摘要
目的:月球表面尘埃因长期受宇宙辐射及陨石撞击作用的影响,形成具有高静电吸附特性的纳米级棱角结构,这些尘埃具有吸入风险,但其对肺部的毒理机制尚不清楚。针对月球基地建设和资源开发过程中的粉尘暴露防护需求,本研究通过构建月球土壤模拟物(lunar soil simulant,LSS)急性暴露模型,并以地球土壤(Earth soil,ES;陕西黄土)为对照,旨在探究粉尘吸入对肺损伤的作用机制。方法:将C57BL/6小鼠随机分为3组:磷酸盐缓冲液(phosphate buffered saline,PBS)组、LSS组和ES组,每组5~7只小鼠。LSS组和ES组分别使用LSS和ES建立急性吸入模型,采用单次呛咳法进行暴露。实验过程中动态监测小鼠体重变化,持续28 d,并于暴露后第3、7、14、28天分批处死小鼠取材,采集外周血、肺泡灌洗液(bronchoalveolar lavage fluid,BALF)及肺组织样本。通过流式细胞术分析BALF中免疫细胞亚群的动态变化;采用HE染色评估肺组织结构及炎症程度,过碘酸-雪夫(periodic acid-Schiff,PAS)染色评估气道黏液分泌,Masson染色判断胶原沉积情况。通过实时反转录聚合酶链反应(real-time reverse transcription PCR,real-time RT-PCR)检测炎症因子(IL-1β、IL-6、TNF-α)及上皮屏障相关基因(Occludin、Cadherin-1、Zo-1)的mRNA表达水平。选取暴露后第7天的肺组织样本进行转录组学测序,并结合免疫浸润分析和功能通路富集分析,探讨LSS和ES对肺部免疫微环境的调控效应。结果:ES组小鼠在暴露18 d后体重逐渐下降(均P<0.05),而LSS组小鼠的体重与PBS组的差异无统计学意义。HE染色结果显示:LSS与ES均可诱发气道及血管周围的炎症细胞浸润,且炎症反应在28 d内持续存在,但随着时间延长有逐渐减轻的趋势。PAS染色提示LSS组小鼠在第3天出现明显的气道黏液分泌增加,之后逐渐恢复;ES组未见显著变化。Masson染色结果未见LSS组或ES组在28 d内出现明显的肺纤维化改变。Real-time RT-PCR结果显示:LSS和ES处理均可显著上调肺组织中IL-1β及TNF-α的表达,尤其在暴露第7天时炎症反应最为显著,同时伴随Occludin、Cadherin-1、Zo-1等上皮屏障标志基因表达下调(均P<0.05)。转录组分析结果显示:LSS与ES处理均显著激活与趋化因子相关的通路,富集到多条白细胞迁移、中性粒细胞募集等免疫相关通路。进一步验证发现LSS组肺组织中CXCL2及MMP12表达上调;ES组则以CXCL3及MMP12表达上调为主。结论:LSS和ES均可诱导小鼠持续性肺损伤和中性粒细胞浸润,但所依赖的分子机制存在差异。与ES相比,LSS暴露还会引发短暂的嗜酸性粒细胞应答,提示LSS颗粒具有更强的免疫激活潜能和生物毒性。
Objective:Due to prolonged exposure to cosmic radiation and meteorite impacts,lunar surface dust forms nanoscale angular particles with strong electrostatic adsorption properties.These dust particles pose potential inhalation risks,yet their pulmonary toxicological mechanisms remain unclear.Given the need for dust exposure protection in future lunar base construction and resource development,this study established an acute exposure model using lunar soil simulant(LSS)and used Earth soil(ES;Loess from Shaanxi,China)as a comparison to investigate lung injury mechanisms.Methods:C57BL/6 mice were randomly assigned to 3 groups:Phosphate buffered saline(PBS),LSS,and ES,with 5 to 7 mice per group.Mice in the LSS and ES groups received a single intratracheal instillation to induce acute inhalation exposure.Body weight was monitored for 28 days.Mice were euthanized at days 3,7,14,and 28 post-exposure,and peripheral blood,bronchoalveolar lavage fluid(BALF),and lung tissues were collected.Immune cell subsets in BALF were analyzed using flow cytometry.Hematoxylin-eosin(HE)staining assessed lung structure and inflammation;periodic acid-Schiff(PAS)staining evaluated airway mucus secretion;Masson staining examined collagen deposition.Realtime reverse transcription PCR(real-time RT-PCR)was used to measure the mRNA expression of inflammatory cytokines(IL-1β,IL-6,and TNF-α)and epithelial barrier genes(Occludin,Cadherin-1,and Zo-1).Lung tissues at day 7 were subjected to transcriptomic sequencing,followed by immune infiltration and pathway enrichment analyses to determine immunoregulatory mechanisms.Results:Body weight in the ES group progressively declined after day 18(all P<0.05),while the LSS group showed no significant changes compared with the control group.HE staining showed both LSS and ES induced inflammatory cell infiltration around airways and vasculature,which persisted for 28 days but gradually lessened over time.PAS staining revealed marked mucus hypersecretion in the LSS group at day 3,followed by gradual recovery;no significant mucus changes were observed in the ES group.Masson staining indicated no obvious pulmonary fibrosis in either group within 28 days.Real-time RT-PCR demonstrated significant upregulation of IL-1βand TNF-αin both LSS and ES groups,peaking on day 7,accompanied by downregulation of epithelial barrier genes(Occludin,Cadherin-1,and Zo-1)(all P<0.05).Transcriptomic analysis showed that both LSS and ES activated chemokine-related pathways and enriched leukocyte migration and neutrophil recruitment pathways.Further validation revealed upregulation of CXCL2 and MMP12 in the LSS group,whereas CXCL3 and MMP12 were predominantly elevated in the ES group.Conclusion:Both LSS and ES can induce sustained lung injury and neutrophil infiltration in mice,though the underlying molecular mechanisms differ.Compared with ES,exposure to LSS additionally triggers a transient eosinophilic response,suggesting that lunar dust particles possess stronger immunostimulatory potential and higher biological toxicity.
作者
龚晓晓
何诗月
陈一霄
刘祎伟
程琦云
陈娅
胡新月
王振兴
谢辉
GONG Xiaoxiao;HE Shiyue;CHEN Yixiao;LIU Yiwei;CHENG Qiyun;CHEN Ya;HU Xinyue;WANG Zhenxing;XIE Hui(Department of Respiratory and Critical Care Medicine,National Key Clinical Specialty,Branch of National Clinical Research Center for Respiratory Disease,Xiangya Hospital,Central South University,Changsha 410008;Department of Orthopedics,Xiangya Hospital,Central South University,Changsha 410008;Movement System Injury and Repair Research Center,Xiangya Hospital,Central South University,Changsha 410008;Department of Pathology,Xiangya School of Basic Medical Sciences,Central South University,Changsha 410078;Hunan Key Laboratory of Angmedicine,Changsha 410008;National Clinical Research Center for Geriatric Disorders(Xiangya Hospital),Changsha 410008,China)
出处
《中南大学学报(医学版)》
北大核心
2025年第8期1306-1319,共14页
Journal of Central South University (Medical Science)
基金
湖南省科技厅湖湘青年英才项目(2023RC3075)。
