【摘要】： 研究背景及目的：喉鱗狀細(xì)胞癌(laryngeal squamous cell carcinoma, LSCC)是頭頸部最常見(jiàn)的惡性腫瘤之一,腫瘤浸潤(rùn)和轉(zhuǎn)移是造成喉癌患者死亡的主要原因之一,喉癌發(fā)生侵襲轉(zhuǎn)移是一個(gè)多環(huán)節(jié)、多步驟的復(fù)雜過(guò)程,包括腫瘤細(xì)胞粘附、移動(dòng)、酶降解和血管生成等多種腫瘤細(xì)胞的生物學(xué)行為,轉(zhuǎn)移的確切機(jī)制目前尚不十分明確。 整合素連接激酶(integrin-linked kinase,ILK)是一個(gè)59KD的細(xì)胞內(nèi)信號(hào)蛋白,具有絲氨酸／蘇氨酸蛋白激酶活性。ILK不僅在正常發(fā)育中起著重要作用,其在腫瘤轉(zhuǎn)移過(guò)程中也有著與其它已知的癌基因一樣的生物學(xué)效應(yīng)。ILK介導(dǎo)細(xì)胞與細(xì)胞外基質(zhì)的連接,調(diào)節(jié)肌動(dòng)蛋白細(xì)胞骨架的運(yùn)動(dòng),參與細(xì)胞粘附的形成,促進(jìn)腫瘤細(xì)胞的粘附和轉(zhuǎn)移。 上皮-間充質(zhì)轉(zhuǎn)型(epithelial-mesenchymal transition, EMT)是多細(xì)胞生物形態(tài)學(xué)發(fā)生中的一個(gè)基本生理過(guò)程。是指上皮細(xì)胞向間質(zhì)細(xì)胞轉(zhuǎn)分化的現(xiàn)象,在此過(guò)程中,上皮細(xì)胞極性喪失,遷移能力增強(qiáng),同時(shí)逐漸獲得間質(zhì)表型。上皮-間充質(zhì)轉(zhuǎn)型參與了上皮源性腫瘤的發(fā)生及發(fā)展,尤其在癌的侵襲和轉(zhuǎn)移方面。因此,對(duì)EMT在腫瘤形成和侵襲轉(zhuǎn)移中的作用及其調(diào)控機(jī)制的研究正越來(lái)越受到人們的關(guān)注。目前,ILK作為一種EMT的誘導(dǎo)因子,在喉癌方面的研究幾乎是一片空白。因此,我們以人喉鱗狀細(xì)胞癌細(xì)胞株Hep-2為研究對(duì)象,采用反義ILK寡核苷酸對(duì)其進(jìn)行轉(zhuǎn)染,就ILK對(duì)人喉鱗狀細(xì)胞癌Hep-2細(xì)胞上皮-間充質(zhì)轉(zhuǎn)型的影響及其在腫瘤侵襲、轉(zhuǎn)移過(guò)程中的作用和機(jī)制進(jìn)行了研究。 方法：采用免疫組織化學(xué)法研究臨床喉鱗狀細(xì)胞癌標(biāo)本中ILK的表達(dá)與臨床病理因素的相關(guān)性,并且進(jìn)一步以人喉鱗狀細(xì)胞癌細(xì)胞株Hep-2為研究對(duì)象,采用ILK反義寡核苷酸對(duì)其進(jìn)行轉(zhuǎn)染,應(yīng)用MTT、流式細(xì)胞術(shù)分析、Western Blot以及Transwell小室等分子生物學(xué)技術(shù)和方法,深入研究了ILK對(duì)人喉鱗狀細(xì)胞癌Hep-2細(xì)胞上皮-間充質(zhì)轉(zhuǎn)型相關(guān)基因表達(dá)的影響及其在腫瘤侵襲、轉(zhuǎn)移過(guò)程中的作用和機(jī)制。 結(jié)果：ILK在56例喉鱗狀細(xì)胞癌患者癌組織標(biāo)本中的陽(yáng)性表達(dá)率(66.07%)明顯高于ILK在28例癌旁組織中的陽(yáng)性表達(dá)率(21.43%),統(tǒng)計(jì)學(xué)分析結(jié)果顯示二者存在顯著性差異(p0.05)。并且,ILK的高表達(dá)與喉癌患者病理分級(jí)及是否存在淋巴結(jié)轉(zhuǎn)移密切相關(guān)。ILK在人喉鱗狀細(xì)胞癌Hep-2細(xì)胞中亦呈高表達(dá),免疫細(xì)胞化學(xué)顯示ILK在人喉鱗狀細(xì)胞癌Hep-2細(xì)胞內(nèi)的表達(dá)以胞質(zhì)為主,胞核僅見(jiàn)少量弱表達(dá)。反義ILK干擾降低了人喉鱗狀細(xì)胞癌Hep-2細(xì)胞的ILK的表達(dá),抑制了細(xì)胞的增殖并增加了細(xì)胞的凋亡。EMT相關(guān)基因的表達(dá)亦出現(xiàn)了相應(yīng)的變化：間充質(zhì)標(biāo)志物Vimentin蛋白表達(dá)水平降低,上皮性標(biāo)志物E-cadherin的表達(dá)顯著增加。這些基因蛋白水平的改變也同時(shí)導(dǎo)致了人喉鱗狀細(xì)胞癌Hep-2細(xì)胞異質(zhì)粘附能力、遷移能力以及侵襲能力的降低。 結(jié)論：ILK可促進(jìn)人喉鱗狀細(xì)胞癌發(fā)生轉(zhuǎn)移,反義ILK干擾可誘導(dǎo)人喉鱗狀細(xì)胞癌Hep-2細(xì)胞間充質(zhì)-上皮細(xì)胞轉(zhuǎn)型的發(fā)生,并因此而顯著降低其侵襲和轉(zhuǎn)移潛力。
[Abstract]:The research background and purpose: Laryngeal square cell carcinoma (LSCC) is one of the most common malignant tumors in the head and neck, and the invasion and metastasis of the tumor is one of the main causes of the death of the patients with laryngeal carcinoma. The invasion and metastasis of the laryngeal carcinoma is a multi-link and multi-step complex process. Including the biological behavior of various tumor cells such as tumor cell adhesion, movement, enzyme degradation and angiogenesis, the exact mechanism of the transfer is not yet very clear. integrin-linked kinase (ILK) is a 59KD intracellular signal protein with serine/ threonine protein The activity of ILK plays an important role not only in the normal development, but also in the process of tumor metastasis. Study effect. ILK mediates the connection of the cell to the extracellular matrix, regulates the movement of the actin cytoskeleton, participates in the formation of cell adhesion, promotes the adhesion of the tumor cells, Epithelium-Mesenchymal