本文選題：免疫球蛋白M + 鼻咽癌　； 參考：《瀘州醫(yī)學(xué)院》2011年碩士論文

【摘要】：目的：盡管鼻咽癌的診斷和治療方法都在不斷進(jìn)步,但對(duì)其早期診斷與預(yù)后判斷還缺乏有效的手段,患者的5年生存率還有待進(jìn)一步提高；因而我們需要在鼻咽癌發(fā)病機(jī)制理論研究領(lǐng)域取得突破性進(jìn)展,尋找新的靶點(diǎn),以提高其早期診斷水平和整體治療效果。本實(shí)驗(yàn)通過(guò)研究抗人IgM抗體對(duì)鼻咽癌HNE-1細(xì)胞裸鼠移植瘤生長(zhǎng)、腫瘤細(xì)胞凋亡以及免疫球蛋白M(IgM)、糖蛋白gp96 (Glycoprotein96, gp96)表達(dá)的影響,以期探討IgM在鼻咽癌中異位表達(dá)的生物學(xué)活性以及抗人IgM抗體影響裸鼠成瘤的可能機(jī)制。方法：對(duì)鼻咽癌HNE-1細(xì)胞進(jìn)行傳代培養(yǎng),制成細(xì)胞懸液,接種于Balb/c-nu/nu裸鼠右側(cè)背腹部皮下,建立鼻咽癌移植瘤模型；15只荷瘤裸鼠隨機(jī)分為實(shí)驗(yàn)組(腹腔注射抗人IgM抗體,1.5mg/裸鼠,分5次,每隔3天1次)、IgG對(duì)照組(腹腔注射IgG抗體)及PBS對(duì)照組(腹腔注射PBS),同時(shí)觀察裸鼠移植瘤生長(zhǎng)情況,定期(每四天)稱(chēng)量裸鼠體重并以游標(biāo)卡尺測(cè)量移植瘤的長(zhǎng)徑及短徑,繪制腫瘤生長(zhǎng)曲線,計(jì)算腫瘤體積=腫瘤長(zhǎng)徑×腫瘤短徑2/2。5次腹腔給藥后觀察1周,處死裸鼠,取瘤體,稱(chēng)量移植瘤重量,計(jì)算抑瘤率。常規(guī)HE染色觀察移植瘤光鏡下改變,并取裸鼠主要內(nèi)臟行HE染色。采用TUNEL技術(shù)檢測(cè)鼻咽癌HNE-1細(xì)胞裸鼠移植瘤組織中細(xì)胞凋亡情況,采用圖像分析系統(tǒng)計(jì)算凋亡指數(shù)；同時(shí)利用免疫組化SP法檢測(cè)裸鼠移植瘤組織中IgM、gp96蛋白的表達(dá)情況,同樣采用圖像分析系統(tǒng)作平均光密度值測(cè)定。結(jié)果：(1)成功構(gòu)建出人鼻咽癌裸鼠移植瘤模型,三組裸鼠均有鼻咽癌原位移植瘤生長(zhǎng),成瘤率100%；其瘤組織經(jīng)病理切片檢查證實(shí)為鼻咽癌,且在實(shí)驗(yàn)期內(nèi),三組動(dòng)物全部存活。(2)三組裸鼠飲食正常,活動(dòng)自如,精神狀態(tài)良好,無(wú)1只死亡；其心、肝、脾、肺、腎組織形態(tài)觀察無(wú)明顯差異。(3)抗人IgM抗體干預(yù)組裸鼠移植瘤體積為(26.73±16.51)mm3,較IgG對(duì)照組(204.97±151.88)mm3和PBS對(duì)照組(230.16±167.78)mm3明顯減小,三組間差異有統(tǒng)計(jì)學(xué)意義(P0.05)；抗人IgM抗體干預(yù)組裸鼠體重為(21.10±2.35)g,而IgG對(duì)照組為(20.70±3.29)g, PBS對(duì)照組(20.01±2.22)g,三組間差異無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)；抗人IgM抗體干預(yù)組裸鼠移植瘤重量(0.09±0.07)g,較IgG對(duì)照組(0.35±0.10)g和PBS對(duì)照組(0.38±0.08)g明顯減小,經(jīng)統(tǒng)計(jì)學(xué)分析差異有統(tǒng)計(jì)學(xué)意義(P0.05)；同時(shí)計(jì)算抑瘤率分別為74.45%和76.84%。(4)TUNEL法檢測(cè)鼻咽癌HNE-1細(xì)胞裸鼠移植瘤組織中細(xì)胞凋亡水平,抗人IgM抗體干預(yù)組腫瘤細(xì)胞平均凋亡指數(shù)為(1.50±0.76)%,較IgG對(duì)照組(0.49±0.15)%和PBS對(duì)照組(0.42±0.20)%明顯升高,三組間差異經(jīng)統(tǒng)計(jì)學(xué)分析有統(tǒng)計(jì)學(xué)意義(P0.05)；(5)免疫組織化學(xué)SP法檢測(cè)裸鼠移植瘤組織中IgM、gp96蛋白表達(dá),抗人IgM抗體干預(yù)組IgM蛋白表達(dá)的平均光密度值為(0.01±0.01),與IgG對(duì)照組(0.06±0.03)和PBS對(duì)照組(0.05±0.03)相比均顯著下降,三組間差異有統(tǒng)計(jì)學(xué)意義(P0.05)；抗人IgM抗體干預(yù)組gp96蛋白表達(dá)顯著下降(0.05±0.01),經(jīng)統(tǒng)計(jì)學(xué)分析與IgG對(duì)照組(0.10±0.03)和PBS對(duì)照組(0.11±0.02)比較差異有統(tǒng)計(jì)學(xué)意義(P0.05)。結(jié)論：(1)抗人IgM抗體能顯著抑制人鼻咽癌HNE-1細(xì)胞裸鼠移植瘤生長(zhǎng),提示鼻咽癌異位表達(dá)的IgM具有生長(zhǎng)因子樣作用。(2)抗人IgM抗體能顯著抑制IgM蛋白表達(dá),通過(guò)阻斷其生物學(xué)活性來(lái)抑制腫瘤生長(zhǎng)。(3)抗人IgM抗體可抑制gp96蛋白表達(dá),推測(cè)其機(jī)制可能與抑制IgM表達(dá)有關(guān)。(4)抗人IgM抗體能促進(jìn)裸鼠移植瘤中鼻咽癌HNE-1細(xì)胞凋亡,其機(jī)制可能與抑制IgM、gp96蛋白表達(dá)有關(guān)。
