Antonini J M, Hemenway D R, Davis G S
Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, West Virginia, USA.
Exp Lung Res. 2000 Mar;26(2):71-88. doi: 10.1080/019021400269880.
Pulmonary fibrosis is a disabling consequence of many lung diseases but is difficult to quantify. Lucifer yellow CH fluorescent dye (LY) appears to stain connective tissue matrix macromolecules selectively. Laser scanning confocal microscopy can quantify the intensity of fluorescence and determine the area of fluorescent material. We hypothesized that the abundance of lucifer yellow-stained matrix macromolecules in lung tissue sections could be measured by laser scanning confocal microscopy, would reflect differences between varying degrees of pulmonary fibrosis, and could be compared directly to biochemical measurements of lung collagen. We exposed C57B1/6 and 129 strains of mice by aerosol to cristobalite silica (70 mg/m3, 12 days, 5 hours/day) or sham-air and examined them 2 and 16 weeks after exposure. The area of LY-stained matrix in tissue sections was quantitated by laser scanning confocal microscopy, and total lung collagen was measured biochemically as hydroxyproline (OH-proline). The LY-stained connective tissue matrix appeared as bright linear bands in the alveolar septae, and was increased significantly by image analysis in C57B1/6 and 129 mice with silicosis 16 weeks after exposure. Total lung OH-proline was significantly increased in silica-exposed mice from both stains at both time points. Comparing all 8 groups, there was a significant linear correlation between the average area of connective tissue measured by LY stain and the total OH-proline per lung measured by chemical analysis (r = .72, P = .042). LY staining and confocal microscopy with image analysis offers a rapid technique for quantitative measurements of the extent of pulmonary fibrosis.
肺纤维化是许多肺部疾病导致的一种致残后果，但难以进行量化。路西法黄CH荧光染料（LY）似乎能选择性地对结缔组织基质大分子进行染色。激光扫描共聚焦显微镜可以量化荧光强度并确定荧光物质的面积。我们推测，通过激光扫描共聚焦显微镜可以测量肺组织切片中路西法黄染色的基质大分子的丰度，它能反映不同程度肺纤维化之间的差异，并且可以直接与肺胶原蛋白的生化测量结果进行比较。我们通过气溶胶让C57B1/6和129品系的小鼠暴露于方石英（70毫克/立方米，12天，每天5小时）或假空气环境中，并在暴露后2周和16周对它们进行检查。通过激光扫描共聚焦显微镜对组织切片中LY染色的基质面积进行定量，并用生化方法将总肺胶原蛋白测量为羟脯氨酸（OH - 脯氨酸）。LY染色的结缔组织基质在肺泡间隔中呈现为明亮的线性条带，在暴露16周后患矽肺的C57B1/6和129小鼠中，通过图像分析其明显增加。在两个时间点，来自两种品系的暴露于二氧化硅的小鼠的总肺羟脯氨酸均显著增加。比较所有8组，LY染色测量的结缔组织平均面积与化学分析测量的每肺总OH - 脯氨酸之间存在显著的线性相关性（r = 0.72，P = 0.042）。LY染色以及结合图像分析的共聚焦显微镜提供了一种快速技术，可用于定量测量肺纤维化的程度。
