本文選題：Fibulin-5　切入點(diǎn)：IgG4-ROD　出處：《石河子大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：目的探討基質(zhì)蛋白Fibulin-5與眼眶IgG4相關(guān)性疾病纖維化、炎癥、硬化的關(guān)系。方法選取2014年3月至2016年6月在解放軍第474醫(yī)院收治的患者因眼眶IgG4相關(guān)性疾病(IgG4-related disease、IgG4-RD)(眼眶淚腺腫物摘除術(shù)中采集)的淚腺組織標(biāo)本20例20眼(IgG4-ROD組),收集同期的20例20眼正常的淚腺組織標(biāo)本(眼眶開放性粉碎性骨折清創(chuàng)術(shù)中采集)作為對照組。所有患者術(shù)前均未接受免疫和化學(xué)治療,且在年齡方面具有可比性(P0.05)。在獲得醫(yī)院倫理委員會同意并取得患者的知情同意后,在術(shù)中所有組織均在行眼眶淚腺腫物摘除中留取。把淚腺組織標(biāo)本分為大小均為約0.3cm×0.3cm×0.3cm三塊,生理鹽水充分沖洗干凈。取一塊組織放入4%多聚甲醛固定24h,然后取出放入自動組織脫水機(jī),隨后用石蠟包埋,制成蠟塊備用,另外兩塊放入錫箔紙包裹和裝有Trizol EP管中,放入-80℃冰箱保存?zhèn)溆谩?對經(jīng)-80℃冰箱凍存的標(biāo)本采用實(shí)時(shí)熒光定量PCR檢測Fibulin-5及mRAN表達(dá)和采用Western blot法檢測Fibulin-5及蛋白表達(dá),數(shù)據(jù)采用SPSS19.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)學(xué)分析,數(shù)據(jù)資料經(jīng)Shapiro-Wilk檢驗(yàn)呈正態(tài)分布,結(jié)果進(jìn)行t檢驗(yàn),均數(shù)±標(biāo)準(zhǔn)差(X±s)表示,組間均數(shù)經(jīng)Levenne檢驗(yàn),以P0.05定為有顯著性差異。2對經(jīng)甲醛固定的標(biāo)本采用免疫組織化學(xué)兩步法檢測Fibulin-5及蛋白的表達(dá),其表達(dá)情況根據(jù)細(xì)胞染色面積和染色強(qiáng)度進(jìn)行綜合評分,所得結(jié)果采用Fisher確切概率法,相關(guān)性檢驗(yàn)采用Sperman相關(guān)性分析法,以P0.05定為有顯著性差異。結(jié)果1.實(shí)時(shí)熒光定量PCR檢測:眼眶IgG4相關(guān)性疾病與正常淚腺的患者相比,眼眶IgG4相關(guān)性疾病淚腺組織中Fibulin-5mRNA較正常淚腺明顯增高。IgG4-ROD組中Fibulin-5mRNA的表達(dá)水平為3.69±1.72,而對照組中1.69±0.45,依據(jù)P0.05的檢驗(yàn)水準(zhǔn),兩組比較差異有統(tǒng)計(jì)學(xué)意義(P0.05)。2 Western印跡法檢測:Fibulin-5蛋白與Fibulin-5mRNA表達(dá)變化趨勢較為一致。眼眶IgG4相關(guān)性疾病與正常淚腺的患者相比,眼眶IgG4相關(guān)性疾病淚腺組織中Fibulin-5蛋白較正常淚腺明顯增高。IgG4-ROD組中Fibulin-5蛋白的表達(dá)為0.55±0.12,而對照組中Fibulin-5蛋白表達(dá)為0.19±0.11,按照P0.05的檢驗(yàn)水準(zhǔn),兩組比較差異有統(tǒng)計(jì)學(xué)意義(P0.01)。3免疫組織化學(xué)染色檢測:Fibulin-5蛋白主要分布于細(xì)胞胞漿,陽性染色為黃色或棕黃色呈束狀彌漫分布。免疫組化結(jié)果顯示,IgG4-ROD組中淚腺細(xì)胞紊亂、部分破壞,染色為黃色、棕黃色的纖維化細(xì)胞排列紊亂,對照組中淚腺腺管規(guī)則,細(xì)胞規(guī)則排列,核染藍(lán)色。IgG4-ROD組中Fibulin-5蛋白陽性率80%(16/20),對照組中Fibulin-5蛋白陽性率22.5%(9/40),按照P0.05的檢驗(yàn)水準(zhǔn),兩組比較差異有統(tǒng)計(jì)學(xué)意義,IgG4-ROD組Fibulin-5蛋白的陽性表達(dá)顯著高于對照組。結(jié)論1在Fibulin-5mRNA和蛋白水平的檢測結(jié)果中均顯示,眼眶IgG4相關(guān)性疾病與正常淚腺的患者相比,眼眶IgG4相關(guān)性疾病淚腺組織中Fibulin-5mRNA和蛋白表達(dá)較正常淚腺顯著升高。
[Abstract]:Objective to study the expression of matrix protein Fibulin-5 and IgG4 orbital diseases associated with fibrosis, inflammation, atherosclerosis. Methods from March 2014 to June 2016 in the 474th Hospital of PLA for patients with orbital IgG4 related disease (IgG4-related disease (IgG4-RD) of lacrimal gland tumor excision by collecting) lacrimal gland tissue specimens of 20 cases of 20 eyes (IgG4-ROD group). Collected in the same period in 20 cases 20 eyes of normal lacrimal gland tissue specimens (eyes open comminuted fracture debridement in the acquisition) as the control group. All patients were not receiving preoperative immune and chemical treatment, and is comparable in terms of age (P0.05). The hospital ethics committee and informed consent were obtained agree, in all the tissues in the lacrimal gland tumor in the lacrimal gland tissue specimens. The specimens were divided into size were about 0.3cm * 0.3cm * 0.3cm three, saline flush Clean. Take a piece of tissue were fixed in 4% paraformaldehyde 24h, then take out into the automatic tissue dehydration machine, followed by paraffin, paraffin spare, another two pieces into