Calcium-activated potassium channel KCa3.1 in lung dendritic cell migration

Zhifei Shao, Toluwalope O. Makinde, Devendra K. Agrawal

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

Migration to draining lymph nodes is a critical requirement for dendritic cells (DCs) to control T-cell-mediated immunity. The calcium-activated potassium channel KCa3.1 has been shown to be involved in regulating cell migration in multiple cell types. In this study, KCa3.1 expression and its functional role in lung DC migration were examined. Fluorescence-labeled antigen was intranasally delivered into mouse lungs to label lung Ag-carrying DCs. Lung CD11c highCD11b low and CD11c lowCD11b high DCs from PBS-treated and ovalbumin (OVA)-sensitized mice were sorted using MACS and FACS. Indo-1 and DiBAC4(3) were used to measure intracellular Ca2+ and membrane potential, respectively. The mRNA expression of KCa3.1 was examined using real-time PCR. Expression of KCa3.1 protein and CCR7 was measured using flow cytometry. Migration of two lung DC subsets to lymphatic chemokines was examined using TransWell in the absence or presence of the KCa3.1 blocker TRAM-34. OVA sensitization up-regulated mRNA and protein expression of KCa3.1 in lung DCs, with a greater response by the CD11c highCD11b low than CD11c lowCD11b high DCs. Although KCa3.1 expression in Ag-carrying DCs was higher than that in non - Ag-carrying DCs in OVA-sensitized mice, the difference was not as prominent. However, Ag-carrying lung DCs expressed significantly higher CCR7 than non - Ag-carrying DCs. CCL19, CCL21, and KCa3.1 activator 1-EBIO induced an increase in intracellular calcium in both DC subsets. In addition, 1-EBIO - induced calcium increase was suppressed by TRAM-34. In vitro blockade of KCa3.1 with TRAM-34 impaired CCL19/CCL21-induced transmigration. In conclusion, KCa3.1 expression in lung DCs is up-regulated by OVA sensitization in both lung DC subsets, and KCa3.1 is involved in lung DC migration to lymphatic chemokines.

Original languageEnglish
Pages (from-to)962-968
Number of pages7
JournalAmerican Journal of Respiratory Cell and Molecular Biology
Volume45
Issue number5
DOIs
StatePublished - Nov 1 2011

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Calcium-Activated Potassium Channels
Dendritic Cells
Cell Movement
Lung
Ovalbumin
Chemokines
Calcium
Intracellular Membranes
Messenger RNA
T-cells
Flow cytometry

All Science Journal Classification (ASJC) codes

  • Cell Biology
  • Pulmonary and Respiratory Medicine
  • Molecular Biology
  • Clinical Biochemistry

Cite this

Calcium-activated potassium channel KCa3.1 in lung dendritic cell migration. / Shao, Zhifei; Makinde, Toluwalope O.; Agrawal, Devendra K.

In: American Journal of Respiratory Cell and Molecular Biology, Vol. 45, No. 5, 01.11.2011, p. 962-968.

Research output: Contribution to journalArticle

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abstract = "Migration to draining lymph nodes is a critical requirement for dendritic cells (DCs) to control T-cell-mediated immunity. The calcium-activated potassium channel KCa3.1 has been shown to be involved in regulating cell migration in multiple cell types. In this study, KCa3.1 expression and its functional role in lung DC migration were examined. Fluorescence-labeled antigen was intranasally delivered into mouse lungs to label lung Ag-carrying DCs. Lung CD11c highCD11b low and CD11c lowCD11b high DCs from PBS-treated and ovalbumin (OVA)-sensitized mice were sorted using MACS and FACS. Indo-1 and DiBAC4(3) were used to measure intracellular Ca2+ and membrane potential, respectively. The mRNA expression of KCa3.1 was examined using real-time PCR. Expression of KCa3.1 protein and CCR7 was measured using flow cytometry. Migration of two lung DC subsets to lymphatic chemokines was examined using TransWell in the absence or presence of the KCa3.1 blocker TRAM-34. OVA sensitization up-regulated mRNA and protein expression of KCa3.1 in lung DCs, with a greater response by the CD11c highCD11b low than CD11c lowCD11b high DCs. Although KCa3.1 expression in Ag-carrying DCs was higher than that in non - Ag-carrying DCs in OVA-sensitized mice, the difference was not as prominent. However, Ag-carrying lung DCs expressed significantly higher CCR7 than non - Ag-carrying DCs. CCL19, CCL21, and KCa3.1 activator 1-EBIO induced an increase in intracellular calcium in both DC subsets. In addition, 1-EBIO - induced calcium increase was suppressed by TRAM-34. In vitro blockade of KCa3.1 with TRAM-34 impaired CCL19/CCL21-induced transmigration. In conclusion, KCa3.1 expression in lung DCs is up-regulated by OVA sensitization in both lung DC subsets, and KCa3.1 is involved in lung DC migration to lymphatic chemokines.",
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