本文選題：食管鱗狀細(xì)胞癌 + miR-193a-5p　； 參考：《鄭州大學(xué)》2017年博士論文

【摘要】：1背景及目的食管癌(esophageal cancer)是目前發(fā)病率、死亡率都較高的消化道惡性腫瘤之一,目前全球癌癥發(fā)病率中排名第9位,總死亡率占到第6位。在發(fā)達(dá)國家,食管癌的發(fā)病率排名第20位,但其死亡排第11位。在發(fā)展中國家,食管癌發(fā)病率排名第8位,死亡率排在第5位。食管癌已從1990年的第七大癌癥上升至2013年的第六大癌癥,衰老和不斷增長的人群是食管癌病例增加的驅(qū)動因素。在食管癌中,鱗狀細(xì)胞癌和腺癌為主要病理類型,其中鱗狀細(xì)胞癌在我國占90%以上。世界每年約有40萬人死于食管癌,而我國是世界上食管癌的高發(fā)地區(qū)之一,平均每年因該疾病的死亡人數(shù)大于15萬。由于食管癌的早期癥狀不明顯并且缺乏早期診斷,多數(shù)患者在首次診斷時(shí)已處于疾病晚期,食管癌切除術(shù)后5年生存率9.5-45%,腫瘤侵襲和轉(zhuǎn)移是導(dǎo)致食管癌患者死亡的主要因素,侵襲和轉(zhuǎn)移主要依賴于原癌基因的激活和腫瘤抑制基因的失活,以及細(xì)胞增殖和凋亡等各種因素協(xié)作的共同結(jié)果。如果我們可以從遺傳學(xué)和表觀遺傳學(xué)的角度研究食管癌的發(fā)生和發(fā)展,它將為食管癌的診斷和治療提供新的靶點(diǎn)以及新的策略。人類表皮生長因子受體2(human epidermal growth factor receptor 2,ERBB2)是人表皮生長因子受體家族的成員之一,位于染色體17q12-21.32,編碼相對分子質(zhì)量為185,000跨膜受體樣蛋白,屬于跨膜酪氨酸激酶受體蛋白,具有酪氨酸激酶活性,該家族成員的過表達(dá)與乳腺癌、前列腺癌等腫瘤疾病的不良后果有關(guān)。已明確證明ERBB2的擴(kuò)增已經(jīng)顯示在某些侵襲型乳腺癌的進(jìn)展中起重要作用,ERBB2蛋白已成為30%乳腺癌患者的重要生物標(biāo)志物和治療靶標(biāo)。ERBB2在細(xì)胞生長、分化和遷移中起重要作用,其過表達(dá)促進(jìn)腫瘤生長、轉(zhuǎn)移和腫瘤血管生成。在食管鱗狀細(xì)胞癌(esophageal squamous cell carcinoma,ESCC)中,已報(bào)道有ERBB2過表達(dá),ERBB2被認(rèn)為是食管癌的潛在治療靶點(diǎn)。然而,ERBB2如何調(diào)節(jié)ESCC發(fā)生的分子機(jī)制仍然不明確。Micro RNA(miRNA)是近年來發(fā)現(xiàn)的小的非編碼RNA分子,含有約22個(gè)核苷酸,廣泛分布于真核生物中。雖然大多數(shù)miRNA位于細(xì)胞內(nèi),但也已經(jīng)在細(xì)胞外環(huán)境中發(fā)現(xiàn)了一些miRNA,稱之為循環(huán)miRNA或細(xì)胞外miRNA。目前最常見對靶基因的調(diào)控是miRNA通過與一些蛋白結(jié)合形成miRNA誘導(dǎo)沉默復(fù)合物,進(jìn)而引導(dǎo)與信使RNA(Messenger RNA,m RNA)的3'非編碼區(qū)(3'-UTR)完全或不完全配對,識別特異的m RNA后使其降解或阻遏其轉(zhuǎn)錄后翻譯,即基因表達(dá)的轉(zhuǎn)錄后抑制靶基因的表達(dá)。miRNA通過調(diào)節(jié)靶基因的表達(dá),在細(xì)胞增殖、凋亡、分化等過程中起著重要作用,并且在維持細(xì)胞的正常功能中起重要作用。研究人員已經(jīng)發(fā)現(xiàn),miRNA失調(diào)與許多疾病有關(guān),如與腫瘤密切相關(guān)。本課題首先采用免疫組化方法探討ERBB2在食管鱗狀細(xì)胞癌中的表達(dá)及其與臨床病理特征之間的關(guān)系;闡明miR-193a-5p抑制ERBB2翻譯的分子機(jī)制揭示ERBB2的全新調(diào)控方式;進(jìn)一步檢測食管癌組織及患者血清中miR-193a-5p的表達(dá)水平,評價(jià)miR-193a-5p作為食管癌預(yù)后指標(biāo)的價(jià)值。2方法及結(jié)果2.1免疫組化檢測ERBB2在食管鱗癌中的表達(dá)及意義我們采用免疫組化探討ERBB2在食管鱗狀細(xì)胞癌中的表達(dá)及其與臨床病理特征之間的關(guān)系,食管鱗癌組織中的ERBB2表達(dá)率(35/68,51.5%)高于正常食管組織8.8%(6/68)(χ2=33.03 P0.01)。ERBB2的表達(dá)與腫瘤淋巴結(jié)轉(zhuǎn)移和腫瘤分期相關(guān),差異有統(tǒng)計(jì)學(xué)意義(P0.01),與年齡、性別無統(tǒng)計(jì)學(xué)意義(P0.05)。2.2 miR-193a-5p在翻譯水平抑制ERBB2表達(dá)我們通過QRT-PCR及Western blot兩種方法檢測了ERBB2在正常食管粘膜細(xì)胞Het-1A及食管癌細(xì)胞株KYSE150、KYSE410、ECA-109、TE-2、TE-13中的mRNA和蛋白表達(dá)情況。發(fā)現(xiàn)ERBB2的mRNA水平和蛋白表達(dá)水平在食管癌細(xì)胞系中的表達(dá)與正常細(xì)胞不同。我們推斷在食管癌中ERBB2存在轉(zhuǎn)錄后水平的調(diào)控機(jī)制來抑制其表達(dá),通過三個(gè)主要micro RNA預(yù)測軟件,Targetscan,miRanda和Micro Inspector,預(yù)測與ERBB2 3'URT區(qū)域結(jié)合的潛能,發(fā)現(xiàn)miR-193a-5p與ERBB2的3'URT區(qū)有一個(gè)結(jié)合位點(diǎn),位于轉(zhuǎn)錄終止密碼子下游的41nt。通過轉(zhuǎn)染miR-193a-5p mimics來上調(diào)miR-193a-5p的表達(dá)水平,然后檢測ERBB2的蛋白表達(dá)水平和m RNA表達(dá)水平,發(fā)現(xiàn)miR-193a-5p能夠顯著下調(diào)ERBB2蛋白水平而對RNA水平無明顯影響。結(jié)果提示miR-193a-5p能夠通過與ERBB2 3'URT區(qū)結(jié)合進(jìn)而抑制ERBB2翻譯下調(diào)其表達(dá)。2.3 miR-193a-5p在組織及血清中的表達(dá)及臨床指標(biāo)關(guān)系通過QRT-PCR檢測35例中miR-193a-5p的表達(dá)水平,通過配對T檢驗(yàn)比較癌組織和癌旁組織中miR-193a-5p的表達(dá)水平,結(jié)果表明食管鱗狀細(xì)胞癌組織中的miR-193a-5p表達(dá)水平顯著低于癌旁正常組織。根據(jù)臨床資料,我們將標(biāo)本分為淋巴結(jié)轉(zhuǎn)移組和非淋巴結(jié)轉(zhuǎn)移組,通過Wilcoxon秩和檢驗(yàn),發(fā)現(xiàn)淋巴結(jié)轉(zhuǎn)移組中miR-193a-5p的表達(dá)水平顯著低于無轉(zhuǎn)移組(P0.05),結(jié)果表明miR-193a-5p可能參與抑制食管癌的轉(zhuǎn)移過程。將ERBB2的表達(dá)水平分類為ERBB2高表達(dá)組和ERBB2低表達(dá)組,通過Wilcoxon秩和檢驗(yàn),發(fā)現(xiàn)ERBB2高表達(dá)組中miR-193a-5p的表達(dá)水平顯著低于ERBB2低表達(dá)組(P0.05),提示在食管鱗狀細(xì)胞癌組織中ERBB2的表達(dá)水平與miR-193a-5p的表達(dá)水平呈負(fù)相關(guān)。通過對數(shù)秩檢驗(yàn)比較miR-193a-5p高表達(dá)組和低表達(dá)組的存活情況,結(jié)果表明miR-193a-5p高表達(dá)組的預(yù)后顯著高于miR-193a-5p低表達(dá)組(P0.05)。