小鼠变应性鼻炎合并急性细菌性鼻窦炎的初步研究

Preliminary investigation into the allergic rhinitis complicated with acute bacterial sinusitis in mice

目的探讨变应性鼻炎动物模型合并急性细菌性鼻窦炎的方法。方法清洁级C57BL6/J 小鼠40只,随机分为4组:A 组(卵清蛋白+肺炎链球菌);B 组(卵清蛋白+生理盐水);C 组[磷酸盐缓冲溶液(phosphate buffered solution,PBS)+肺炎链球菌];D 组(PBS+生理盐水),每组各10只。①实验第1～9天,A 组和 B 组每天腹腔注射200μl 10%卵清蛋白,第10～17天A、B 组用6%卵清蛋白鼻腔局部激发建立变应性鼻炎模型,C 组和 D 组用等量 PBS 替代,步骤同前;②于实验第13天 A 组和 C 组鼻腔接种肺炎链球菌 ATCC 49619 200μl;B 组和 D 组用等量生理盐水代替。接种后第6天处死动物,处死前各组动物行内眦静脉采血,以间接酶联免疫吸附试验检测血清白介素5水平。取鼻面骨,石蜡包埋,连续切片,行 HE 染色和0.5%甲苯胺蓝染色,计算机辅助显微镜下观察肥大细胞脱颗粒情况,并计算每平方毫米鼻窦黏膜中多形核中性粒细胞和嗜酸粒细胞的数量。结果A组和 B 组分别有9只和8只动物变应性鼻炎建模成功,鼻部症状、黏膜水肿和黏膜下微血管明显扩张,C 组症状轻微,D 组无任何症状。A 组鼻窦黏膜多形核中性粒细胞密度(±s)为(139.3±26.5)个/mm^2,高于 B 组(70.7±16.7)个/mm^2、C 组(63.0±14.7)个/mm^2和 D 组(40.2±14.1)个/mm^2(P值均<0.01);A 组和 B 组嗜酸粒细胞密度和白介素5水平(±s)分别为(134.6±25.5)个/mm^2、(48.2±13.9)pg/ml 和(116.2±25.2)个/mm^2、(40.8±7.8)pg/ml,均高于 C 组(16.7±2.7)个/mm^2、(23.9±8.7)pg/ml (P 值均<0.05)和 D 组(13.4±4.9)个/mm^2、(24.6±6.5)pg/ml(P 值均<0.05)。结论变应性鼻炎合并细菌性鼻窦炎造模成功。

Objective To develop a mouse model of bacterial rhinosinusitis superposed on allergic rhinitis （AR）, and to explore whether ongoing allergic rhinitis enhance the acute sinus infection and inflammation associated with Streptococcus pneumoniae（SP）. Methods Fourty mice of C57BL6/J were randomly divided on average into 4 groups ： A [ ovalbumin（OVA） ＋ SP], B[ OVA ＋ normal saline（NS） ], C[phosphate buffered solution（PBS） ＋ SP] and D （PBS ＋ NS） . ①Group A and B were sensitized by intraperitoneal injection with 200 μl （10%）OVA on days 1 through 9, and exposed to OVA （6%） intranasally on days 10 through 17, to induce allergic inflammation. OVA was replaced with PBS in group C and D in the same way. ②Subsequently, group A and C were inoculated with SP intranasally on day 13, and NS was used in group B and D. On the 6th day after inoculation, mice were killed. Blood was collected from the orbital venous sinus after anesthesia. The heads were embedded with paraffin and serial sections were followed and stained with hematoxlin-eosin and toluidine blue （0. 5% ）for histological analysis and inflammation cells count. The number of polymorphounclear neutrophils （ PMN ） and eosinophils （EOS） per square millimeter of sinus mucosa were calculated by using a computer-aided special software under microscope. Results AR models were successfully established in 9 mice from group A and 8 from group B. Histologic examination of the sinus from group A and B revealed significant mucosal edema and dilated venules. The symptoms were mild in group C, and no symptom was observed in group D. PMN （x± s） in group A （ 139. 3 ± 26. 5 ）/mm^2 was significantly higher than that in group B （ 70. 7 ± 16. 7 ）/mm^2, C （63. 0 ± 14.7）/mm^2 and D（40. 2 ±14. 1 ）/mm^2 respectively （P 〈 0. 01 ）; EOS and serous IL-5 level in group A （ 134. 6 ± 25.5 ）/mm^2, （48. 2 ± 13.9 ） pg/ml and B （ 116. 2 ± 25.2 ）/mm^2, （40. 8 ± 7.8 ） pg/ml, were higher than that in group C （ 16. 7 ± 2. 7 ）/mm^2, （ 23. 9 ± 8. 7 ） pg/ml （ P 〈 0. 05 ） and D （ 13.4 ± 4. 9 ）/mm^2, （ 24. 6 ± 6.5 ） pg/ml （ P 〈 0. 05 ）. Conclusions The data demonstrate that an ongoing local allergic response augments bacterial infection in mice, and allergic sensitization alone without SP does not induce the sinus infection.