本文選題：分泌性中耳炎　切入點(diǎn)：鼓室積液　出處：《瀘州醫(yī)學(xué)院》2011年碩士論文

【摘要】：目的：分泌性中耳炎(Otitis media with effusion, OME)是以鼓室積液為特征的慢性非化膿性炎癥,其發(fā)病機(jī)制目前尚不明確,通常認(rèn)為與咽鼓管機(jī)械性阻塞、感染、免疫反應(yīng)有關(guān)。本實(shí)驗(yàn)通過建立OME大鼠模型,研究TNF-α與CD44在SD大鼠聽泡粘膜上皮中的表達(dá)變化,探討TNF-α與CD44在OME發(fā)生、發(fā)展中所起的作用。方法：成年健康SD大鼠40只,設(shè)立正常對(duì)照組、手術(shù)對(duì)照(PBS)組、OME模型組三個(gè)實(shí)驗(yàn)組。正常對(duì)照組選SD大鼠4只,不作任何處理,直接取聽泡表面粘膜及聽泡灌洗液檢測(cè)；手術(shù)對(duì)照(PBS)組選用SD大鼠16只分別在中耳腔內(nèi)注射磷酸緩沖液(PBS),按2天、4天、7天、15天四個(gè)不同時(shí)間點(diǎn)處死SD大鼠后取聽泡灌洗液和聽泡粘膜組織檢測(cè)；OME模型組在中耳腔注射脂多糖(LPS)后,與手術(shù)對(duì)照(PBS)組相對(duì)應(yīng),按四個(gè)不同時(shí)間點(diǎn)取聽泡粘膜組織及大鼠聽泡灌洗液行檢測(cè)(各時(shí)間段均多選入一只SD大鼠,以防出現(xiàn)造模過程中動(dòng)物死亡所致實(shí)驗(yàn)動(dòng)物數(shù)量不夠之情況)。聽泡灌洗液運(yùn)用ELISA法檢測(cè)各組中腫瘤壞死因子(TNF-α)濃度。聽泡粘膜組織通過固定、包埋制成切片,運(yùn)用HE染色法查看各組大鼠聽聽泡粘膜組織細(xì)胞的形態(tài)學(xué)改變,采用免疫組化SP法檢測(cè)聽泡粘膜中粘附分子CD44及TNF-α在不同實(shí)驗(yàn)組中的陽性表達(dá),用Image-Pro Plus 6.0多媒體彩色病理圖像分析系統(tǒng)進(jìn)行分析,統(tǒng)計(jì)學(xué)分析得出實(shí)驗(yàn)結(jié)論。結(jié)果：1、采用ELISA法對(duì)正常對(duì)照組聽泡灌洗液中的TNF-α濃度進(jìn)行檢測(cè),其平均濃度為：91.831ng/ml。手術(shù)對(duì)照(PBS)組聽泡灌洗液中平均TNF-α濃度在術(shù)后2天、4天、7天、15天時(shí)為148.858ng/ml、169.898ng/ml、135.753ng/ml、101.830ng/ml。OME實(shí)驗(yàn)組,聽泡滲液中的TNF-α平均濃度分別為201.020ng/ml、235.774ng/ml、230.681ng/ml、116.958ng/ml。OME實(shí)驗(yàn)組灌洗液的TNF-α濃度在受LPS影響后的七天左右最高,其后逐漸減輕,在15天時(shí),炎癥因子降至最低點(diǎn)。以a=0.05為檢驗(yàn)標(biāo)準(zhǔn),做組間配對(duì)t檢驗(yàn),OME實(shí)驗(yàn)組與正常對(duì)照組和手術(shù)對(duì)照(PBS)組比較有明顯差異(P0.05)。提示在OME大鼠模型,TNF-α的濃度在OME實(shí)驗(yàn)組顯著高于對(duì)照組。2、HE染色切片可見,正常對(duì)照組聽泡粘膜無炎癥改變；手術(shù)對(duì)照(PBS)組術(shù)后2天、4天、7天、15天均未見聽泡粘膜上皮細(xì)胞有明顯改變,僅粘膜下有少量炎癥細(xì)胞浸潤(rùn)；OME實(shí)驗(yàn)組術(shù)后兩天可見聽泡粘膜水腫,炎性細(xì)胞浸潤(rùn),其中有大量中性粒細(xì)胞。術(shù)后四天組見毛細(xì)血管擴(kuò)張,炎性細(xì)胞浸潤(rùn)增強(qiáng),以大量中性粒細(xì)胞為主。術(shù)后七天組可見上皮下間隙增寬,更多的炎性細(xì)胞浸潤(rùn),仍以大量中性粒細(xì)胞為主。術(shù)后15天組出現(xiàn)成纖維細(xì)胞增生,新生血管增多,并可見到單核巨噬細(xì)胞浸潤(rùn),中耳粘膜明顯增厚的改變。3、免疫組化SP染色,正常對(duì)照組TNF-α和CD44均在粘膜上皮及上皮下結(jié)締組織僅少量表達(dá)。手術(shù)對(duì)照(PBS)組TNF-α和CD44染色陽性的細(xì)胞表達(dá)部位與正常對(duì)照組無變化,且四個(gè)時(shí)間點(diǎn)之間表達(dá)強(qiáng)度和陽性細(xì)胞數(shù)無明顯差異(P0.05),無統(tǒng)計(jì)學(xué)意義。OME實(shí)驗(yàn)組聽泡粘膜上皮細(xì)胞和粘膜下結(jié)締組織中均有TNF-α和CD44陽性表達(dá),尤其在杯狀細(xì)胞及粘膜下混合腺?gòu)?qiáng)烈表達(dá),其術(shù)后15天強(qiáng)烈表達(dá)。經(jīng)Image-Pro Plus 6.0檢測(cè)雙側(cè)聽泡粘膜陽性細(xì)胞灰度值,使用SPSS13.0采用組間t檢驗(yàn)、相關(guān)性分析。OME實(shí)驗(yàn)組與手術(shù)對(duì)照(PBS)組和正常對(duì)照組TNF-α和CD44陽性表達(dá)經(jīng)統(tǒng)計(jì)學(xué)分析發(fā)現(xiàn)：OME實(shí)驗(yàn)組與對(duì)照組間兩種因子的表達(dá)強(qiáng)度有明顯差異(P0.05)。