對35例冰凍食管鱗狀細(xì)胞癌組織和血清提取了miRNA,通過QRT-PCR檢測miR-193a-5p的表達(dá)水平,并通過Pearson相關(guān)性分析血清中miR-193a-5p和組織中miR-193a-5p之間的相關(guān)性,結(jié)果表明血清中miR-193a-5p的表達(dá)水平與組織中miR-193a-5p的表達(dá)水平呈正相關(guān)。3結(jié)論3.1 ERBB2免疫組化研究結(jié)果,ERBB2的表達(dá)水平增高則預(yù)示食管癌的預(yù)后較差,參考學(xué)者在乳腺癌中的相關(guān)報(bào)道,ERBB2可能參與了腫瘤的侵襲及轉(zhuǎn)移等相關(guān)機(jī)制,ERBB2上調(diào)是食管癌預(yù)后不良的危險(xiǎn)因素之一。3.2在食管鱗狀細(xì)胞癌中ERBB2在轉(zhuǎn)錄后水平的表達(dá)被抑制,通過分析可能的調(diào)控miRNA,miR-193a-5b結(jié)合ERBB2 3'URT區(qū)域并抑制ERBB2的翻譯和沉默其表達(dá),揭示ERBB2的新調(diào)節(jié)機(jī)制。3.3食管鱗狀細(xì)胞癌組織中ERBB2的表達(dá)水平與miR-193a-5p的表達(dá)水平呈負(fù)相關(guān),miR-193a-5p表達(dá)水平與患者轉(zhuǎn)移和預(yù)后不良呈負(fù)相關(guān)。血清中miR-193a-5p表達(dá)水平可以反映組織中miR-193a-5p的表達(dá)水平,提示我們血清中miR-193a-5p的水平可作為食管癌預(yù)后的非侵入性診斷指標(biāo)。
[Abstract]:1 background and objective esophageal cancer (esophageal cancer) is one of the most prevalent digestive tract malignant tumors with high morbidity and mortality. At present, the incidence of cancer is ranked ninth in the world, and the total mortality is sixth. In developed countries, the incidence of esophageal cancer ranks twentieth, but the death row is eleventh. In developing countries, the incidence of esophageal cancer is in row. The mortality rate is fifth. Esophageal cancer has risen from the seventh largest cancer in 1990 to the sixth major cancer in 2013. Aging and growing population are the driving factors for the increase of esophageal cancer. In esophageal cancer, squamous cell carcinoma and adenocarcinoma are the main pathological types, and squamous cell carcinoma accounts for more than 90% in our country. The world is about eighth in the world. 400 thousand people died of esophageal cancer, and China is one of the most frequent areas of esophageal cancer in the world. The average death toll of the disease is more than 150 thousand per year. Because of the early symptoms of esophageal cancer and the lack of early diagnosis, most patients are in the advanced stage of the first diagnosis. The 5 year survival rate after esophagectomy is 9.5-45%, and the tumor invades the tumor. Attack and metastasis are the main causes of death in patients with esophageal cancer. Invasion and metastasis are mainly dependent on the activation of proto oncogenes and the inactivation of tumor suppressor genes, and the cooperation of cell proliferation and apoptosis. If we can study the occurrence and development of esophageal cancer from the perspective of genetics and epigenetics, it is possible to study the occurrence and development of esophageal cancer. It will provide new targets and new strategies for the diagnosis and treatment of esophageal cancer. Human epidermal growth factor receptor 2 (human epidermal growth factor receptor 2, ERBB2) is one of the members of the human epidermal growth factor receptor family, located in the chromosome 17q12-21.32, encoding the relative sub mass of the 185000 transmembrane receptor like protein, belonging to the transmembrane cheese. Tyrosine kinase receptor protein, which has tyrosine kinase activity, is associated with adverse consequences of breast cancer and prostate cancer. It has been shown that ERBB2 amplification has been shown to play an important role in the progression of invasive breast cancer, and ERBB2 egg white has become an important biomarker for 30% breast cancer patients. Target.ERBB2 plays an important role in cell growth, differentiation and migration, and its overexpression promotes tumor growth, metastasis and tumor