荧蒽和萘暴露对哮喘小鼠速发相反应的作用及其机制的研究

发布时间：2018-06-02 00:11

本文选题：荧蒽、萘 + 哮喘反应　；参考：《延边大学》2012年硕士论文

【摘要】：目的：探讨不同浓度的荧蒽和萘暴露对哮喘小鼠速发相反应的作用及其机制。方法：选择雄性小鼠120只(Balb/c),随机分组为12组,10只/组；哮喘模型组(C组),利用新鲜配置卵清白蛋白(OVA)20ug+氢氧化铝2mg共0.2ml于第1,8,15天给予小鼠腹腔注射致敏,第22天开始给予3%卵清白蛋白(OVA)+生理盐水溶液雾化吸入,1次/天,30min/d,连续雾化吸入7天。正常对照组(A组),以生理盐水代替卵清白蛋白分别进行腹腔注射和雾化吸入。酒精对照组(B组),以95%乙醇溶液代替卵清白蛋白,步骤同A组。D组为荧蒽低浓度组,在制备哮喘模型前暴露于荧蒽低浓度(50ug/m3)；E组为荧蒽中浓度组,在制备哮喘模型前暴露于荧蒽中浓度(100ug/m3)；F组为荧蒽高浓度组,在制备哮喘模型前暴露于荧蒽高浓度(200ug/m3)；G组为萘低浓度组,在制备哮喘模型前暴露于萘低浓度(50mg/m3)；H组为萘中浓度组,在制备哮喘模型前暴露于萘中浓度(250mg/m3)；Ⅰ组为萘高浓度组,在制备哮喘模型前暴露于萘高浓度(500mg/m3)；J组为混合低浓度组,在制备哮喘模型前暴露于荧蒽+萘低浓度(荧蒽50ug/m+萘50mg/m3)；K组为混合中浓度组,在制备哮喘模型前暴露于荧蒽+萘中浓度(荧蒽100ug/m3+萘250mg/m3)；L组为混合高浓度组,在制备哮喘模型前暴露于荧蒽+萘高浓度(荧蒽200ug/m3+萘500mg/m3)。暴露染毒时在玻璃熏气箱中进行动式染毒,4小时/日,连续2周。同时A,B,C组也在同样大小的玻璃箱中接受新鲜空气,4小时/日,连续2周。D、E、F、G、H、I、J、K、L组暴露染毒后,再以卵清白蛋白致敏为哮喘模型。各组攻击结束一周后,再给予3%卵清白蛋白激发气道30分钟,并利用多道信号采集处理系统测定气道阻力；计数肺泡灌洗液中嗜酸性粒细胞数：采用免疫组化方法检测肺组织中血管内皮生长因子(VEGF).基质金属蛋白酶-9(MMP-9)、基质金属蛋白酶抑制剂-1(TIMP-1)阳性表达；并观察肺组织病理学形态的变化。结果：①A、B组的小鼠吸入组胺(MCH)后气道阻力比值相似,无统计学差异(P0.05)。C-L组小鼠气道阻力比值均高于A、B组,与A、B组比较差异C-L组均有统计学意义(P0.01)。与C组相比较,D-L组小鼠气道阻力比值均有所升高,但仅有E、F、H、I、K、L组与C组比较差异有统计学意义(P0.05)。②支气管肺泡灌洗液中A、B组未见或见少量的嗜酸性粒细胞(EOS),A组与B组比较无统计学意义(P0.05)。C-L组支气管肺泡灌洗液中EOS数占细胞总数的百分比明显高于A组及B组,比较差异有统计学意义(P0.01)。D-L组中EOS数占细胞总数的百分比有所升高,且随吸入的荧蒽和萘浓度的增加,嗜酸性粒细胞数也逐渐增多,但与C组比较差异无统计学意义(P0.05)。③A、B组肺组织中的血管内皮生长因子(VEGF)、基质金属蛋白酶-9(MMP-9)、基质金属蛋白酶抑制剂-1(TIMP-1)的阳性表达相似,无统计学差异(P0.05)。A组及B组小鼠血管内皮细胞未见明显VEGF、MMP-9、TIMP-1表达；D-L组小鼠VEGF、MMP-9、TIMP-1在肺组织细胞浆内见不同程度的阳性表达,可见胞核着色,与A组及B组比较有统计学意义(P0.01)。VEGF混合高浓度组(L组)及MMP-9混合高浓度组(L组)比哮喘模型组(C组)阳性表达明显,比较差异有统计学意义(P0.05)；TIMP-1混合高浓度组(L组)比哮喘模型组(C组)阳性表达明显,比较差异有统计学意义(P0.05)。D-L组阳性表达比哮喘模型组(C组)不同程度增高,但混合中、低浓度组,以及荧葸和萘的高、中、低浓度组阳性表达与哮喘模型组比较无统计学意义(P0.05)。混合高浓度组MMP-9与TIMP-1的比值明显升高,MMP-9与VEGF呈正相关。④A、B组肺组织病理切片中可以看到支气管上皮细胞完整,无变性、坏死,间质内未见炎症细胞的浸润,血管周围未见水肿等改变。C组可见气道周围较多炎症细胞浸润。D、G、J组见部分气道周围有大量炎性细胞浸润,周围血管轻度水肿。E、H、K组上皮细胞脱落,较多炎性细胞浸润,周围血管水肿较明显。F、I、L组上皮细胞明显脱落,大量炎性细胞浸润,周围血管水肿明显。病变程度与浓度呈正相关。结论：1、荧蒽和萘的暴露可以加重哮喘小鼠气道高反应性。 2、高浓度(荧蒽+萘)使VEGF、MMP-9的表达上调,MMP-9/TIMP-1的平衡失调,导致气道炎症、气道反应性增高。 3、荧蒽和萘的吸入可以加重哮喘小鼠速发相反应。
[Abstract]:Objective: To investigate the effects of different concentrations of fluoranthene and naphthalene on the rapid phase response in asthmatic mice and its mechanism.
