發(fā)布時(shí)間：2018-06-22 11:19

  本文選題：肝細(xì)胞肝癌 + 性別差異　； 參考：《第二軍醫(yī)大學(xué)》2016年博士論文

【摘要】：原發(fā)性肝細(xì)胞肝癌(Hepatocellular carcinoma,HCC),作為最常見(jiàn)的肝癌類(lèi)型,是一種世界范圍內(nèi)的致死性惡性腫瘤,具有高復(fù)發(fā)、高轉(zhuǎn)移的特點(diǎn)。流行病學(xué)調(diào)查資料顯示,肝癌具有顯著的性別差異,其中男性與女性的肝癌發(fā)病比例約為2:1~4:1,并且在不同原因?qū)е碌母伟┲?男性患者的比例也均高于女性患者。不僅在發(fā)病率上,男性明顯高于女性,在預(yù)后方面,男性的生存時(shí)間與生存率也顯著低于女性。對(duì)于肝癌性別差異的解釋,目前主要集中于性激素和細(xì)胞因子,然而針對(duì)雌雄激素受體通路和炎癥干預(yù)的治療方案在臨床上并未取得滿(mǎn)意的結(jié)果。一方面歸因于現(xiàn)有的很多研究成果來(lái)源于化學(xué)誘導(dǎo)的動(dòng)物模型,并不能完全模擬臨床上的肝癌,尤其是由于HBV和HCV感染導(dǎo)致的肝癌;另一方面是由于肝癌性別差異的原因和機(jī)制并不完全清楚,尤其是基因組層面的變化并不了解�？截悢�(shù)變異(Copy number variations,CNVs)作為最重要的基因組變異之一,與各種遺傳性和發(fā)育性疾病密切相關(guān),是腫瘤形成的重要原因�？截悢�(shù)變異可以通過(guò)影響變異區(qū)域內(nèi)重要的癌基因和抑癌基因的基因組含量,導(dǎo)致這些重要調(diào)控基因的表達(dá)異常,從而導(dǎo)致腫瘤的發(fā)生,而拷貝數(shù)變異在肝癌發(fā)生發(fā)展中所發(fā)揮的重要作用也越來(lái)越受到人們的重視。然而在肝癌的性別差異方面,對(duì)于男女基因組層面的差異卻很少有研究涉及。對(duì)于性別相關(guān)的基因組變異區(qū)域的確定,對(duì)性別相關(guān)基因組變異的意義研究,以及對(duì)基因組變異的性別差異原因的探索,將會(huì)增加我們對(duì)于肝癌性別差異的認(rèn)識(shí),補(bǔ)充完善肝癌性別差異的機(jī)制,為肝癌的預(yù)防和治療提供新的思路。在本研究中,我們重新分析了網(wǎng)上公共數(shù)據(jù)庫(kù)中一組大規(guī)模的肝癌基因組芯片數(shù)據(jù),將肝癌的基因組數(shù)據(jù)與病人的性別資料進(jìn)行了匹配,篩選得到性別特異性的拷貝數(shù)變異區(qū)域,首先確定了肝癌中確實(shí)存在基因組變異的性別差異。我們進(jìn)一步將性別相關(guān)的拷貝數(shù)變異區(qū)域與我們的一組RNA測(cè)序結(jié)果進(jìn)行了聯(lián)合分析,篩選得到了性別相關(guān)的拷貝數(shù)變異基因,作為我們研究拷貝數(shù)變異性別差異意義的候選基因集。之后我們?cè)谝唤M肝癌隊(duì)列中對(duì)芯片結(jié)果進(jìn)行了q PCR的驗(yàn)證,發(fā)現(xiàn)其中UBE2D1在基因組水平和轉(zhuǎn)錄水平均存在顯著的性別差異。因此我們靶定UBE2D1作為研究拷貝數(shù)變異性別差異的關(guān)鍵基因。我們首先確定了UBE2D1的臨床意義,發(fā)現(xiàn)UBE2D1在肝癌中存在基因組的復(fù)制增加,并且基因組的復(fù)制增加導(dǎo)致了其在肝癌組織中的高表達(dá)。UBE2D1的高表達(dá)與肝癌病人較差的總體生存率相關(guān)。進(jìn)一步探索UBE2D1的生物學(xué)功能發(fā)現(xiàn),UBE2D1可以促進(jìn)正常肝細(xì)胞和肝癌細(xì)胞的增殖,增強(qiáng)克隆形成能力,抑制細(xì)胞凋亡,并且促進(jìn)肝癌細(xì)胞的體內(nèi)成瘤能力。機(jī)制研究發(fā)現(xiàn),UBE2D1是通過(guò)增強(qiáng)p53的泛素化,降低了p53的蛋白水平,進(jìn)而抑制了p53依賴(lài)的細(xì)胞衰老過(guò)程。在確定肝癌中性別相關(guān)拷貝數(shù)變異的意義基礎(chǔ)上,我們進(jìn)一步探索了拷貝數(shù)變異性別差異的形成原因。我們發(fā)現(xiàn)IL-6、IL-8、IL-1β和TNF-α在肝癌的癌旁組織中存在顯著的性別差異。之后我們利用復(fù)制壓力和染色體不穩(wěn)定性作為評(píng)判拷貝數(shù)變異形成的指標(biāo),發(fā)現(xiàn)IL-6可以最為明顯的誘導(dǎo)細(xì)胞的復(fù)制壓力反應(yīng)和染色體不穩(wěn)定性,而持續(xù)的IL-6刺激可以導(dǎo)致染色體不穩(wěn)定性的顯著增強(qiáng),并增加UBE2D1的基因組含量。不同于短期IL-6刺激,我們發(fā)現(xiàn)長(zhǎng)期IL-6刺激會(huì)導(dǎo)致不依賴(lài)于外源細(xì)胞因子刺激的自分泌增加,說(shuō)明一定時(shí)間的IL-6刺激對(duì)于這一過(guò)程是必須的。機(jī)制研究發(fā)現(xiàn),長(zhǎng)期IL-6刺激是通過(guò)Stat3抑制了Rad51b的轉(zhuǎn)錄激活,進(jìn)而維持了復(fù)制壓力反應(yīng),積累了染色體不穩(wěn)定性。長(zhǎng)期的IL-6刺激可以顯著促進(jìn)細(xì)胞增殖,抑制細(xì)胞凋亡,增強(qiáng)肝癌細(xì)胞的體內(nèi)成瘤能力和正常肝細(xì)胞的惡性轉(zhuǎn)化。而Stat3和Rad51b的干預(yù)會(huì)逆轉(zhuǎn)對(duì)復(fù)制壓力和染色體不穩(wěn)定的影響,逆轉(zhuǎn)長(zhǎng)期IL-6的生物學(xué)功能。我們進(jìn)一步探討了IL-6-Stat3-Rad51b這一調(diào)控軸的臨床意義,發(fā)現(xiàn)Rad51b在肝癌中顯著低表達(dá),并且這種低表達(dá)在癌前病變就已出現(xiàn)。根據(jù)病人血清中IL-6水平進(jìn)行分組,發(fā)現(xiàn)高的血清IL-6水平組的病人Rad51b下調(diào)更為明顯,而UBE2D1的基因組復(fù)制增加也更為顯著。而Rad51b在癌旁組織中低表達(dá)是肝癌病人預(yù)后的危險(xiǎn)因素,其表達(dá)量與Stat3顯著負(fù)相關(guān),與UBE2D1的表達(dá)水平也存在負(fù)相關(guān)。通過(guò)我們的研究,我們得到了以下的結(jié)論:首先,在肝癌中確實(shí)存在著基因組層面的性別差異,確定了男女在拷貝數(shù)變異水平的不同;其次,UBE2D1作為性別相關(guān)拷貝數(shù)變異的關(guān)鍵基因,在男性中的基因組水平顯著高于女性,并且可以通過(guò)調(diào)控p53泛素化下調(diào)p53蛋白水平,進(jìn)而促進(jìn)肝癌發(fā)生發(fā)展;第三,肝癌中,拷貝數(shù)變異的性別差異是由于腫瘤微環(huán)境中IL-6的差異,長(zhǎng)期IL-6刺激可以通過(guò)Stat3抑制Rad51b,維持復(fù)制壓力反應(yīng),增強(qiáng)染色體不穩(wěn)定性,導(dǎo)致了UBE2D1基因組水平的增加,并且這一過(guò)程是個(gè)長(zhǎng)期作用結(jié)果,較長(zhǎng)時(shí)間的IL-6刺激和Rad51b的作用是這一過(guò)程所必需的。通過(guò)對(duì)肝癌中拷貝數(shù)變異性別差異的研究,可以加深我們對(duì)于肝癌性別差異在基因組層面的認(rèn)識(shí),并且完善了肝癌中拷貝數(shù)變異性別差異的形成機(jī)制,作為性別相關(guān)拷貝數(shù)變異的關(guān)鍵基因UBE2D1和形成這一差異的關(guān)鍵因子Rad51b,可以作為診斷肝癌的標(biāo)志物和治療肝癌的潛在藥物靶定。
