Over the past decade, evidence has accumulated describing a rapid and non-genomic steroid hormone response (seconds-minutes) that involves novel membrane-associated receptors and fast activation of signal transduction pathways. One of the physiological roles of the non-genomic response is to increase electrolyte absorption and inhibit secretion in pluripotential epithelia. Steroid hormones such as glucocorticoids, mineralocorticoids and estradiol are involved in this anti-secretory effect. In upper airway epithelia, aldosterone and dexamethasone reduce intracellular calcium mobilization, whereas in the distal colon, estradiol inhibits basolateral K⁺ channels to affect an anti-secretory response. All three steroid hormones activate the cAMP-dependent signal transduction pathway. Previous studies have demonstrated that aldosterone and estradiol down-regulate trans-epithelial chloride secretion and inhibit the action of secretagogues such as carbachol and forskolin via PKC, PKA, COX, Ca²⁺ and pH_i dependent signalling mechanisms in distal colon and lung. (Doolan et al. 2000).

The aim of this study was to compare the rapid effects of aldosterone, dexamethasone and estradiol on PKA and Ca²⁺ signalling response in lung epithelia.

The PepTag Assay^TM for non-radioactive detection of cAMP-dependent protein kinase (PKA) was optimised to detect steroid-induced rapid up regulation of PKA activity in cultured human bronchial (16HBE14o-) and tracheal (CFTE-29o-) epithelia. Intracellular calcium concentration ([Ca²⁺]_i) changes were measured in single cells using Fura-2 spectrofluorescence.

In CFTE cells, low concentrations of aldosterone (1 nM) and dexamethasone (10 nM) caused a rapid reduction of [Ca²⁺]_i. We show for the first time that estradiol (1 nM) similarly caused a rapid reduction of [Ca²⁺]_i in CFTE cells. Rp-cAMP[S], the cAMP antagonist, inhibited the aldosterone, dexamethasone and estradiol effects on [Ca²⁺]_i. Aldosterone caused decrease in basal Ca²⁺ as measured by a reduction in the F340/ F380 ratio of 0.44 ± 0.06, n = 3, mean ± S.E.M., P < 0.001. [Fishers t test]. Similar reductions in calcium were found for dexamethasone and estradiol. In both bronchial and tracheal epithelial cells PKA activity was up-regulated 3-fold over basal levels in less than 5 mins in response to low concentrations of aldosterone (1 nM) and glucocorticoids (10 nM). RP-cAMP[S] inhibited the dexamethasone activation of PKA in both epithelia.

In conclusion, we have shown rapid reduction in calcium and activation of PKA in cultured epithelia from normal human bronchial and CF tracheal airway. Intracellular calcium is a potent regulator of secretion in the lung and the anti-secretory effects of steroid hormones may be explained, in part, by a decreased calcium mobilization. Rapid non-genomic effects of estrogen on protein kinase and Ca²⁺ signalling in human lung epithelia may have relevance for the progression of cystic fibrosis lung disease, as it has been shown that female CF patients have a more severe lung disease progression than male patients (Stallings, 2003). It has been reported that during puberty, female CF patients have a marked exasperation of lung disease. (Fredriksen, 2001) which may be related to elevated estrogen levels.