性激素及其受体在鼻咽纤维血管瘤发病机制中的作用

发布时间：2018-06-23 01:25

本文选题：鼻咽纤维血管瘤 + 性激素　；参考：《复旦大学》2013年博士论文

【摘要】：鼻咽纤维血管瘤,又称青少年鼻咽纤维血管瘤(juvenile nasopharyngeal angiofibroma, JNA),是一种几乎只发生于青少年男性的鼻咽部良性肿瘤。JNA发病率较低,肿瘤极易出血,手术彻底切除难度较大。组织学上,JNA没有包膜,由未成熟的内皮细胞和成纤维细胞组成,虽然为良性肿瘤,但JNA具有局部侵犯的特征,易侵犯鼻窦、翼腭窝、颞下窝、翼板、眶上裂,甚至颅内。 JNA特定的发病年龄和性别很早就受到学者们的关注,众多学者期望从中找到研究JNA发病机制的相关线索。早期的研究认为JNA患者垂体-性腺轴发生混乱导致肿瘤的发生,但随后的研究表明JNA患者体内的性激素水平正常,且并没有第二性征的发育异常。近期的研究则关注肿瘤本身性激素受体的表达与疾病的关系,但至今为止并没有统一的结论。本研究收集了40例JNA患者的血清标本,对血清中的睾酮,游离睾酮,雌激素,孕激素的水平进行elisa检测,对比JNA患者的血清激素水平与对照组相比是否存在异常；将70例JNA的肿瘤组织和10例中甲组织制作成组织芯片,检测雄激素受体(androgen receptor, AR)、雌激素受体α (estrogen receptor α, ER-α)、雌激素受体β (estrogen receptor α, ER-β)、孕激素受体(progesterone receptor, PR)及血管内皮生长因子(vascular endothelial growth factor receptor, VEGF)在JNA中的表达及其相互关系；对5例高侵袭JNA组织、4例低侵袭JNA组织和5例正常中甲黏膜组织进行了表达谱基因芯片的研究,试图发现参与JNA发病的关键基因及信号通路。 JNA易侵犯邻近组织甚至颅内,术中极易出血,侵犯范围较广的肿瘤手术常常无法切除干净,且存在较大风险,药物及靶向治疗一直是研究的重点。JNA肿瘤组织中性激素受体的表达以及表达谱芯片的研究为进一步探究JNA治疗的新方法提供了基础。第一部分鼻咽纤维血管瘤患者血清中性激素的表达目的：鼻咽纤维血管瘤(JNA)是一种几乎只发生于青少年男性的良性肿瘤,但至今病因不明。JNA特殊的发病年龄和性别提示性激素在肿瘤发病过程的可能起作用,本研究旨在探明JNA患者是否存在青春期性激素水平异常以及性激素水平是否与肿瘤分期相关。方法：采集40例JNA患者(平均年龄18.3岁,均为术后病理证实)及14例正常对照组患者(平均年龄19.7岁)的血清,对照组患者为同时段、同年龄段、同性别的鼻中隔、骨瘤及视神经减压的患者。用Elisa法检测JNA组与对照组患者血清中的睾酮、游离睾酮、雌二醇及孕酮的水平,比较两组间性激素的水平以及雌雄激素的比值是否存在差异。收集40例JNA患者的肿瘤分期数据,分析其与血清中性激素的水平是否存在关联。结果：实验数据表明,JNA组与对照组相比,血清中的睾酮、游离睾酮、雌二醇及孕酮的水平并没有显著异常,且血清中睾酮与雌二醇的比值、游离睾酮和雌二醇的比值也无明显差异。肿瘤分期与JNA患者血清中的性激素水平没有显著相关性。结论：虽然JNA是一种特发于青少年男性的鼻咽部良性肿瘤,但是患者循环血清中的性激素水平以及雌雄激素的比值并无异常,性激素的水平与肿瘤分期也无相关性。这说明JNA患者本身性激素水平正常,也可能并不存在垂体-性腺轴的失平衡,肿瘤的发生可能是由于原位肿瘤组织在正常性激素水平的刺激下所产生的。第二部分鼻咽纤维血管瘤组织中性激素受体和VEGF的表达及其意义目的：众多学者对雄激素及雌激素受体在鼻咽纤维血管瘤(JNA)中的表达进行了研究,但目前为止并没有统一的结论。本研究试图通过高通量的免疫组织芯片技术对性激素受体及血管内皮生长因子(VEGF)的表达及其与肿瘤预后的关系进行研究,并进行RNA水平及蛋白水平的验证。方法：收集70例JNA患者(平均年龄17.0岁,术后病理证实)的肿瘤组织及10例对照组患者的中甲黏膜组织(平均年龄21.5岁),制作成组织芯片。对照组患者为同时段、同年龄段、同性别的鼻中隔、骨瘤及额筛囊肿的患者。