transition (EMT) is one of the multi-cell biological morphology. A basic physiological process. It refers to the phenomenon of the differentiation of the epithelial cells to the mesenchymal cells. In this process, the polarity of the epithelial cells is lost, the migration ability is enhanced, and at the same time, The formation and development of epithelial-derived tumors are involved in the transformation of epithelial-mesenchymal transition, especially in the case of cancer. Therefore, the role of EMT in tumor formation and invasion and metastasis and its regulation and control mechanism are becoming more and more important At present, ILK, as an induction factor for EMT, has been studied in the field of laryngeal cancer. The effect of ILK on the transformation of human laryngeal squamous cell carcinoma (Hep-2) cells and its role in the invasion and metastasis of human laryngeal squamous cell carcinoma (Hep-2) was studied by using human laryngeal squamous cell carcinoma cell line Hep-2 as the research object. Methods: The relationship between the expression of ILK and the clinicopathological factors in the clinical specimens of laryngeal squamous cell carcinoma was studied by immunocytochemical method, and the human laryngeal squamous cell carcinoma cell line Hep-2 was used as the research object, and the ILK antisense oligonucleotide was used. The effects of ILK on the expression of human laryngeal squamous cell carcinoma (Hep-2) and the expression of mesenchymal transition-related genes in human laryngeal squamous cell carcinoma (Hep-2) and its tumor invasion were studied by means of MTT, flow cytometry, Western Blot and Transwell. Results: The positive expression rate of ILK in 56 patients with laryngeal squamous cell carcinoma (66.07%) was significantly higher than that of ILK in 28 patients with carcinoma (21.43%). There was a significant difference in the expression of ILK in the patients with laryngeal carcinoma (p0.05). The expression of ILK in the Hep-2 cells of human laryngeal squamous cell carcinoma (Hep-2) and the expression of ILK in the human laryngeal squamous cell carcinoma (Hep-2) were highly correlated with the presence or absence of lymph node metastasis. The expression of ILK in human laryngeal squamous cell carcinoma (Hep-2) cells was reduced by the expression of ILK in human laryngeal squamous cell carcinoma (Hep-2). The proliferation of the cells and the increase of the apoptosis of the cells. The expression of the EMT-related genes also showed a corresponding change: the level of the expression of the mesenchymal marker Vimentin, the epithelial marker E- There was a significant increase in the expression of cadherin. The changes in the level of these genes also led to the heterogeneous adhesion of the Hep-2 cells in the human laryngeal squamous cell carcinoma. Conclusion: ILK can promote the transfer of human laryngeal squamous cell carcinoma, and the anti-sense ILK interference can induce the transformation of human laryngeal squamous cell carcinoma Hep-2 cell-derived mesenchymal-epithelial cells.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R739.65

【共引文獻(xiàn)】

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