腫瘤巨細(xì)胞與口腔鱗癌臨床病理因素相關(guān)性研究及其相關(guān)機(jī)制初步探討
發(fā)布時(shí)間：2019-01-04 17:34
【摘要】：目的 探究腫瘤巨細(xì)胞(giant cancer cell,GCC)在口腔鱗癌(oral squamous cell carcinoma,OSCC)組織中的存在情況及其數(shù)目與OSCC臨床病理因素的關(guān)系。并初步探究腫瘤巨細(xì)胞產(chǎn)生的可能機(jī)制及其對OSCC增殖、耐藥和遷移能力的影響。方法 1、腫瘤巨細(xì)胞在OSCC中的存在情況及其數(shù)目與臨床病理因素的相關(guān)性研究:收集2005年3月至2014年12月天津醫(yī)科大學(xué)腫瘤醫(yī)院76例OSCC患者的石蠟切片及臨床病理資料。HE染色下觀察GCC與普通腫瘤細(xì)胞的形態(tài)差異,并計(jì)數(shù)GCC。免疫組織化學(xué)染色下觀察GCC與普通腫瘤細(xì)胞Ki-67表達(dá)差異。分析GCC數(shù)目與OSCC臨床生物學(xué)行為及預(yù)后的關(guān)系。使用SPSS18.0對收集的數(shù)據(jù)進(jìn)行分析,認(rèn)為P0.05為差異具有統(tǒng)計(jì)學(xué)意義。2、腫瘤巨細(xì)胞產(chǎn)生的可能機(jī)制及其對OSCC增殖、耐藥和遷移能力的影響:通過CCK-8實(shí)驗(yàn)確定細(xì)胞乏氧的Co Cl2濃度,用Co Cl2對OSCC細(xì)胞系進(jìn)行誘導(dǎo)處理,顯微鏡下觀察GCC的形成及其產(chǎn)生子代細(xì)胞的過程。通過real-time PCR檢測誘導(dǎo)前后細(xì)胞干性標(biāo)記物mi RNA水平表達(dá)的變化;通過western blotting檢測乏氧誘導(dǎo)因子HIF-1α和干性標(biāo)記物的蛋白水平表達(dá)變化;通過CCK-8實(shí)驗(yàn)檢測誘導(dǎo)前后細(xì)胞增殖能力及耐化療藥(順鉑)能力的變化;通過劃痕實(shí)驗(yàn)檢測細(xì)胞遷移能力的變化。Western blotting檢測遷移相關(guān)蛋白的表達(dá)變化。結(jié)果 1、GCC存在于口腔鱗癌組織中,且其細(xì)胞核體積大或含有多個(gè)細(xì)胞核,細(xì)胞核形態(tài)不規(guī)則,可呈啞鈴型、分葉狀等多種不規(guī)則形態(tài),可見多個(gè)核仁。Ki-67在OSCC胞核中部分陽性表達(dá),在GCC細(xì)胞核中均呈陽性表達(dá)。GCC細(xì)胞數(shù)與臨床分期、組織學(xué)分級、化療及復(fù)發(fā)相關(guān)。臨床分期、淋巴結(jié)轉(zhuǎn)移、組織學(xué)分級、局部復(fù)發(fā)及遠(yuǎn)處轉(zhuǎn)移和OS相關(guān);臨床分期、淋巴結(jié)轉(zhuǎn)移、局部復(fù)發(fā)及遠(yuǎn)處轉(zhuǎn)移和DFS相關(guān)。術(shù)前誘導(dǎo)化療不能改善OS和DFS。GCC數(shù)目3個(gè)/HP組OS和DFS明顯較GCC數(shù)目≤3個(gè)/HP組者差,GCC細(xì)胞可能是影響OSCC預(yù)后的重要因素。臨床分期、局部復(fù)發(fā)和遠(yuǎn)處轉(zhuǎn)移是影響OSCC患者OS的獨(dú)立預(yù)后因素;臨床分期、局部復(fù)發(fā)和遠(yuǎn)處轉(zhuǎn)移是影響OSCC患者DFS的獨(dú)立預(yù)后因素。2、以UM1為細(xì)胞模型,通過Co Cl2多次處理誘導(dǎo)純化形成GCC。乏氧后,大部分普通大小的腫瘤細(xì)胞被殺死,留下純化的GCC或者經(jīng)細(xì)胞融合形成的GCC。GCC細(xì)胞體積明顯較普通腫瘤細(xì)胞大,其內(nèi)可含有多個(gè)細(xì)胞核,其形態(tài)多樣,通過不均分裂(出芽)產(chǎn)生子代細(xì)胞。Real-time PCR結(jié)果顯示GCC細(xì)胞及其子代細(xì)胞的干性標(biāo)記物NANOG、SOX-2在mi RNA水平表達(dá)明顯升高。Western blotting結(jié)果顯示GCC及其子代細(xì)胞的HIF-1α及SOX-2在蛋白水平表達(dá)明顯升高。誘導(dǎo)形成GCC后,CCK-8結(jié)果顯示腫瘤細(xì)胞的增殖能力上升,耐藥能力上升;劃痕實(shí)驗(yàn)結(jié)果顯示腫瘤細(xì)胞的遷移能力上升;Western blotting檢測遷移相關(guān)蛋白,E-cadherin的表達(dá)下調(diào),N-cadherin、Vimentin的表達(dá)上調(diào)。結(jié)論 1、腫瘤巨細(xì)胞細(xì)胞數(shù)與口腔鱗癌臨床分期、組織學(xué)分級、誘導(dǎo)化療和局部復(fù)發(fā)相關(guān),可能是影響口腔鱗癌預(yù)后的重要因素。腫瘤巨細(xì)胞可能在口腔鱗癌的發(fā)展及耐藥過程中發(fā)揮作用。2、乏氧可誘導(dǎo)口腔鱗癌細(xì)胞產(chǎn)生腫瘤巨細(xì)胞,具有腫瘤干細(xì)胞的部分特性,腫瘤巨細(xì)胞及其子代細(xì)胞干性標(biāo)記物表達(dá)上調(diào)并發(fā)生EMT,其增殖、耐藥和遷移能力均增強(qiáng)。
[Abstract]:Objective To investigate the presence of tumor giant cell (GCC) in oral squamous cell carcinoma (OSCC) and its relationship with the clinicopathological factors of OSCC. The possible mechanism of the tumor giant cell production and its effect on the proliferation, drug resistance and migration ability of OSCC were investigated. Method 1. The presence of tumor giant cells in OSCC and its correlation with the clinical and pathological factors were studied. The paraffin sections and clinical pathological data of 76 patients with OSCC from March 2005 to December 2014 were collected. The morphological differences of GCC and normal tumor cells were observed with HE staining and the GCC was counted. The expression of Ki-67 in GCC and normal tumor cells was observed with immunohistochemical staining. The relationship between the number of GCC and the clinical biological behavior and prognosis of OSCC was analyzed. The collected data was analyzed using SPSS18. 0, which was considered to have a statistical significance on the difference of P0. 