应用组织芯片分别对雄激素受体、雌激素受体α、雌激素受体β、孕激素受体及VEGF进行免疫组化染色,分析其在肿瘤组织中的表达与对照组相比是否存在差异,以及几种性激素受体之间、性激素受体与VEGF之间是否存在表达关联性。采集70例JNA患者的临床病理数据,随访无瘤生存时间,随访时间为2-98个月,分析JNA患者的发病年龄、肿瘤分期、术中出血、无瘤生存时间与肿瘤组织中性激素受体、VEGF表达水平之间的关系。采用Real time PCR法检测12例JNA肿瘤组织及4例JNA患者中甲黏膜组织中雄激素受体、雌激素受体α、雌激素受体p、孕激素受体及VEGF mRNA的表达水平,统计分析实验组与对照组之间的差异,以及几种性激素受体之间、性激素受体与VEGF之间是否存在表达关联性。采用Western blot检测6例JNA肿瘤组织及3例JNA患者中甲黏膜组织中雄激素受体、雌激素受体α、雌激素受体β、孕激素受体的表达水平,验证组织芯片及PCR的实验结果。结果：统计结果显示,与中甲黏膜相比,雄激素受体、雌激素受体α、雌激素受体β、孕激素受体及VEGF在JNA肿瘤组织中有较高表达,统计学差异明显,p值小于0.05。雄激素受体、雌激素受体a、雌激素受体β、孕激素受体表达量之间彼此存在正相关,p值小于0.05。雌激素受体α、雌激素受体β与VEGF的表达量呈正相关(p值均为0.006)。雌激素受体α高表达组的无瘤生存时间较短(p值均为0.031),其余性激素受体的高表达虽然提示无瘤生存时间缩短的趋势,但并无明显的统计学差异。Real time PCR及Western blot检测的实验结果与组织芯片一致。结论：JNA肿瘤组织中高表达雄激素受体、雌激素受体α、雌激素受体β、孕激素受体及VEGF。性激素本身存在彼此调节的关系,雌激素受体和血管性生长因子可能参与肿瘤的发生和发展。生存分析显示高表达雌激素受体α提示术后易复发。第三部分鼻咽纤维血管瘤基因表达谱及相关基因研究目的：表达谱芯片是近期较为成熟的分子生物学技术,本研究试图通过表达谱芯片找到参与JNA发病的主要基因和信号通路,分析差异表达基因,为下一步的研究提供前期数据。方法：本研究应用包含30000条人类全长基因的PCR产物为靶基因制成的寡聚核苷酸基因芯片,检测5例高侵袭的JNA组织、4例低侵袭的JNA组织以及5例JNA患者中甲黏膜组织的基因表达谱,分析差异表达基因,旨在通过研究JNA基因表达谱的改变,从基因水平探讨JNA的发病机制。结果：高侵袭组与低侵袭组相比,基因芯片筛选出83条上调表达基因和122条下调表达基因；高侵袭组与对照组相比,基因芯片筛选出1553条上调表达基因和1640条下调表达基因；低侵袭组与对照组相比,基因芯片共筛选出1181条上调表达基因和1239条下调表达基因。其中部分差异表达基因是与性激素结合、细胞凋亡,内皮细胞侵袭转移等密切相关的基因。结论：通过基因表达谱芯片检测及相关分析,发现JNA存在多个基因表达及调控机制的异常,这些差异表达基因为研究JNA的发病机制提供了线索,有望为JNA的基础研究和临床治疗提供新的方向。
[Abstract]:Nasopharyngeal fibroangioma, also known as juvenile nasopharyngeal angiofibroma (JNA), is a relatively low incidence of.JNA in nasopharyngeal benign tumors that only occur in young men. The tumor is very easy to bleed and the operation is difficult to remove. In group learning, JNA has no capsule, from immature endothelial cells and Although fibroblast is a benign tumor, JNA is characterized by local invasion. It is easy to invade the paranasal sinuses, pterygopalatine fossa, infratemporal fossa, wing plate, supraorbital fissure, and even intracranial.
