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 acut e renal dysfunction seen in acute tubular necrosis. Mechanisms of deta chment of tubular cells are unknown but must involve changes in tubula r cell adhesion. To begin to define mechanisms of altered cell adhesio n, cultured human proximal tubular cells were made hypoxic by nitrogen gassing. Cells were monitored (blinded) for cell retraction and round ing over 90 minutes of N-2. Hypoxia caused gradual alterations in cell shape, with 37.9% +/- 5.2% retracted-rounded cells by 90 minutes; con trol monolayers showed no significant change. Fluorescence confocal mi croscope imaging revealed that hypoxia caused displacement of actin fi laments to basal margins of the retracted cells and produced a perinuc lear 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 h ypoxia to <5% by 90 minutes (p < 0.01). Viability, as assessed by Tryp an blue dye exclusion, was well maintained (90% to 98% at 90 minutes) and did not correlate with shape changes. In separate experiments, cyt ochalasin (10(-6) mol/L)-which depolymerizes microfilaments-but not no codazole-which disrupts microtubules-produced cell shape change in non -hypoxic monolayers. Disruption of microfilaments appears to play a ro le 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 v itro observations may be relevant to renal proximal tubular cell detac hment in in vivo renal injury.