本文關(guān)鍵詞： 鼻咽癌 清鼻方 CNE細(xì)胞 免疫力低下　出處：《廣州中醫(yī)藥大學(xué)》2012年博士論文　論文類型：學(xué)位論文

【摘要】：鼻咽癌是我國常見的惡性腫瘤之一,在頭頸部腫瘤中占首位,發(fā)病率有明顯的地區(qū)分布。我國及東南亞地區(qū)較多見,其中廣東中西部發(fā)病率最高。近年來NPC發(fā)病率逐年攀升,因其具體發(fā)病機(jī)制尚不明確,聯(lián)合化療近期效果顯著,但毒副反應(yīng)嚴(yán)重,容易導(dǎo)致不同程度的后遺癥,嚴(yán)重影響患者的生活質(zhì)量及預(yù)后。近年來中醫(yī)藥治療鼻咽癌的優(yōu)勢備受國內(nèi)外關(guān)注,已有實(shí)驗(yàn)和臨床資料表明,中醫(yī)藥在提高腫瘤病人生活質(zhì)量,延長生存時(shí)間,降低腫瘤復(fù)發(fā)和轉(zhuǎn)移等方面具有獨(dú)特的優(yōu)勢。 “清鼻方”是陳銳深教授臨床治療鼻咽癌的臨床經(jīng)驗(yàn)方,廣泛用于晚期鼻咽癌的治療及鼻咽癌放療的輔助治療,在臨床使用多年,具有良好的臨床療效。 本課題將探討“清鼻方”對(duì)免疫力低下小鼠非特異性免疫功能的影響及通過體外實(shí)驗(yàn)探討“清鼻方”對(duì)CNE細(xì)胞增殖的影響,旨在從不同層面、不同靶點(diǎn)探討“清鼻方”對(duì)鼻咽癌的作用機(jī)理,為臨床使用提供科學(xué)的依據(jù)。 目的： 在建立免疫力低下模型小鼠基礎(chǔ)上,應(yīng)用清鼻方干預(yù)模型小鼠,研究清鼻方在增強(qiáng)機(jī)體免疫力的作用機(jī)理；在己分離出的鼻咽癌細(xì)胞株(CNE細(xì)胞)及血清藥理學(xué)的基礎(chǔ)上,通過細(xì)胞生物學(xué)、分子生物學(xué)技術(shù),探討清鼻方對(duì)CNE細(xì)胞的抑制作用。 方法： (1)清鼻方對(duì)環(huán)磷酰胺誘導(dǎo)所致免疫力低下小鼠非特異性免疫功能的影響取健康雌雄各半的BALB/c種小白鼠70只,按隨機(jī)區(qū)組法分為5組：模型對(duì)照組、清鼻方高劑量組、清鼻方低劑量組、干擾素陽性藥對(duì)照藥組及正常對(duì)照組。除正常對(duì)照組外,其余各組皮下注射環(huán)磷酰胺,按80mg/kg體重給藥,連續(xù)給藥3d造模。正常對(duì)照組、模型對(duì)照組、清鼻方高劑量組、清鼻方低劑量組均采取每天灌胃給藥,干擾素組則采用隔天肌注給藥,試驗(yàn)周期均為30天。以小鼠體重、脾臟稱重、脾臟懸液細(xì)胞計(jì)數(shù)、血清IgA、IgG、IgM含量、T細(xì)胞亞群CD3/4/8表達(dá)水平進(jìn)行評(píng)價(jià)。(2)清鼻方治療效應(yīng)對(duì)大鼠鼻咽上皮細(xì)胞增殖的影響研究制備中藥復(fù)方清鼻方濃縮煎液。取健康雌雄各半SD大鼠12只,隨機(jī)分為模型組6只和實(shí)驗(yàn)組6只。模型組采用生理鹽水灌胃,而實(shí)驗(yàn)組采用中藥清鼻方濃縮煎液灌胃,處理時(shí)長為5天。在大鼠腹主動(dòng)脈取血,離心取上清制成含藥血清直接作用于CNE細(xì)胞,并采用MTT法、流式細(xì)胞術(shù),電子顯微鏡觀察細(xì)胞生長曲線來研究清鼻方分別作用于CNE細(xì)胞后對(duì)腫瘤細(xì)胞生長的抑制情況,以及清鼻方誘導(dǎo)腫瘤細(xì)胞發(fā)生凋亡的情況。 結(jié)果： (1)與空白組比較,各組小鼠體重、脾質(zhì)量、脾臟指數(shù)、脾細(xì)胞總數(shù)均明顯下降(P0.01),其中模型組各數(shù)值均最低。骨髓有核細(xì)胞數(shù)各組與空白組比較,差異無統(tǒng)計(jì)學(xué)意義(P0.05)。模型組IgA水平明顯降低,干擾素組、低劑量組和高劑量組IgA水平明顯升高(P0.01); IgG水平方面,模型組、干擾素組均明顯降低(P0.01),清鼻方低劑量組和高劑量組變化均不明顯(P0.05)；模型組、干擾素組、低劑量組IgM水平均升高,高劑量組反而降低,但均無統(tǒng)計(jì)學(xué)差異(P0.05)。(2)與模型組比較,各用藥組骨髓有核細(xì)胞數(shù)變化不明顯(P0.05)。低劑量組在體重、脾質(zhì)量、脾臟指數(shù)、脾細(xì)胞總數(shù)方面與之比較,兩者均無統(tǒng)計(jì)學(xué)意義(P0.05)；高劑量組在小鼠體重、脾質(zhì)量、脾臟指數(shù)方面明顯高于模型組(P0.01)；干擾素組體重、脾質(zhì)量方面與模型組對(duì)比,差異均明顯具有統(tǒng)計(jì)學(xué)意義(P0.01)。干擾素組、低劑量組和高劑量組IgA水平均明顯升高(P0.01)；干擾素組IgG變化不明顯,清鼻方低劑量組和高劑量組IgG水平俊明顯升高(P0.05)；干擾素組和低劑量組均無統(tǒng)計(jì)學(xué)差異(P0.05),高劑量組明顯低于模型組,兩者相比差異有統(tǒng)計(jì)學(xué)意義(P0.05)。(3)小鼠外周血CD3、CD4、CD8的整體表達(dá)水平相對(duì)脾臟較低。小鼠造模后外周血CD3、 CD4、CD8表達(dá)水平無明顯下降,而脾臟CD3、CD4、CD8表達(dá)水平則明顯下降；給予干擾素和清鼻方低高劑量后CD3、CD4、CD8表達(dá)水平均有一定程度的提升,其中干擾素組接近正常組水平,清鼻方低、高劑量組則均未達(dá)到正常組水平。(4)MTT法比色結(jié)果顯示,在作用24h后,不同濃度清鼻方含藥血清對(duì)CNE細(xì)胞的抑制作用不明顯。作用48h后,血清濃度5%、10%、15%對(duì)CNE細(xì)胞的抑制作用明顯；作用72h后,血清濃度5%、10%、對(duì)CNE細(xì)胞的抑制作用明顯。