Effects of cytokines on mechanical and epithelial bioelectric responses to methacholine and hyperosmolarity in guinea-pig airways: An in vitro study

Ismailoglu, Ugur B.; Scott, Michael R. Van; Fedan, Jeffrey S.

doi:10.1016/j.ejphar.2009.04.025

Cited by 8 publications

(8 citation statements)

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“…Amiloride-sensitive short-circuit current is inhibited by LPS in the mouse trachea in the Ussing chamber (35). Although the ionic basis of the effect was not examined, we observed that incubation of guinea pig trachea in vitro with LPS reduced V t (32).…”

Section: Discussionmentioning

confidence: 83%

“…Therefore, we earlier investigated the in vitro effects of LPS and cytokines on the reactivity of the airway smooth muscle and epithelium to methacholine and hyperosmolarity in the absence of LPS binding protein in the IPT and the Ussing chamber (32). Incubating tracheas with LPS, IL-1␤, IL-4, IL-13, IFN␥, and TNF␣ alone or in combination caused effects similar to those that in vivo LPS treatment had on mechanical responses of the airways to methacholine and hyperosmolar solutions, but had varying effects on V t and bioelectric responses of the epithelium that did not entirely reproduce the effects of LPS given in vivo (32).…”

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confidence: 99%

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Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na⁺ channel and the Na⁺-K⁺ pump

Dodrill

Fedan

2010

American Journal of Physiology-Lung Cellular and Molecular Phys

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Earlier, we found that systemic administration of lipopolysaccharide (LPS; 4 mg/kg) hyperpolarized the transepithelial potential difference (V(t)) of tracheal epithelium in the isolated, perfused trachea (IPT) of the guinea pig 18 h after injection. As well, LPS increased the hyperpolarization component of the response to basolateral methacholine, and potentiated the epithelium-derived relaxing factor-mediated relaxation responses to hyperosmolar solutions applied to the apical membrane. We hypothesized that LPS stimulates the transepithelial movement of Na(+) via the epithelial sodium channel (ENaC)/Na(+)-K(+) pump axis, leading to hyperpolarization of V(t). LPS increased the V(t)-depolarizing response to amiloride (10 μM), i.e., offset the effect of LPS, indicating that Na(+) transport activity was increased. The functional activity of ENaC was measured in the IPT after short-circuiting the Na(+)-K(+) pump with basolateral amphotericin B (7.5 μM). LPS had no effect on the hyperpolarization response to apical trypsin (100 U/ml) in the Ussing chamber, indicating that channel-activating proteases are not involved in the LPS-induced activation of ENaC. To assess Na(+)-K(+) pump activity in the IPT, ENaC was short-circuited with apical amphotericin B. The greater V(t) in the presence of amphotericin B in tracheas from LPS-treated animals compared with controls revealed that LPS increased Na(+)-K(+) pump activity. This finding was confirmed in the Ussing chamber by inhibiting the Na(+)-K(+) pump via extracellular K(+) removal, loading the epithelium with Na(+), and observing a greater hyperpolarization response to K(+) restoration. Together, the findings of this study reveal that LPS hyperpolarizes the airway epithelium by increasing the activities of ENaC and the Na(+)-K(+) pump.

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Section: Discussionmentioning

confidence: 83%

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confidence: 99%

Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na⁺ channel and the Na⁺-K⁺ pump

Dodrill

Fedan

2010

American Journal of Physiology-Lung Cellular and Molecular Phys

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“…Lipopolysaccharide treatment in vivo (Dodrill and Fedan, 2010) or exposure to cytokines in vitro (Ismailoglu et al, 2009) potentiated hyperosmolarity-induced relaxation. Lipopolysaccharide treatment in vivo also increased transepithelial potential difference ( V t ) and potentiated depolarization responses to elevations in osmolarity (Johnston et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

Tracheal epithelium cell volume responses to hyperosmolar, isosmolar and hypoosmolar solutions: relation to epithelium-derived relaxing factor (EpDRF) effects

Fedan¹,

Thompson²,

Ismailoglu³

et al. 2013

Front. Physiol.

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In asthmatic patients, inhalation of hyperosmolar saline or D-mannitol (D-M) elicits bronchoconstriction, but in healthy subjects exercise causes bronchodilation. Hyperventilation causes drying of airway surface liquid (ASL) and increases its osmolarity. Hyperosmolar challenge of airway epithelium releases epithelium-derived relaxing factor (EpDRF), which relaxes the airway smooth muscle. This pathway could be involved in exercise-induced bronchodilation. Little is known of ASL hyperosmolarity effects on epithelial function. We investigated the effects of osmolar challenge maneuvers on dispersed and adherent guinea-pig tracheal epithelial cells to examine the hypothesis that EpDRF-mediated relaxation is associated with epithelial cell shrinkage. Enzymatically-dispersed cells shrank when challenged with ≥10 mOsM added D-M, urea or NaCl with a concentration-dependence that mimics relaxation of the of isolated perfused tracheas (IPT). Cells shrank when incubated in isosmolar N-methyl-D-glucamine (NMDG) chloride, Na gluconate (Glu), NMDG-Glu, K-Glu and K2SO4, and swelled in isosmolar KBr and KCl. However, isosmolar challenge is not a strong stimulus of relaxation in IPTs. In previous studies amiloride and 4,4′-diisothiocyano-2,2′-stilbenedisulfonic acid (DIDS) inhibited relaxation of IPT to hyperosmolar challenge, but had little effect on shrinkage of dispersed cells. Confocal microscopy in tracheal segments showed that adherent epithelium is refractory to low hyperosmolar concentrations that induce dispersed cell shrinkage and relaxation of IPT. Except for gadolinium and erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), actin and microtubule inhibitors and membrane permeabilizing agents did not affect on ion transport by adherent epithelium or shrinkage responses of dispersed cells. Our studies dissociate relaxation of IPT from cell shrinkage after hyperosmolar challenge of airway epithelium.