JNA's specific age and sex are very early in the attention of scholars. Many scholars expect to find a clue to study the pathogenesis of JNA. Early studies suggest that the disorder of the pituitary gonadal axis in JNA patients leads to cancer, but subsequent studies have shown that the levels of sex hormones in the JNA patients are normal and have no secondary sex. Recent studies have focused on the relationship between the expression of sex hormone receptors and diseases, but so far no conclusion has been reached.
The serum samples of 40 patients with JNA were collected, and the serum levels of testosterone, free testosterone, estrogen and progestin were detected by ELISA. The serum hormone levels in JNA patients were compared with those in the control group. 70 cases of JNA tumor tissue and 10 cases of middle thyroid tissue were made into tissue chips to detect androgen receptor (Andro). Gen receptor, AR), estrogen receptor alpha (estrogen receptor alpha, ER- alpha), estrogen receptor beta (estrogen receptor A, ER- beta), progesterone receptor (progesterone receptor, PR) and vascular endothelial growth factor (VEGF) in 5 cases of highly invasive tissue, 4 cases of low invasive JNA tissues and 5 normal middle thyroid tissues were studied by gene chip. The key genes and signaling pathways involved in the pathogenesis of JNA were found.
JNA is easy to invade adjacent tissues and even intracranial. It is very easy to bleed in the operation, and the tumor operation with a wide range of invasion is often unresectable and there is a great risk. Drug and target therapy has been the focus of research on the expression of neutral hormone receptor in.JNA tumor tissue and the study of expression spectrum chip to provide a new method for further exploring the treatment of JNA. The foundation.
Part one
Expression of serum sex hormones in patients with nasopharyngeal fibroangioma
Objective: nasopharyngeal angioma (JNA) is a benign tumor that is almost only occurring in young men. But to date, the specific age of pathogenesis of.JNA and sex suggestive hormone may play a role in the process of cancer. The purpose of this study is to determine whether there is abnormal sex hormone levels in puberty and sex hormone levels in JNA patients. It is not associated with tumor staging.
Methods: the serum of 40 patients with JNA (average age 18.3 years, confirmed by postoperative pathology) and 14 normal controls (average age 19.7 years old) were collected. The control group was at the same time, the same age, the same sex other nasal septum, the osteoma and the optic nerve decompression. The serum testosterone in JNA and the control group was detected by Elisa method. The levels of testosterone, estradiol and progesterone were compared between the two groups of sex hormones and whether there were differences in the ratio of male androsteroid. The tumor staging data of 40 patients with JNA were collected to determine whether there was a correlation with the level of serum neutral hormones.
Results: the experimental data showed that there was no significant difference in serum testosterone, free testosterone, estradiol and progesterone in serum, and there was no significant difference in the ratio of testosterone to estradiol in serum and the ratio of estradiol and estradiol in serum, and there was no significant correlation between the tumor staging and the level of sex hormone in serum of patients with JNA.
Conclusion: Although JNA is a benign tumor of the nasopharynx of adolescent male, there is no abnormal sex hormone level and the ratio of sex hormone in the circulating serum, and there is no correlation between the level of sex hormone and tumor staging. This indicates that the level of sex hormone in patients with JNA is normal and may not have the loss of the pituitary gonadal axis. The occurrence of tumor may be due to the stimulation of normal tumor hormones at normal sex hormone levels.
The second part
Expression and significance of sex hormone receptor and VEGF in nasopharyngeal fibroangioma tissue
Objective: many scholars have studied the expression of androgen and estrogen receptor in nasopharyngeal angioma (JNA), but so far there is no unified conclusion. This study attempts to express the expression of sex hormone receptor and vascular endothelial growth factor (VEGF) by high throughput immuno tissue microarray and its relationship with the prognosis of the tumor. Conduct research and verify the RNA level and protein level.
