Remodeling of the actin cytoskeleton of target hepatocytes and NK cells during induction of apoptosis

Natural Killer cells are immune cells that recognize and eliminate altered and non-self cells from the circulation. To study the interaction between N K cells and target cells, we set up an experimental system consisting of ra t Interleukin-2 activated Natural Killer cells (A-NK cells) and rat hepatoc ytes with a masked Major Histocompatibility Complex (MHC). The masking of t he MHC induces recognition of the hepatocytes by the NK cells as non self. We showed that in vitro apoptosis is rapidly induced in the hepatocytes [Bl om et al., 1999] after co-incubation with A-NK cells. Now we describe the m orphological changes that occur during and after interaction of A-NK cells with hepatocytes. Confocal laser scanning microscopy showed that the actin cytoskeleton of the NK cells was remodeled during attack of hepatocytes. So me NK cells were in close contact with the hepatocytes while others had for med actin-containing dendrites of varying length that made contact with the hepatocytes. However, dendrite formation is not obligatory for induction o f apoptosis because cells that were unable to form these did induce FAS-dep endent apoptosis in hepatocytes. Apparently both direct as well as distant contact resulted in apoptosis. Formation of the dendrites was calcium-depen dent as EGTA largely prevented it. Importantly, chelation of the calcium al so suppressed killing of the hepatocytes. Within 1 h after addition of the A-NK cells, morphological changes in hepatocytes that are characteristic of apoptosis, such as the formation of apoptotic bodies and fragmented nuclei , became apparent. Specifically, the actin cytoskeleton of the hepatocytes was remodeled resulting in the formation of the apoptotic bodies. Inhibitio n of caspase activity by z-Val-Ala-DL-Asp-fluoromethylketone (100 muM) part ly protected against the rearrangement of the actin filaments in the hepato cytes. Cell Motil. Cytoskeleton 49.78-92, 2001. (C) 2001 Wiley-Liss, Inc.