Methods: 120 male mice (Balb/c) were randomly divided into 12 groups, 10 rats and 12 groups. The asthma model group (group C) was treated by intraperitoneal injection of OVA 20ug+ aluminum hydroxide 2mg altogether on day 1,8,15, and 3% ovum albumin (OVA) + physiological saline solution inhalation, 1 times / day, 30min/d began on the day of twenty-second days. In the normal control group (group A), the normal control group (group A) replaced ovalbumin by intraperitoneal injection and atomization inhalation with normal saline instead of ovalbumin. The alcohol control group (group B) replaced ovalbumin with 95% ethanol solution, and the.D group of group A was the low concentration fluoranthene concentration group before making asthma model (50ug/m3), and E group was fluoranthene. In the medium concentration group, the concentration of fluoranthene was exposed to the concentration of fluoranthene (100ug/m3) before the preparation of the asthma model; the F group was exposed to high fluoranthene concentration (200ug/m3) before the preparation of the asthma model, and the low concentration of naphthalene in the G group was exposed to the low concentration of naphthalene (50mg/m3) before the preparation of the asthma model; the group H was the concentration group of naphthalene, and was exposed to the model of asthma before preparing the model of asthma. Concentration (250mg/m3) in naphthalene, group I was high concentration of naphthalene, exposed to high concentration of naphthalene (500mg/m3) before preparing asthma model, group J was mixed low concentration group and exposed to fluoranthene + naphthalene 50mg/m3 (fluoranthene 50ug/m+ naphthalene 50mg/m3) before preparing asthma model; K group was mixed medium concentration group, before preparing asthma model, the concentration of fluoranthene + naphthalene (fluoranthene) 100ug/m3+ naphthalene 250mg/m3), L group was a mixed high concentration group, and was exposed to fluoranthene + naphthalene high concentration (fluoranthene 200ug/m3+ naphthalene 500mg/m3) before preparing the asthma model. Exposure was carried out in a glass smoked gas box for 4 hours for 2 weeks, while A, B, C group also received fresh air in the same small glass box, 4 hours per day, 2 weeks in a continuous period of.D. E, F, G, H, I, J, K, and L group were exposed to an asthma model after exposure. After one week of attack, 3% ovum albumin was given to stimulate the airway for 30 minutes, and the airway resistance was measured by a multichannel signal acquisition and processing system, and the number of eosinophils in the alveolar lavage fluid was counted: the lung group was detected by immunohistochemical method. Vascular endothelial growth factor (VEGF), matrix metalloproteinase -9 (MMP-9) and matrix metalloproteinase inhibitor -1 (TIMP-1) were expressed in the tissue, and the pathological changes in lung tissue were observed.
