Polymorphonuclear leukocytes (PMNL) are a major carrier of human cytomegalovirus (CMV) in viremic immunodepressed patients. We transmitted infectious virions and viral components to PMNL by coculturing these cells with infected human embryonic lung fibroblasts (HELF) or human umbilical vein endothelial cells (HUVEC). Quantitative time-course analysis of viral DNA and protein expression in PMNL, after functional separation from infected donor cells, indicated the initiation of viral cycling, with immediate-early protein expression. No viral replication or early or late gene expression was observed, but infected PMNL were able to infect naive fibroblasts more than 48 h after the end of co-culture. PMNL apoptosis was significantly delayed during co-culture with infected or uninfected HUVEC, and this phenomenon did not require contact between the two cell populations. The increased production of IL-8 in the same culture conditions that protect PMNL from apoptosis, associated with the reversion of this protection by inhibiting or depleting this factor in the culture media, targets this cytokine as a likely candidate for this protective effect. These data suggest that PMNL play a key role in virus dissemination in vivo, through their interactions with infected endothelial cells.