Evaluation of basement membrane degradation during TNF-α-induced increase in epithelial permeability

Jean Claude Lacherade, Andry Van De Louw, Emmanuelle Planus, Estelle Escudier, Marie Pia D'Ortho, Chantal Lafuma, Alain Harf, Christophe Delclaux

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Abstract

We evaluated whether tumor necrosis factor (TNF)-α induces an increase in permeability of an alveolar epithelial monolayer via gelatinase secretion and basement membrane degradation. Gelatinase secretion and epithelial permeability to radiolabeled albumin under unstimulated and TNF-α-stimulated conditions of an A549 human epithelial cell line were evaluated in vitro. TNF-α induced both upregulation of a 92-kDa gelatinolytic activity (pro form in cell supernatant and activated form in extracellular matrix) and an increase in the epithelial permeability coefficient compared with the unstimulated condition (control: 1.34 ± 0.04 × 10-6 cm/s; 1 μg/ml TNF-α: 1.47 ± 0.05 × 10-6 cm/s, P < 0.05). The permeability increase in the TNF-α-stimulated condition involved both paracellular permeability, with gap formation visualized by actin cytoskeleton staining, and basement membrane permeability, with an increase in the basement membrane permeability coefficient (determined after cell removal; control: 2.58 ± 0.07 × 10-6 cm/s; 1 μg/ml TNF-α: 2.82 ± 0.02.10-6 × cm/s, P < 0.05). Because addition of gelatinase inhibitors [tissue inhibitor of metalloproteinase (TIMP)-1 or BB-3103] to cell supernatants failed to inhibit the permeability increase, the gelatinase-inhibitor balance in the cellular microenvironment was further evaluated by cell culture on a radiolabeled collagen matrix. In the unstimulated condition, spontaneous collagenolytic activity inhibited by addition to the matrix of 1 μg/ml TIMP-1 or 10-6 M BB-3103 was found. TNF-α failed to increase this collagenolytic activity because it was associated with dose-dependent upregulation of TIMP-1 secretion by alveolar epithelial cells. In conclusion, induction by TNF-α of upregulation of both the 92-kDa gelatinase and its inhibitor TIMP-1 results in maintenance of the gelatinase-inhibitor balance, indicating that basement membrane degradation does not mediate the TNFα-induced increase in alveolar epithelial monolayer permeability.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
Volume281
Issue number1 25-1
StatePublished - Oct 16 2001

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Basement Membrane
Permeability
Tumor Necrosis Factor-alpha
Tissue Inhibitor of Metalloproteinase-1
Matrix Metalloproteinase Inhibitors
Gelatinases
Up-Regulation
Alveolar Epithelial Cells
Cellular Microenvironment
Matrix Metalloproteinase 9
Actin Cytoskeleton
Extracellular Matrix
Albumins
Collagen
Cell Culture Techniques
Epithelial Cells
Maintenance
Staining and Labeling
Cell Line

All Science Journal Classification (ASJC) codes

  • Physiology
  • Pulmonary and Respiratory Medicine
  • Physiology (medical)
  • Cell Biology

Cite this

Lacherade, Jean Claude ; Van De Louw, Andry ; Planus, Emmanuelle ; Escudier, Estelle ; D'Ortho, Marie Pia ; Lafuma, Chantal ; Harf, Alain ; Delclaux, Christophe. / Evaluation of basement membrane degradation during TNF-α-induced increase in epithelial permeability. In: American Journal of Physiology - Lung Cellular and Molecular Physiology. 2001 ; Vol. 281, No. 1 25-1.
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abstract = "We evaluated whether tumor necrosis factor (TNF)-α induces an increase in permeability of an alveolar epithelial monolayer via gelatinase secretion and basement membrane degradation. Gelatinase secretion and epithelial permeability to radiolabeled albumin under unstimulated and TNF-α-stimulated conditions of an A549 human epithelial cell line were evaluated in vitro. TNF-α induced both upregulation of a 92-kDa gelatinolytic activity (pro form in cell supernatant and activated form in extracellular matrix) and an increase in the epithelial permeability coefficient compared with the unstimulated condition (control: 1.34 ± 0.04 × 10-6 cm/s; 1 μg/ml TNF-α: 1.47 ± 0.05 × 10-6 cm/s, P < 0.05). The permeability increase in the TNF-α-stimulated condition involved both paracellular permeability, with gap formation visualized by actin cytoskeleton staining, and basement membrane permeability, with an increase in the basement membrane permeability coefficient (determined after cell removal; control: 2.58 ± 0.07 × 10-6 cm/s; 1 μg/ml TNF-α: 2.82 ± 0.02.10-6 × cm/s, P < 0.05). Because addition of gelatinase inhibitors [tissue inhibitor of metalloproteinase (TIMP)-1 or BB-3103] to cell supernatants failed to inhibit the permeability increase, the gelatinase-inhibitor balance in the cellular microenvironment was further evaluated by cell culture on a radiolabeled collagen matrix. In the unstimulated condition, spontaneous collagenolytic activity inhibited by addition to the matrix of 1 μg/ml TIMP-1 or 10-6 M BB-3103 was found. TNF-α failed to increase this collagenolytic activity because it was associated with dose-dependent upregulation of TIMP-1 secretion by alveolar epithelial cells. In conclusion, induction by TNF-α of upregulation of both the 92-kDa gelatinase and its inhibitor TIMP-1 results in maintenance of the gelatinase-inhibitor balance, indicating that basement membrane degradation does not mediate the TNFα-induced increase in alveolar epithelial monolayer permeability.",
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Evaluation of basement membrane degradation during TNF-α-induced increase in epithelial permeability. / Lacherade, Jean Claude; Van De Louw, Andry; Planus, Emmanuelle; Escudier, Estelle; D'Ortho, Marie Pia; Lafuma, Chantal; Harf, Alain; Delclaux, Christophe.

