發(fā)布時(shí)間：2019-02-15 08:57

【摘要】：背景和目的:胰腺癌是一種病死率極高的消化系統(tǒng)惡性腫瘤。侵襲轉(zhuǎn)移是胰腺癌的常見(jiàn)現(xiàn)象與局部復(fù)發(fā)和疼痛有著重要聯(lián)系。一氧化氮(NO)可調(diào)控胰腺癌細(xì)胞的生長(zhǎng)及侵襲等生物學(xué)行為,在維持胰腺癌的惡性生物學(xué)行為起到重要作用。NO可上調(diào)癌細(xì)胞自噬水平為侵襲轉(zhuǎn)移供應(yīng)能量。神經(jīng)浸潤(rùn)(PNI)是胰腺癌侵襲轉(zhuǎn)移的主要方式,也是胰腺癌轉(zhuǎn)移的特殊通道。研究目的:(1)觀察NO相關(guān)蛋白在胰腺癌中的表達(dá)及其臨床意義,探討NO相關(guān)蛋白與胰腺癌侵襲轉(zhuǎn)移尤其是神經(jīng)浸潤(rùn)(PNI)的相關(guān)性;(2)觀察NO供體對(duì)胰腺癌細(xì)胞增殖、遷移及侵襲的影響及其可能的分子機(jī)制。方法:(1)免疫組織化學(xué)染色技術(shù)檢測(cè)E-cadherin、iNOS、NQO1及LC-3在胰腺癌組織中的表達(dá),分析其與胰腺癌臨床病理特征及PNI之間的相關(guān)性。(2)用MTT方法檢測(cè)不同濃度DETA/NO對(duì)胰腺癌細(xì)胞株增殖的影響。(3)通過(guò)細(xì)胞劃痕實(shí)驗(yàn)及Transwell侵襲實(shí)驗(yàn)檢測(cè)DETA/NO對(duì)胰腺癌細(xì)胞株遷移及侵襲能力的影響。(4)應(yīng)用Real-time PCR技術(shù)檢測(cè)DETA/NO干預(yù)胰腺癌細(xì)胞株后其侵襲相關(guān)蛋白mRNA水平的變化。(5)Western blot檢測(cè)DETA/NO對(duì)胰腺癌細(xì)胞株自噬相關(guān)蛋白表達(dá)變化的影響。結(jié)果:(1)E-cadherin、iNOS、NQO1、及LC-3在胰腺癌組織中表達(dá)率分別是68.9%、63.9%、55.7%、57.4%,E-cadherin表達(dá)與LC-3具有相關(guān)性,LC-3與胰腺癌局部浸潤(rùn)發(fā)生具有相關(guān)性,E-cadherin、LC-3表達(dá)與胰腺癌神經(jīng)浸潤(rùn)具有顯著相關(guān)性。(2)不同濃度DETA/NO對(duì)胰腺癌細(xì)胞株增殖影響有差異,低濃度促進(jìn)癌細(xì)胞增殖水平,高濃度抑制增殖。(3)低濃度DETA/NO對(duì)胰腺癌細(xì)胞株進(jìn)行干預(yù),可促進(jìn)胰腺癌細(xì)胞株遷移及侵襲能力。(4)50μM濃度的DETA/NO干預(yù)Bx Pc-3細(xì)胞株可上調(diào)相關(guān)基因iNOS、TNF、MMP-2、MMP-9的mRNA水平,其中MMP-2的mRNA水平顯著增高;(5)用100 uM的DETA/NO干預(yù)1h后,LC-3表達(dá)水平即開(kāi)始升高,而隨干預(yù)時(shí)間延長(zhǎng)至8小時(shí)即又降至接近正常,而NQO1和Beclin 1無(wú)明顯差異。結(jié)論:NO代謝相關(guān)蛋白的表達(dá)與胰腺癌惡性生物學(xué)特性密切相關(guān),在胰腺癌進(jìn)展中可能扮演重要角色。低濃度NO促使胰腺癌細(xì)胞遷移侵襲能力增強(qiáng),可能與胰腺癌神經(jīng)浸潤(rùn)發(fā)生相關(guān);NO上調(diào)侵襲相關(guān)蛋白表達(dá)水平可能是改變胰腺癌細(xì)胞侵襲能力的分子機(jī)制;NO對(duì)細(xì)胞自噬蛋白表達(dá)水平的影響可能與癌細(xì)胞維持增殖活性及侵襲能力有關(guān)。本研究可為NO代謝途徑導(dǎo)致胰腺癌神經(jīng)浸潤(rùn)研究提供前期基礎(chǔ)。
[Abstract]:Background & objective: pancreatic cancer is a malignant tumor of digestive system with high mortality. Invasion and metastasis is a common phenomenon of pancreatic cancer and has an important relationship with local recurrence and pain. Nitric oxide (NO) can regulate the growth and invasion of pancreatic cancer cells and play an important role in maintaining the malignant biological behavior of pancreatic cancer. NO can up-regulate the level of autophagy of cancer cells to supply energy for invasion and metastasis. Neuroinvasive (PNI) is the main invasion and metastasis of pancreatic cancer, and it is also a special channel for pancreatic cancer metastasis. Objective: (1) to investigate the expression and clinical significance of NO related proteins in pancreatic carcinoma, and to explore the relationship between NO related proteins and invasion and metastasis of pancreatic carcinoma, especially (PNI). (2) to observe the effect of NO donor on the proliferation, migration and invasion of pancreatic cancer cells and its possible molecular mechanism. Methods: (1) Immunohistochemical staining