05. The possible mechanism of the tumor giant cell production and its effect on the proliferation, drug resistance and migration ability of the OSCC were determined by the CCK-8 experiment. The OSCC cell line was induced with Co Cl2, and the formation of GCC and the process of producing subcell were observed under the microscope. The changes of the expression of the expression of mi-RNA in the cell-dry marker before and after induction were detected by the real-time PCR, and the changes of the expression of the protein in the HIF-1 and the dry markers were detected by western blotting, and the ability of the cell proliferation and the resistance to chemotherapy (cisplatin) was detected by CCK-8. The change of cell migration ability was detected by a scratch test. The expression of the related protein was detected by Western blotting. Results 1, GCC was present in the oral squamous cell carcinoma, and its nuclear volume is large or contains a plurality of nuclei, and the nuclear form is irregular, and can be made into a plurality of irregular shapes such as a dumbbell type, a lobule and the like, and a plurality of nucleoli can be seen. The positive expression of Ki-67 in the core of the OSCC was positive in the nucleus of the GCC. The number of GCC cells was related to clinical stage, histological grade, chemotherapy and recurrence. Clinical stage, lymph node metastasis, histological grade, local recurrence and distant metastasis and OS related; clinical stage, lymph node metastasis, local recurrence and distant metastasis and DFS correlation. The pre-operative induction of chemotherapy could not improve OS and DFS. The number of GCC 3/ HP group OS and DFS was significantly lower than that of the 3/ HP group, and GCC cells may be an important factor in the prognosis of OSCC. Clinical staging, local recurrence and distant metastasis are independent prognostic factors that affect the OS in OSCC patients; clinical staging, local recurrence and distant metastasis are independent prognostic factors that affect DFS in OSCC patients. After spent oxygen, most of the normal size of the tumor cells are killed, leaving purified GCC or the GCC formed by cell fusion. Real-time PCR results showed that the expression of NNOG and SOX-2 in the dry markers of GCC cells and its subcells increased significantly at the level of mi RNA. The results of Western blotting showed that the expression of HIF-1 and SOX-2 in GCC and its sub-cells increased significantly at the level of protein. After the formation of GCC, the results of CCK-8 showed that the proliferation ability of the tumor cells increased and the resistance to drug resistance increased; the results of the scratch test showed that the migration ability of the tumor cells increased; the expression of E-cadherin was down-regulated by Western blotting, and the expression of N-cadherin and Vimentin was up-regulated. Conclusion 1. The number of the tumor cells in the tumor is related to the clinical stage, the histological grade, the induction of chemotherapy and the local recurrence of the oral squamous cell carcinoma, which may be an important factor in the prognosis of oral squamous cell carcinoma. the tumor giant cell can play a role in the development and the drug resistance of the oral squamous cell carcinoma. Both resistance and migration were enhanced.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R739.8
本文編號：2400580
[Abstract]:Objective To investigate the presence of tumor giant cell (GCC) in oral squamous cell carcinoma (OSCC) and its relationship with the clinicopathological factors of OSCC. The possible mechanism of the tumor giant cell production and its effect on the proliferation, drug resistance and migration ability of OSCC were investigated. Method 1. The presence of tumor giant cells in OSCC and its correlation with the clinical and pathological factors were studied. The paraffin sections and clinical pathological data of 76 patients with OSCC from March 2005 to December 2014 were collected. The morphological differences of GCC and normal tumor cells were observed with HE staining and the GCC was counted. The expression of Ki-67 in GCC and normal tumor cells was observed with immunohistochemical staining. The relationship between the number of GCC and the clinical biological behavior and prognosis of OSCC was analyzed. The collected data was analyzed using SPSS18. 