IEX-1的UVB诱导性及其与HIF-1α、NF-κB在宫颈癌组织中表达的相关性

发布时间：2018-03-19 09:57

本文选题：即刻早期反应基因-1　切入点：辐射诱导性　出处：《郑州大学》2014年硕士论文　论文类型：学位论文

【摘要】：背景和目的子宫颈癌是妇科三大常见的恶性肿瘤之一,严重威胁着女性的健康。随着对肿瘤的发生、发展机制的不断深入，基因治疗成为继手术、放疗、化疗等传统治疗方式后的新的治疗手段。肿瘤基因-放射治疗也成为肿瘤治疗研究的新热点。即刻早期反应基因-1(Immediate Early Response Gene X-1，IEX-1),属于即刻早期基因（IEG）家族之一，定位于人6号染色体短臂(6p21.3)，是一种能在生长因子、紫外线、电离辐射及缺血缺氧等诱导下快速表达,调控细胞的生长、增殖、分化和凋亡等各方面的基因。IEX-1的在不同组织中的过表达可正性或负性影响细胞的生存和细胞进程。有研究表明IEX-1mRNA的表达在受到紫外线照射之后显著增加，具有时间和剂量依赖性。IEX-1的UVB诱导性使得其成为肿瘤基因治疗的靶点具有可能性,也为其带来了潜在的临床应用性。研究证实IEX-1在多种肿瘤组织中低表达或表达消失；但其在宫颈癌组织中的表达情况尚罕有报道。本研究采用紫外线照射方法照射宫颈癌HeLa细胞，分析宫颈癌细胞中IEX-1基因的辐射可诱导性，同时应用免疫组化的方法从蛋白水平分别检测IEX-1、HIF-1α、NF-κB在宫颈癌组织,宫颈CIN组织及正常宫颈组织的表达情况及三者间的关系，进一步探讨IEX-1与宫颈癌发生发展的联系，为其作为临床治疗宫颈癌的靶点基因提供依据。方法 1．采用不同剂量紫外线照射人宫颈癌HeLa细胞，RT-PCR检测照射后细胞IEX-1mRNA的表达情况，选出最佳紫外线剂量。 2．以最佳紫外线照射剂量照射人宫颈癌HeLa细胞，RT-PCR检测照射后不同时间点细胞中IEX-1mRNA的表达情况。 3．应用免疫组织化学SP法检测50例宫颈癌患者、40例CIN及26例正常宫颈组织中IEX-1、HIF-1α、NF-κB蛋白的表达情况，并分析三者之间的关系。 4．应用SPSS16.0进行统计学分析，χ2检验和Spearman等级相关分析,检验水准α=0.05,以P＜0.05为差异有统计学意义。结果 1．分别应用不同等剂量的UVB照射人宫颈癌HeLa细胞后，，IEX-1mRNA表达量均有明显增高（P0.05）。以40mJ/cm2的UVB剂量照射细胞后IEX-1mRNA表达增高幅度最大，为最佳紫外线照射剂量。 2．以最佳紫外线剂量40mJ/cm2照射人宫颈癌HeLa细胞，培养不同时间后检测IEX-1mRNA表达的表达量，培养4小时后IEX-1mRNA表达量最高（P0.05）。 3．IEX-1在宫颈癌组织中的阳性表达率明显低于宫颈CIN和正常宫颈组织,差异具有统计学意义(P0.05)；HIF-1α和NF-κB在宫颈癌组织中的阳性表达率均明显高于宫颈CIN和正常宫颈组织,差异具有统计学意义(P0.05)。 4．宫颈癌组织中IEX-1、HIF-1α、 NF-κB的表达均与组织学分级及有无淋巴结转移相关(P0.05),与患者年龄、肿瘤的组织学类型及FIGO分期无关(P0.05)。 5．宫颈癌组织中HIF-1α和NF-κB的表达呈正相关（r=0.398,P0.05）,两者同IEX-1的表达呈负相关(r1=-0.322,P0.05；r2=-0.517,P0.05)。结论 1．人宫颈癌HeLa细胞中IEX-1具有辐射诱导性，可被UVB诱导表达，且具有时间和剂量依赖性。 2．IEX-1在宫颈癌组织中低表达，与NF-κB、HIF-1α表达呈负相关，三者共同调节肿瘤细胞的生长和凋亡。
[Abstract]:Background and purpose
Cervical cancer is one of the three most common gynecologic malignant tumor, a serious threat to women's health. With the tumor occurrence, development mechanism deeply, gene therapy has become following surgery, radiotherapy, chemotherapy and other new treatment after conventional treatment. Tumor gene radiotherapy has become a new hotspot in cancer treatment.
Immediate early response gene -1 (Immediate Early Response Gene X-1, IEX-1), which belongs to the family of immediate early genes (IEG), located on the short arm of chromosome 6 (6p21.3), is a growth factor in the ultraviolet, ionizing radiation, hypoxia and ischemia induced the rapid expression, regulation of cell growth the proliferation in different tissues. The overexpression of positive or negative effects of cell survival and cell process of.IEX-1 gene in all aspects of differentiation and apoptosis. The study showed that the expression of IEX-1mRNA increased significantly after exposure to ultraviolet light, a time and dose dependent induction of.IEX-1 UVB has become a target for tumor gene therapy has the possibility, also brings the potential for its clinical application.
Study confirmed that low expression of IEX-1 in tumor tissue or expression disappeared; but its expression in cervical cancer tissues of Shanghan have been reported. This study used the irradiation of cervical cancer HeLa cells induced by ultraviolet irradiation method, analysis of radiation IEX-1 expression in cervical cancer cells, and immunohistochemistry at protein level were detected IEX-1, HIF-1 alpha, NF- kappa B in cervical cancer, the relationship between the expression of CIN in cervical tissue and normal cervical tissue and between the three, to further explore the occurrence and development of IEX-1 and cervical cancer, and provide the basis for the clinical treatment of cervical cancer as a target gene.
