Centrosome-directed translocation of Weibel-Palade bodies is rapidly induced by thrombin, calyculin A, or cytochalasin B in human aortic endothelial cells

To examine the possible role of the cytoskeleton in exocytosis of Weibel-Pa lade bodies (WPBs), we used double immunofluorescence and electron microsco py to study the spatial relationships between WPBs and main cytoskeletal el ements in endothelial cells treated with secretagogue, such as thrombin, or cytoskeleton-damaging agents. Unexpectedly, we have found that WPBs underg o rapid translocation towards the centrosome both in cells treated with thr ombin and in those treated with cytochalasin B or calyculin A. Typically, 3 or 5 min after agent addition compact cluster of WPBs became visible near the microtubule-oganizing center (MTOC) in most endothelial cells in which a fivefold increase in WPBs localized in close proximity to the mother cent riole had been detected. In both thrombin- and cytochalasin-treated cells t hat exhibit a noticeable depletion in WPBs compared to control cells, WPBs located at the cell periphery were found to colocalize with vimentin interm ediate filaments, but not with microtubules. In contrast, there was precise colocalization observed between WPBs and microtubules in calyculin-treated cells in which all WPBs undergo centrosome-directed translocation within 1 5 min after the agent addition. When vimentin filaments were induced to col lapse to a perinucle ar location by the microtubule-disrupting agent demeco lcine, WPBs also translocated to the perinuclear region, where numerous WPB s were found to be localized within the bundles of intermediate-sized filam ents. The data provide the first direct evidence that secretory granules ut ilize microtubule-based transport system to move in retrograde direction, i .e., away from the plasma membrane, towards the centrosome. We suggest that anterograde movement of WPBs is primarily dependent on their interaction w ith vimentin intermediate filaments. (C) 2000 Wiley-Liss, Inc.