[Abstract]:Objective: Although the diagnosis and treatment methods of nasopharyngeal carcinoma are constantly improving, the early diagnosis and prognosis are still lack of effective means. The 5 year survival rate of the patients still needs to be further improved. Therefore, we need to make a breakthrough in the theoretical research field of nasopharyngeal cancer pathogenesis and find new targets in order to improve their early diagnosis. The effect of anti human IgM antibody on the growth of xenografts in nude mice of nasopharyngeal carcinoma HNE-1 cells, the apoptosis of tumor cells and the expression of immunoglobulin M (IgM) and glycoprotein gp96 (Glycoprotein96, gp96) in nude mice of nasopharyngeal carcinoma, in order to explore the biological activity of ectopic expression of IgM in nasopharyngeal carcinoma and the effect of anti human IgM antibody on the effect of anti human anti human antibody on the growth of nude mice in nasopharyngeal carcinoma cells. Methods: the possible mechanism of tumor formation in nude mice. Methods: the HNE-1 cells of nasopharyngeal carcinoma were cultured and made into cell suspension and inoculated into the right dorsal abdomen of Balb/c-nu/nu nude mice to establish the model of nasopharyngeal carcinoma transplantation tumor. 15 tumor bearing nude mice were randomly divided into experimental group (intraperitoneal injection of anti human IgM antibody, 1.5mg/ nude mice, 5 times every 3 days), IgG control group (1 times every 3 days). Intraperitoneal injection of IgG antibody and PBS control group (intraperitoneal injection of PBS), and observing the growth of transplanted tumor in nude mice, weighing the body weight of nude mice regularly (every four days) and measuring the length and short diameter of the transplanted tumor with vernier caliper, plotting the tumor growth curve, and calculating the tumor volume = the tumor length and the short diameter of the tumor for 1 weeks after the intraperitoneal administration, and killing nude mice. The tumor body was taken and weighed and the tumor suppressor rate was calculated. Conventional HE staining was used to observe the changes of the transplanted tumor and the main viscera of nude mice was stained with HE. The apoptosis in the transplanted tumor tissues of nude mice of nasopharyngeal carcinoma HNE-1 cells was detected by TUNEL technique, and the apoptosis index was calculated by the image analysis system, and the nude was detected by the immunohistochemical SP method. The expression of IgM and gp96 protein in the transplanted tumor tissues of the mice was also measured by the mean optical density value of the image analysis system. Results: (1) the transplanted tumor model of human nasopharyngeal carcinoma was successfully constructed. The three groups of nude mice had the growth of nasopharyngeal carcinoma in situ and the tumor rate was 100%. The tumor tissue was confirmed by pathological section to be nasopharyngeal carcinoma and the experiment was in the experiment. During the period, all the three groups of animals survived. (2) the three groups of nude mice had normal diet, good activity, good mental state, no 1 death, and the heart, liver, spleen, lung and kidney tissue