In vivo intravital endoscopic confocal fluorescence microscopy of normal and acutely injured rat lungs
In vivo intravital endoscopic confocal fluorescence microscopy of normal and acutely injured rat lungs
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DOI：
10.1038/labinvest.2010.76
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发表时间：
2010-06-01
影响因子：
5
通讯作者：
Lesur, Olivier
中科院分区：
文献类型：
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作者：
Chagnon, Frederic;Fournier, Clement;Lesur, Olivier
我们提出了一种新的基于内窥镜共聚焦荧光显微镜（ECFM）的肺成像技术，这是一种新的方法，能够通过直接接触内脏胸膜的小共聚焦探针提供活组织的细胞和结构评估。为了观察正常和损伤肺（高氧和博来霉素挑战）的远端空域结构和细胞状况，我们使用荧光特异性标记对比和ECFM。使用fitc标记物或天然荧光染料在488 nm激发下对肺泡空间ECFM进行光谱分析。将正常肺与患病肺进行比较，我们的体内成像实验与相应的体外常规检测结果相吻合。我们专门研究了与急性肺损伤/急性呼吸窘迫综合征（ALI/ARDS）病理生理相关的四个主要因素和确定的早期关键事件：肺泡上皮膜表型、肺细胞凋亡、中性粒细胞募集和水肿。ECFM可以可视化(i)微调超微结构凝集素（RCA-1）和唾液糖蛋白（RTI40）上皮细胞膜表达，（ii） yo - pro -1相关DNA连接肺细胞凋亡，（iii） PKH2绿色荧光细胞连接物标记的中性粒细胞在肺微循环网络和空气空间中的跟踪，（iv） fitc -葡聚糖血浆对比和水肿形成的外渗。ECFM为相应的离体荧光方法提供了可靠的结果。ECFM使用微创5 -1(R)光学仪器和特异性荧光标记物，能够提供实时的潜在有用的活体未固定正常和损伤肺组织成像，在改善急性肺损伤患者的床边诊断和决策治疗策略方面有很好的发展。实验室调查（2010）90,824-834；doi: 10.1038 / labinvest.2010.76;2010年4月12日在线发布
We present a new lung imaging technique based on endoscopic confocal fluorescence microscopy (ECFM), which is a new method that is able to provide cellular and structural assessment of living tissue using a small confocal probe in direct contact with the visceral pleura. To observe distal airspace structure and cellular condition in normal and injured lungs (hyperoxic and bleomycin challenged), we used fluorescent-specific marker contrast and ECFM. Alveolar space ECFM with spectral analyses were performed at 488-nm excitation using FITC-labeled markers or naturally fluorescent dyes. The normal lung was compared with the sick lung, where our in vivo imaging experiments correlated well with results obtained with corresponding ex vivo conventional assays. Four main elements pertaining to the acute lung injury/acute respiratory distress syndrome (ALI/ARDS) pathophysiology and established early key events were specifically studied: alveolar epithelial membrane phenotype, lung cell apoptosis, neutrophil recruitment, and edema. ECFM allowed visualization of (i) fine-tuned ultrastructural lectin (RCA-1) and sialoglycoprotein (RTI40) epithelial cell membrane expression, (ii) YO-PRO-1-related DNA linking of lung cell apoptosis, (iii) PKH2 green fluorescent cell linker-labeled neutrophil tracking in lung microcirculatory network and airspaces, (iv) FITC-dextran plasma contrast and extravasation with edema formation. ECFM provides reliable results to corresponding ex vivo fluorescent methods. ECFM, using the minimally invasive Five-1(R) optical instrument and specific fluorescent markers, is able to provide real-time potentially useful imaging of live unfixed normal and injured lung tissue with promising developments for improving bedside diagnostic and decision-making therapeutic strategy in patients with ALI. Laboratory Investigation (2010) 90, 824-834; doi:10.1038/labinvest.2010.76; published online 12 April 2010