纳米氧化铟锡诱导大鼠肺损伤模型的建立及病理特征的探讨

Establishment of Nano-ITO induced rat model of indium lung disease and exploration of its pathological characteristics

目的研究纳米氧化铟锡(Nano-ITO)致大鼠肺损伤模型中的动态病理学特征,指导临床和基础科研进一步探索肺间质损伤和肺泡蛋白沉积症(PAP)的机制。方法建立大鼠非暴露式气管灌注染毒Nano-ITO模型,进行剂量反应(1.2、3、6 mg·kg^(-1)·bw^(-1))和时间效应(3、7、14、28、56、84 d)研究,分析支气管肺泡灌洗液(BALF)中炎症因子和氧化应激指标水平,HE、PAS、Masson、油红O、天狼星红染色观察肺组织病理形态的改变,透射电镜观察肺组织细胞超微结构,免疫组化测定IL^(-1)β、HO^(-1)、SP-A蛋白变化,免疫荧光检测α-SMA蛋白表达情况,PAS染色BALF细胞蜡块切片。结果气管内灌注Nano-ITO,通过在暴露后第3天诱发急性炎症、第14天诱发肉芽肿(结节)形成和胶原增生、第56天诱发PAP和脂滴积累引起肺毒性。肺组织病理学特征包括典型的肺泡内渗出物、纤维细胞性结节、肺泡脂肪滴融合变大、胆固醇结晶肉芽肿和肺泡蛋白沉积症。Nano-ITO暴露第84天组大鼠肺组织的透射电镜发现,肺泡内嗜酸性物质(多层、格子状和髓鞘样结构)显示异常的板层体(肺泡Ⅱ型上皮细胞的特征)和丰富的粗面内质网和线粒体(成纤维细胞的特征)。细胞病理学光镜观察可见,Nano-ITO暴露28 d后,BALF中出现大量无定形PAS染色阳性物质,肺泡巨噬细胞内可见粉红色颗粒蛋白样物质。结论急性炎症、肉芽肿(结节)形成和胶原增生、肺泡蛋白沉积是Nano-ITO诱导大鼠肺损伤过程中的3个特征性发展阶段,为铟肺病的致病机理研究提供了可参考的特征模型。

ObjectiveTo study the dynamic pathological characteristics of lung tissue in a Nano-ITO induced rat model of indium lung disease and to guide clinical and basic scientific research to further explore the mechanisms of pulmonary interstitial injury and pulmonary alveolar proteinosis(PAP).MethodsDose-response(three divided doses)and time-course studies(six exposure periods)were performed to investigate the pulmonary toxicity induced by Nano-ITO.At the end of the experiment,cytokine levels and oxidative stress were analyzed in the bronchoalveolar lavage fluid.Rat lung tissues were also collected for staining with H&E,PAS,Masson′s,Oil Red O,and Sirius Red.Ultrastructure of lung tissue cells was observed by transmission electron microscopy.Expression of IL^(-1)β,HO^(-1),SP-A was observed by immunohistochemistry,and the expression ofα-SMA was observed by immunofluorescence.ResultsNano-ITO intratracheal instillation caused pulmonary toxicity by inducing acute inflammation at 3 days,granuloma(nodule)formation and collagen hyperplasia at 14 days,and alveolar proteinosis at 56 days post-exposure.Pathological features of lung tissue included typical alveolar exudates,cellular fibrous nodules,enlarged alveolar fat droplet fusion,cholesterol crystal granuloma and pulmonary alveolar proteinosis.The intra-alveolar eosinophilic material(multilamellated,lattice-shaped,and myelin-like structure)showed abnormal lamellar bodies(features of alveolar typeⅡepithelial cells)and abundant rough endoplasmic reticulum and mitochondria(features of fibroblasts)on transmission electron microscopy of the lung tissue from rats exposed to Nano-ITO on the 84th day.Cellular pathology revealed that a large amount of amorphous PAS stain-positive substances appear in BALF at 28 days post-exposure,and pink granular protein-like substances can be seen in alveolar macrophages.ConclusionsThere are three characteristic developmental stages in Nano-ITO induced pulmonary injury in rats,acute inflammation,granuloma(nodule)formation and collagen proliferation,and pulmonary alveolar proteinosis,which provide a reference feature model for the pathogenesis of indium lung disease.