Methods: 70 cases of JNA patients (average age 17 years old, pathologically confirmed) tumor tissue and 10 cases of control group (mean age 21.5 years old) were made into tissue chips. The control group was the same age group, the same age group, the same sex other nasal septum, the osteoma and the frontal sieving cyst. The hormone receptor, estrogen receptor alpha, estrogen receptor beta, progesterone receptor and VEGF were immunohistochemical staining. The difference between the expression of the tumor tissue and the control group was analyzed, and the correlation between the sex hormone receptor and the VEGF was found. The clinicopathological data of 70 patients with JNA were collected. No tumor survival time was followed up for 2-98 months. The relationship between the onset age of JNA, tumor stage, intraoperative hemorrhage, tumor free survival time and the level of VEGF expression of neutrophilic hormone receptor and tumor tissue were analyzed.
The Real time PCR method was used to detect the expression of androgen receptor, estrogen receptor alpha, estrogen receptor P, progesterone receptor and VEGF mRNA in 12 JNA tumor tissues and 4 JNA patients. The difference between the experimental group and the control group was statistically analyzed, and there were several sex hormone receptors between the sex hormone receptors and VEGF. In the expression of relevance.
The expression levels of androgen receptor, estrogen receptor alpha, estrogen receptor beta and progesterone receptor were detected by Western blot in 6 cases of JNA tumor tissue and 3 cases of JNA patients. The experimental results of tissue microarray and PCR were verified.
Results: the results showed that androgen receptor, estrogen receptor alpha, estrogen receptor beta, progesterone receptor and VEGF were highly expressed in JNA tumor tissues compared with middle thyroid mucosa, and the statistical difference was obvious, P was less than 0.05. androgen receptor, estrogen receptor A, estrin receptor beta, and progesterone receptor expression existed in positive phase. The p value was less than 0.05. estrogen receptor alpha, the expression of estrogen receptor beta was positively correlated with the expression of VEGF (P value was 0.006). The tumor free survival time of the estrogen receptor alpha high expression group was shorter (P value was 0.031), while the high expression of the other sex hormone receptors indicated the trend of shortening the tumor free survival time, but there was no significant statistical difference of.Real time. The results of PCR and Western blot test were consistent with tissue microarray.
Conclusion: the high expression of androgen receptor, estrogen receptor alpha, estrogen receptor beta, progesterone receptor and VEGF. sex hormone itself are related to each other in JNA tumor tissue. Estrogen receptor and vascular growth factor may participate in the occurrence and development of tumor. Survival analysis shows that high expression of estrogen receptor alpha suggests recurrence after operation.
The third part
Study on gene expression profile and related genes of nasopharyngeal fibroangioma
Objective: the expression chip is a relatively mature molecular biology technique in the near future. This study attempts to find the main genes and signal pathways involved in the pathogenesis of JNA by expressing the spectral chip, and analyze the differentially expressed genes, and provide the preliminary data for the next step of research.
Methods: in this study, the oligonucleotide microarray of the PCR products containing 30000 human full-length genes was used as the target gene. The gene expression profiles of 5 highly invasive JNA tissues, 4 low invasive JNA tissues and 5 JNA patients were detected and the differentially expressed genes were analyzed. The aim of this study was to study the modification of JNA gene expression profiles. Change, explore the pathogenesis of JNA from the gene level.
Results: compared with the low invasion group, 83 up-regulated genes and 122 down regulated genes were screened by gene chip compared with the low invasion group. Compared with the control group, the high invasion group screened 1553 up-regulated genes and 1640 down regulated genes, and the low invasion group screened 1181 up-regulated genes compared with the control group. Genes and 1239 down regulated expression genes, some of which are differentially expressed genes, are closely related to sex hormone binding, apoptosis, endothelial cell invasion and metastasis.
Conclusion: through the detection and correlation analysis of gene expression spectrum, it is found that there are many abnormal genes expression and regulation mechanism in JNA. These differentially expressed genes provide clues for the study of the pathogenesis of JNA. It is expected to provide a new direction for the basic and clinical treatment of JNA.
【学位授予单位】：复旦大学
【学位级别】：博士
【学位授予年份】：2013
【分类号】：R739.63

【共引文献】