Confocal imaging was conducted after immunocytochemical staining for acetylated [alpha]-tubulin, PLC[beta]3, TRPV4, and PAR-2.
After stringency washing, complexes were investigated by Western blotting using antibodies specific for TRPV4, PAR-2, or calmodulin.
Using MMP-1 reporter assays, we found that TRPV4-specific siRNA decreased MMP-1 transcriptional activation, thus implying that TRPV4 is critical for DEP/ OE-evoked [Ca.
TRPV4 activation by 4[alpha]-PDD or hypotonicity strongly increased MMP-1 secretion, indicating that in airway epithelia, TRPV4 activation is sufficient to up-regulate MMP-1 (Figure 3D).
Taken together, these findings point toward critical functioning of TRPV4 in [Ca.
TRPV4 signaling complex is located on motile cilia of primary human airway epithelia.
Because TRPV4 channels have been found in primary motile cilia of mouse tracheal epithelia (Lorenzo et al.
i]) via disinhibited TRPV4 and/or activation of PLC[beta]3, both of which have previously been shown to bind calmodulin.
2+] influx via TRPV4 causes MMP-1 activation rationalizes airway injury by MMP-1 as caused by TRPV4 channel activity, not by DN channels, we attempted to resolve these seemingly contradictory concepts.
2+] influx mediated by TRPV4 channels, via membrane phospholipid signaling involving PLC[beta]3 and PI3-K.
The critical participants of the DEP response, PAR-2, PLC[beta]3, and TRPV4, were localized to motile cilia of differentiated primary human respiratory epithelia.
Two possible causes for the slow kinetics are a) DEP-dependent PLC[beta]3 phosphorylation, and b) DEP-enhanced calmodulin binding to a receptor-signaling multiplex containing TRPV4 and PLC[beta]3 (Figures 5, 7), both of which attenuate signaling via known properties of the modified phospholipase or channel.