VTCN1基因在神经胶质瘤发生发展中作用机制的研究
发布时间：2018-07-29 08:55
【摘要】：研究目的： 一些肿瘤细胞系呈组成性表达VTCNl。在肺癌、卵巢癌、肾细胞癌均有VTCN1蛋白的表达；乳腺癌和子宫内膜癌几乎普遍存在VTCN1的表达,且不依赖肿瘤的分级和分期,而且这种VTCN1高表达是明显增加神经胶质瘤病例死亡风险的独立因素。但VTCN1在神经胶质瘤肿瘤组织表达的确切临床意义还有待进一步验证,VTCN1是否可以作为神经胶质瘤早期诊断和预后评价的指标也值得进一步深入研究。因此,本部分拟探讨VTCN1在神经胶质瘤组织中的表达及其与临床病理及预后的关系。 研究方法： 以酶联免疫反应(ELISA)检测神经胶质瘤患者外周血中VTCN1蛋白的含量,以逆转录聚合酶链反应(RT-PCR)技术检测神经胶质瘤患者手术切除的肿瘤组织标本中VTCN1基因表达水平,以免疫组织化学染色(IHC)检测神经胶质瘤患者手术切除的肿瘤组织标本中VTCN1蛋白质表达水平。 研究结果： VTCN1蛋白在神经胶质瘤患者外周血中的含量(44.80±8.20gg/L)明显高于脑部良性肿瘤患者(35.10±6.33μg/L)和正常人群(32.17±5.96μg/L)(P0.05)。随后检测VTCN1mRNA在30例神经胶质瘤组织和癌旁组织中均有表达,神经胶质瘤组织中表达明显高于癌旁组织(P0.01),且癌旁组织中的表达明显高于脑部良性肿瘤组织(P0.01)。进一步检测70例神经胶质瘤组织中VTCN1蛋白的表达,其阳性表达率68.57%,其中77.08%为高表达,明显高于脑部良性肿瘤组织(P0.01)。VTCN1蛋白的表达与神经胶质瘤组织学分级、肿瘤大小及肿瘤复发或转移密切相关(P0.05),而与患者年龄、性别和组织学类型等无关(P0.05)。VTCN1高表达患者生存率明显低于低表达患者(P0.05)。 研究结论： VTCN1在神经胶质瘤组织表达状态与神经胶质瘤的恶性程度、进展及预后关系密切,可以作为反映神经胶质瘤生物学行为和判断预后的有效指标。 研究目的： VTCN1基因家族负性分子在肿瘤的异常表达提示其在肿瘤发生发展中发挥了重要作用,这其中涉及的机制将为干预肿瘤的发生发展提供有效途径,具有较大的潜在应用价值。越来越多的临床研究已发现VTCN1分子与肿瘤免疫调控密切相关,但在机体肿瘤免疫应答中的作用机理还有待进一步研究。因此本部分通过RNA干扰技术下调VTCN1基因在人神经胶质瘤U251细胞株中的表达,探讨VTCN1基因表达下调后对神经胶质瘤细胞生物学行为的影响。 研究方法： 将针对VTCNI基因的siRNA序列重组入真核表达载体中构建pU-VTCN1-siRNA,转染至神经胶质瘤U251细胞株中,采用逆转录PCR(RT-PCR)、免疫印迹法检测转染pU-VTCN1-siRNA后的U251细胞中VTCN1基因在基因表达水平和蛋白质表达水平的变化。采用流式细胞仪分析转染pU-VTCN1-siRNA后的U251细胞的细胞周期变化,应用Transwell侵袭小室模型观察转染pU-VTCN1-siRNA后U251细胞侵袭力的变化。 研究结果： 转染pU-VTCN1-siRNA后的U251细胞中,VTCN1基因的基因表达水平和蛋白质表达水平显著下降(P0.05),细胞周期被阻滞在G1期,细胞生长缓慢,体外侵袭能力明显下降(P0.05)。研究结论： 通过下调VTCN1基因的表达,能有效抑制U251细胞的生长、增殖和侵袭能力,提示VTCN1基因在神经胶质瘤的发生、发展过程中起重要作用。
[Abstract]:The purpose of the study is:
Some tumor cell lines are composed of VTCNl. in the expression of VTCN1 protein in lung cancer, ovarian cancer and renal cell carcinoma; the expression of VTCN1 is almost common in breast and endometrial cancer, and does not depend on the classification and staging of the tumor. Moreover, this high expression of VTCN1 is an independent factor to increase the risk of death in the cases of glioma. But VTC The exact clinical significance of the expression of N1 in glioma tumor tissue remains to be further verified. Whether VTCN1 can be used as an indicator of early diagnosis and prognostic evaluation of glioma is also worth further study. Therefore, this part is to explore the expression of VTCN1 in glioma tissue and its relationship with clinicopathology and prognosis.
