【摘要】：研究目的水通道蛋白(Aquaporins, AQPs)是一類介導(dǎo)水、甘油、尿素等分子跨膜轉(zhuǎn)運(yùn)的疏水性跨膜蛋白,廣泛分布于動(dòng)、植物及微生物,迄今為止在哺乳動(dòng)物體內(nèi)已經(jīng)發(fā)現(xiàn)了13個(gè)亞型,即AQP0-12。AQP1表達(dá)于多種上皮組織和內(nèi)皮細(xì)胞。血管內(nèi)皮生長(zhǎng)因子(vascularendothelial growth factor,VEGF)是已知最強(qiáng)的血管通透因子,其通透作用強(qiáng)于組胺1×106倍,對(duì)血管生長(zhǎng)具有強(qiáng)烈的誘導(dǎo)作用,可增生形成微血管,同時(shí)可作用于血管內(nèi)皮細(xì)胞調(diào)節(jié)小血管和毛細(xì)血管的通透作用,這種作用使血漿及部分大分子物質(zhì)(如纖維蛋白原)滲到血管外基質(zhì)中致使組織發(fā)生水腫。研究發(fā)現(xiàn),兩者的異常表達(dá)和調(diào)節(jié)失控與某些疾病的炎性、水腫有關(guān)系,但在變應(yīng)性鼻炎(allergic rhinitis, AR)中鮮有研究,且相關(guān)的變態(tài)反應(yīng)機(jī)制尚不清楚。本實(shí)驗(yàn)旨在通過(guò)對(duì)照AQP1和AEGF在AR大鼠、正常大鼠以及地塞米松干預(yù)后AR大鼠鼻黏膜中的表達(dá)水平和兩者的相關(guān)性,從而進(jìn)一步探討AQP1、AEGF在AR發(fā)病機(jī)制中的作用。研究方法選取8周齡健康Wistar大鼠30只,雌雄各半,體重160g-200g。正常飼養(yǎng)一周后隨機(jī)分為三組,對(duì)照組,模型組和干預(yù)組,每組10只,以卵清蛋白(Ovalbumin,OVA)經(jīng)典致敏法制作模型組和干預(yù)組Wistar大鼠AR模型,對(duì)照組以生理鹽水代替OVA,干預(yù)組大鼠造模成功后腹腔注射地塞米松,干預(yù)一周,模型組以等量生理鹽水代替。用組織化學(xué)方法觀察變應(yīng)性鼻炎的組織形態(tài)學(xué)特征,運(yùn)用免疫組織化學(xué)方法(SP法)和real-time PCR兩種方法分別測(cè)定各組鼻腔黏膜AQP1與VEGF的表達(dá)。實(shí)驗(yàn)結(jié)果以x±s表示,采用SPSS13.0統(tǒng)計(jì)分析軟件對(duì)三組數(shù)據(jù)進(jìn)行分析,數(shù)據(jù)符合正態(tài)分布且方差齊的進(jìn)行方差分析及多組均數(shù)差別的多重比較(LSD法)。偏態(tài)或方差不齊的數(shù)據(jù)進(jìn)行多組數(shù)據(jù)的Kruskal-Wallis檢驗(yàn)及Bonferroni法兩兩比較平均秩和,AQP1與VEGF雙變量符合正態(tài)分布,二者之間的相關(guān)性采用Pearson分析,反之采用Spearman秩相關(guān)。以P0.05為有統(tǒng)計(jì)學(xué)意義。研究結(jié)果按行為學(xué)評(píng)分標(biāo)準(zhǔn),模型組和干預(yù)組大鼠均被激發(fā)出典型的癥狀,示造模成功。干預(yù)組經(jīng)過(guò)地塞米松干預(yù)后,偶有流涕,噴嚏和撓鼻現(xiàn)象基本消失。造模成功后上皮杯狀細(xì)胞明顯增多,上皮下層組織水腫,炎性細(xì)胞浸潤(rùn)及嗜酸性細(xì)胞明顯增多,腺體增生,小血管增加、充血。干預(yù)組組織水腫,血管擴(kuò)張、腺體增生及炎性細(xì)胞浸潤(rùn)程度均減輕。三組大鼠的癥狀學(xué)分析模型組評(píng)分高于對(duì)照組和干預(yù)組。AQP1主要表達(dá)于大鼠鼻黏膜被覆上皮基底膜、血管內(nèi)皮細(xì)胞膜和腺體細(xì)胞內(nèi),VEGF主要表達(dá)于小動(dòng)、靜脈血管內(nèi)皮細(xì)胞和腺體細(xì)胞內(nèi)。AQP1與VEGF在模型組中經(jīng)免疫組織化學(xué)和real-time PCR法檢測(cè)均高表達(dá),干預(yù)組表達(dá)低于模型組。差異均具有統(tǒng)計(jì)學(xué)意義,且兩者具有正相關(guān)性。結(jié)論AQP1、VEGF在實(shí)驗(yàn)性變應(yīng)性鼻炎大鼠鼻粘膜的強(qiáng)表達(dá),表明AQP1、VEGF參與了變應(yīng)性鼻炎的發(fā)病過(guò)程,與組織的水腫、腺體的過(guò)度分泌可能有關(guān)。糖皮質(zhì)激素可以明顯下調(diào)AQP1、VEGF的表達(dá)水平,提示糖皮質(zhì)激素可能是通過(guò)改變AQP1、VEGF的表達(dá),減少腺體的分泌和改善鼻塞癥狀。且AQP1與VEGF之間具有正相關(guān)性,但具體機(jī)制尚不明確。
[Abstract]:Aquaporins (AQPs) are a class of hydrophobic transmembrane proteins that mediate the transmembrane transport of water, glycerol, urea and other molecules. AQPs are widely distributed in animals, plants and microorganisms. So far, 13 subtypes have been found in mammals, namely AQP 0-12. AQP 1 is expressed in many kinds of epithelial tissues and endothelial cells. Vascular endothelial growth factor (VEGF) is the strongest known vascular permeability factor. Its