Abstract
Beta-adrenergically-stimulated sweat rates determined by evaporimetry or by sweat bubble imaging are useful for measuring CFTR function because they provide a near-linear readout across almost the full range of CFTR function. They differentiate cystic fibrosis (CF) subjects from CF carriers and carriers from controls. However, evaporimetry, unlike bubble imaging, appears to be unable to detect improved levels of CFTR function in G551D subjects taking the CFTR modulator ivacaftor. Here, we quantify the sensitivity of evaporimetry and bubble imaging methods for assessing low levels of CFTR-dependent sweat rates. To establish sensitivity, we did dose-ranging studies using intradermally injected [cAMP]i–elevating cocktails. We reduced isoproterenol/aminophylline levels while maintaining a high level of atropine to block muscarinic elevation of [Ca2+]i. We stimulated the same sets of glands for both assays and recorded responses for 20 min. In response to a 3-log dilution of the stimulating cocktail (0.1%), bubble responses were detected in 12/12 tests (100%), with 49% ± 3% of glands secreting to produce an aggregate volume of 598 nl across the 12, 20-min tests. This was ~5% of the response to full cocktail. Evaporimetry detected responses in 3/12 (25%) tests with an aggregate secretion volume of 175 nl. After stimulation with a still more dilute cocktail (0.03%), bubble imaging detected 15 ± 13% of glands secreting at a rate ~0.9% of the response to full cocktail, while zero responding was seen with evaporimetry. The bubble imaging method detected secretion down to aggregate rates of <0.2 nl/(cm2·min), or ~1/30th of the average basal transepithelial water loss (TEWL) in the test subject of 4 g/m2·hr or 6.7 nl/(cm2·min). The increased sensitivity of bubble imaging may be required to detect small but physiologically important increases in secretion rates produced by CFTR modulators.
Highlights
MethodsTwo Methods for Measuring SweatRate several million simple, tubular glands that express CFTR in both the absorptive duct and secretory coil
The eccrine sweat gland is a near-ideal organ for assessing the function of CFTR—the anion channel that causes cystic fibrosis (CF) when absent or defective [1]
The sweat chloride level is a sensitive indicator for detecting improved CFTR function in response to CFTR modulators [4], and recent exhaustive work to track sources of variation in the sweat test determined that most variation is due to variation of CFTR [5]
Summary
Two Methods for Measuring SweatRate several million simple, tubular glands that express CFTR in both the absorptive duct and secretory coil. Elevated NaCl concentration in sweat was the earliest consistent method for diagnosing CF [3] and is buttressed by decades of experience in virtually all CF centers worldwide. Sweat chloride values change as a log function of CFTR activity, providing a highly sensitive indicator of very low levels of CFTR function, but becoming increasingly less sensitive at higher values [6]. We used a single male heterozygote subject (EB01) for dose-ranging doi:10.1371/journal.pone.0165254.g001 experiments. This subject has tattooed sites (~1 mm dots) to facilitate gland identification and has been repeatedly tested in the past so that extensive reference data are available for the population of glands selected. Cystic fibrosis: lessons from the sweat gland.
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