SARS冠状病毒N蛋白致大鼠肺部炎症及糖皮质激素对其的作用

A study of pulmonary inflammatory reaction induced by N-protein of SARS-CoV in rat models and effects of glucocorticoids on it

目的探讨严重急性呼吸综合征(SARS)冠状病毒N蛋白所致大鼠肺部炎症反应及糖皮质激素(以下简称激素)对其的调节作用。方法24只SD大鼠随机分为4组,每组6只;A组大鼠气管内滴入无菌生理盐水0.2ml,B组(6h)、C组(24h)大鼠气管内滴入SARS病毒N蛋白溶液0.2ml,D组大鼠气管内滴入SARS病毒N蛋白溶液0.2ml的同时腹腔内注射10mg/kg地塞米松。测4组大鼠外周血、支气管肺泡灌洗液(BALF)中WBC总数及分类;测4组大鼠肺组织湿/干(W/D)比值;观察4组大鼠肺组织病理学变化;ELISA测4组大鼠血清及BALF中IL6、IL10、转化生长因子(TGF)β1水平。结果(1)外周血淋巴细胞比例C组较A组低(P<0.05),D组较A、C组均低(P值均小于0.01);D组WBC总数较A、C组均低(P<0.01)。BALF中WBC总数C组较A组高(P<0.05),D组较C组低(P<0.05);B、C组BALF中肺泡巨噬细胞占98%～99%。(2)肺脏W/D比值B、C组较A组高(P<0.05),D组较C组低(P<0.01)。(3)肺组织病理学变化:B、C组大鼠肺泡间隔明显增宽,有较多的炎性细胞渗出,包括中性粒细胞、淋巴细胞、单核细胞、成纤维细胞等,血管充血、淤血,有的支气管腔内有炎性细胞渗出,C组变化较B组无明显加重;D组大鼠肺组织炎性反应较B、C组减轻,肺泡间隔变薄,炎性细胞渗出减少。(4)血清及BALF中IL6、IL10、TGFβ1水平B组较A组高(P<0.01),C组进一步升高(P<0.01),D组较C组低(P<0.01)。结论SARS冠状病毒N蛋白具有致病性,能够引起大鼠肺部炎症反应和(或)急性肺损伤,肺损伤与促炎性细胞因子、抗炎性细胞因子的升高及失衡有关;激素可有效地减轻SARS冠状病毒N蛋白所致的肺部炎症反应。

Objective To study the pulmonary inflammatory reaction induced by N-protein of SARS-CoV in rat models and the effects of glucocorticoids on the inflammatory reaction. Methods The pulmonary inflammatory reaction in rat models were induced by intratracheal instillation of N-protein of SARS-CoV with a dose of 0. 2 mg/kg. The rats were randomly divided into four groups： normal saline control group （Nc group） , N-protein group 1 （P1 group, 6 h） , N-protein group 2（ P2 group, 24 h） , and N-protein ＋ dexamethasone group （ P ＋ D group, dexamethasone 10 mg/kg intraperitoneally ）. The blood samples, bronchial alveolar lavage fluid （BALF） and lung tissue were collected after challenge. The cytological and histopathologic changes of lung tissues were observed and the wet/dry ratios （W/D） of lung tissue were determined. The interleukin（IL） -6, IL-10 and transforming growth factor-β1 （ TGF-β1 ） of serum and BALF were measured by ELISA. Results （ 1 ） Compared with the percentage of peripheral blood lymphocytes in Nc group [ （ 68.42 ± 13.07 ） % ], that in P2 group [ （ 50.50 ± 14. 36 ） % ] was significantly decreased （ P 〈 0. 05 ） ; compared with Nc group and P2 group, that in P ＋ D group was furthermore significantly decreased（ P 〈0.01 ）. Compared with the total WBC of peripheral blood in Nc group[ （5.86 ±2.25） ×10^9] and P2 group [（4.83 ± 1.49） × 10^9] , that in P ＋ D group[ （1.96 ± 1.30） × 10^9] was significantly decreased（P 〈0. 01）. （2） Compared with the total WBC of BALF in Nc group[ （95 ±29） × 10^7 ] , that in P2 group [ （ 160 ± 60） × 10^7 ] was significantly increased （ P 〈 0.05 ） ; but compared with P2 group, that in P ＋ D group [ （ 62 ± 23 ） × 10^7 ] was significantly decreased （ P 〈 0. 05 ）. Analysis of BALF differential cell counts showed that the majority of cells were alveolar macrophages in all groups. （ 3 ） The W/D ratios of lung tissue in both P1 and P2 group[ （5.18 ±0. 29） and （5.19 ±0. 34） ,respectively] after N-protein challenge were significantly increased than that in Nc groups [ （ 4. 77 ± 0.27 ） , P 〈 0. 05 ] ; the W/D ratio in P ＋ D group（4. 70 ± 0. 18） was significantly decreased than that in P2 group（ P 〈 0. 01 ）. （4） Compared with Nc group, the levels of IL-6.IL-10,TGF-β1 in both serum and BALF of PI group were significantly increased （P 〈0. 01 ）, and the levels of these cytokines in P2 group were significantly higher than those in P1 group（P 〈 0. 01 ）, but significantly lower in P ＋ D group compared with P2 group（ P 〈 0. 01 ）. Conclusions The N-protein of SARS-CoV had pathogenicity and could induce obvious pulmonary inflammatory reaction and acute lung injury, which were related to the increase and imbalance of pro-inflammatory and anti-inflammatory cytokines. Glucocorticoids could effectively alleviate the pulmonary inflammatory reaction induced by N-protein of SARS-CoV.