Elevated [Ca2+]i Leads to Altered Cytoskeletal Architecture in a Mouse Model of ARPKD

Although it has been suggested that there are alterations in cytoskeletal architecture in polycystic kidney disease (PKD), there is a paucity of detailed morphological descriptions of the actin cytoskeleton in PKD. Cilium‐deficient cortical collecting duct cells from Oak Ridge Polycystic Kidney mouse ( orpk cilia(−)) have increased apical Ca 2+ permeability which results in elevated sub‐apical Ca 2+ levels when compared to their cilium‐competent counterparts ( orpk cilia(+)). We postulate that elevated Ca 2+ levels in orpk cilia(−) cells contribute to alterations in actin cytoskeletal organization. For these studies, actin was labeled in cultured monolayers of orpk cilia(+) and cilia(−) cells with ALEXA‐conjugated phalloidin and observed with confocal microscopy. In untreated cilia(+) cells, we observed an abundance of well organized, bundled stress fibers, as well as a prominent adhesion belt. Stress fibers in untreated cilia(−) cells were much less abundant and organized and the adhesion belt was poorly defined. To verify the effects of elevated [Ca 2+ ] i on actin organization, we treated cells apically with 5 μM ionomycin. After 10 min. treatment with ionomycin, cilia(+) cells stress fibers were nearly ablated and abundant perinuclear actin staining was observed. Cilia(−) cells, under the same conditions, displayed a further decrease in stress fibers and some perinuclear actin staining. In summary, these findings suggest that elevated [Ca 2+ ] i levels appear to disrupt actin cytoskeletal organization in orpk cilia(−) cells, which may contribute to the pathogenesis of PKD.