal carcinoma cell proliferation and apoptosis.
Methods: using the technique of lentivirus infection, miR-421 was transfected into CNE2, and the proliferation of CNE2 was observed by miR-421 in vitro, and the cell apoptosis was detected by Annexin V and TUNEL.
Results: the soft agar cloning experiments showed that the proliferation rate of miR-421 overexpressed cells was 3 times as high as that of the control group, indicating that up regulation of miR-421 could significantly enhance the growth of nasopharyngeal carcinoma cells. In addition, Annexin V and TUNEL tests showed that the overexpression of miR-421 could enhance the apoptosis resistance of nasopharyngeal carcinoma cells to cisplatin. The results showed that miR-421 played a role in regulating cell genes in nasopharyngeal carcinoma in vitro.
Conclusion: upregulation of miR-421 can promote the proliferation of nasopharyngeal carcinoma cells and enhance the ability of cell apoptosis.
2, inhibition of miR-421 reduces proliferation and induces apoptosis in nasopharyngeal carcinoma cells.
Objective: To investigate the effect of inhibiting miR-421 on proliferation and apoptosis of nasopharyngeal carcinoma cells.
Methods: using the technique of lentivirus infection, CNE2 was transfected by miR-421, and the proliferation of CNE2 was observed by miR-421 in vitro, and the cell apoptosis was detected by Annexin V and TUNEL.
Results: the soft agar clone formation experiment showed that the proliferation rate of miR-421 cells decreased after inhibition of miR-421, and the control group was 3 times as high as that of the inhibition group. It showed that inhibition of miR-421 could significantly reduce the growth of nasopharyngeal carcinoma cells. In addition, Annexin V and TUNEL detection showed that inhibition of miR-421 could enhance the apoptosis promoting energy of nasopharyngeal cancer cells to cisplatin. These results indicate that miR-421 plays a role in regulating cell genes in nasopharyngeal carcinoma in vitro.
Conclusion: inhibition of miR-421 can reduce the proliferation and promote apoptosis of NPC cells.
The second part
1, to explore the effect of miR-421 on the FOXO4 signal pathway
Objective: To investigate whether miR-421 promotes the proliferation and anti apoptosis of nasopharyngeal carcinoma cells through FOXO4 signaling pathway.
Methods: the luciferase reporter gene vector for the rake gene FOXO4 was constructed and the fluorescence values of the normal group, the miR-421 overexpression group and the miR-421 inhibition group were measured by the fluorescent illuminometer. The expression of the mRNA and protein of the FOXO4 target gene p21, p27, Bim and FasL was detected by real-time quantitative PCR and Western blot.
Results: 1. in the activity detection of luciferase activity, the luciferase activity of FOXO4 decreased in miR-421 overexpressed cells, and the expression of.2. in miR-421 inhibited cells was consistent with this phase. In the normal group, the miR-421 overexpression group and the miR-421 inhibition group, we found that the FOXO4 four classic target genes p21, p27, Bim and FasL mRNA. The protein expression was downregulated in the miR-421 overexpression group, while in the miR-421 inhibited cells, the above genes and proteins were upregulated.
Conclusion: by constructing luciferase, overexpression and inhibition of miR-421, the results show that miR-421 can inhibit FOXO4 signaling pathway.
2, study on the relationship between miR-421 and FOXO4
Objective: To explore the regulatory role of miR-421 in raking gene FOXO4 in gene regulation.
Methods: the TargetScan software tool mir-421 target prediction analysis showed that FOXO4 was a potential target of mir-421. The expression of FOXO4 in the miR-421 overexpressed group and the miR-421 inhibition group was significantly different in the normal group, and the expression of the overexpressed histone expression was lower than that in the normal group, and the expression of the inhibitory histone expression was higher than that in the normal group. Normal group.
In order to further verify the relationship between miR-421 and FOXO4, we cloned FOXO43'UTR fragment, psiCHECK-2 luciferase carrier containing miR-421 binding site, and we cloned the FOXO43'UTR mutant vector and mutated two bases.
The luciferase activity assay showed that the miR-421 overexpression group was lower than the normal group, while the miR-421 inhibition group was higher than the normal group.
In addition, in order to further analyze the interaction between miR-421 and FOXO4 in apoptosis and inhibition of miR-421 at the time of inhibiting the FOXO4 as control, the experiment showed that after the inhibition of the FOXO4 gene, the apoptosis was lower than that of the only inhibition of the miR-421 group.
Results: 1, the expression of gene FOXO4 in miR-421 overexpression and miR-421 inhibition group was different. 2, fluorescein detection showed that miR-421 overexpression was different from that in miR-421 inhibition group, but there was no difference in the mutant group; 3, at the same time inhibition of miR-421, the selective down gene FOXO4 was selected. The results showed that the apoptotic cells were lower than simple cells. Inhibition of group miR-421.
Conclusion: gene FOXO4 is the target gene for the regulation of miR-421.
3. FOXO4 is a key gene for cell growth and anti apoptosis
Objective: To explore the regulatory effect of FOXO4 on cell growth and anti apoptosis.
Methods: in order to further analyze the role of miR-421 and FOXO4 in apoptosis, we chose to inhibit miR-421 and then only inhibit gene FOXO4 to control, and the detection data showed that the number of apoptotic cells decreased after the inhibition of miR-421 at the same time.
Results: when miR-421 was inhibited, the FOXO4 gene was downregulated. The results showed that the apoptotic cells were lower than those of miR-421 alone.
Conclusion: FOXO4 is a key gene for cell growth and anti apoptosis.
4, the clinical correlation between miR-421 and FOXO4 in nasopharyngeal carcinoma tissues.
Objective: To study the relationship between miR-421 and FOXO4 in clinical specimens of nasopharyngeal carcinoma.
Methods: 7 samples of freshly collected human nasopharyngeal carcinoma samples were collected. The expression of miR-421 and FOXO4 was detected by QPCR technique. The expression of FOXO4 was detected by Western blot, and the relationship between them was studied.
Results: in 7 samples of nasopharyngeal carcinoma, the expression level of miR-421 was high, and the expression level of FOXO4 was low, and there was a negative correlation between them (r=-0.798, P=0.032).
Conclusion: the expression of miR-421 is negatively correlated with the expression of FOXO4 in nasopharyngeal carcinoma tissues.
In conclusion, up regulation of miR-421 can inhibit the expression of FOXO4, which leads to the proliferation and apoptosis resistance of nasopharyngeal carcinoma cells. Conversely, the inhibition of miR-421 expression in the experiment can inhibit the proliferation of nasopharyngeal carcinoma cells and promote its apoptosis.
【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2013
【分類(lèi)號(hào)】：R739.63

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