不同剂量脂多糖在不同作用时间下诱导小鼠急性肺损伤的效果评价

An evaluation of effects on induction of acute lung injury in mice by different doses of lipopolysaccharide and different durations

目的：观察脂多糖（LPS）诱导急性肺损伤（ALI）小鼠的炎症因子变化，探讨不同剂量LPS在ALI发生发展过程中不同时间点对肺损伤的影响。方法将210只C57BL/6小鼠按气管内滴注LPS剂量分为2.5、5.0、7.5、10.0 mg/kg组，采用气管内滴注LPS的方法复制ALI模型；并设正常对照组、生理盐水对照组，每组10只。LPS损伤后1、2、4、8 h观察小鼠的肺组织湿/干质量（W/D）比值、肺损伤评分及肺组织病理学的变化，同时检测血清和支气管肺泡灌洗液（BALF）中去甲肾上腺素（NE）、肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）和蛋白含量。结果①LPS所致肺损伤呈剂量依赖性增加和时间依赖性增加。②随LPS剂量增加和作用时间延长，LPS组肺组织W/D比值、肺损伤评分增加，血清和BALF中NE、TNF-α、IL-6及BALF中蛋白含量均显著增加，LPS 5.0 mg/kg组作用4 h是出现急性呼吸窘迫综合征（ARDS）显著特征的剂量和时间点〔肺W/D比值：4.97±0.41，肺损伤评分（分）：5.60±1.52；血清NE（ng/L）：379.99±27.65， TNF-α（ng/L）：159.15±20.62，IL-6（ng/L）：177.15±29.13；BALF 中 NE（mg/kg）：105.85±13.66，TNF-α（mg/kg）：227.22±48.01，IL-6（mg/kg）：251.55±54.08，总蛋白含量（g/L）：1.59±0.37〕，但上述指标的变化以LPS 10.0 mg/kg组作用后8 h较LPS 7.5、5.0、2.5 mg/kg组相同时间点更显著〔肺组织W/D比值：5.10±0.18比5.01±0.43、5.01±0.19、4.91±0.30，肺损伤评分（分）：9.20±1.48比8.00±1.00、6.00±1.22、4.40±0.89；血清NE（ng/L）：447.43±34.63比419.23±30.62、391.16±54.91、372.59±51.52，TNF-α（ng/L）：205.99±31.31比181.01±25.11、161.01±13.98、138.83±28.95，IL-6（ng/L）：233.76±34.84比206.21±26.68、186.58±26.54、156.99±28.83；BALF 中 NE（mg/kg）：190.82±41.75比153.30±35.42、122.64±25.15、80.23±13.69，TNF-α（mg/kg）：305.24±72.99比292.77±38.07、249.60±35.20、193.63±10.83，IL-6（mg/kg）：354.81±67.79比303.02±54.24、272.43±32.34、197.64±12.35，BALF总蛋白含量（g/L）：2.31±0.30比2.02±0.26、1.62±0.19、1.10±0.24，P＜0.05或P＜0.01〕。结论 LPS诱导的小鼠ALI程度与剂量和时间呈正相关。LPS 5.0 mg/kg作用4 h是出现ARDS显著特征的剂量和时间点。

Objective To observe the changes of inflammatory factors in acute lung injury （ALI） in mice induced by lipopolysaccharide （LPS）, and to explore the influence of different doses of LPS on ALI onset and progress at different time points. Methods Intratracheally, LPS at the dosages of 2.5, 5.0, 7.5 and 10.0 mg/kg were administered to a total of 210 C57BL/6 mice, and according to the difference in dosage, they were divided into four groups. The ALI model was replicated by intratracheally dropping of LPS. And a normal control group and a normal saline control group were established （each, n=10）. The changes of index of pathological lung tissue and lung tissue wet/dry （W/D） ratio were observed at 1, 2, 4, and 8 hours after injury, and simultaneously, the levels of norepinephrine （NE）, tumor necrosis factor-α （TNF-α）, interleukin-6 （IL-6） and protein in serum and bronchoalveolar lavage fluid （BALF） were detected. Results ①The degree of lung injury induced by LPS was dose-and time-dependent.②With the increase of LPS dosage and prolongation of time, in LPS group, the lung W/D ratio and the index of pathological lung tissue were increased;additionally, the levels of NE, TNF-α, IL-6 and protein in serum or BALF were also significantly increased. The critical occurrence point of acute respiratory distress syndrome （ARDS） with specific characteristics was at 5.0 mg/kg of LPS acting for 4 hours [lung W/D ratio： 4.97±0.41, index of pathological changes of lung tissue （score）： 5.60±1.52; serum NE （ng/L）： 379.99±27.65, TNF-α （ng/L）： 159.15±20.62, IL-6 （ng/L）： 177.15±29.13;BALF NE （mg/kg）：105.85±13.66, TNF-α（mg/kg）：227.22±48.01, IL-6 （mg/kg）：251.55±54.08, total protein （g/L）：1.59±0.37]. The injury induced by LPS acting for 8 hours in the dosage group 10.0 mg/kg was the most significant in comparisons with other groups of dosages at the same time points [lung W/D ratio：5.10±0.18 vs. 5.01±0.43, 5.01±0.19, 4.91±0.30; index of pathological changes of lung tissue （score）： 9.20±1.48 vs. 8.00±1.00, 6.00±1.22, 4.40±0.89;serum NE （ng/L）： 447.43±34.63 vs. 419.23±30.62, 391.16±54.91, 372.59±51.52; TNF-α（ng/L）： 205.99±31.31 vs. 181.01±25.11, 161.01±13.98, 138.83±28.95; IL-6 （ng/L）： 233.76±34.84 vs. 206.21±26.68, 186.58±26.54, 156.99±28.83;BALF NE （mg/kg）：190.82±41.75 vs. 153.30±35.42, 122.64±25.15, 80.23±13.69;TNF-α（mg/kg）：305.24±72.99 vs. 292.77±38.07, 249.60±35.20, 193.63±10.83; IL-6 （mg/kg）： 354.81±67.79 vs. 303.02±54.24, 272.43±32.34, 197.64±12.35;total protein （g/L）：2.31±0.30 vs. 2.02±0.26, 1.62±0.19, 1.10±0.24, P〈0.05 or P〈0.01]. Conclusions The severity of ALI induced by LPS in mice was positively correlated to LPS dosage and duration of its action. After administration of LPS 5 mg/kg for 4 hours, remarkable characteristic manifestations of ARDS occur in mice, reaching the critical point.