Hyperosmolar Solutions Stimulate Mucus Secretion in the Ferret Trachea: Statistical Analysis

Morphologic Evaluation of the Tracheal Epithelium
Tracheal segments (n = 4 each) were taken before and after immersion in 3.69 g/dL of saline solution (1,192 mOsm), 10.69 g/dL of saline solution (3,612 mOsm), or in KHS (288 mOsm). Tissue segments were immersed in 10% formalin and processed for light (wide-field) microscopy. Paraffin-embedded tissue was cut to 4-^m thickness, and slides were stained with hematoxylin and eosin. These were examined by a trained investigator masked as to treatment group (J.S.K.). He randomly selected eight fields for evaluation of epithelial damage.
Statistical analysis of data were performed using the StatView 5 statistics package (SAS Institute; Cary, NC). Data were analyzed by analysis of variance Scheffe F test or Mann-Whitney U test to assess secretory response and the effect of inhibitors. Studies were powered to detect a 20% difference in measured secretion with a power of 0.85. Conventionally, p values < 0.05 were considered statistically significant. All data are presented as means ± SEM. more

Results
Mucin and Lysozyme Secretion

There was a dose-related increase in mucin secretion compared to KHS (0.69 g/dL of saline solution [288 mOsm/L], 3,572 ± 291 ng/g secreted mucin, n = 16) after incubation with 1.69 g/dL of saline solution (597 mOsm/L) [6,264 ± 520 ng/g, 77% increase, n = 7; p < 0.01 for SI]; 3.69 g/dL of saline solution (1,192 mOsm) [7,227 ± 516 ng/g, 106% increase, n = 16; p < 0.0001]; 5.69 g/dL of saline solution (1,823 mOsm) [7,440 ± 490 ng/g, 116% increase, n = 11; p < 0.0001]; and 10.69 g/dL of saline solution (3,612 mOsm) [7,695 ± 389 ng/g, 130% increase, n = 10; p < 0.0001]. Mucin secretion was also significantly increased after exposure to 15 g/dL of mannitol (1,040 mOsm) [8,415 ± 606 ng/g, 135% increase, n = 4; p < 0.005; Fig 1].

Fig1
Figure 1. The effect of saline or mannitol solutions on mucin secretion. After 2 h of resting in KHS, ferret tracheal segments were incubated for 20 min with KHS (288 mOsm/L) [period 1], and then incubated for another 20 min in a hyperosmolar or KHS solution (period 2). An SI expressing the relative increase in mucin was calculated for each tracheal segment as the concentration of period 2 divided by that of period 1. The increased mucin secretion is presented as percentage increase compared to KHS. Data are presented as mean ± SEM. There was a dose-related increase in mucin secretion after incubation with 1.69 g/dL (597 mOsm/L) [n = 7, p < 0.01], 3.69 g/dL (1,192 mOsm/L) [n = 16, p < 0.0001], 5.69 g/dL (1,823 mOsm/L) [n = 11, p < 0.0001], and 10.69 g/dL (3,612 mOsm/L) [n = 10, p < 0.0001] of saline solution, and with 15 g/dL (1,040 mOsm/L) of mannitol (n = 4, p < 0.005) when compared to KHS control.

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