本文選題：哮喘 + 基因芯片��； 參考：《南華大學(xué)》2013年碩士論文

【摘要】：目的 利用全基因表達(dá)譜芯片技術(shù)觀察哮喘大鼠和應(yīng)用三氧化二砷干預(yù)后不同時(shí)間點(diǎn)的基因差異表達(dá)的變化，闡明哮喘的發(fā)病機(jī)制和三氧化二砷治療哮喘的分子機(jī)制。 方法 健康雄性清潔級(jí)SD大鼠70只，，按隨機(jī)表隨機(jī)分成3組：正常對(duì)照組A組10只，哮喘模型組B組30只(分別為二周B1組、四周B2組、八周B3組，每組各10只），三氧化二砷干預(yù)組C組30只(分別為二周C1組、四周C2組、八周C3組，每組各10只）。以腹腔內(nèi)注射聯(lián)合霧化吸入卵蛋白建立哮喘模型，末次激發(fā)后24h處死大鼠檢測(cè)支氣管肺泡灌洗液（BALF）中的白細(xì)胞數(shù)及嗜酸性粒細(xì)胞數(shù)；左肺行染色、病理圖像分析；再切取右肺提取肺組織總RNA行全基因組表達(dá)譜芯片檢測(cè)，以基因表達(dá)倍數(shù)值2.0和基因表達(dá)倍數(shù)值－2.0為閾值來(lái)確定差異表達(dá)基因，然后用GO及Pathway分析對(duì)差異表達(dá)基因進(jìn)行功能分類分析，并對(duì)基因芯片結(jié)果進(jìn)行實(shí)時(shí)定量PCR驗(yàn)證。 結(jié)果 1、哮喘模型組BALF中白細(xì)胞總數(shù)、嗜酸性粒細(xì)胞（Eos）比例較正常組均有明顯增高，三氧化二砷組較哮喘模型組明顯降低，差異有顯著性(P 0.05)。 2、哮喘模型組大鼠氣道支氣管及血管周圍存在大量炎癥細(xì)胞浸潤(rùn)，以嗜酸性粒細(xì)胞、淋巴細(xì)胞為主，氣道平滑肌增厚，基底膜增厚，管腔狹窄，粘膜下細(xì)胞外基質(zhì)沉積，且隨著時(shí)間的延長(zhǎng)，以上變化逐漸加重；三氧化二砷組較哮喘模型組上述改變減輕，正常組無(wú)上述改變。 3、在24358個(gè)基因表達(dá)譜的篩選中，哮喘模型組對(duì)比正常組共有438個(gè)差異基因表達(dá)，其中348個(gè)上調(diào)，90個(gè)下調(diào)；三氧化二砷組對(duì)比哮喘模型組共有236個(gè)基因表達(dá)差異，其中96個(gè)上調(diào)，140個(gè)下調(diào)。其中GZMA、MAL基因在哮喘模型組中表達(dá)下調(diào)，三氧化二砷干預(yù)組后表達(dá)上調(diào)，通過(guò)實(shí)時(shí)定量PCR驗(yàn)證后與基因芯片篩選結(jié)果表達(dá)方向一致。 結(jié)論 1、GZMA、MAL等基因可能參與了哮喘的發(fā)生發(fā)展過(guò)程。 2、三氧化二砷治療哮喘的機(jī)理可能與上調(diào)GZMA、MAL基因在肺組織中的表達(dá)有關(guān)。
[Abstract]:Purpose In order to elucidate the pathogenesis of asthma and the molecular mechanism of arsenic trioxide treatment, the changes of gene differential expression at different time points after arsenic trioxide intervention in asthmatic rats were observed by whole gene expression microarray technique. Method Seventy healthy male SD rats of clean grade were randomly divided into three groups: normal control group (n = 10), asthma model group (B group) (n = 30) (group B _ 1 for two weeks, group B _ 2 for four weeks, group B _ 3 for eight weeks). There were 10 rats in each group, and 30 rats in group C in arsenic trioxide intervention group (10 rats in each group) (C1 group for two weeks, C 2 group for four weeks and C 3 group for eight weeks). The asthmatic model was established by intraperitoneal injection and atomization inhalation of ovalbumin. The white blood cells and eosinophils in bronchoalveolar lavage fluid (BALF) were measured 24 hours after the last stimulation, and the left lung was stained and analyzed by pathological images. The total RNA of lung tissue was extracted from the right lung and detected by whole genome expression microarray. The differentially expressed genes were determined by the threshold of gene expression doubling value 2.0 and gene expression doubling value -2.0. Then go and Pathway analysis were used to analyze the function of differentially expressed genes, and real-time quantitative PCR was used to verify the results of gene chip. Result 1. The percentage of total white blood cells and eosophils in BALF of asthmatic model group was significantly higher than that of normal group, and that of arsenic trioxide group was significantly lower than that of asthmatic model group (P 0.05). 2. In the asthma model group, there were a large number of inflammatory cells infiltrating around the airway bronchus and blood vessels, mainly eosinophils and lymphocytes, airway smooth muscle thickening, basement membrane thickening, lumen stenosis, and extracellular matrix deposition. The above changes were gradually aggravated with the prolongation of time, and the above changes in arsenic trioxide group were lighter than those in asthma model group, but no such changes were found in normal group. 3. In the screening of 24358 gene expression profiles, there were 438 differentially expressed genes in asthmatic model group compared with normal group, 348 of which were up-regulated and 90 were down-regulated, and 236 genes were differentially expressed in arsenic trioxide group compared with asthmatic model group. Of these, 96 were up-regulated and 140 downgraded. The expression of GZMA-MAL gene was down-regulated in asthmatic model group and up-regulated in arsenic trioxide intervention group. The expression direction of GZMA-MAL gene was consistent with the result of gene chip screening by real-time quantitative PCR. Conclusion Genes such as GZMA-MAL may be involved in the occurrence and development of asthma. The mechanism of arsenic trioxide in the treatment of asthma may be related to the up-regulation of GZMA-MAL gene expression in lung tissue.
【學(xué)位授予單位】：南華大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2013
【分類號(hào)】：R562.25

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