Intratracheal administration of liposomal clodronate accelerates alveolar macrophage reconstitution following fetal liver transplantation
Intratracheal administration of liposomal clodronate accelerates alveolar macrophage reconstitution following fetal liver transplantation
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DOI：
10.1189/jlb.1203647
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发表时间：
2005-02-01
影响因子：
5.5
通讯作者：
Blackwell, TS
中科院分区：
文献类型：
--
作者：
Everhart, MB;Han, W;Blackwell, TS
为了促进肺泡巨噬细胞在体内的研究，我们开发了一种方法，快速，有效地取代居民肺泡巨噬细胞与不同的（供体）基因型的巨噬细胞。通过致死性照射产生嵌合小鼠，然后使用绿色荧光蛋白（GFP）转基因报告小鼠作为供体进行胎肝移植（FLT）。外周血单核细胞（PBM）和肺泡巨噬细胞重建的动力学在FLT后4周和10周通过定量GFP+细胞的百分比来确定。为了增强FLT后供体单核细胞向肺中的募集，在FLT后6周用脂质体氯膦酸盐的气管内施用处理小鼠以消耗宿主肺泡巨噬细胞。在FLT后4周发生PBM重建（85.7 +/-1.6%CD11b +/Gr-1+单核细胞为GFP+），并且观察到最小肺泡巨噬细胞再增殖（9.5%GFP+）。FLT后10周，通过肺组织切片上巨噬细胞的免疫染色，48%的肺泡巨噬细胞为GFP+，通过荧光素活化细胞分选仪分析，55.1 +/- 1.6%的肺灌洗液巨噬细胞为GFP+。氯膦酸盐治疗导致FLT后10周GFP+肺泡巨噬细胞显著增加。通过免疫染色，肺组织切片上90%的巨噬细胞为GFP+，肺灌洗液中87.5 +/- 1.1%为GFP+（与GFP转基因对照相比）。新招募的肺泡巨噬细胞清除铜绿假单胞菌和激活核因子-κ B的能力，以响应土方大肠杆菌脂多糖证明正常的巨噬细胞功能。优化这种方法提供了一个重要的工具，研究特定的基因和它们的贡献肺泡巨噬细胞功能在体内。
To facilitate study of alveolar macrophages in vivo, we developed a method to rapidly and efficiently replace resident alveolar macrophages with macrophages of a different (donor) genotype. Chimeric mice were generated by lethal irradiation followed by fetal liver transplantation (FLT) using green fluorescent protein (GFP) transgenic reporter mice as donors. Kinetics of peripheral blood monocyte (PBM) and alveolar macrophage reconstitution was determined 4 and 10 weeks post-FLT by quantifying the percentage of GFP+ cells. To enhance the recruitment of donor monocytes into the lung after FLT, mice were treated with intratracheal administration of liposomal clodronate to deplete host alveolar macrophages at 6 weeks post-FLT. PBM reconstitution occurred by 4 weeks after FLT (85.7 +/- 1.6% of CD11b+/Gr-l+ monocytes were GFP+), and minimal alveolar macrophage repopulation was observed (9.5% GFP+). By 10 weeks following FLT, 48% of alveolar macrophages were GFP+ by immuno-staining of macrophages on lung tissue sections, and 55.1 +/- 1.6% of lung lavage macrophages were GFP+ by fluorescein-activated cell sorter analysis. Clodronate treatment resulted in a significant increase in GFP+ alveolar macrophages 10 weeks after FLT. By immunostaining, 90% of macrophages were GFP+ on lung tissue sections and 87.5 +/- 1.1% GFP+ in lung lavage (compared,with GFP-transgenic controls). The ability of newly recruited alveolar macrophages to clear Pseudomonas aeruginosa and activate nuclear factor-kappaB in response to Eschericia coli lipopolysaccharide demonstrated normal macrophage function. Optimizing this methodology provides an important tool for the study of specific genes and their contribution to alveolar macrophage function in vivo.