Effect of reversible ATP depletion on tight-junction integrity in LLC-PK1 cells

P. Ellen Canfield, Alison M. Geerdes, Bruce Molitoris

Research output: Contribution to journalArticle

112 Citations (Scopus)

Abstract

To further understand and investigate how ischemia affects the tight junction we have developed a 2-h model of rapidly reversible ATP depletion and cellular injury in confluent LLC-PK1 monolayers. ATP depletion was achieved utilizing substrate-free medium containing 0.1 μM antimycin A (AA). Cellular ATP levels dropped rapidly to 3H]mannitol increased from a control level of 7 to 13.5% during ATP depletion. Recovery of tight-junction integrity was demonstrated by a slowing of [3H]mannitol transfer from the basolateral to the apical medium. The transfer rate in control monolayers was 0.0126%/min. During the initial 120 min of cellular recovery from 2 h of ATP depletion, the transfer rate was 0.0789%/min, but this decreased to 0.0045%/min between 2 and 4 h of recovery. In summary, physiology, biochemical, and morphological evidence indicates that reversible ATP depletion results in rapid opening of cellular tight junctions. After ATP-repletion physiological studies indicate a recovery of tight-junction integrity.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Renal Fluid and Electrolyte Physiology
Volume261
Issue number6 30-6
StatePublished - 1991
Externally publishedYes

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LLC-PK1 Cells
Tight Junctions
Adenosine Triphosphate
Mannitol
Antimycin A
Ischemia
Wounds and Injuries

Keywords

  • Electrical resistance across monolayers
  • Paracellular permeability
  • Ruthenium red

ASJC Scopus subject areas

  • Physiology

Cite this

Effect of reversible ATP depletion on tight-junction integrity in LLC-PK1 cells. / Ellen Canfield, P.; Geerdes, Alison M.; Molitoris, Bruce.

In: American Journal of Physiology - Renal Fluid and Electrolyte Physiology, Vol. 261, No. 6 30-6, 1991.

Research output: Contribution to journalArticle

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AU - Geerdes, Alison M.

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N2 - To further understand and investigate how ischemia affects the tight junction we have developed a 2-h model of rapidly reversible ATP depletion and cellular injury in confluent LLC-PK1 monolayers. ATP depletion was achieved utilizing substrate-free medium containing 0.1 μM antimycin A (AA). Cellular ATP levels dropped rapidly to 3H]mannitol increased from a control level of 7 to 13.5% during ATP depletion. Recovery of tight-junction integrity was demonstrated by a slowing of [3H]mannitol transfer from the basolateral to the apical medium. The transfer rate in control monolayers was 0.0126%/min. During the initial 120 min of cellular recovery from 2 h of ATP depletion, the transfer rate was 0.0789%/min, but this decreased to 0.0045%/min between 2 and 4 h of recovery. In summary, physiology, biochemical, and morphological evidence indicates that reversible ATP depletion results in rapid opening of cellular tight junctions. After ATP-repletion physiological studies indicate a recovery of tight-junction integrity.

AB - To further understand and investigate how ischemia affects the tight junction we have developed a 2-h model of rapidly reversible ATP depletion and cellular injury in confluent LLC-PK1 monolayers. ATP depletion was achieved utilizing substrate-free medium containing 0.1 μM antimycin A (AA). Cellular ATP levels dropped rapidly to 3H]mannitol increased from a control level of 7 to 13.5% during ATP depletion. Recovery of tight-junction integrity was demonstrated by a slowing of [3H]mannitol transfer from the basolateral to the apical medium. The transfer rate in control monolayers was 0.0126%/min. During the initial 120 min of cellular recovery from 2 h of ATP depletion, the transfer rate was 0.0789%/min, but this decreased to 0.0045%/min between 2 and 4 h of recovery. In summary, physiology, biochemical, and morphological evidence indicates that reversible ATP depletion results in rapid opening of cellular tight junctions. After ATP-repletion physiological studies indicate a recovery of tight-junction integrity.

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