Alterations in human proximal tubule cell attachment in response to hypoxia: role of microfilaments.

Detachment of viable renal proximal tubular cells is seen in clinical and experimental acute tubular necrosis and may contribute to the acute renal dysfunction seen in acute tubular necrosis. Mechanisms of detachment of tubular cells are unknown but must involve changes in tubular cell adhesion. To begin to define mechanisms of altered cell adhesion, cultured human proximal tubular cells were made hypoxic by nitrogen gassing. Cells were monitored (blinded) for cell retraction and rounding over 90 minutes of N2. Hypoxia caused gradual alterations in cell shape, with 37.9% +/- 5.2% retracted-rounded cells by 90 minutes; control monolayers showed no significant change. Fluorescence confocal microscope imaging revealed that hypoxia caused displacement of actin filaments to basal margins of the retracted cells and produced a perinuclear aggregation of short filaments. Phalloidin (10(-6) mol/L), which stabilizes microfilaments and is able to penetrate these hypoxic cells, decreased the percentage of cells showing morphologic changes with hypoxia to < 5% by 90 minutes (p < 0.01). Viability, as assessed by Trypan blue dye exclusion, was well maintained (90% to 98% at 90 minutes) and did not correlate with shape changes. In separate experiments, cytochalasin (10(-6) mol/L)--which depolymerizes microfilaments--but not nocodazole--which disrupts microtubules--produced cell shape change in non-hypoxic monolayers. Disruption of microfilaments appears to play a role in loss of cell-to-cell and cell-to-substrate adhesion and loss of epithelial integrity in hypoxic injury to the renal tubule. These in vitro observations may be relevant to renal proximal tubular cell detachment in in vivo renal injury.