Research methods:
The content of VTCN1 protein in peripheral blood of glioma patients was detected by enzyme linked immunosorbent assay (ELISA), and the expression of VTCN1 gene expression in the tumor tissue specimens of patients with glioma was detected by reverse transcriptase polymerase chain reaction (RT-PCR), and the surgical resection of glioma patients was detected by immunohistochemical staining (IHC). The expression level of VTCN1 protein in tissue specimens.
The results of the study:
The content of VTCN1 protein in peripheral blood of glioma patients (44.80 + 8.20gg/L) was significantly higher than that of benign tumors in the brain (35.10 + 6.33 g/L) and normal people (32.17 + 5.96 g/L) (P0.05). Then VTCN1mRNA was detected in 30 cases of glioma tissue and para cancerous tissue, and the expression of glioma tissues was significantly higher than that of the para cancerous tissue. The expression of VTCN1 in the tissue (P0.01) was significantly higher than that of the benign tumor tissue (P0.01). The expression of VTCN1 protein in 70 cases of glioma was further detected. The positive expression rate was 68.57%, of which 77.08% was high, which was significantly higher than the expression of.VTCN1 protein in the benign tumor tissue of the brain and the histological grade of glioma. The tumor size and tumor recurrence or metastasis were closely related (P0.05), but the survival rate of patients with high expression of.VTCN1 was significantly lower than that of low expression patients (P0.05), which was not related to age, sex and histology type (P0.05).
The conclusions are as follows:
The expression of VTCN1 in glioma tissue is closely related to the malignancy, progression and prognosis of glioma. It can be used as an effective indicator to reflect the biological behavior of glioma and to judge the prognosis.
The purpose of the study is:
The abnormal expression of negative molecules in the VTCN1 gene family suggests that it plays an important role in the development of tumor. The mechanism involved will provide an effective way to interfere with the development of tumor, and it has great potential application value. More and more clinical studies have shown that VTCN1 molecules are closely related to the immunoregulation of tumor. However, the mechanism of action in the immune response of the body remains to be further studied. Therefore, this part reduces the expression of VTCN1 gene in human glioma U251 cell lines by RNA interference technique, and explores the effect of down regulation of VTCN1 gene expression on the biological behavior of glioma cells.
Research methods:
The siRNA sequence of VTCNI gene was reorganized into the eukaryotic expression vector to construct pU-VTCN1-siRNA and transfected into the glioma U251 cell line. Reverse transcriptase PCR (RT-PCR) was used to detect the change of VTCN1 gene in the gene expression level and protein expression level in U251 cells transfected with pU-VTCN1-siRNA. Flow cytometry was used. The cell cycle changes of U251 cells after transfection of pU-VTCN1-siRNA were analyzed by means of the instrument, and the invasiveness of U251 cells after transfection of pU-VTCN1-siRNA was observed by Transwell invasion model.
The results of the study:
In the U251 cells transfected with pU-VTCN1-siRNA, the gene expression level and protein expression level of VTCN1 gene decreased significantly (P0.05), the cell cycle was blocked in the G1 stage, the cell growth was slow and the invasion ability in vitro decreased significantly (P0.05).
Down regulation of the expression of VTCN1 gene can effectively inhibit the growth, proliferation and invasion of U251 cells, suggesting that the VTCN1 gene plays an important role in the development of glioma.
