ISSN:
1432-2013
Schlagwort(e):
αhENaC
;
Airway epithelial cells Amiloride
;
CFTR
;
Patch clamp
;
RT-PCR
Quelle:
Springer Online Journal Archives 1860-2000
Thema:
Medizin
Notizen:
Abstract According t0 previous studies, amiloride-sensitive (Amil+) Na+ channels are present in apical membranes of airway epithelial cells. When isolated from intact tissue and grown in primary culture 0r as immortalized cell lines, these cells tend t0 lose these Amil+ Na+ channels. The present study examines this issue in immortalized human bronchial epithelial cells (16HBE140- cell line). The mRNA of one subunit of the Na+ channel (αhENaC) was semi-quantified by polymerase chain reaction of reverse transcribed RNA. Transcripts were significantly increased when cells were exposed t0 aldosterone and dexamethasone irrespective of whether grown on permeable supports 0r plastic. When grown on plastic dishes 16HBE140-cells showed cAMP-dependent Cl− currents in whole-cell (WC) patch-clamp experiments, corresponding t0 expression of the cystic fibrosis transmembrane conductance regulator (CFTR). Na+ currents could not be detected although cells expressed significant amounts ofαhENaC as demonstrated by Northern blot analysis. In contrast, when cells were grown on permeable supports 0r cultured in the presence of butyrate (5 mmol/l, plastic 0r permeable support) 0r aldosterone and dexamethasone (both 1 μol/l, plastic 0r permeable support), amiloride (10 μmol/1) hyperpolarized the membrane voltage (ΔVm) by 2–9 mV paralleled by small reductions of WC conductances (ΔGm) of 0.4-4.0 nS. The effects of amiloride on ΔVm were generally more pronounced (up t0 12 mV) when cells were grown on permeable supports. The amiloride effect (ΔVm) was concentration dependent with an inhibitory constant, Ki, of about 0.1 μmol/l. We further examined whether the induction of an Amil+ Na+ conductance was paralleled by additional changes in membrane conductance. In fact, the cAMP-activated Cl− conductance was significantly attenuated by approximately 80% (n = 35) in cells responding t0 amiloride, whilst the ATP-activated K+ conductance remained unaffected. The present data suggest that cellular mechanisms determining differentiation control the functional expression of Na+ and Cl− conductances in human airway epithelial cells.
Materialart:
Digitale Medien
URL:
http://dx.doi.org/10.1007/BF02191906
