P13K在细胞外基质分泌细胞因子表达中的作用

目的 探讨纤维连接蛋白(FN)、Ⅰ型胶原蛋白(Col Ⅰ)两种细胞外基质(ECM)成分对被动致敏的人气道平滑肌细胞(HASMCs)免疫功能的影响及磷脂酰肌醇-3激酶(PDK)在此调控中的作用.方法 将HASMCs接种于FN、Col Ⅰ包被的培养板和空白培养板中,用10%哮喘患者血清被动致敏HASMCs,以10%非哮喘者血清为对照,在加入血清前用PDK抑制剂(LY294002)预处理HASMCs 30 min.采用RT-PCR法检测HASMCs RANTES(regulated upon activation.normal T cell ex-pressed and secreted)、Eotaxin、TGF·β1 mRNA表达;ELISA法测定HASMCs培养上清中RANTES、Eotax-in、TGF-β1蛋白水平.结果 与单纯对照血清组比较,单纯哮喘血清组、对照血清+FN组、对照血清+Col Ⅰ组HASMCs RANTES、Eotaxin、TGF-β1 mRNA和HASMCs培养上清中蛋白的表达均增高(P<0.05).哮喘血清+FN、哮喘血清+Col Ⅰ处理HASMC后,RANTES、Eotaxin、TGF-β1mRNA和HASMCs培养上清中蛋白的表达均高于单纯哮喘血清组,且LY294002干预后上述各指标均下降(P<0.05).结论 细胞外基质对被动致敏的HASMCs免疫功能具有调控作用,PI3可能参与细胞外基质对被动致敏的HASMCs的免疫功能调控.

The role of PDK in regulating cytokine secretion by extracellular matrixes

Objective To investigate the regulatory effects of extracellular matrixes, including fi-bronectin(FN), and collagen Ⅰ (ColⅠ ) on the immunologic function of human airway smooth muscle cells (HASMCs) passively sensitized with asthmatic serum, and the role of phosphoinosifide 3-kinase (PI3K). Methods Primarily cultured HASMCs were inoculated on the blank plates or on the plates coated with difference matrix proteins, added 10% asthmatic serum to passively sensitized non-asthmatic HASMCs and 10% non-asthmatic serum treated HASMCs as control, cell pretreated with PI3K inhibitor LY294002 for 30 min. The expressions of RANTES, Eotaxin, TGF-β1 mRNA were observed by RT-PCR and RANTES, Eotaxin, TGF-β1 protein in the ceil culture supernatunts was detected by enzyme-linked immtmosorbent assay (ELISA). Results Compared with the control serum group, the expressions of RANTES, Eotaxin, TGF-β1 mRNA of HASMCs and those protein in HASMCs culture supematants were significantly increased in the asthmatic serum group and the control serum + FN group and the control serum + Col Ⅰ group ( P < 0.05 ). The expressions of RANTES, Eotaxin, TGF-β1 mRNA of HASMCs and protein in HASMCs culture superna-tants were significantly increased in the asthmatic serum + FN group and the asthmatic serum + Col Ⅰ group. 50 μmoL/L LY294002 could significantly inhibit the expressions of RANTES, Eotaxin, TGF-β1 mRNA of HASMCs and protein in HASMCs culture supematants(P < 0.05). Conclusion These results suggest ex-tracellular matrixe may regulate immunomodulatory function of HASMCs passively sensitized with asthmatic serum and PI3K signaling pathway may play an important role in the process.