【学位授予单位】：武汉大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R739.41
本文编号：2152160
[Abstract]:The purpose of the study is:
Some tumor cell lines are composed of VTCNl. in the expression of VTCN1 protein in lung cancer, ovarian cancer and renal cell carcinoma; the expression of VTCN1 is almost common in breast and endometrial cancer, and does not depend on the classification and staging of the tumor. Moreover, this high expression of VTCN1 is an independent factor to increase the risk of death in the cases of glioma. But VTC The exact clinical significance of the expression of N1 in glioma tumor tissue remains to be further verified. Whether VTCN1 can be used as an indicator of early diagnosis and prognostic evaluation of glioma is also worth further study. Therefore, this part is to explore the expression of VTCN1 in glioma tissue and its relationship with clinicopathology and prognosis.
Research methods:
The content of VTCN1 protein in peripheral blood of glioma patients was detected by enzyme linked immunosorbent assay (ELISA), and the expression of VTCN1 gene expression in the tumor tissue specimens of patients with glioma was detected by reverse transcriptase polymerase chain reaction (RT-PCR), and the surgical resection of glioma patients was detected by immunohistochemical staining (IHC). The expression level of VTCN1 protein in tissue specimens.
The results of the study:
The content of VTCN1 protein in peripheral blood of glioma patients (44.80 + 8.20gg/L) was significantly higher than that of benign tumors in the brain (35.10 + 6.33 g/L) and normal people (32.17 + 5.96 g/L) (P0.05). Then VTCN1mRNA was detected in 30 cases of glioma tissue and para cancerous tissue, and the expression of glioma tissues was significantly higher than that of the para cancerous tissue. The expression of VTCN1 in the tissue (P0.01) was significantly higher than that of the benign tumor tissue (P0.01). The expression of VTCN1 protein in 70 cases of glioma was further detected. The positive expression rate was 68.57%, of which 77.08% was high, which was significantly higher than the expression of.VTCN1 protein in the benign tumor tissue of the brain and the histological grade of glioma. The tumor size and tumor recurrence or metastasis were closely related (P0.05), but the survival rate of patients with high expression of.VTCN1 was significantly lower than that of low expression patients (P0.05), which was not related to age, sex and histology type (P0.05).
The conclusions are as follows:
The expression of VTCN1 in glioma tissue is closely related to the malignancy, progression and prognosis of glioma. It can be used as an effective indicator to reflect the biological behavior of glioma and to judge the prognosis.
The purpose of the study is:
The abnormal expression of negative molecules in the VTCN1 gene family suggests that it plays an important role in the development of tumor. The mechanism involved will provide an effective way to interfere with the development of tumor, and it has great potential application value. More and more clinical studies have shown that VTCN1 molecules are closely related to the immunoregulation of tumor. However, the mechanism of action in the immune response of the body remains to be further studied. Therefore, this part reduces the expression of VTCN1 gene in human glioma U251 cell lines by RNA interference technique, and explores the effect of down regulation of VTCN1 gene expression on the biological behavior of glioma cells.
Research methods:
The siRNA sequence of VTCNI gene was reorganized into the eukaryotic expression vector to construct pU-VTCN1-siRNA and transfected into the glioma U251 cell line. Reverse transcriptase PCR (RT-PCR) was used to detect the change of VTCN1 gene in the gene expression level and protein expression level in U251 cells transfected with pU-VTCN1-siRNA. Flow cytometry was used. The cell cycle changes of U251 cells after transfection of pU-VTCN1-siRNA were analyzed by means of the instrument, and the invasiveness of U251 cells after transfection of pU-VTCN1-siRNA was observed by Transwell invasion model.
The results of the study:
In the U251 cells transfected with pU-VTCN1-siRNA, the gene expression level and protein expression level of VTCN1 gene decreased significantly (P0.05), the cell cycle was blocked in the G1 stage, the cell growth was slow and the invasion ability in vitro decreased significantly (P0.05).
Down regulation of the expression of VTCN1 gene can effectively inhibit the growth, proliferation and invasion of U251 cells, suggesting that the VTCN1 gene plays an important role in the development of glioma.
【学位授予单位】：武汉大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R739.41
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