A method to enable the investigation of murine bronchial immune cells, their cytokines and mediators

Abstract

Innovative therapies for severe lung diseases (such as allergic and chronic asthma, chronic obstructive pulmonary disease or any type of lung cancer) require a detailed understanding of the cellular and immune processes in the lung. This protocol details a method to obtain the immune cells of the bronchi as well as the cytokines and mediators produced by these cells for further investigation. The broncho-alveolar lavage fluid (BALF) is taken by injecting physiological solution through the tracheal tube into the murine airways and carefully regained by winding up the connected syringe. After centrifugation, the resulting BALF supernatant can be stored for detection of cytokines or other mediators by enzyme-linked immunosorbent assay or other methods; the resuspended cell pellet can also be used for flow cytometric analyses, to check cell viability and the level of apoptosis, as well as other applications. In addition, CD4+ T cells isolated from wild-type and genetically modified mice alone or along with other immunologically important cells such as T regulatory cells, which can be used to reconstitute immunodeficient mice, may be retrieved from the airways with this method. This protocol can be completed within 35 min.

Dual Role of Interleukin-1α in Delayed-Type Hypersensitivity and Airway Hyperresponsiveness

“…Next, BAL fluid was obtained after OVA aerosol challenge of IL-1α-treated and control mice on day 30 [20]. Both the cellular composition (fig.…”

“…After assessment of the AHR, bronchoalveolar lavage (BAL) was performed [20]. Mice were euthanized by an intraperitoneal injection of 5% ketamine-HCl (Ratiopharm) and 2% xylacin (Rompun®, Bayer) at a ratio of 1:1.…”

Protective role of nuclear factor of activated T cells 2 in CD8+ long-lived memory T cells in an allergy model

“…BALF supernatants were frozen and subsequently analyzed by ELISA and cell pellet resuspended in PBS and used for fluorescence-activated cell sorting (FACS) analysis as previously described. 8 Histology Lungs were removed, fixed in 10% formalin-PBS solution, and embedded in paraffin. Sections 5 mm thick were stained either with hematoxylin and eosin for quantification of inflammation or with Goldner stain for the quantification of peribronchial collagen deposition.…”

Aspiration techniques for bronchoalveolar lavage in translational respiratory research: Paving the way to develop novel therapeutic moieties

“…Maxeiner and co-workers standardized a method in murine model of lung carcinoma and melanoma which assists in understanding and analyzing various cells and released mediators in natural state [26]. Analyzing BALf is one of the prime parameters to understand the underlying disease pathology in prominent mice models of major respiratory diseases, including asthma [27][28][29], COPD [30,31], and pulmonary fibrosis [32].…”