Here, for the first time, we report the KCa3.1 expression in two functionally distinct lung DC subsets in PBS-treated and OVA-sensitized and OVA-challenged mice. In PBS-treated mice, the immunogenic CD11clowCD11bhigh DCs had significantly lower expression of KCa3.1 as compared with the regulatory CD11chighCD11blow DCs. OVA sensitization differentially up-regulated KCa3.1 expression in two lung DCs subsets, with the greatest up-regulation observed in immunogenic CD11clowCD11bhigh DCs. However, the final expression levels of KCa3.1 in two lung DC subsets in OVA-sensitized mice are similar. This explains why the blockade of KCa3.1 by its specific marker TRAM-34 did not demonstrate a differential impairing effect on lung DC migration in the two DC subsets. Additionally, although Ag-carrying DCs expressed higher levels of KCa3.1 than non–Ag-carrying DCs, the difference was not as prominent as the difference between PBS- and OVA-sensitized mice. This suggests that, under the condition of allergic airway inflammation, KCa3.1 expression in lung DCs is probably regulated by multiple factors other than antigen loading. Other factors that are present in the microenvironment, such as cytokines and growth factors, contribute to the consequential up-regulation of KCa3.1 expression.
The implications of these findings help to explain the different patterns of migration that we reported previously. OVA sensitization rapidly recruits immunogenic CD11clow
DCs, significantly enhances their migration to draining lymph node, and largely boosts their antigen uptake activity (28
). This is well correlated with a greater up-regulation of KCa3.1 by OVA sensitization in this DC subset, providing strong evidence that KCa3.1 is involved in OVA sensitization–mediated DC activation, with a greater effect in immunogenic CD11clow
The linkage between KCa3.1 and lung DC migration appears to be CCR7-induced intracellular calcium release and the following store-operated calcium entry. This is evidenced by the coexpression of CCR7 and KCa3.1, CCR7 activation–induced calcium influx, and 1-EBIO–induced membrane hyperpolarization in mouse lung DCs. The blockade of KCa3.1 by TRAM-34 disrupted the temporal coupling between KCa3.1 and calcium influx and subsequently impaired CCR7-induced chemotaxis.
KCa3.1 activity in regulating cell proliferation, activation, and migration is calcium dependent. Only a small increase in intracellular Ca2+
is required to activate KCa3.1 and allow K+
efflux, which counteracts the depolarizing effect of Ca2+
). The net result of a CCR7 activation–induced Ca2+
influx is membrane depolarization, provided that no other ion channel is involved. However, if potassium efflux couples with calcium influx temporally, the overall consequence could be depolarization or hyperpolarization, depending on the kinetics and conductance of the involved calcium and potassium channels. It has been recently reported that store-operated calcium influx leads to cell membrane hyperpolarization in human monocyte–derived macrophages and that the outward cationic current is carried by KCa3.1 (31
). In the context of cell proliferation, calcium influx permits cells in the G1 phase to pass through the G1-S checkpoint (32
). The role of KCa3.1 channels in cell migration is more complicated. Because KCa3.1 channels are not sensitive to voltage stimulation, they seem perfectly designed to maintain the resting membrane potential in the cells without excitable membranes, such as immune cells. In addition, KCa3.1 is responsible for concerting with calcium oscillation in migratory cells by acting in a fluctuating pattern to keep pace with cell protrusion and retraction (34
). KCa3.1 is also a swelling-activated potassium channel in many cell types, and the activation of KCa3.1 can orchestrate the volume, regulate cytoskeleton organization, and facilitate transmembrane water transport (35
). However, sustained opening of KCa3.1 disrupts the coupling between calcium influx and the calcium-activated potassium channel and thus impairs the temporal dynamics of the calcium signal necessary for cell migration. In transformed renal epithelial cells, 1-EBIO reduces migration rate (26
). Finally, TRAM-34 is a highly specific small molecule blocker of KCa3.1 that does not affect cytochrome P450. TRAM-34 has low toxicity and causes minimal cell death and apoptosis (10
). If the role of KCa3.1 in DC migration under inflammatory condition is further established in an in vivo
study, TRAM-34 could be a potential drug that targets KCa3.1.
KCa3.1 seems to be preferentially involved in cell biology under pathological conditions. In the case of OVA allergen–induced acute airway inflammation, KCa3.1 regulates DC migration at two levels. First, CCR7 activation is linked to KCa3.1 activation via CCL19/CCL21-induced intracellular calcium release (1
). The high CCR7 expression in the immunogenic lung DC subset or under inflammation conditions creates a favorable condition for KCa3.1 activation, which facilitates further calcium influx for a rapid DC migration. Second, a higher KCa3.1 expression in lung DCs under allergic inflammation conditions warrants its greater involvement in DC migration. Knowing this will help define a new role of ion channels in the regulation of DC migration.
In conclusion, our data suggest that antigen sensitization up-regulates KCa3.1 expression, which may contribute to enhancing cell migration in response to lymphatic chemokines, particularly in the immunogenic lung DC subset.