In this issue of The Journal,
The use of fluorescent techniques to infer the composition of ASL is a tremendous leap forward in technology, avoiding for the first time the pitfalls associated with techniques based on harvesting ASL or on in situ measurements. The mainstay of ASL studies over the years has been the application of filter paper to the surface of the epithelium (
The elegant but simple system of osmotically sensitive liposomes reported in the present study seems to be well suited to measurements of ASL osmolality. The fluorophore used is stable, and the liposomes are sufficiently small so as not to disturb the ASL while being able to distribute homogeneously though it. The authors have demonstrated that the method is sufficiently sensitive to detect the effects of evaporation or addition of salt on ASL osmolality. The results appear to be both reproducible and consistent in cell culture and in vivo models. A key advantage over previous techniques based on harvesting ASL, salt-sensitive electrodes, and studies of rapidly frozen airways, is the ability to conveniently follow changes in ASL osmolality over time. For the first time, the rate at which intact epithelium responds to stresses like changes in osmolality can be measured directly, thereby providing important insights into the dynamic properties of the airway epithelium. Jayaraman and co-workers (2001) have exploited this capability to estimate the permeability of epithelial cells to water, yielding an independent estimate of epithelial water permeability that is in general agreement with previous results (
The results of greatest interest in this study are those carried out in vivo. The authors have taken advantage of the small size of mice to extend fluorescence microscopy to the airway epithelium of intact animals, which is an important advance. Nevertheless, results from these studies must be interpreted cautiously. The fluorescent techniques developed in the Verkman lab have been described as a noninvasive approach to measure ASL composition in mice. Certainly this approach perturbs the epithelium less than techniques involving direct contact with the epithelial surface; however, it is probably better characterized as minimally invasive rather than noninvasive. The current method involves a midline incision in the neck, isolation of the trachea, as well as the oral insertion of a tracheal microcatheter for instillation of liposomes. One possible consequence of these manipulations is the triggering of an epithelial leak from the submucosa, leading to the introduction of plasmalike material into the ASL. This possibility was effectively excluded by experiments in which a small window in the trachea was created and no discernible effects on the results were detected. Similarly, the authors report that no leak was detected after injection of FITC-labeled macromolecules, which has been used previously to detect leak in the airways of allergen-challenged animals (
Although there has been some interest in the composition and regulation of ASL in other diseases (
The first hypothesis, termed the "isosmotic volume hypothesis" (
The alternate hypothesis emphasizes the role of ASL composition in the passive defense against bacterial infection.
Recently, the high salt hypothesis has been challenged on several levels. Despite the consistent finding of low salt concentrations in ASL under normal circumstances, the finding of higher [Na+] in the ASL of CF patients (
Although the above suggests that the high salt theory is on the ropes, does this mean that the low volume hypothesis is now confirmed? The work of Jayaraman and colleagues raises challenges for the low volume hypothesis as well. Although the new finding of near isotonic osmolality of ASL in both CF knockout mice and controls, as well as the previous observation that ASL [Na+], [Cl-], and pH are similar in normal and CF (
If the findings of Jayaraman are inconsistent with the two most popular hypotheses to explain CF lung disease, what alternatives are suggested by these data? First, it is important to stress that the in vivo fluorescence studies so far have been limited to mice. It remains possible that conditions in human airways are different, although the results using bronchial segments (
One option is to revisit the importance of CFTR in airway secretion, rather than absorption (
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