(5)CNE細(xì)胞的增殖曲線結(jié)果顯示：未滅活的正常大鼠血清或含藥血清(10%和15%)均具有具有抑制細(xì)胞增殖的效果。其中15%未滅活含藥血清較10%未滅活含藥血清具有更強(qiáng)的抑制CNE細(xì)胞增殖的作用。滅活的正常大鼠血清或含藥血清(10%和15%)也出現(xiàn)類似的情況,但整體效應(yīng)低于未滅活血清組。(6)細(xì)胞凋亡率方面,用10%含藥血清和正常大鼠血清處理CNE細(xì)胞后,正常大鼠血清出現(xiàn)凋亡細(xì)胞(23%),含藥血清出現(xiàn)更高的凋亡率31.0%；用15%含藥血清和正常大鼠血清處理CNE細(xì)胞后,正常大鼠血清也出現(xiàn)凋亡細(xì)胞(25.3%),而15%含藥血清凋亡率達(dá)33.9%。 結(jié)論： 清鼻方能減輕化療藥物所致的免疫抑制狀態(tài),調(diào)節(jié)機(jī)體的細(xì)胞免疫與體液免疫,使紊亂免疫機(jī)能趨于正常,這將于鼻咽癌患者恢復(fù)；清鼻方含藥血清能抑制CNE細(xì)胞的增殖,誘導(dǎo)細(xì)胞凋亡,這可能是其重要作用機(jī)理。
[Abstract]:Nasopharyngeal carcinoma is one of the most common malignant tumors in China, occupies the first place in head and neck cancer incidence has obvious district distribution. Many in China and Southeast Asia, the western Guangdong highest incidence rate. The incidence of NPC increased year by year in recent years, because of its specific pathogenesis is still not clear, short-term effect of combined chemotherapy significantly. But serious side effects, easily lead to different degrees of disability, seriously affect the quality of life of patients and prognosis. In recent years the advantages of traditional Chinese medicine in the treatment of nasopharyngeal carcinoma has attracted attention at home and abroad, the experimental and clinical data indicate that Chinese medicine in improving the quality of life of patients, prolong the survival time, has a unique advantage to reduce tumor recurrence and metastasis and so on.
"Clear nose formula" is a clinical experience of Professor Chen Ruishen in the treatment of nasopharyngeal carcinoma. It is widely used in the treatment of advanced nasopharyngeal carcinoma and adjuvant therapy of nasopharyngeal carcinoma. It has been used for many years and has good clinical effect.
This paper will discuss the influence of "Qing Bi" on non specific immune function of immunosuppressed mice and in vitro effects of "Qing Bi" on the proliferation of CNE cells, to from different levels, different targets to investigate the action mechanism of "Qing Bi Fang" of nasopharyngeal carcinoma, for the clinical use of scientific on the basis of.
Objective:
In the establishment of low immunity mice model based on the application of clear nasal party intervention model mice, study the nose cleaning in the reinforcement mechanism of immunity; in nasopharyngeal carcinoma cell line has been isolated (CNE cells) and serum pharmacology, through cell biology, molecular biology technology, to explore the inhibitory effect of nasal cleaning on the CNE cells.
Method錛，

本文編號(hào)：1450434