Biliary secretion and actin-cytokeratin filament distribution in rat hepatocytes during phalloidin-induced cholestasis.

Abstract

The relationship between bile secretion (bile flow, bile acids, phospholipids, and cholesterol) and distribution of actin microfilaments (MFs) and cytokeratin (CK) intermediate filaments (IFs) was examined in hepatocytes of rats injected with a single low dose of phalloidin. This treatment induced a transient cholestasis characterized by a rapid development period (0-90 min postinjection) and a slow recovery period (24 h and 5 days postinjection). No significant changes were observed in bile acid secretion during the 5-day period. The phospholipid output dropped to less than 25% at 90 min and was back to the normal value at 24 h postinjection. In a parallel way, the cholesterol secretion dropped to 30% but came back to only 60% of the control level. Nile Red staining demonstrated a concomitant accumulation of lipids both in the cytoplasm and at the surface membrane. Immunostaining of the actin MFs and CK IFs showed that, in contrast with controls where both cytoskeletal networks were preferentially and uniformly localized at the surface membrane (i.e., sinusoidal, basolateral, and canalicular regions), the toxin treatment led to a major targeting of actin to the pericanalicular region at 24 h and a massive accumulation of well-preserved CK IFs in the cytoplasm at 5 days. Interestingly, this accumulation of CK IFs was not linked to any significant variations in CK isoforms. Together, these data indicate that a selective binding of the toxin to sinusoidal membrane actin at the time of injection triggers a sequence of events that culminate in delayed accumulation of actin MFs at the canalicular pole and of CK IFs in the cytoplasm. Moreover, the reversible perturbation of the bile secretory activity implies a functional adaptation of the hepatocytes that parallels the phalloidin-induced reorganization of both cytoskeleton networks.