angiogenesis. In the esophageal squamous cell carcinoma (ESCC), ERBB2 overexpression has been reported. ERBB2 is considered as a potential therapeutic target for esophageal cancer. However, ERBB2 is to be adjusted. The molecular mechanism of the occurrence of ESCC is still not clear that.Micro RNA (miRNA) is a small non coding RNA molecule found in recent years, containing about 22 nucleotides, widely distributed in eukaryotes. Although most miRNA is in the cell, it has also found some miRNA in the extracellular environment, known as circulating miRNA or extracellular miRNA. at present most. The regulation of target genes is that miRNA combines with some proteins to form miRNA induced silencing complex, and then leads to complete or incomplete pairing of 3'non coding region (3'-UTR) with messenger RNA (Messenger RNA, m RNA), to identify specific m RNA and to degrade or deter its transtranscriptional translation after the transcription of gene expression, that is, the gene expression after the target gene is suppressed. Expression of.MiRNA by regulating the expression of target genes plays an important role in cell proliferation, apoptosis and differentiation, and plays an important role in maintaining the normal function of cells. Researchers have found that miRNA disorders are related to many diseases, such as cancer, which are closely related to tumors. The expression of miR-193a-5p in squamous cell carcinoma and its relationship with the clinicopathological features; to elucidate the molecular mechanism of inhibiting ERBB2 translation to reveal the new regulation of ERBB2, to further detect the expression level of miR-193a-5p in the sera of the esophageal cancer and the patients, and to evaluate the value of miR-193a-5p as a prognostic indicator of esophageal cancer and to evaluate the value of miR-193a-5p. The expression and significance of ERBB2 in the detection of ERBB2 in squamous cell carcinoma of the esophagus by immunohistochemistry. We used immunohistochemistry to investigate the expression of ERBB2 in esophageal squamous cell carcinoma and its relationship with the clinicopathological features. The expression rate of ERBB2 (35/68,51.5%) in esophageal squamous cell carcinoma (35/68,51.5%) is higher than that of normal esophageal tissue (6/68) (x 2=33.03 P0.01).ERBB2 Tumor lymph node metastasis and tumor staging were correlated with statistical significance (P0.01). There was no statistical significance (P0.01) with age and sex (P0.05). The expression of.2.2 miR-193a-5p at the translation level inhibited ERBB2 expression. We detected ERBB2 in normal esophageal mucosa cells Het-1A and esophageal cancer cell strain KYSE150, KYSE410, and Western miR-193a-5p at the translation level. 09, the expression of mRNA and protein in TE-2, TE-13. It is found that the mRNA level and protein expression level of ERBB2 are different from normal cells in esophageal cancer cell lines. We infer that there is a post transcriptional regulation mechanism for ERBB2 in esophageal cancer to inhibit its expression by using the three main micro RNA prediction software, Targetscan, miRanda, and Micro. Inspector, predicting the potential of combining with the ERBB2 3'URT region, it is found that there is a binding site in the 3'URT region of miR-193a-5p and ERBB2, and the 41nt. in the downstream of the transcriptional termination codon up-regulated the expression level of miR-193a-5p by transfection of miR-193a-5p mimics, and then detects the level of ERBB2 protein and the level