[Abstract]:Primary hepatocellular carcinoma (Hepatocellular carcinoma, HCC), as the most common type of liver cancer, is a worldwide fatal malignant tumor, characterized by high recurrence and high metastasis. Epidemiological data show that liver cancer has significant gender differences, and the incidence of liver cancer in males and females is about 2:1~4:1, and The proportion of male patients in different causes of liver cancer is also higher than that of female patients. Not only in the incidence rate, men are obviously higher than women, but also in the prognosis, the survival time and survival rate of men are also significantly lower than that of women. No satisfactory results have been achieved in the treatment of the vegetarian receptor pathway and inflammatory intervention. One reason is that many of the existing research results are derived from chemical induced animal models and can not fully simulate the clinical liver cancer, especially the liver cancer caused by HBV and HCV infection, and the other is the origin of the gender differences in liver cancer. Copy number variations (CNVs), one of the most important genomic variations, is closely related to various genetic and developmental diseases, and is an important cause of tumor formation. The genomic content of oncogene and tumor suppressor gene leads to the abnormal expression of these important regulatory genes, which leads to the occurrence of tumor, and the important role of the copy number variation plays a more and more important role in the development and development of liver cancer. However, there is little difference in gender difference between male and female genomes. There are studies involved. The identification of genomes related genomes, the significance of sex related genome variation and the reasons for gender differences in genome variation will increase our understanding of the gender differences in liver cancer, complement the mechanisms of the gender differences in liver cancer, and provide new prevention and treatment for the liver cancer. In this study, we reanalyzed a group of massive liver cancer genome chip data in the public database of the Internet, matching the genome data of the liver cancer with the patient's sex data, screening the sex specific copy number variation region, and first determined the gender difference of the genomic variation in the liver cancer. We further analyzed the sex related copy number variation region and our group of RNA sequencing results, and screened the sex related copy number variant gene, as a candidate gene for our study of the gender differences in the copy number variation. After that, we conducted a Q PC for the results of the chip in a group of liver cancer groups. R proved that there was a significant gender difference between the genomic level and the transcriptional level of UBE2D1. Therefore, we targeted UBE2D1 as the key gene to study the gender differences in the copy number variation. We first identified the clinical significance of UBE2D1, and found that the replication of the gene group in the liver cancer was increased in UBE2D1 and the replication of the genome was increased. The high expression of the high expression of.UBE2D1 in the liver cancer tissues is associated with the poor overall survival rate of the liver cancer patients. Further exploring the biological function of UBE2D1, UBE2D1 can promote the proliferation of normal hepatocytes and hepatoma cells, enhance the clone formation ability, inhibit the apoptosis of the cell and promote the tumor formation in the liver of the liver cancer cells. Ability. The mechanism study found that UBE2D1, by enhancing the ubiquitination of p53, reduced the protein level of p53 and then inhibited the cell aging process of p53 dependent. On the basis of determining the significance of the mutation of