Nrf2和HO-1在肾癌组织中的表达及临床意义

发布时间：2018-07-03 02:24

本文选题：肾癌 + Nrf2　；参考：《河北医科大学》2014年硕士论文

【摘要】：目的：肾癌是泌尿系肿瘤中常见的肿瘤，，70%-80%为透明细胞癌,近年发病有增加趋势。寻找肾癌的新靶点对于肾癌的早期发现及预后并制定治疗方案有着非常重要的意义。在肿瘤形成和细胞增殖过程中，氧化应激及炎症反应与之密切相关，已成为国内外学者的研究热点，与氧化应激相关途径的有关基因有可能成为潜在的药物治疗肿瘤的作用靶点。目前研究发现很多药物是通过调节Nrf2/HO-1信号通路来发挥抗氧化及抗炎作用的。为探讨Nrf2和HO-1与肾癌发生、发展的关系，本实验通过免疫组化SABC和RT-PCR法检测Nrf2和HO-1在肾癌和癌旁正常组织中的表达情况，探讨Nrf2和HO-1在肾癌发生、发展中的作用。方法：选取河北医科大学第二医院外科手术切除的肾癌组织标本51例。选其中23例患者中的癌旁组织（经病理证实为正常肾组织）为对照组。所有病例术前均未进行过放疗和（或）化疗。所有标本分为两份，一份经4%甲醛溶液固定，石蜡块包埋，另一份保存于液氮之中。免疫组织化学方法检测：应用免疫组化技术，从蛋白水平检测石蜡包埋组织中Nrf2和HO-1的表达，其中肾癌组织标本51例，正常肾组织标本23例。采用双评分半定量法来进行评分，综合统计染色阳性率及染色强度，运用SPSS17.0统计软件处理数据，比较肾癌组织和正常肾组织中Nrf2及HO-1的阳性表达率之间的差异。采用χ2检验，把具有统计学意义显著性差异的临界值定为P0.05。Nrf2与HO-1关系采用Pearson相关分析。 RT-PCR方法检测：根据Trizol试剂盒说明提取冰冻组织中的总RNA，加入RT反应体系管，进行逆转录反应合成cDNA，设立β-actin基因片段为Nrf2和HO-1基因表达的内参照，在RT-PCR仪上进行扩增，取每个标本的扩增产物行琼脂糖凝胶电泳，以DNA Marker（DL2000）作为标准片段标记，电泳后用紫外透射仪观察，应用Quantity One凝胶图象分析软件对目的电泳条带进行分析，以β-actin电泳条带作为参照，结果以两者吸光度的比值表示。两组间吸光度比值采用t检验进行统计分析。P0.05认为差异有统计学意义。结果： 1免疫组织化学染色示：Nrf2主要表达于肿瘤细胞的细胞浆中，在肾癌组织中的阳性表达率为82.4%，在癌旁正常组织中表达的阳性率为34.8%，两者差异有统计学意义（P=0.00，P0.05）。HO-1主要位于肿瘤细胞的胞浆中，其在肾癌组织中的阳性表达率为76.5%，在癌旁正常组织中的阳性表达率为39.1%，二者阳性表达率差异有统计学意义（P=0.02，P0.05）。Nrf2和HO-1蛋白的表达呈正相关（r=0.515P0.05）。 2RT-PCR结果显示：Nrf2在肾癌和癌旁正常组织的表达量分别为0.747±0.101和0.374±0.071，二者差异有统计学意义（P0.05）。HO-1在肾癌和癌旁正常组织的表达量分别为0.853±0.078和0.410±0.085，二者差异有统计学意义（P0.05）。结论： 1Nrf2和HO-1蛋白在肾癌组织中的表达高于癌旁正常组织中的表达。 2Nrf2、HO-1蛋白在肾癌组织中的表达呈正相关。 3Nrf2、HO-1蛋白可能参与了肾癌的发生、发展，并在演变过程中起到了重要作用。
[Abstract]:Objective: renal cancer is a common tumor in urological tumors. 70%-80% is a clear cell carcinoma. It has an increasing trend in recent years. The new target for searching for renal cancer is of great significance for the early detection and prognosis of renal cancer and the formulation of treatment scheme. In the process of tumor formation and cell proliferation, oxidative stress and inflammatory reaction are closely related. The relevant genes related to oxidative stress may become potential targets for the potential drug treatment of tumor. Many drugs have been found to play antioxidation and anti-inflammatory effects by regulating the Nrf2/HO-1 signaling pathway. To explore the relationship between the development of Nrf2 and HO-1 and the development of renal carcinoma, In this experiment, the expression of Nrf2 and HO-1 in renal carcinoma and adjacent normal tissues was detected by immunohistochemical SABC and RT-PCR method, and the role of Nrf2 and HO-1 in the development of renal carcinoma was discussed.
Methods: 51 cases of renal carcinoma tissue excised by surgical operation in Second Hospital of Hebei Medical University were selected and 23 of them were selected as control group. All cases were not treated with radiotherapy and / or chemotherapy before operation. All the specimens were divided into two parts, one with 4% Formaldehyde Solution and paraffin paraffin. Buried, the other in liquid nitrogen.
