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目的通过观察骨髓间充质干细胞(MSC)对博来霉素致大鼠肺纤维化动物模型的影响,探讨治疗肺纤维化的新方法。方法体外分离、培养雄性6周龄 SD 大鼠的骨髓 MSC。将48只雌性 SD 大鼠随机分为6组,第1～5组经气管注入5.0 mg/kg 博来霉素0.3 ml,第1和第3组分别于博来霉素注射第1天和第7天经尾静脉注入雄性大鼠 MSC 液0.2 ml(细胞数为2.5×10~6个);第2和第4组分别于博来霉素注射第1天和第7天经尾静脉注入等量的磷酸缓冲液0.2 ml;第5组作为模型阳性对照,注射博来霉素后未给予其他处理;第6组作为模型阴性对照,经气管注入等量生理盐水0.3ml 代替博来霉素,未给予其他处理。于实验第28天统一处死大鼠,留取肺组织行病理学检查、羟脯氨酸含量测定;提取肺组织的 DNA,通过聚合酶链反应(PCR)-琼脂糖电泳法检测雄鼠的性别决定基因(sry 基因),以判断外源给予的 MSC 是否在雌性大鼠肺组织中存在。结果在博来霉素致肺损伤第1天和第7天给予 MSC 干预治疗后的大鼠肺脏病理改变均较对照组减轻,肺纤维化程度评分分别为(1.0±0.2)分、(2.5±0.5)分和(1.6±0.5)分、(2.3±0.8)分;肺组织羟脯氨酸含量分别为(83±17)μg/mg、(96±20)μg/mg 和(123±32)μg/mg、(127±34)μg/mg,在博来霉素致肺损伤第1天较第7天给予 MSC 其作用更明显。PCR 检测结果显示,在博来霉素致肺损伤第1天给予 MSC 组的大鼠肺组织可以检测到 sry 基因。结论外源给予的 MSC 可以减轻肺纤维化的形成,在肺损伤早期给予 MSC 干预的疗效更好。
Objective To observe the effect of bone marrow mesenchymal stem cells (MSCs) on bleomycin-induced pulmonary fibrosis in rats and to explore a new method for the treatment of pulmonary fibrosis. Methods The bone marrow MSCs from male 6-week-old SD rats were isolated and cultured in vitro. Forty eight female Sprague-Dawley rats were randomly divided into 6 groups. Groups 1 to 5 were injected with 0.3 ml bleomycin 5.0 mg / kg intratracheally. Groups 1 and 3 were injected with bleomycin on days 1 and 7 days after injection into the male rat MSC solution 0.2 ml (cell number of 2.5 × 10 ~ 6); the second and the fourth group were bleomycin injection on the first day and the seventh day via the tail vein injection Volume of phosphate buffer 0.2 ml; Group 5 as a model positive control, injection of bleomycin did not give any other treatment; Group 6 as a model negative control, tracheal injection of equal amount of normal saline 0.3ml instead of bleomycin, No other treatment was given. The rats were sacrificed on the 28th day of the experiment, and the lung tissue was collected for pathological examination and hydroxyproline content determination. The lung tissue DNA was extracted and the male sex was detected by polymerase chain reaction (PCR) - agarose electrophoresis The gene (sry gene) is determined to determine whether exogenously administered MSC is present in the lungs of female rats. Results The pathological changes in the lungs of rats treated with MSC were relieved on the 1st day and the 7th day after bleomycin-induced lung injury. The lung fibrosis scores were (1.0 ± 0.2), (2.5 ± (96 ± 20) μg / mg and (123 ± 32) μg / mg respectively, while the levels of hydroxyproline in lung tissue were (83 ± 17) μg / μg / mg and (127 ± 34) μg / mg, respectively. The effect was more pronounced on the first day of bleomycin-induced lung injury than on the seventh day. The results of PCR showed that the sry gene was detected in the lung tissue of rats that were given MSC on day 1 of bleomycin-induced lung injury. Conclusion Exogenous administration of MSC can reduce the formation of pulmonary fibrosis, and the effect of MSC intervention in the early stage of lung injury is better.