Introduction Idiopathic pulmonary fibrosis (IPF) is a common, progressive interstitial lung disease. Current treatments are ineffective. Ion channels are emerging as attractive therapeutic targets and in particular, the Ca2+-activated K+ channel KCa3.1 has been shown to modulate the activity of several structural and inflammatory cells which play important roles in model diseases characterised by tissue remodelling and fibrosis. We hypothesise that KCa3.1-dependant cell processes are a common denominator in IPF.
Aims The aim of the study was to determine whether the KCa3.1 channel is expressed in human lung derived (myo)fibroblasts, key effector cells in IPF.
Methods Human lung (myo)fibroblasts derived from non-fibrotic lobectomy specimens were grown in vitro, and characterised by immunofluorescence. RT-PCR was used to examine KCa3.1 mRNA expression. Western blot was used to confirm the presence of KCa3.1 channel protein. Patch clamp electrophysiology was performed to demonstrate the presence of functional KCa3.1 channels. To elicit KCa3.1 currents the KCa3.1 opener 1-EBIO (0.1 mM) was used.
Results Human lung (myo)fibroblasts, express the KCa3.1 channel at the mRNA level. Western blot demonstrated that the KCa3.1 protein is also present in human lung (myo)fibroblasts. (Myo)fibroblast cell lysates contained immunoreactive protein of approximately 48kD molecular weight, consistent with the reported size of the KCa3.1 channel. Patch clamp electrophysiology demonstrated the presence of ion currents typical of those carried by KCa3.1 channels. These increased in magnitude from passage two through passage seven. Overall, KCa3.1 currents were elicited in 62% of cells tested. In those cells, baseline currents of (mean±SEM) 53.76±7.95 pA at +40 mV increased to 1375±195.1 pA following addition of 1-EBIO (n=40 cells, p<0.0001), and were blocked by the selective KCa3.1 blocker, TRAM-34 (200 nM). There was an accompanying negative shift in cell reversal potential from −13.11±2.011 to −42.60±2.061 with addition of 1-EBIO (p<0.0001), which was reversed by TRAM-34.
Conclusion Human lung-derived (myo)fibroblasts express functional KCa3.1, K+ channels. These findings raise the possibility that blocking the KCa3.1 channel may inhibit pathological myo(fibroblast) function in IPF, and thus offer a novel approach to therapy.
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