Uptake of pp65 in in vitro Generated pp65-Positive Polymorphonuclear Cells Mediated by Phagocytosis and Cell Fusion?

Abstract

Objective: The cytomegalovirus (CMV) antigenemia consists of the detection of CMV pp65 in the nucleus of polymorphonuclear granulocytes (PMN), but it is unclear where and how PMN pick up virus particles or proteins. In an in vitro model for CMV antigenemia we investigated the mechanism of pp65 uptake by PMN that results in its expression in the nucleus. Methods: A series of inhibitors of different mechanisms was used to study the uptake of pp65 by PMN during coculture with CMV-infected endothelial cells and we performed a morphological analysis by light and transmission electron microscopy. Results: Nocodazole and cytochalasin B inhibited uptake of pp65 by PMN with 59.4 ± 14.1 and 73.3 ± 12.7%, respectively. The presence of anti-CMV hyperimmune globulin or lactoferrin during coculture reduced the number of pp65-positive PMN with 45.8 ± 7.0 or 40.6 ± 3.2%. Furthermore, a small number of the pp65-positive PMN obtained after coculture had fused to large cells with multilobed nuclei. PMN were observed that enclosed viral particles as well as free viral particles containing PMN in the cytoplasm. Conclusion: Fusion of viral particles with PMN and phagocytosis are both involved in the uptake of pp65.

Betaherpesviruses in Adult Liver Transplant Recipients

“…Moreover, neutropenia can raise difficulties to perform this technique due to the fact that antigenemia requires a sufficient number of neutrophils to detect CMV viral antigen. CMV causes an abortive replication within neutrophils and leads to uptake of antigen in perinuclear area (Kas-Deelen et al, 2001) which can be detected by use of monoclonal antibodies against pp65-antigen. Either fluorescent or enzyme labelled conjugate can be used to reveal the reaction.…”

Dissemination of Rat Cytomegalovirus through Infected Granulocytes and Monocytes In Vitro and In Vivo

“…We previously postulated that HCMV might employ MNC and PMN as a vehicle for its dissemination through the body (15,21). Using clinical isolates of HCMV, we and others have shown that PMN and MNC can acquire infective HCMV in vitro by coculture on infected cells.…”

“…Using clinical isolates of HCMV, we and others have shown that PMN and MNC can acquire infective HCMV in vitro by coculture on infected cells. Subsequently, these leukocytes were able to retransmit the virus to uninfected cells (10,14,15,19,25). The presence of intact viral particles in these leukocytes could also be demonstrated with electron microscopy (10,15,19).…”

“…Subsequently, these leukocytes were able to retransmit the virus to uninfected cells (10,14,15,19,25). The presence of intact viral particles in these leukocytes could also be demonstrated with electron microscopy (10,15,19).We hypothesized that infected MNC and PMN facilitate the dissemination of CMV in vivo. To test our hypothesis, we transferred rat CMV (RCMV) by coculturing from infected fibroblasts to phagocytes.…”

“…Generation of HCMV-infected MNC and PMN can be accomplished only when low-passage-number clinical isolates are used (14,15,19). Comparison of genomes of clinical isolates and laboratory strains of HCMV revealed the presence of approximately 20 additional open reading frames in clinical isolates, which have been only partially characterized so far (6).…”