Method
1. the HeLa cells of human cervical cancer were irradiated with different doses of ultraviolet radiation. The expression of IEX-1mRNA in the cells after irradiation was detected by RT-PCR, and the best ultraviolet dose was selected.
2. the human cervical cancer HeLa cells were irradiated with the best ultraviolet radiation dose, and the expression of IEX-1mRNA in the cells at different time points after irradiation was detected by RT-PCR.
3. immunohistochemical SP method was used to detect the expression of IEX-1, HIF-1, NF- and kappa B protein in 50 cases of cervical cancer, 40 cases of CIN and 26 cases of normal cervical tissue, and the relationship between the three groups was analyzed.
4. SPSS16.0 was used for statistical analysis, x 2 test and Spearman grade correlation analysis, the level of alpha =0.05 was tested, and the difference was statistically significant with the difference of P < 0.05.
Result
1. the expression level of IEX-1mRNA was significantly increased after different doses of UVB irradiation on human cervical cancer HeLa cells respectively (P0.05). The highest expression of IEX-1mRNA was observed after irradiation with UVB dose of 40mJ/cm2, which was the best ultraviolet radiation dose.
2., we irradiated human cervical cancer HeLa cells with the best ultraviolet dose of 40mJ/cm2, and detected the expression level of IEX-1mRNA after different time. After 4 hours of culture, the expression level of IEX-1mRNA was the highest (P0.05).
The positive expression of 3.IEX-1 in cervical cancer tissues was significantly lower than that in normal cervical tissue and cervical CIN, the difference was statistically significant (P0.05); the positive expression of HIF-1 kappa B alpha and NF- in cervical cancer tissues was significantly higher in cervical CIN and normal cervical tissue, the difference has statistical significance (P0.05).
4. the expression of IEX-1, HIF-1 and NF- kappa B in cervical cancer tissues was correlated with histological grade and lymph node metastasis (P0.05), which was not related to the age of patients, histological type and FIGO stage of tumor (P0.05).
5. the expression of HIF-1 alpha and NF- kappa B in cervical cancer tissues was positively correlated (r=0.398, P0.05), and they were negatively correlated with the expression of IEX-1 (r1=-0.322, P0.05; r2=-0.517, P0.05).
conclusion
In 1. human cervical cancer HeLa cells, IEX-1 is radiation-induced, which can be induced by UVB and has time and dose dependence.
The low expression of 2.IEX-1 in cervical cancer tissues is negatively correlated with the expression of NF- kappa B and HIF-1 alpha, and the three together regulate the growth and apoptosis of tumor cells.

【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R737.33

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