A new technique of bronchial microsampling and proteomic analysis of epithelial lining fluid in a rat model of acute lung injury

作者：Kamiyama Ikuo; Kohno Mitsutomo^*; Kamiya Kazunori; Nakamura Hidetoshi; Sawafuji Makoto; Kobayashi Koichi; Watanabe Masazumi

来源：Molecular Immunology, 2014, 59(2): 217-225.

DOI：10.1016/j.molimm.2014.02.013

• Rat bronchial microsampling
• Epithelial lining fluid
• Bronchoalveolar lavage
• Lipopolysaccharide stimulation
• Proteomic analysis

The standard technique to evaluate the proteins present in epithelial lining fluid (ELF) is bronchoalveolar lavage (BAL). Bronchoscopic microsampling (BMS) method has been developed for humans as a less invasive alternative. We establish the usefulness of a rat bronchial microsampling (rBMS) to evaluate various proteins in ELF in lipopolysaccharide (LPS)-induced lung injury models in rats. In the first experiment of this study, we validate that whether the rBMS can obtain information from ELF in place of BAL. Tumor necrosis factor (TNE)-alpha concentrations were increased in the rBMS samples similar to BAL 1 and 3 h after LPS instillation. In the second part of this study, a proteomic analysis of the rBMS, using the Protein Chip(R) system, revealed the presence of proteins whose molecular weights corresponded to TNF-related proteins in the LPS-treated rats. In rats treated with a TNF-alpha converting enzyme inhibitor, the concentrations of these proteins in rBMS decreased or disappeared. In the third experiment, rBMS was performed without tracheostomy at 6 and 24h after instillation of LPS, and a rat multiple cytokines assay system detected heterogeneous variations in the concentrations of interleukin (IL)-1 alpha, IL-1 beta, IL-2, IL-4, IL-6, IL-10, TNF-alpha and interferon (IFN)-gamma in ELF. The cytokine profile was significantly modified by pre-treatment with dexamethasone. This new rBMS technique could be used to measure TNF-alpha. in LPS-induced acute lung injury (ALI) as well as for proteomic analysis, without sacrificing the rats. Furthermore, this procedure enables the serial collection of ELF, which would allow the examination of time-dependent cytokine variations in rat ALI model.

• 出版日期2014-6