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“…Incubating tracheas with LPS, IL-1β, IL-4, IL-13, IFN-γ, and TNF-α alone or in combination caused effects similar to those that in vivo LPS treatment had on mechanical responses of the airways to methacholine and hyperosmolar solutions, but had varying effects on V t and bioelectric responses of the epithelium that did not entirely reproduce the effects of LPS given in vivo (32).…”

Section: Introductionmentioning

confidence: 91%

“…The effects of systemic LPS require the presence of LPS binding protein, needed for the activation of TLR4 by LPS (61). Therefore, we earlier investigated the in vitro effects of LPS and cytokines on the reactivity of the airway smooth muscle and epithelium to methacholine and hyperosmolarity in the absence of LPS binding protein in the IPT and the Ussing chamber (32).…”

Section: Introductionmentioning

confidence: 99%

Systemic lipopolysaccharide stimulates airway transepithelial Na+ transport by increasing ENaC and Na+,K+ -pump activity

Dodrill¹

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Systemic lipopolysaccharide stimulates airway transepithelial Na + transport by increasing ENaC and Na + ,K +-pump activity Michael W. Dodrill Our laboratory found that systemic administration of lipopolysaccharide (LPS; 4 mg/kg) hyperpolarized the transepithelial potential difference (V t) of tracheal epithelium in the isolated, perfused trachea (IPT) of the guinea pig 18 h after injection. We hypothesized that LPS stimulates the transepithelial movement of Na + via the epithelial sodium channel (ENaC)/Na + ,K +-pump axis, leading to hyperpolarization of V t. LPS increased the V t response to amiloride, i.e., offset the effect of LPS, indicating that Na + transport was increased. The functional activity of ENaC was measured in the IPT after short-circuiting the Na + ,K +-pump with basolateral amphotericin B. LPS had no effect on the hyperpolarization response to apical trypsin in the Ussing chamber, indicating that channel activating proteases are not involved in the LPS-induced activation of ENaC. To assess Na + ,K +-pump activity in the IPT, ENaC was short-circuited with apical amphotericin B. The greater V t in the presence of amphotericin B in tracheas from LPS-treated animals compared to controls revealed that LPS increased Na + ,K +-pump activity. This finding was confirmed in the Ussing chamber by inhibiting the Na + ,K +-pump via extracellular K + removal, loading the epithelium with Na + , and observing a greater hyperpolarization response to K + restoration. Using qPCR, the effects of LPS on the transcription of αENaC, α 1 Na + ,K +pump, COX-2, eNOS, iNOS, IL-1β, and TNF-α were measured at 3 and 18 h. In the epithelium, LPS increased the transcription of COX-2, IL-1β, and, to nonsignificant extent, TNF-α at 3 h, but not at 18 h. In alveolar macrophages, TNF-α, and, to a nonsignificant extent, COX-2 and IL-1β were up-regulated at 3 h, but not at 18 h. Even though LPS stimulated the transcription of some genes, αENaC and α 1 Na + ,K +-ATPase transcription were not affected. The expression of α-, β-, and γ-ENaC and α 1 Na + ,K +-pump from tracheal epithelium and kidney cortex/medulla were investigated by western blotting. All three ENaC subunits were detected as cleavage fragments, yet LPS had no effect on their expression. LPS increased the expression of the α 1 subunit and the α 1-, α 2-, and α 3-subunits, collectively, of the Na + ,K +-pump. Taken together, the findings of this study reveal that LPS hyperpolarizes the airway epithelium by increasing the activities of ENaC and the Na + ,K +-pump. ENaC activation by LPS is not accomplished via a change in ENaC regulation involving proteolytic cleavage. LPS increases Na + transport downstream of the genetic level, in part, by stimulating the expression of the Na + ,K +-pump.

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Effects of cytokines on mechanical and epithelial bioelectric responses to methacholine and hyperosmolarity in guinea-pig airways: An in vitro study

Cited by 8 publications

References 31 publications

Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na⁺ channel and the Na⁺-K⁺ pump

Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na⁺ channel and the Na⁺-K⁺ pump

Tracheal epithelium cell volume responses to hyperosmolar, isosmolar and hypoosmolar solutions: relation to epithelium-derived relaxing factor (EpDRF) effects

Systemic lipopolysaccharide stimulates airway transepithelial Na+ transport by increasing ENaC and Na+,K+ -pump activity

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Effects of cytokines on mechanical and epithelial bioelectric responses to methacholine and hyperosmolarity in guinea-pig airways: An in vitro study

Cited by 8 publications

References 31 publications

Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na+ channel and the Na+-K+ pump

Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na+ channel and the Na+-K+ pump

Tracheal epithelium cell volume responses to hyperosmolar, isosmolar and hypoosmolar solutions: relation to epithelium-derived relaxing factor (EpDRF) effects

Systemic lipopolysaccharide stimulates airway transepithelial Na+ transport by increasing ENaC and Na+,K+ -pump activity

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Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na⁺ channel and the Na⁺-K⁺ pump

Lipopolysaccharide hyperpolarizes guinea pig airway epithelium by increasing the activities of the epithelial Na⁺ channel and the Na⁺-K⁺ pump