of M expression. The level of ERBB2 protein has no significant effect on the level of RNA. The results suggest that miR-193a-5p can inhibit the expression of.2.3 miR-193a-5p in the tissue and serum by binding to the ERBB2 3'URT region and the expression of the expression of.2.3 miR-193a-5p in the tissue and serum and the relationship between the expression level of miR-193a-5p in 35 cases by QRT-PCR, by paired T test ratio. The expression level of miR-193a-5p in the carcinoma tissue and adjacent tissue showed that the expression of miR-193a-5p in the squamous cell carcinoma of the esophagus was significantly lower than that of the normal tissue adjacent to the carcinoma. According to the clinical data, we divided the specimens into lymph node metastasis group and non lymph node metastasis group. By the Wilcoxon rank sum test, we found the miR-19 in the lymph node metastasis group. The expression level of 3a-5p was significantly lower than that in the non metastasis group (P0.05). The results showed that miR-193a-5p might be involved in inhibiting the metastasis of esophageal cancer. The expression level of ERBB2 was classified into ERBB2 high expression group and ERBB2 low expression group. The expression level of miR-193a-5p in the high expression group of ERBB2 was significantly lower than that of the ERBB2 low expression group (P0) (P0). .05) showed a negative correlation between the expression level of ERBB2 in the squamous cell carcinoma of the esophagus and the expression level of miR-193a-5p. The survival of the miR-193a-5p high expression group and the low expression group was compared by the logarithmic rank test. The results showed that the prognosis of the high expression group of miR-193a-5p was significantly higher than that of the miR-193a-5p low expression group (P0.05). 35 cases of frozen esophageal squamous cells were compared. MiRNA was extracted from the tissue and serum of the cell carcinoma, and the expression level of miR-193a-5p was detected by QRT-PCR. The correlation between the serum miR-193a-5p and the miR-193a-5p in the tissues was analyzed by Pearson correlation. The results showed that the expression level of miR-193a-5p in serum was positively correlated with the level of miR-193a-5p expression in the tissue and the.3 conclusion 3.1 ERBB2 exempts. The higher expression level of ERBB2 indicates that the prognosis of esophageal cancer is poor, the related reports of reference scholars in breast cancer, ERBB2 may be involved in the related mechanisms of tumor invasion and metastasis, and the up-regulation of ERBB2 is one of the risk factors for the poor prognosis of esophageal cancer,.3.2 in the post transcriptional level of ERBB2 in the squamous cell carcinoma of the esophagus Expression was suppressed, and the possible regulation of miRNA, miR-193a-5b combined with ERBB2 3'URT region and inhibition of ERBB2 translation and silence expression, the new regulatory mechanism of ERBB2 revealed the negative correlation between the expression level of ERBB2 in the squamous cell carcinoma of.3.3 and the expression level of miR-193a-5p, and the expression level of miR-193a-5p and the metastasis and prognosis of the patients. The level of miR-193a-5p expression in the serum can reflect the level of miR-193a-5p expression in the tissue, suggesting that the level of miR-193a-5p in the serum can be used as a noninvasive diagnostic index for the prognosis of esophageal cancer.
【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R735.1

