Functional evidence for the expression of P2X1, P2X4 and P2X7 receptors in human lung mast cells.
OBJECTIVE
P2X receptors are widely expressed in cells of the immune system with varying functions. This study sought to characterize P2X receptor expression in the LAD2 human mast cell line and human lung mast cells (HLMCs).


METHODS
Reverse transcriptase polymerase chain reaction (RT-PCR) and patch clamp studies were used to characterize P2X expression in mast cells using a range of pharmacological tools.


RESULTS
RT-PCR revealed P2X1, P2X4 and P2X7 transcripts in both cell types; mRNA for P2X6 was also detected in LAD2 cells. Under whole-cell patch clamp conditions, rapid application of ATP (1-1000 microM) to cells clamped at -60 mV consistently evoked inward currents in both types of cells. Brief application of ATP (1 s) evoked a rapidly desensitizing P2X1-like current in both cell types. This current was also elicited by alphabetamethylene ATP (10 microM, 94% cells, n= 31) and was antagonized in LAD2 cells by NF 449 (1 microM) and pyridoxal phosphate-6-azo(benzene-2,4-disulphonic acid) (1-10 microM). A P2X7-like non-desensitizing current in response to high concentrations of ATP (1-5 mM) was also seen in both cell types (96% LAD2, n= 24; 54% HLMCs, n= 24) which was antagonized by AZ11645373 (1 microM). P2X7-like responses were also evoked in LAD2 cells by 2'(3')-0-(4-benzoylbenzoyl)ATP (300 microM). A P2X4-like current was evoked by 100 microM ATP (80% LAD2, n= 10; 21% HLMCs, n= 29), the amplitude and duration of which was potentiated by ivermectin (3 microM).


CONCLUSIONS
Our data confirmed the presence of functional P2X1, P2X4 and P2X7 receptors in LAD2 cells and HLMCs.
Wareham
K
K
Department of Biomedical Science, University of Sheffield, Western Bank, Sheffield, UK.
Vial
C
C
Wykes
R C E
RC
Bradding
P
P
Seward
E P
EP
D000255:Adenosine Triphosphate / Q000378:metabolism; D002412:Cations; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D005110:Extracellular Space / Q000378:metabolism; D006801:Humans; D007473:Ion Channels / Q000502:physiology; D008168:Lung / Q000378:metabolism*; D008407:Mast Cells / Q000166:cytology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D012333:RNA, Messenger / Q000096:biosynthesis; D018048:Receptors, Purinergic P2 / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D058469:Receptors, Purinergic P2X; D058848:Receptors, Purinergic P2X4; D058486:Receptors, Purinergic P2X7; D020133:Reverse Transcriptase Polymerase Chain Reaction
D002412:Cations; D007473:Ion Channels; C548226:P2RX4 protein, human; C546468:P2RX7 protein, human; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D058848:Receptors, Purinergic P2X4; D058486:Receptors, Purinergic P2X7; D000255:Adenosine Triphosphate