【摘要】：目的：通過(guò)檢測(cè)中性粒細(xì)胞彈性蛋白酶（HNE）、腫瘤壞死因子-α轉(zhuǎn)化酶（TACE）與黏蛋白MUC5AC在CRS患者中的表達(dá)情況，以及研究體外細(xì)胞培養(yǎng)試驗(yàn)中HNE通過(guò)TACE誘導(dǎo)MUC5AC表達(dá)的作用，提供HNE通過(guò)TACE誘導(dǎo)CRS中杯狀細(xì)胞增生的證據(jù)，為進(jìn)一步了解CRS的發(fā)病機(jī)制提供幫助。 方法：取伴或不伴息肉CRS患者鼻竇黏膜各20例，另外取10例正常鼻腔鼻竇黏膜作為對(duì)照，用HE染色和PAS染色觀察各樣本病理改變,用免疫組化的方法分別檢測(cè)HNE、TACE和MUC5AC在粘膜中的表達(dá)情況，并且用熒光定量PCR的方法分別檢測(cè)組織中HNE、TACE和MUC5AC mRNA的表達(dá)情況。建立體外原代鼻黏膜上皮細(xì)胞培養(yǎng)模型，并且用細(xì)胞免疫化學(xué)的方法進(jìn)行鑒定，穩(wěn)定傳代后給予HNE刺激，并且以腫瘤壞死因子轉(zhuǎn)換酶抑制劑-1(TAPI-1)為干預(yù)因素，用熒光定量PCR的方法檢測(cè)處理后細(xì)胞中MUC5ACmRNA的表達(dá)情況。 結(jié)果：HE染色結(jié)果顯示，在伴或不伴息肉CRS患者鼻竇黏膜中主要的病理特征是杯狀細(xì)胞，炎性細(xì)胞及黏膜下腺體增生。兩組CRS患者鼻竇粘膜上皮層PAS染色均可見(jiàn)強(qiáng)陽(yáng)性表達(dá)，而正常對(duì)照組鼻竇粘膜層為弱陽(yáng)性表達(dá)。免疫組化結(jié)果顯示MUC5AC，TACE和HNE在兩組CRS患者的鼻竇黏膜中的表達(dá)均高于對(duì)照組,且差別均有顯著統(tǒng)計(jì)學(xué)意義(P0.05)，其中，MUC5AC和TACE主要表達(dá)于黏膜上皮杯狀細(xì)胞中,HNE在黏膜上皮和黏膜下腺體中均有表達(dá)；兩組CRS患者間各指標(biāo)的表達(dá)無(wú)差異（P0.05)。熒光定量PCR結(jié)果表明伴息肉與不伴息肉CRS患者的鼻竇組織中MUC5AC，TACE和HNE mRNA的表達(dá)均增加，且與正常對(duì)照組相比有統(tǒng)計(jì)學(xué)差異(P0.05)，而兩組之間并無(wú)明顯差異(P0.05)。體外培養(yǎng)的鼻粘膜上皮細(xì)胞，，經(jīng)細(xì)胞角蛋白AE1抗體染色后證實(shí)其為上皮源性，熒光定量PCR檢測(cè)表明給予HNE刺激后，其MUC5ACmRNA表達(dá)水平高于未刺激組，且差異有統(tǒng)計(jì)學(xué)意義(P0.05);而用腫瘤壞死因子轉(zhuǎn)換酶抑制劑TAPI-1預(yù)處理組及單純使用TAPI-1組，其MUC5ACmRNA表達(dá)水平明顯下調(diào),并且與對(duì)照組及單純HNE刺激組相比差異均有統(tǒng)計(jì)學(xué)意義(P0.01)。 結(jié)論：在伴與不伴息肉CRS中MUC5AC，TACE和HNE的表達(dá)均上調(diào)，且HNE通過(guò)TACE可誘導(dǎo)鼻黏膜上皮細(xì)胞中MUC5AC的增多，表明HNE-TACE信號(hào)通路在CRS中杯狀細(xì)胞增生的過(guò)程中具有重要的作用。
[Abstract]:Aim: to investigate the expression of neutrophil elastase (HNE), tumor necrosis factor- 偽 converting enzyme (TACE) and mucin MUC5AC (MUC5AC) in patients with CRS and to study the role of HNE in TACE induced MUC5AC expression in vitro. To provide evidence of TACE induced goblet cell proliferation in CRS by HNE, and to further understand the pathogenesis of CRS. Methods: 20 cases of nasal sinus mucosa with or without polyps, 20 cases of nasal sinus mucosa and 10 cases of normal nasal cavity and sinus mucosa were taken as control. HE staining and PAS staining were used to observe the pathological changes of each sample, and HNE, was detected by immunohistochemical method. The expression of TACE and MUC5AC in mucous membrane and the expression of HNE,TACE and MUC5AC mRNA in tissues were detected by fluorescence quantitative PCR. The primary nasal epithelial cell culture model was established in vitro, and identified by cell immunocytochemistry. After stable passage, HNE was stimulated, and tumor necrosis factor converting enzyme inhibitor 1 (TAPI-1) was used as the intervention factor. The expression of MUC5ACmRNA was detected by fluorescence quantitative PCR. Results: HE staining showed that the main pathological features of nasal sinus mucosa with or without polyp CRS were goblet cells, inflammatory cells and hyperplasia of submucosal glands. Strong positive expression was found in the epithelial layer of nasal sinuses in both groups of CRS patients, while weak positive expression was found in the mucosal layer of nasal sinuses in the normal control group. The results of immunohistochemistry showed that the expression of MUC5AC,TACE and HNE in nasal sinus mucosa of CRS patients was higher than that of control group, and the difference was statistically significant (P0.05). Among them, MUC5AC and TACE were mainly expressed in mucosal goblet cells. HNE was expressed in mucosal epithelium and submucosal gland. There was no difference in the expression of each index between the two groups of CRS patients (P0.05). The results of fluorescence quantitative PCR showed that the expression of MUC5AC,TACE and HNE mRNA in paranasal sinus tissues of patients with polyps and without polyps CRS increased significantly compared with the normal control group (P0.05), but there was no significant difference between the two groups (P0.05). The epithelial cells of nasal mucosa cultured in vitro were confirmed to be epithelial-derived by cytokeratin AE1 antibody staining. Fluorescence quantitative PCR assay showed that the expression of MUC5ACmRNA was higher than that of unstimulated cells after HNE stimulation. The difference was statistically significant (P0.05). The expression of MUC5ACmRNA in TAPI-1 pretreated with TNF- converting enzyme inhibitor and TAPI-1 alone was significantly decreased compared with control group and HNE stimulation group (P0.01). Conclusion: the expression of MUC5AC,TACE and HNE is up-regulated in CRS with and without polyps, and HNE can induce the increase of MUC5AC in nasal epithelial cells through TACE, indicating that HNE-TACE signaling pathway plays an important role in the proliferation of goblet cells in CRS.
【學(xué)位授予單位】：南昌大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類(lèi)號(hào)】：R765.41

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