矽尘对大鼠肺表面活性蛋白影响的研究

The change of pulmonary surfactant protein of rat following silica exposure

目的探讨不同矽尘暴露时间对于大鼠肺表面活性蛋白（SP）A、B、C、D的影响，为矽肺的早期诊断和临床治疗提供理论依据。方法将60只大鼠随机分为染尘组和相应对照组，每组30只。染尘组大鼠经气管灌注50mg／ml的粉尘1ml，左右两肺各0．5ml；对照组灌注等量的生理盐水。在染尘后第3、7、14、21、28天分别获取血清和支气管肺泡灌洗液（BALF），应用酶联免疫法（ELISA）检测大鼠血清和BALF中SP—A、SP—B、SP-C、SP-D的含量。同时检测肺组织中总抗氧化能力（T-AOC）和羟脯氨酸（HYP）的含量，对肺组织进行HE染色观察病理变化。结果与对照组比较，3、14、21、28d染尘组大鼠BALF中SP—A含量及各时点染尘组大鼠BALF中SP—D含量均明显降低，差异均有统计学意义（P〈0．05）；与对照组比较，7、14、21、28d染尘组大鼠BALF中SP-B含量及14、21、28d染尘组大鼠BALF中SP—C含量明显升高，差异有统计学意义（P〈0．05）。与对照组比较，14、21、28d染尘组大鼠血清中SP—A含量及7、14、21d染尘组大鼠血清中SP-B含量及7、14、21、28d染尘组大鼠血清中SP-C含量明显增加，差异均有统计学意义（P〈0．05），且随染矽尘时间的延长，血清中SP—C含量呈增加趋势，有时间-效应关系（r=0．618，P=0．042）。与对照组比较，7、14、21、28d染尘组大鼠血清中SP—D含量降低，差异有统计学意义（P〈0．05）；随着染尘时间的增加，血清中SP—D含量呈下降的趋势，有时间一效应关系（r=-0．731，P=0．016）。与对照组比较，3、7、14、21、28d染尘组大鼠肺组织中HYP含量升高，7、14、21、28d染尘组大鼠肺组织中T—AOC含量下降，差异均有统计学意义（P〈0．05）。染尘大鼠BALF中SP—C的含量与肺组织中HYP呈正相关（r=0．803，P=0．045）；BALL中SP-D的含量与肺组织中HYP呈负相关（r=-0．867，P=0．033）。BALF中SP-A、SP-B含量与肺组织中HYP无相关（r=0．416，P=0．28；r=0．592，P=-0．071）。BALF与血清中SP-D含量的均呈下降趋势，呈正相关（r=0．870，P=-0．034）。BALF与血清中SP—C含量均增加呈正相关（r=0．539，P=0．046）。染尘组大鼠肺组织病理观察可见，肺泡间隔逐渐增厚，尘细胞浸润，有少量胶原纤维分布，至28d时，出现尘细胞结节。结论矽尘导致大鼠BALF中SP含量改变，血清中SP-C、SP—D的水平可作为矽肺早期效应标志物。

Objective To investigate the change of lung surfactant protein （SP） A,B,C,D of rats following silica dust exposure in order to provide the evidences for the early diagnosis indices or therapy of silicosis. Methods 60 male SD rats were randomly divided into silica group, and corresponding controls group. Rats in silica group were administrated 1 ml silica solution by intratracheal instillation at dose of 50mg/ml. Rats in control group were administrated the same amount saline. At 3rd, 7th, 14th, 21st, 28th after silica exposure, serum and bronchoalveolar lavage fluid （BALF） samples were obtained. The concentration of SP-A,SP-B,SP-C, SP-D in serum and BALF were measured by using enzyme immunoassay （ELISA）. Meanwhile the levels of total anti-oxidative activity （T-AOC） and hydroxyproline （HYP） in lung tissue were also detected. The pathology of lung tissue was conducted. Results Compared with control group, SP-A concentration in BALF of silica exposed rat for 3, 14, 21, 28 d was significant lower and SP-D concentration in BALF of silica exposed rat for all time points was also lower. The differences were significant （P〈0.05）. Meanwhile SP-B level in 7,14,21, 28d silica exposed rats BALF and SP-C level in 14,21,28d silica exposed rats markedly decreased （P〈0.05）. In addition compared with control group, SP-A, SP-B and SP-C concentration in serum of silica exposed rat were higher when SP-A for 1d, 21, 28 d silica exposure, SP-B for 7, 14, 21 d silica exposure and Sp-C for 7, 14, 21, 28 d exposure. And all difference were significant （P〈0.05）. As silica exposure time increased, SP-C concentra- tion in serum showed an increase trend, which showed a time-response relationship （r=-0.618, P=-0.042）. How- ever,SP-D concentration in serum of rat for 7, 14, 21, 28 d silica exposure were significant lower than that of control group （P〈0.05）. And there was a decrease trend with time point exposure regarding of SP-D （r=-0.731, P=-0.016）. The HYP content in lung tissue of experiment rats increased at 3rd,7th, 14th, 21st and 28th day time point and The T-AOC activity in lung tissue decrease at ,7th, 14th, 21st and 28th day time point. The differ- ences were significant （P〈0.05）. There was a positive correlation （P=0.803, P=0.045） between SP-C in BALF and HYP of silica exposed rats and a negative correlation between SP-D in BALF and HYP （r=-0.867, P= 0.033）. No significant correlation were seen between SP-A,SP-B BALF and HYP（y=0.416, P=-0.28; r=-0.592, P= 0.071 ）. SP-C concentration in BALF and serum all showed an increased trend and a positive correlation was seen （r=0.539, P=0.046）. The same decrease trend was seen between SP-D in BALF and serum and correlation value was 0.870 （P=0.034）. Conclusion The silica exposure did cause the change of SP content both in BALF and serum. The SP-C and SP-D content in serum might be served as an early effective biomarker of silicosis.