In: American Journal of Physiology - Lung Cellular and Molecular Physiology, Vol. 281, No. 1 25-1, 16.10.2001.

Research output: Contribution to journalArticle

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T1 - Evaluation of basement membrane degradation during TNF-α-induced increase in epithelial permeability

AU - Lacherade, Jean Claude

AU - Van De Louw, Andry

AU - Planus, Emmanuelle

AU - Escudier, Estelle

AU - D'Ortho, Marie Pia

AU - Lafuma, Chantal

AU - Harf, Alain

AU - Delclaux, Christophe

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N2 - We evaluated whether tumor necrosis factor (TNF)-α induces an increase in permeability of an alveolar epithelial monolayer via gelatinase secretion and basement membrane degradation. Gelatinase secretion and epithelial permeability to radiolabeled albumin under unstimulated and TNF-α-stimulated conditions of an A549 human epithelial cell line were evaluated in vitro. TNF-α induced both upregulation of a 92-kDa gelatinolytic activity (pro form in cell supernatant and activated form in extracellular matrix) and an increase in the epithelial permeability coefficient compared with the unstimulated condition (control: 1.34 ± 0.04 × 10-6 cm/s; 1 μg/ml TNF-α: 1.47 ± 0.05 × 10-6 cm/s, P < 0.05). The permeability increase in the TNF-α-stimulated condition involved both paracellular permeability, with gap formation visualized by actin cytoskeleton staining, and basement membrane permeability, with an increase in the basement membrane permeability coefficient (determined after cell removal; control: 2.58 ± 0.07 × 10-6 cm/s; 1 μg/ml TNF-α: 2.82 ± 0.02.10-6 × cm/s, P < 0.05). Because addition of gelatinase inhibitors [tissue inhibitor of metalloproteinase (TIMP)-1 or BB-3103] to cell supernatants failed to inhibit the permeability increase, the gelatinase-inhibitor balance in the cellular microenvironment was further evaluated by cell culture on a radiolabeled collagen matrix. In the unstimulated condition, spontaneous collagenolytic activity inhibited by addition to the matrix of 1 μg/ml TIMP-1 or 10-6 M BB-3103 was found. TNF-α failed to increase this collagenolytic activity because it was associated with dose-dependent upregulation of TIMP-1 secretion by alveolar epithelial cells. In conclusion, induction by TNF-α of upregulation of both the 92-kDa gelatinase and its inhibitor TIMP-1 results in maintenance of the gelatinase-inhibitor balance, indicating that basement membrane degradation does not mediate the TNFα-induced increase in alveolar epithelial monolayer permeability.

AB - We evaluated whether tumor necrosis factor (TNF)-α induces an increase in permeability of an alveolar epithelial monolayer via gelatinase secretion and basement membrane degradation. Gelatinase secretion and epithelial permeability to radiolabeled albumin under unstimulated and TNF-α-stimulated conditions of an A549 human epithelial cell line were evaluated in vitro. TNF-α induced both upregulation of a 92-kDa gelatinolytic activity (pro form in cell supernatant and activated form in extracellular matrix) and an increase in the epithelial permeability coefficient compared with the unstimulated condition (control: 1.34 ± 0.04 × 10-6 cm/s; 1 μg/ml TNF-α: 1.47 ± 0.05 × 10-6 cm/s, P < 0.05). The permeability increase in the TNF-α-stimulated condition involved both paracellular permeability, with gap formation visualized by actin cytoskeleton staining, and basement membrane permeability, with an increase in the basement membrane permeability coefficient (determined after cell removal; control: 2.58 ± 0.07 × 10-6 cm/s; 1 μg/ml TNF-α: 2.82 ± 0.02.10-6 × cm/s, P < 0.05). Because addition of gelatinase inhibitors [tissue inhibitor of metalloproteinase (TIMP)-1 or BB-3103] to cell supernatants failed to inhibit the permeability increase, the gelatinase-inhibitor balance in the cellular microenvironment was further evaluated by cell culture on a radiolabeled collagen matrix. In the unstimulated condition, spontaneous collagenolytic activity inhibited by addition to the matrix of 1 μg/ml TIMP-1 or 10-6 M BB-3103 was found. TNF-α failed to increase this collagenolytic activity because it was associated with dose-dependent upregulation of TIMP-1 secretion by alveolar epithelial cells. In conclusion, induction by TNF-α of upregulation of both the 92-kDa gelatinase and its inhibitor TIMP-1 results in maintenance of the gelatinase-inhibitor balance, indicating that basement membrane degradation does not mediate the TNFα-induced increase in alveolar epithelial monolayer permeability.

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