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Connexin 43 expression on peripheral blood eosinophils: role of gap junctions in transendothelial migration.

Vliagoftis H, Ebeling C, Ilarraza R, Mahmudi-Azer S, Abel M, Adamko D, Befus AD, Moqbel R - Biomed Res Int (2014)

Bottom Line: Gap junctions mediate direct communication between adjacent cells and may represent a new way of communication between immune cells distinct from communication through cytokines and chemokines.Cx43 is localized not only in the cytoplasm but also on the plasma membrane.The gap junction inhibitors 18-a-glycyrrhetinic acid and octanol did not have a significant effect on dye transfer but reduced dye exit from eosinophils.

View Article: PubMed Central - PubMed

Affiliation: Pulmonary Research Group, Department of Medicine, University of Alberta, 550 HMRC, Edmonton, AB, Canada T6G 2S2.

ABSTRACT
Eosinophils circulate in the blood and are recruited in tissues during allergic inflammation. Gap junctions mediate direct communication between adjacent cells and may represent a new way of communication between immune cells distinct from communication through cytokines and chemokines. We characterized the expression of connexin (Cx)43 by eosinophils isolated from atopic individuals using RT-PCR, Western blotting, and confocal microscopy and studied the biological functions of gap junctions on eosinophils. The formation of functional gap junctions was evaluated measuring dye transfer using flow cytometry. The role of gap junctions on eosinophil transendothelial migration was studied using the inhibitor 18-a-glycyrrhetinic acid. Peripheral blood eosinophils express Cx43 mRNA and protein. Cx43 is localized not only in the cytoplasm but also on the plasma membrane. The membrane impermeable dye BCECF transferred from eosinophils to epithelial or endothelial cells following coculture in a dose and time dependent fashion. The gap junction inhibitors 18-a-glycyrrhetinic acid and octanol did not have a significant effect on dye transfer but reduced dye exit from eosinophils. The gap junction inhibitor 18-a-glycyrrhetinic acid inhibited eosinophil transendothelial migration in a dose dependent manner. Thus, eosinophils from atopic individuals express Cx43 constitutively and Cx43 may play an important role in eosinophil transendothelial migration and function in sites of inflammation.

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Cx43 expression by peripheral blood eosinophils: (a) RT-PCR for Cx43 expression in peripheral blood eosinophils from atopic donors. (b) Western blot analysis of Cx43 expression by freshly isolated eosinophils and eosinophils cultured for 24 h in the presence or absence of IL-5 (10 ng/mL).
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fig1: Cx43 expression by peripheral blood eosinophils: (a) RT-PCR for Cx43 expression in peripheral blood eosinophils from atopic donors. (b) Western blot analysis of Cx43 expression by freshly isolated eosinophils and eosinophils cultured for 24 h in the presence or absence of IL-5 (10 ng/mL).

Mentions: Peripheral blood eosinophils expressed Cx43 mRNA (Figure 1(a)) but all individuals tested showed no expression of Cx32 (data not shown). We verified that peripheral blood eosinophils express Cx43 protein using Western blotting (Figure 1(b)). Incubation of eosinophils with IL-5 for up to 24 h did not change the level of expression of Cx43 protein (Figure 1(b)).


Connexin 43 expression on peripheral blood eosinophils: role of gap junctions in transendothelial migration.

Vliagoftis H, Ebeling C, Ilarraza R, Mahmudi-Azer S, Abel M, Adamko D, Befus AD, Moqbel R - Biomed Res Int (2014)

Cx43 expression by peripheral blood eosinophils: (a) RT-PCR for Cx43 expression in peripheral blood eosinophils from atopic donors. (b) Western blot analysis of Cx43 expression by freshly isolated eosinophils and eosinophils cultured for 24 h in the presence or absence of IL-5 (10 ng/mL).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109672&req=5

fig1: Cx43 expression by peripheral blood eosinophils: (a) RT-PCR for Cx43 expression in peripheral blood eosinophils from atopic donors. (b) Western blot analysis of Cx43 expression by freshly isolated eosinophils and eosinophils cultured for 24 h in the presence or absence of IL-5 (10 ng/mL).
Mentions: Peripheral blood eosinophils expressed Cx43 mRNA (Figure 1(a)) but all individuals tested showed no expression of Cx32 (data not shown). We verified that peripheral blood eosinophils express Cx43 protein using Western blotting (Figure 1(b)). Incubation of eosinophils with IL-5 for up to 24 h did not change the level of expression of Cx43 protein (Figure 1(b)).

Bottom Line: Gap junctions mediate direct communication between adjacent cells and may represent a new way of communication between immune cells distinct from communication through cytokines and chemokines.Cx43 is localized not only in the cytoplasm but also on the plasma membrane.The gap junction inhibitors 18-a-glycyrrhetinic acid and octanol did not have a significant effect on dye transfer but reduced dye exit from eosinophils.

View Article: PubMed Central - PubMed

Affiliation: Pulmonary Research Group, Department of Medicine, University of Alberta, 550 HMRC, Edmonton, AB, Canada T6G 2S2.

ABSTRACT
Eosinophils circulate in the blood and are recruited in tissues during allergic inflammation. Gap junctions mediate direct communication between adjacent cells and may represent a new way of communication between immune cells distinct from communication through cytokines and chemokines. We characterized the expression of connexin (Cx)43 by eosinophils isolated from atopic individuals using RT-PCR, Western blotting, and confocal microscopy and studied the biological functions of gap junctions on eosinophils. The formation of functional gap junctions was evaluated measuring dye transfer using flow cytometry. The role of gap junctions on eosinophil transendothelial migration was studied using the inhibitor 18-a-glycyrrhetinic acid. Peripheral blood eosinophils express Cx43 mRNA and protein. Cx43 is localized not only in the cytoplasm but also on the plasma membrane. The membrane impermeable dye BCECF transferred from eosinophils to epithelial or endothelial cells following coculture in a dose and time dependent fashion. The gap junction inhibitors 18-a-glycyrrhetinic acid and octanol did not have a significant effect on dye transfer but reduced dye exit from eosinophils. The gap junction inhibitor 18-a-glycyrrhetinic acid inhibited eosinophil transendothelial migration in a dose dependent manner. Thus, eosinophils from atopic individuals express Cx43 constitutively and Cx43 may play an important role in eosinophil transendothelial migration and function in sites of inflammation.

Show MeSH
Related in: MedlinePlus