permeability is stronger than that of histamine by 1 65 It was found that the abnormal expression and regulation of fibrinogen were related to the inflammation and edema of some diseases, but little research was done in allergic rhinitis (AR), and the mechanism of allergic reaction was not clear. The expression of AQP1 and AEGF in the nasal mucosa of AR rats, normal rats and AR rats after dexamethasone intervention were compared, so as to further explore the role of AQP1 and AEGF in the pathogenesis of AR. Three groups, control group, model group and intervention group, 10 rats in each group, Ovalbumin (OVA) classical sensitization model group and intervention group Wistar rats AR model, control group with normal saline instead of OVA, intervention group rats after successful modeling intraperitoneal injection of dexamethasone, intervention for a week, model group with the same amount of normal saline instead. Methods The histomorphological features of allergic rhinitis were observed and the expressions of AQP1 and VEGF in nasal mucosa were detected by immunohistochemical method (SP) and real-time PCR respectively. Variance analysis and multiple comparisons of mean differences (LSD). Kruskal-Wallis test and Bonferroni method were used to compare the mean rank sums of skewed or uneven data. AQP1 and VEGF bivariates were in normal distribution. Pearson analysis was used to analyze the correlation between the two variables, and Spearman rank correlation was used instead. Results According to the behavioral scoring criteria, the rats in the model group and the intervention group were all stimulated with typical symptoms, indicating that the model was successful. The degree of edema, vasodilation, glandular hyperplasia and inflammatory cell infiltration were alleviated in the intervention group. The scores in the model group were higher than those in the control group and intervention group. AQP1 was mainly expressed in the basement membrane of nasal mucosa and vascular endothelial cells. AQP1 and VEGF were highly expressed in the model group by immunohistochemistry and real-time PCR. The expression of AQP1 and VEGF in the intervention group was lower than that in the model group. The strong expression of AQP1 and VEGF in nasal mucosa of rats with rhinitis suggests that AQP1 and VEGF are involved in the pathogenesis of allergic rhinitis and may be related to tissue edema and excessive secretion of glands. There is a positive correlation between AQP1 and VEGF, but the specific mechanism is not yet clear.
【學(xué)位授予單位】：山西醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R765.21

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