Rickettsial interactions with human endothelial cells in vitro: adherence and entry

Rickettsia prowazekii, Madrid E strain, was assessed for its ability to enter endothelial cells derived from the veins of human umbilical cord in vitro. Rickettsial entry increased linearly with multiplicity of infection up to a multiplicity of 500; thereafter, additional rickettsiae adhered, but without a concomitant increase in the number of intracellular rickettsiae. Rickettsial entry required participation both of rickettsiae and endothelial cells; inactivation of rickettsiae with N-ethylmaleimide or Formalin, or of endothelial cells with cytochalasin B or D or NaF greatly reduced rickettsial entry. Because rickettsiae adhered to inactivated endothelial cells, adherence could be examined in the absence of entry. Rickettsial adherence was inhibited by poisons that inhibited rickettsial hemolysis. Calcium ionophore A23187, which did not inhibit endothelial pinocytosis, stimulated rickettsial adherence to endothelial cells, but inhibited rickettsial entry. These results indicated that typhus rickettsiae entered endothelial cells via induced phagocytosis, and that the signal for entry, which was dependent upon rickettsial energy, probably involved formation of a calcium gradient.