【參考文獻(xiàn)】

相關(guān)期刊論文 前2條

1 Xin Wang;Su-hua Shi;Hai-jiang Yao;Quan-kai Jing;Yu-ping Mo;Wei Lv;Liang-yu Song;Xiao-chen Yuan;Zhi-gang Li;Li-na Qin;;Electroacupuncture at Dazhui(GV14) and Mingmen(GV4) protects against spinal cord injury:the role of the Wnt/β-catenin signaling pathway[J];Neural Regeneration Research;2016年12期

2 Juan Xie;Zhi-Hui Tan;Xia Tang;Ming-Shu Mo;Yan-Ping Liu;Run-Liang Gan;Yi Li;Li Zhang;Guo-Qing Li;;miR-374b-5p suppresses RECK expression and promotes gastric cancer cell invasion and metastasis[J];World Journal of Gastroenterology;2014年46期

相關(guān)博士學(xué)位論文 前3條

1 王濤;食管鱗癌中miR-451和miR-21表達(dá)及對生長、侵襲和凋亡的影響[D];鄭州大學(xué);2014年

2 卜芳芳;miR-205下調(diào)ASPP2表達(dá)促進(jìn)食管癌轉(zhuǎn)移及EMT的作用及機(jī)制研究[D];第二軍醫(yī)大學(xué);2013年

3 余茜穎;miR-203調(diào)控食管癌干細(xì)胞自我更新及食管癌相關(guān)基因ECRG1轉(zhuǎn)錄調(diào)控的研究[D];北京協(xié)和醫(yī)學(xué)院;2011年

相關(guān)碩士學(xué)位論文 前1條

1 葛曉晴;磷酯酶CE1在肺鱗狀細(xì)胞癌組織中的表達(dá)及意義[D];鄭州大學(xué);2012年

，

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