sIL-1RAP在低级别胶质瘤中的作用机制研究

发布时间：2018-06-18 14:42

本文选题：IL-1RAP + sIL-1RAP　；参考：《河北联合大学》2014年硕士论文

【摘要】：目的神经胶质瘤（glioma）在人类中枢神经系统的肿瘤中较为多见。其中，脑肿瘤中的40%～60%具有广泛的侵袭性以及由低级别向高级别转化的趋势。相关研究显示，低级别胶质瘤（low grade gliomas, LGGs）占全部中枢神经系统肿瘤的33%，在成人中枢神经系统肿瘤中LGGs占25%，，在儿童中枢神经系统肿瘤中LGGs占35%。研究表明，病理级别相同，且生长部位也相同（脑干除外）的低级别胶质瘤病人，成人的临床进展和预后却远不及儿童。成年患者低级别星形胶质瘤（除毛细胞型星形细胞瘤外）往往更容易恶变为恶性程度更高的胶质母细胞瘤，相比之下，儿童发生恶变的机率较低，其预后也较好。儿童与成人低级别胶质瘤预后的不同，提示二者的生物学特性不同。我们已有的研究表明sIL-1RAP在儿童和成人低级别胶质瘤中的明显差异表达，且与STAT3共定位于U251细胞的细胞核，但已有的相关研究表明该蛋白并非定位于细胞核，那么研究STAT3与sIL-1RAP的细胞核共定位机制,将有助于我们揭示sIL-1RAP在儿童和成人低级别胶质瘤预后差异的分子机制。方法（1）提取收集的儿童和成人低级别胶质瘤组织的总RNA，并通过RT-PCR及qRT-PCR的方法进一步验证先前的实验结果。（2）同时，利用Western Blot方法检测儿童和成人低级别胶质瘤中sIL-1RAP的表达情况，进一步验证上述（1）的结果。（3）利用细胞转染的方法，将STAT3的siRNA转染胶质瘤细胞系U251，用Western Blot方法检测STAT3在胶质瘤细胞系U251中的表达。（4）利用细胞转染的方法，将STAT3的siRNA转染胶质瘤细胞系U251，在RNA的水平，通过RT-PCR及qRT-PCR的方法检测STAT3的siRNA对胶质瘤细胞系U251中STAT3的抑制程度。（5）培养U251细胞，同时转染STAT-3的siRNA和sIL-1RAP的质粒，利用免疫荧光的方法检测抑制STAT-3的表达对sIL-1RAP进入细胞核的抑制情况。结果本研究在RNA干扰实验的同时也操作免疫荧光实验，验证STAT3对sIL-1RAP的入核的影响，但抑制STAT3后未能影响sIL-1RAP入核。结论未能影响sIL-1RAP入核的原因可能是抑制STAT3的程度不够或者除外STAT3有其他代偿机制协助sIL-1RAP入核，sIL-1RAP的入核对胶质瘤的发生、发展等具有显著影响，所以抑制sIL-1RAP的入核可以作为一种潜在药物靶点用于治疗神经胶质瘤，我们将继续深入研究抑制sIL-1RAP入核的方法。
[Abstract]:Objective glioma is more common in human central nervous system (CNS) tumors. Among them, 40% of brain tumors have extensive invasiveness and the trend from low grade to high grade. Low grade gliomas (LGGs) account for 33% of all central nervous system tumors, 25% of adult central nervous system tumors and 35% of children's central nervous system tumors. Studies have shown that patients with lower grade gliomas with the same pathological grade and the same growth site (except brainstem) have far less clinical progress and prognosis than children. Adult patients with low-grade astrocytoma (other than hairy astrocytoma) are more likely to become malignant glioblastoma with higher degree of malignancy. In contrast, children have a lower incidence of malignancy and better prognosis. The prognosis of low grade gliomas in children and adults is different, suggesting that their biological characteristics are different. Our previous studies have shown that sIL-1RAP is significantly differentially expressed in low-grade gliomas in children and adults, and co-located with STAT3 in the nucleus of U251 cells, but related studies have shown that the protein is not located in the nucleus. Therefore, studying the nuclear colocalization mechanism of STAT3 and sIL-1RAP will be helpful to reveal the molecular mechanism of the prognosis difference of sIL-1RAP in children and adults. Methods 1) Total RNAs were extracted from low grade gliomas in children and adults, and the results were further verified by RT-PCR and qRT-PCR. Meanwhile, the expression of sIL-1RAP in children and adults was detected by Western blot. To further verify the results of the above 1), we used cell transfection method to transfect STAT3 siRNA into glioma cell line U251. Western Blot was used to detect STAT3 expression in glioma cell line U251. STAT3 siRNA was transfected into U251 glioma cell line U251. At the level of RNA, the inhibition of STAT3 siRNA on U251 cell line U251 was detected by RT-PCR and qRT-PCR. U251 cells were transfected with STAT-3 siRNA and sIL-1RAP plasmid at the same time. Immunofluorescence was used to detect the inhibition of the expression of STAT-3 on the entry of sIL-1RAP into the nucleus. Results the effect of STAT3 on the nucleation of sIL-1RAP was verified by immunofluorescence assay as well as RNA interference, but no effect of STAT3 on the entry of sIL-1RAP was found after inhibition of STAT3. Conclusion the reason why sIL-1RAP does not enter into the nucleus may be that the degree of inhibition of STAT3 is not enough or that STAT3 has other compensatory mechanisms to assist in the development and development of glioma of sIL-1RAP into the nucleus of sIL-1RAP. Therefore, inhibiting the entry of sIL-1RAP can be used as a potential drug target for the treatment of glioma. We will continue to study the methods of inhibiting the entry of sIL-1RAP.
【学位授予单位】：河北联合大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R739.41

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