the neutrophilic copy number of liver cancer, we further explored the causes of the gender differences in the copy number variation. We found that IL-6, IL-8, IL-1 beta and IL-1 beta. TNF- alpha has significant gender differences in the adjacent tissues of the carcinoma of the liver. Then we use the replication pressure and chromosome instability as an indicator of the copy number variation. It is found that IL-6 can most significantly induce the replicative stress response and chromosome instability of the cells, and the persistent IL-6 stimulation can lead to chromosome instability. A significant increase in sex and an increase in the genomic content of UBE2D1. Unlike short-term IL-6 stimuli, we found that long-term IL-6 stimulation can lead to an autocrine increase that is not dependent on exogenous cytokines stimulation, indicating that a certain time of IL-6 stimulation is necessary for this process. Mechanism research is developed, and long-term IL-6 stimulation is the inhibition of Rad51b through Stat3. The transcriptional activation, which then maintains the replicative stress response, accumulates chromosome instability. Long-term IL-6 stimulation can significantly promote cell proliferation, inhibit cell apoptosis, enhance the tumorigenicity of hepatoma cells and malignant transformation of normal liver cells. Stat3 and Rad51b may reverse the replication pressure and chromosome instability. Effect, reversing the biological function of long-term IL-6. We further explored the clinical significance of the IL-6-Stat3-Rad51b regulation axis, and found that Rad51b was significantly lower in the liver cancer, and this low expression appeared in precancerous lesions. According to the level of IL-6 in the serum of the patients, the patients with high serum IL-6 level were found to be down regulated. It is more obvious that the genomic replication of UBE2D1 is more significant. And the low expression of Rad51b in the para cancerous tissue is a risk factor for the prognosis of the patients with liver cancer, and the expression is negatively correlated with the Stat3, and there is a negative correlation with the expression level of UBE2D1. The gender differences at the genome level determine the difference in the level of copy number variation between men and women. Secondly, UBE2D1 is the key gene of the sex related copy number variation, the genome level of the male is significantly higher than that of the female, and it can reduce the level of p53 protein by regulating the p53 ubiquitination, and then promote the development and development of liver cancer; third, liver cancer. The sex difference of the copy number variation is due to the difference in the IL-6 in the tumor microenvironment. The long-term IL-6 stimulation can inhibit Rad51b through Stat3, maintain the replicative stress response and enhance the chromosome instability, which leads to the increase of the UBE2D1 genome level, and this process is a long-term result, the longer IL-6 stimulation and the role of Rad51b. It is necessary for this process. Through the study of the gender differences in the copy number variation in the liver cancer, we can deepen our understanding of the gender difference at the genome level and improve the formation mechanism of the sex difference of the copy number variation in the liver cancer, as the key gene UBE2D1 of the sex related copy number variation and the difference in the formation of the sex related copy number. The key factor Rad51b can be used as a marker for the diagnosis of liver cancer and a potential drug target for the treatment of liver cancer.
【學(xué)位授予單位】：第二軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類(lèi)號(hào)】：R735.7