通過相關(guān)性分析,發(fā)現(xiàn)OME實(shí)驗(yàn)組內(nèi),TNF-α和CD44表達(dá)明顯相關(guān)。手術(shù)組不同時(shí)間段TNF-α和CD44陽性表達(dá)無明顯差異(P0.05),無統(tǒng)計(jì)學(xué)意義。結(jié)論：1、在OME大鼠模型急性期,鼓室積液的中TNF-α濃度與炎癥的嚴(yán)重程度密切相關(guān)。2、TNF-α介導(dǎo)炎癥反應(yīng)的方式可能是激活中性粒細(xì)胞。在這一過程中,粘附分子CD44參與了中性粒細(xì)胞的粘附,其中TNF-α的表達(dá)依賴中性粒細(xì)胞的粘附分子CD44的表達(dá),提示細(xì)胞因子TNF-α與粘附分子CD44之間存在細(xì)胞間信號(hào)轉(zhuǎn)導(dǎo)。3、TNF-α和CD44在OME大鼠模型鼓室積液發(fā)生、發(fā)展中起重要協(xié)同作用。
[Abstract]:Objective : OME media with effusion ( OME ) is a chronic non - suppurative inflammation characterized by effusion of tympanic cavity . The pathogenesis is not clear . It is generally believed to be related to mechanical obstruction , infection and immune response in SD rats . Methods : Forty - four adult healthy SD rats were established , and three experimental groups were established in normal control group , control group ( PBS ) group and OME model group .
In the control group ( PBS ) , 16 SD rats were injected with phosphate buffer ( PBS ) in the middle ear cavity , and SD rats were sacrificed at 4 days , 4 days , 7 days and 15 days .
After lipopolysaccharide ( LPS ) injection in the middle ear cavity , the OME model group corresponds to the operation control ( PBS ) group , and the auditory bubble mucosa tissues and the rat auditory vesicle washing liquid line detection are taken at four different time points ( each time period is more selected into one SD rat , so that the number of experimental animals caused by animal death in the molding process is not sufficient ) . The levels of TNF - 偽 in the normal control group were measured by means of immunohistochemical SP method . Results : 1 . The concentration of TNF - 偽 in the normal control group was 148.858ng / ml , 239.898ng / ml , 135.753ng / ml , 101.830 ng / ml . The concentration of TNF - 偽 in the experimental group was significantly higher than that in the control group .
There were no obvious changes in the cells of the auditory cells in the two days , 4 days , 7 days and 15 days after the operation control ( PBS ) group , and only a small amount of inflammatory cell infiltration was found in the mucosa .
The expression of TNF - 偽 and CD44 in normal control group was significantly correlated with the expression of TNF - 偽 and CD44 in normal control group .

【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
【分類號(hào)】：R764.21

【引證文獻(xiàn)】

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本文編號(hào)：1694797