0, which was considered to have a statistical significance on the difference of P0. 05. The possible mechanism of the tumor giant cell production and its effect on the proliferation, drug resistance and migration ability of the OSCC were determined by the CCK-8 experiment. The OSCC cell line was induced with Co Cl2, and the formation of GCC and the process of producing subcell were observed under the microscope. The changes of the expression of the expression of mi-RNA in the cell-dry marker before and after induction were detected by the real-time PCR, and the changes of the expression of the protein in the HIF-1 and the dry markers were detected by western blotting, and the ability of the cell proliferation and the resistance to chemotherapy (cisplatin) was detected by CCK-8. The change of cell migration ability was detected by a scratch test. The expression of the related protein was detected by Western blotting. Results 1, GCC was present in the oral squamous cell carcinoma, and its nuclear volume is large or contains a plurality of nuclei, and the nuclear form is irregular, and can be made into a plurality of irregular shapes such as a dumbbell type, a lobule and the like, and a plurality of nucleoli can be seen. The positive expression of Ki-67 in the core of the OSCC was positive in the nucleus of the GCC. The number of GCC cells was related to clinical stage, histological grade, chemotherapy and recurrence. Clinical stage, lymph node metastasis, histological grade, local recurrence and distant metastasis and OS related; clinical stage, lymph node metastasis, local recurrence and distant metastasis and DFS correlation. The pre-operative induction of chemotherapy could not improve OS and DFS. The number of GCC 3/ HP group OS and DFS was significantly lower than that of the 3/ HP group, and GCC cells may be an important factor in the prognosis of OSCC. Clinical staging, local recurrence and distant metastasis are independent prognostic factors that affect the OS in OSCC patients; clinical staging, local recurrence and distant metastasis are independent prognostic factors that affect DFS in OSCC patients. After spent oxygen, most of the normal size of the tumor cells are killed, leaving purified GCC or the GCC formed by cell fusion. Real-time PCR results showed that the expression of NNOG and SOX-2 in the dry markers of GCC cells and its subcells increased significantly at the level of mi RNA. The results of Western blotting showed that the expression of HIF-1 and SOX-2 in GCC and its sub-cells increased significantly at the level of protein. After the formation of GCC, the results of CCK-8 showed that the proliferation ability of the tumor cells increased and the resistance to drug resistance increased; the results of the scratch test showed that the migration ability of the tumor cells increased; the expression of E-cadherin was down-regulated by Western blotting, and the expression of N-cadherin and Vimentin was up-regulated. Conclusion 1. The number of the tumor cells in the tumor is related to the clinical stage, the histological grade, the induction of chemotherapy and the local recurrence of the oral squamous cell carcinoma, which may be an important factor in the prognosis of oral squamous cell carcinoma. the tumor giant cell can play a role in the development and the drug resistance of the oral squamous cell carcinoma. Both resistance and migration were enhanced.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R739.8
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