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

骨髓移植嵌合小鼠哮喘模型的分析方法：用骨髓移植的嵌合C57BL/6小鼠制作小鼠哮喘模型。在卵白蛋白(OVA)致敏后2周，每隔一天暴露于OVA，共1个月。最后一次染毒24小时后，分别于第7天、第14天和第28天采集血和支气管肺泡灌洗液(BAL)，取肺组织。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，观察组织学变化。肺组织用甲醛固定，石蜡包埋。在组织切片上，苏木精-伊红染色半定量评价气道壁炎性细胞的浸润程度。结果：OVA暴露后第7天，嵌合小鼠的支气管周围和肺泡组织中可见以嗜酸性粒细胞为主的淋巴细胞和单核细胞。第14天和第28天，上皮基底膜增厚，粘膜下层胶原沉积明显。上皮下部分表达GFP的细胞呈梭形，呈成纤维细胞样细胞。免疫组织化学结果显示，G-CSF治疗组小鼠肺组织中GFP表达细胞较对照组明显增多，弹性蛋白-马森染色结果显示，G-CSF治疗组小鼠肺组织中胶原沉积明显减少。结论：骨髓来源细胞在变态反应性炎症小鼠的支气管周围组织中广泛存在，可能对呼吸道胶原沉积起到调节作用。

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.