were not significantly different. (3) the volume of the transplanted tumor in the anti human IgM antibody group was (26.73 + 16.51) mm3, compared with the IgG control group (204.97 + 151.88) mm3 and PBS control group (230.16 + 167.78) M The difference between the three groups was statistically significant (P0.05), and the body weight of nude mice in the anti human IgM antibody group was (21.10 + 2.35) g, while the IgG control group was (20.70 + 3.29) g, PBS control group (20.01 + 2.22) g, and there was no statistical difference (P0.05), and the weight of nude mice in the anti human IgM antibody group was (0.09 + 0.07) g, compared with IgG control group (0.35 + 0.). 10) g and PBS control group (0.38 + 0.08) g decreased significantly, and statistically significant (P0.05). At the same time, the tumor suppressor rate was 74.45% and 76.84%. (4) TUNEL method was used to detect the cell apoptosis level in nude mice of nasopharyngeal carcinoma HNE-1 cells. The average apoptosis index of tumor cells in anti human IgM anti body intervention group was (1.50 + 0.76)%, and was more than Ig. G control group (0.49 + 0.15)% and PBS control group (0.42 + 0.20)% increased significantly, the difference between the three groups was statistically significant (P0.05); (5) immunohistochemical SP method was used to detect the expression of IgM, gp96 protein in the transplanted tumor tissues of nude mice, and the average optical density of IgM egg white expression in the anti human IgM antibody group was (0.01 + 0.01), and that of the IgG control group was 0.06 (0.06). Compared with the PBS control group (0.05 + 0.03), the difference between the three groups was statistically significant (P0.05), and the expression of gp96 protein in the anti human IgM antibody group decreased significantly (0.05 + 0.01). The difference between the IgG control group (0.10 + 0.03) and the PBS control group (0.11 + 0.02) was statistically significant (P0.05). Conclusion: (1) anti human IgM resistance (P0.05). Physical ability significantly inhibited the growth of human nasopharyngeal carcinoma HNE-1 cells in nude mice, suggesting that the ectopic expression of IgM in nasopharyngeal carcinoma has a growth factor like effect. (2) anti human IgM antibodies can significantly inhibit the expression of IgM protein and inhibit the growth of tumor by blocking its biological activity. (3) anti human IgM antibody can inhibit the expression of gp96 protein, presumably its mechanism may be inhibited. IgM expression. (4) anti human IgM antibody can promote the apoptosis of nasopharyngeal carcinoma HNE-1 cells in nude mice. The mechanism may be related to the inhibition of IgM and gp96 protein expression.
【學(xué)位授予單位】：瀘州醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2011
【分類(lèi)號(hào)】：R739.63

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