the package with Trizol foil and EP tube, placed in the refrigerator on the standby.1 -80 C by -80 C refrigerator frozen samples real time fluorescence quantitative PCR detection of Fibulin-5 and the expression of mRAN and blot using Western method to detect the expression of Fibulin-5 and protein data were statistically analyzed using SPSS19.0 statistical software, data by Shapiro-Wilk test of normal distribution, the results of t test, mean standard deviation (X + s) said that the two groups were tested by Levenne, to P0.05 is a significant difference of.2 expression by immunohistochemical detection of Fibulin-5 protein in two steps and formalin fixed specimens, the expression of cells according to the staining area and intensity in comprehensive evaluation, The results obtained using the Fisher exact test, correlation analysis using Sperman correlation analysis method, using P0.05 as a significant difference. The results of 1. real time fluorescence quantitative PCR detection: compared with normal orbital IgG4 related diseases of lacrimal gland in patients with orbital IgG4 related diseases of lacrimal gland tissue Fibulin-5mRNA than normal lacrimal gland significantly increased the expression level of Fibulin-5mRNA in the.IgG4-ROD group is 3.69 + 1.72, 1.69 + 0.45 in the control group, according to the P0.05 standard test, there was significant difference between two groups (P0.05): Fibulin-5 Western blot detection of.2 protein and Fibulin-5mRNA expression of the trend is more consistent. Compared with the orbital IgG4 disease patients and normal lacrimal gland, lacrimal gland tissue expression of IgG4 related diseases Fibulin-5 protein Fibulin-5 protein was significantly higher than normal lacrimal gland in.IgG4-ROD group was 0.55 + 0.12, while the control group in Fibulin-5 Protein expression was 0.19 + 0.11, according to the P0.05 standard test, there was significant difference between two groups (P0.01).3 immunohistochemical staining: Fibulin-5 protein mainly distributed in the cytoplasm, the positive staining for yellow or brown bundles of diffuse distribution. Immunohistochemical results show that the lacrimal gland cell disorder, IgG4-ROD group in the part of the damage, was yellow, brown and yellow fibrosis cells arranged in disorder, the control group in the lacrimal gland tube rules, cells arranged regularly, nuclear staining positive rate of Fibulin-5 protein in 80%.IgG4-ROD Blue Group (16/20). The positive rate of the control group in Fibulin-5 protein 22.5% (9/40), according to the P0.05 standard test, the comparison between the two groups there was significant difference in the positive expression of IgG4-ROD protein in Fibulin-5 group was significantly higher than the control group. Conclusion: 1 at the level of Fibulin-5mRNA and protein detection results showed that the orbital IgG4 related diseases and normal lacrimal gland The expression of Fibulin-5mRNA and protein in the lacrimal gland tissue of the orbital IgG4 related disease was significantly higher than that of the normal lacrimal gland.

【學(xué)位授予單位】：石河子大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R777.5

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本文編號：1596809