Results: (1) A, B group mice inhaled histamine (MCH) after the airway resistance ratio is similar, no statistical difference (P0.05).C-L group of mice airway resistance ratio is higher than A, B group, and A, B group difference C-L group have statistical significance (P0.01). The difference was statistically significant (P0.05). A in bronchoalveolar lavage fluid, group B did not see or see a small amount of eosinophil (EOS), A group and B group had no statistical significance (P0.05), the percentage of EOS number of EOS in the bronchoalveolar lavage fluid of.C-L group was significantly higher than that of A group and B group, and the difference was statistically significant (P0.01). The percentage of the total number of cells increased, and the number of eosinophils increased with the increase of the concentration of fluoranthene and naphthalene, but there was no significant difference between the C group (P0.05). (3) A, the vascular endothelial growth factor (VEGF), the matrix metalloproteinase -9 (MMP-9), the matrix metalloproteinase inhibitor -1 (TIMP-1) in the lung tissue of the group B The positive expression was similar, there was no statistical difference (P0.05) in group.A and B group, no significant VEGF, MMP-9, TIMP-1 expression was found in the vascular endothelial cells of group B, and the positive expression of VEGF, MMP-9 and TIMP-1 in the D-L group was found in the cytoplasm of lung tissue. The positive expression of MP-9 mixed high concentration group (group L) was significantly higher than that of the asthma model group (group C), and the difference was statistically significant (P0.05); the positive expression of TIMP-1 mixed high concentration group (L group) was significantly higher than that of the asthma model group (C group), and the difference was statistically significant (P0.05) the positive expression of the.D-L group was higher than the asthma model group (C group), but in the mixture, the low level was lower. In the concentration group, the positive expression of the high, middle and low concentration groups was not significant (P0.05). The ratio of MMP-9 to TIMP-1 in the mixed high concentration group was significantly higher, and the MMP-9 was positively correlated with VEGF. (4) A. In the pathological section of lung tissue of group B, the epithelial cells of the bronchiotracheal epithelium could be seen intact, no denaturation, necrosis and no interstitial tissue. The infiltration of inflammatory cells and no edema around the blood vessels in group.C showed that many inflammatory cells around the airway were infiltrated.D, G, group J showed a large number of inflammatory cells around the airway, the peripheral vascular edema was.E, H, K group epithelial cells fell off, more inflammatory cells infiltrated, peripheral vascular edema was obvious.F, I, L group epithelial cells obviously shedding, A large number of inflammatory cells infiltration and peripheral vascular edema were observed. The degree of lesion was positively correlated with the concentration of inflammatory cells.
Conclusion: 1, exposure to fluoranthene and naphthalene can aggravate airway hyperresponsiveness in asthmatic mice.
2, high concentration (fluoranthene + naphthalene) increased the expression of VEGF and MMP-9, and MMP-9/TIMP-1 imbalance, leading to airway inflammation and increased airway responsiveness.
3, inhalation of fluoranthene and naphthalene can aggravate the rapid phase reaction in asthmatic mice.
【学位授予单位】：延边大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R725.6

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