was used to detect the expression of E-cadherinia iNOS NQO1 and LC-3 in pancreatic carcinoma. (2) the effect of different concentrations of DETA/NO on proliferation of pancreatic cancer cell line was detected by MTT method. (3) Cell scratch test and Transwell invasion assay were used to detect the effect of DETA/NO on the proliferation of pancreatic cancer cell line. Effects of DETA/NO on migration and invasion ability of pancreatic cancer cell line. (4) Real-time PCR technique was used to detect the changes of mRNA level of invasion-associated protein (mRNA) after DETA/NO intervention. (5) DETA/NO was detected by) Western blot. Effect on the expression of autophagy associated protein in pancreatic cancer cell line. Results: (1) the expression rates of E-cadherin and LC-3 in pancreatic carcinoma were 68.9 and 63.9, respectively. The expression of E-cadherin was correlated with LC-3. The expression of E-cadherinon LC-3 was significantly correlated with the neural invasion of pancreatic carcinoma. (2) the effects of different concentrations of DETA/NO on the proliferation of pancreatic cancer cell line were different. Low concentration of DETA/NO can promote the proliferation of pancreatic cancer cells and inhibit the proliferation of pancreatic cancer cells at high concentration. (3) low concentration of DETA/NO intervention on pancreatic cancer cell lines, (4) 50 渭 M concentration of DETA/NO could up-regulate the mRNA level of iNOS,TNF,MMP-2,MMP-9 gene in Bx Pc-3 cell line, and the mRNA level of MMP-2 increased significantly. (5) the expression of LC-3 began to increase 1 h after the treatment with 100 uM DETA/NO, and then decreased to close to normal as the intervention time extended to 8 hours, but there was no significant difference between NQO1 and Beclin 1. Conclusion: the expression of NO metabolism-related proteins is closely related to the malignant biological characteristics of pancreatic cancer and may play an important role in the progression of pancreatic cancer. Low concentration of NO enhanced the migration and invasion of pancreatic cancer cells, which may be related to the neuroinvasion of pancreatic cancer, and up-regulated the expression level of invasive-associated protein in pancreatic cancer cells, which may be the molecular mechanism that changes the invasion ability of pancreatic cancer cells. The effect of NO on the expression of autophagy protein may be related to the ability of cancer cell to maintain proliferation and invasion. This study may provide a preliminary basis for the study of the neural invasion of pancreatic cancer induced by NO metabolic pathway.
【學(xué)位授予單位】：河南科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R735.9

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相關(guān)期刊論文 前2條

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本文編號(hào)：2423173