Analysis of bone-marrow derived fibroblasts involved in asthmatic airway remodeling and its interventional control of function

方法：用骨髓移植嵌合C57BL/6小鼠建立小鼠哮喘模型。在卵清蛋白（OVA）致敏后两周，小鼠在一个月内每隔一天暴露于OVA。在最终暴露于OVA 24小时后，我们采集了这些小鼠的血液和支气管肺泡灌洗液（BAL），并在第7天、第14天和第28天采集了肺组织。此外，我们将这些小鼠分为两组。一组小鼠腹腔注射G-CSF，另一组小鼠腹腔注射生理盐水作为对照。对组织学变化进行估计。肺组织用甲醛固定，石蜡包埋。在组织切片上，我们用苏木精-伊红染色估计浸润到气道壁的炎症细胞作为半定量评分。我们在化学发光显微镜下鉴定了支气管组织中表达GFP的细胞，并用抗GFP抗体免疫组化方法鉴定了GFP阳性细胞。结果：OVA暴露后第7天，嵌合小鼠支气管周围和肺泡组织中以嗜酸性粒细胞为主细胞、淋巴细胞和单核细胞为主。在14^<th>和28^<th>时，支气管上皮基底膜增厚，粘膜下层胶原沉积明显。部分表达gfp的细胞在上皮下呈梭形，呈成纤维细胞样。免疫组化证实。与对照组相比，G-CSF处理小鼠肺中表达gfp的细胞增加更多，而用Elastica-Masson染色估计的G-CSF处理小鼠的胶原沉积与对照组相比减少。结论：骨髓源性细胞可向变应性炎症的支气管周围组织募集，并可能在气道胶原沉积中起调节作用。

Analysis of the asthma model with bone marrow transferred chimeric miceMethod : We made the murine asthma model with bone marrow transferred chimeric C57BL/6 mice. At two weeks after sensitization with Ovalbumin (OVA), the mice were exposed to OVA every other day during one month. Twenty four hours later from final exposure with OVA, we collected blood and broncho-alveolar lavage fluid (BAL) and obtained the lung tissue from these mice on 7^<th> day, 14^<th> day and 28^<th> day. In addition, we divided these mice into two groups. One group of the mice was administered intraperitoneally with G-CSF and the other group with saline as the control.Estimation of histological changes was performed. The pulmonary tissue was fixed with formaldehyde and were embedded in paraffin. On the tissue sections, we estimated infiltrated inflammatory cells into airway wall as a score semi-quantitatively with Hematoxylin-Eosin staining. We identified GFP-expressing cells in bronchial tissues under chemiluminescence microscopy and also GFP positive cells with immunohistochemistry using ant-GFP antibody.Results : On 7^<th> day after OVA exposure, eosinophils as major cells, lymphocytes and monocytes were observed in peribronchial and alveolar tissue in the chimeric mice. On 14^<th> and 28^<th> day, thickening of epithelial basement membrane and deposition of collagen in submucosal layer of bronchi became prominent. Some GFP-expressing cells were spindle-like shape under epithelium and assumed fibroblast like cells. They were also confirmed by immunohistochemistry.GFP-expressing cells increased more in the lung of the mice with G-CSF treatment than control, and collagen deposition estimated with Elastica-Masson staining decreased in G-CSF treated mice compared to control.Conclusion : Bone marrow derived cells were recruited to peribronchial tissue with allergic inflammation and might play a regulatory role in collagen deposition in the airway.