FP228
ROLE OF CELL CYTOSKELETON IN RENAL EPITHELIAL CELL MIGRATION DURING HYPOXIA Free

INTRODUCTION AND AIMS: Acute kidney injury (AKI) is a common and potentially lethal complication in hospitalized patients. Hypoxic injury of kidneys, a major cause of AKI, involves loss of renal tubular epithelial cells which may then be replaced by proliferating migratory neighboring epithelial cells. Even though cell migration is an important mechanism in subsequent epithelial regeneration and wound healing, only scarce data exist on the role of hypoxia on epithelial cell migration and the involvement of the cell cytoskeleton. In the present study, we used an in vitro cell culture system to study the impact of hypoxia on cellular morphology and migratory behavior. We propose a link between cytoskeletal reorganization during hypoxia and cell migration.

METHODS: Here, we used primary isolated human renal tubular cells (hPTEC) as a model to study cell migration and the associated cytoskeletal changes. Hypoxia was induced either by low oxygen pressure or by treatment with dimethyloxalyl glycine (DMOG), an inhibitor of oxygen-sensing prolylhydroxylases (PHD) leading to stabilization of hypoxia-inducible factor (HIF), the central signaling that governs hypoxic cell response.

RESULTS: Using a wound healing assay, we found that hPTEC migrated as cohorts with cells of distal tubular origin moving faster than those of proximal origin. Hypoxia induced by DMOG caused a postponed and disorganized cell migration. Confirmatory, single cell movement tracking via live cell imaging showed significantly reduced migration velocity. In line with reduced movement, cell adhesion visualized by staining of paxilin and focal adhesion kinase (FAK) was reorganized under hypoxia demonstrating a stronger adherence of the tubular cell to the substrate. Compared to non-hypoxic conditions, treatment with DMOG induced rearrangement of actin and keratin filaments while the microtubule structures were not altered. We observed a marked bundling of keratin fibers primarily localized at the perinuclear region. However, keratin expression (K8, K18, K7 and K19) and phosphorylation of K18 (Ser33) remained unchanged under hypoxic conditions in isolated hPTEC.

CONCLUSIONS: Taken together, stabilization of HIF reduced the migratory capacity of hPTEC associated with a marked reorganization of cytoskeletal components, suggesting an important involvement of epithelial migration in kidney regeneration during AKI.