Cytomegalovirus infects human lymphocytes and monocytes: virus expression is restricted to immediate-early gene products.

Abstract

In this investigation, we studied the ability of human cytomegalovirus to infect peripheral blood mononuclear cells. With monoclonal antibody technology, we demonstrated that cytomegalovirus could infect human lymphocytes of T-and B-cell (PBM). Infectious CMV has occasionally been found in buffy-coat preparations (4-6) obtained from patients with clinical CMV infection; only rarely has the virus been isolated from healthy donors (7). Neither the exact cell involved in the leukocyte fraction nor the state of the virus in such cells has been established. When transformed lymphoblastoid and erythroleukemia cell lines were studied for the ability to support CMV replication, some researchers found evidence of short-term virus replication (8, 9), but rarely was infectious virus produced (10, 11). There have been occasional reports of expression of CMV antigens on B-lymphoblastoid cell lines (12). Attempts to infect normal PBM from healthy donors with laboratory-adapted strains of CMV have failed (11, 13).In the cascade regulation of the CMV genome (14), immediate-early protein synthesis precedes early polypeptide synthesis, both of which set the regulatory state for late polypeptide synthesis and production of mature virions. The early class polypeptides appear to have predominantly regulatory functions; the late polypeptides have mainly structural functions. We reasoned that if lymphocytes were abortively infected, techniques such as cocultivation assays and probes for late CMV gene products would be negative. Hence, we looked for CMV expression in peripheral blood lymphocytes (PBL) with monoclonal antibodies specifically produced to detect polypeptides relevant to major epochs in the cascade regulation of the viral genome. Prompted by observations of biological differences between strains of virus recently derived from infected patients and the fibroblastadapted strains of virus (15), we also studied both forms of this virus. MATERIALS AND METHODSLymphocyte Infection. PBM were removed from the blood of healthy human donors by density gradient centrifugation on Ficoll-Paque and infected at a multiplicity of 0.01-1.0 with CMV recently isolated from patients with various CMV syndromes (Table 1), or with stocks of plaque-purified laboratory-strain AD-169. The recent isolates were propagated in human foreskin fibroblasts (Flow Laboratories) for <12 passages. Because low-passage isolates of human virus are associated predominantly with the cell matrix (15), infected or mock-infected fibroblasts were sonicated, and this material was cultured with PBM for up to 6 days. Thereafter, the PBM were washed 3 times and prepared for immunofluorescence studies by air-drying and fixing the cells with acetone on glass slides. In some experiments, T-lymphocytes were positively selected from the PBM cultures by erythrocyterosetting techniques (17) or by sorting on a fluorescence-activated cell sorter (16).Immunofluorescence Techniques. Monoclonal antibodies were raised against CMV-infected fibroblasts with standard techniques...

Monocytes are a major site of persistence of human cytomegalovirus in peripheral blood mononuclear cells

“…This is consistent with previous evidence that clinical isolates can preferentially infect monocytes, albeit at low levels (Rice et al, 1984), and with in situ evidence suggesting monocyte carriage of HCMV in post-transplant kidneys (Gnann et al, 1988). However, our data do not address the question of how HCMV is maintained in an asymptomatic individual.…”

Antibody-Mediated Response of NKG2Cbright NK Cells against Human Cytomegalovirus

“…2 and 3). We did not detect the expression of HCMV immediate early protein IE1/2 in NK cells upon their coculture with viral particles (data not shown), confirming the established resistance of lymphocytes to HCMV infection (36,37).Virus titration experiments showed a dosedependent NK cell activation, with TNF-a produced in response to as few as 0.08 infective viral particles/cell (Supplemental Fig. 2A).…”

Comparison of Restriction Site Polymorphisms among Clinical Isolates and Laboratory Strains of Human Cytomegalovirus

“…Clearly, since we isolated virus strains with identical EcoRI and HindIII sites from multiple body sites from 14 of 20 individuals we have not identified any difference associated with tropism for oral or genitourinary sites. A genomic difference associated with a tropism for lymphocytes may eventually be found, since differences in the ability of clinical HCMV isolates to infect lymphocytes in vitro may relate to strain differences and/or degree of passage in tissue culture (Rice et al, 1984). No specific difference which can be related to length of time in tissue culture passage has been found between the laboratory strains and the clinical isolates in our study.…”