轻度哮喘患者BALF中活化肌成纤维细胞与气道重塑实验研究

Activated mobile fibroblasts in bronchoalveolar lavage from patients with mild asthma:a possible link to airway remodeling

【目的】探讨轻度支气管哮喘患者BALF中活化肌成纤维细胞与气道重塑关系。【方法】对10例支气管哮喘患者及8例正常志愿者行纤支镜支气管黏膜活检及肺泡灌洗,采用Giemsa染色对支气管肺泡灌洗液(bron-choalveolar lavage fuild,BALF)进行分类:中性粒细胞、淋巴细胞、巨噬细胞、嗜酸性粒细胞;培养BALF及活检组织的成纤维细胞,免疫组化方法检测成纤维细胞的特性(脯氨酰-4羟化酶染色,α-平滑肌肌动蛋白染色);采用Western blotting分析其表达胶原蛋白Ⅲ、纤维粘连蛋白的含量、[3H]-脯氨酸渗入评价脯氨酸的含量。【结果】30%的哮喘患者的BALF能分离和培养出成纤维细胞,而对照组中未发现成纤维细胞,发现成纤维细胞的哮喘患者的嗜酸性细胞的比例比对照组增加9.5倍,比未发现成纤维细胞的哮喘患者增加4.6倍;BALF和活检的成纤维细胞均被成纤维细胞标记脯氨酰-4羟化酶染色,而且BALF和活检的成纤维细胞均被肌成纤维细胞标记α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)染色,而且BALF成纤维细胞比活检成纤维细胞表达更多的α-SMA;BALF成纤维细胞较活检成纤维细胞分泌的胶原蛋白Ⅲ增加至2.87倍(P<0.01)、纤维连接蛋白分别增加至1.9倍(P<0.01)。脯氨酸渗入比较增加至4.24倍(P<0.05)。【结论】轻度哮喘患者的BALF中活化肌成纤维细胞参与多炎性反应,与气道重塑有密切的关系。

[ Objectlve] To evaluate the links of activated mobile fibroblasts in bronchoalveolar lavage from patients with mild asthma to airway remodeling. [Methods] Patients with mild asthma （ n = 10） and non-asthmatic healthy subjects （ n = 8, continl） were collected. Cytospin analysis of the cellular profile in the BALF was performed by Giemsa staining. Macrophages, lymphocytes, neutmphils, and eosinophils were counted. Colocalization of prolyl 4-hydroxylase and a-smooth actin （α-SMA） were identified in bronchial biopsies from patients and controls by immunohistochemical technique. Using Western Bloting techniques we measured levels of Collagen I[], Fibronectin in BAL fluid and biopsy from patients with mild asthma. In 33 % of patients with asthma could isolated from the BALF and cultured, whereas in control subjects no fibrubalsts were presents. Patients with asthma with fibrobalsts present in the BALF had 9.5 - fold increases of eosinophils in their cellular profile when compared with the control subjects and a 4.6 - fold higher percentages when compared with patients with asthma with no BALF fibroblast. When compared with fibmblasts from corresponding bronchial biopsies from the same patients, The fibroblasts accompanied by a 2.87- fold increase of Collagen m and a 1.9 - fold increase of Fibronectin, a 4.24- fold increase 4.24 [^3H] - Proline Incorporation, indicating that the increased expression of the proteins are linked to deposition of extracellular matrix. In the formation of peribronchial fibrosis in asthma. [ Conclusions] Activated fibroblasts are suggested to be involved in the deposition of extracellular matrix in the formation of pefibronchial fibrosis in asthma. These findings indicate the presence of activated and mobile fibroblasts accompanied by an induced inflammatory response outside the airway epithelium in patients with mild asthma, results that may play a role in formation of airway fibrosis.