Recurrent aspiration of refluxed material is an important cause of chronic respiratory illness, especially in children. Currently available diagnostic tests such as the lipid-laden macrophage index, milk scan and barium esophagogram lack sensitivity and specificity. Therefore, there is a need for a more accurate diagnostic test. We hypothesized that immunocytochemical detection of pepsin in alveolar macrophages obtained by bronchoalveolar lavage would serve as a useful diagnostic tool for reflux-associated aspiration.
To test our hypothesis we exposed anaesthetized BALB/c mice to a single aspiration of 25 μl of human gastric juice and lavaged their lungs with saline 2, 4, 6, 12, 24, and 48 hours following aspiration. Control animals received same amount of saline aspiration in a similar fashion. Cells obtained by lung lavage was used to prepare cytospin slides. These slides were then stained immunochemically by using goat anti-porcine pepsin antibody and were examined under a light microscope. Three negative controls were used: (1)cells obtained after aspiration of saline; (2)cells obtained after aspiration of human gastric juice and stained without the primary antibody; and (3)cells incubated with isotype-matched goat IgG. Hundred consecutive cells were counted under high-power field and number of positively stained cells was recorded for each time point. Results were expressed in terms of mean percent positive cells (±SD).
At 2, 4, 6, 12, 24 and 48 hours after a single aspiration of gastric juice, mean percent positive staining(±SD) was 4.5±3.0, 2.8±1.3, 16.3±5.1, 16.0±1.4, 24.8±9.0, 33.5±9.0, respectively, (n=4,at each time point). There was no staining in any of the controls(n=6). Differences between the controls and values detected at all time points were statistically significant (p<0.05).
These findings demonstrate that pepsin can be detected in alveolar macrophages by using immunocytochemistry up to, at least, 48 hours after a single aspiration event.
Immunochemical staining of alveolar macrophages for pepsin has the potential to become a simple, routine test to detect reflux-related aspiration accurately, if it’s validity is confirmed in clinical studies.
Haibin Zhang, None.