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6 黃靜　李媛媛;丙肝發(fā)展成肝癌速度或更快[N];人民政協(xié)報(bào);2013年

7 記者 黃才剛;肝癌宜因人施治[N];健康報(bào);2000年

8 鐘肖協(xié);肝癌“偏愛(ài)”哪些人[N];陜西日?qǐng)?bào);2000年

9 黃顯斌邋唐明山;我科學(xué)家在肝癌發(fā)病機(jī)制研究中取得重大進(jìn)展[N];光明日?qǐng)?bào);2007年

10 福星;肝癌如何早發(fā)現(xiàn)[N];民族醫(yī)藥報(bào);2007年

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2 周少來(lái);CXCL5促進(jìn)中性粒細(xì)胞浸潤(rùn)及其在肝癌微環(huán)境中的作用和機(jī)制研究[D];復(fù)旦大學(xué);2014年

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6 上官輝;肝癌易感相關(guān)基因及組織基因表達(dá)譜生物信息學(xué)分析[D];南方醫(yī)科大學(xué);2015年

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8 盧杉;長(zhǎng)鏈非編碼RNA:HOXD-AS1在肝癌中的功能和機(jī)制研究[D];第二軍醫(yī)大學(xué);2015年

9 冀彤;IL6、TNFα和TNFR1在卵圓細(xì)胞增殖和炎癥誘發(fā)肝癌中作用和機(jī)制的研究[D];浙江大學(xué);2015年

10 趙昌林;CD4~+CXCR5~+CD57~+T細(xì)胞在肝癌發(fā)病中的作用及解毒抗癌方干預(yù)的研究[D];暨南大學(xué);2015年

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7 高增法;HACE1在肝癌中的生物學(xué)特性與其臨床病理學(xué)特征及預(yù)后的關(guān)系[D];蘭州大學(xué);2016年

8 張佳楠;MDSC在肝癌中的表達(dá)及與臨床病理分期的關(guān)系[D];吉林大學(xué);2016年

9 馬杰;CD90和CXCR4在肝癌中的表達(dá)及其與術(shù)后早期復(fù)發(fā)的關(guān)系[D];安徽醫(yī)科大學(xué);2016年

10 李培培;TGF-β1誘導(dǎo)肝星狀細(xì)胞表達(dá)PD-L1促進(jìn)肝癌的發(fā)生發(fā)展[D];安徽醫(yī)科大學(xué);2016年

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