Immunohistochemical method detection: using immunohistochemical technique, the expression of Nrf2 and HO-1 in paraffin embedded tissue was detected from protein level, including 51 cases of renal carcinoma tissue and 23 normal renal tissue specimens. The double score and half quantitative method was used to score, and the positive rate and intensity of dyeing were statistically analyzed, and the number of SPSS17.0 statistical software was used. The difference between the positive expression rates of Nrf2 and HO-1 in renal carcinoma and normal renal tissue was compared. The critical value of statistically significant difference was determined to be the relationship between P0.05.Nrf2 and HO-1 by Pearson correlation analysis by the x 2 test.
RT-PCR method detection: according to the Trizol kit, the total RNA in the frozen tissue was extracted, and the RT reaction system tube was added, and cDNA was synthesized by reverse transcriptase. The gene fragment of the beta -actin was set up as the internal reference for the expression of Nrf2 and HO-1 gene, and the amplified product of each specimen was amplified by the agarose gel electrophoresis with DNA Marker (DL). 2000) as a standard fragment mark, after electrophoretic ultraviolet transmittance, Quantity One gel image analysis software was used to analyze the target bands. The ratio of the absorbance between the two groups was expressed with the ratio of the absorbance between the two groups. The two groups of absorbance ratio using t test were statistically analyzed and.P0.05 thought the difference was statistically significant. Learning meaning.
Result:
1 immunohistochemical staining showed that Nrf2 was mainly expressed in the cytoplasm of tumor cells, the positive expression rate in renal carcinoma tissue was 82.4%, and the positive rate was 34.8% in normal tissues adjacent to the cancer. The difference was statistically significant (P=0.00, P0.05).HO-1 was mainly located in the cytoplasm of tumor cells, and the positive expression rate in the renal carcinoma tissue For 76.5%, the positive expression rate in normal tissues adjacent to cancer was 39.1%, and the difference of positive expression rate between the two groups was statistically significant (P=0.02, P0.05) and the expression of.Nrf2 and HO-1 protein was positively correlated (r=0.515P0.05).
The results of 2RT-PCR showed that the expression of Nrf2 in renal carcinoma and adjacent normal tissues was 0.747 + 0.101 and 0.374 + 0.071 respectively. The difference between the two groups was statistically significant (P0.05).HO-1 was 0.853 + 0.078 and 0.410 + 0.085 in the normal tissues of renal carcinoma and adjacent to cancer, and the two differences were statistically significant (P0.05).
Conclusion:
The expression of 1Nrf2 and HO-1 protein in renal cell carcinoma was higher than that in adjacent normal tissues.
The expression of 2Nrf2 and HO-1 protein was positively correlated with renal cell carcinoma.
3Nrf2 and HO-1 proteins may play an important role in the development and progression of renal cell carcinoma.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R737.11

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