Cytomegalovirus (CMV) infection in the period of temporary immunodeficiency after haematoablative treatment and bone marrow transplantation (BMT) is associated with a risk of graft failure and multiple-organ CMV disease. The efficacy of immune system reconstitution is decisive for the prevention of CMV pathogenesis after BMT. Previous data in murine model systems have documented a redundancy in the immune effector mechanisms controlling CMV. CD8 T cells proved to be relevant but not irreplaceable as antiviral effectors. Specifically, in a state of long-term in vivo depletion of the CD8 T-cell subset, CD4 T cells were educed to become deputy effectors controlling CMV by a mechanism involving antiviral cytokines. It is of medical importance to know whether one can trust in this ' flexible defence ' in all clinical settings. It is demonstrated here that reconstitution of CD8 T cells is crucial for the prevention of fatal multipleorgan CMV disease under the specific conditions of BMT.Patients undergoing bone marrow transplantation (BMT) associated with temporary immunodeficiency are at risk of fatal cytomegalovirus (CMV) disease with multiple organ involvement (for reviews see Winston et al., 1990 ; Einsele et al., 1998). Whether or not primary or recurrent CMV infections develop into disease appears to be related to the efficacy of haematopoietic and immune system reconstitution. Specifically, early reappearance of CD8 T cells is of positive prognostic value for recovery (Reusser et al., 1991). Accordingly, restoration of immunity to CMV by adoptive cytoimmunotherapy with antiviral CD8 T-cell lines is in clinical trials (Riddell et al., 1992 ;Walter et al., 1995). In BMT performed across minor or class I major histocompatibility disparities, graft vs. host disease (GvHD) is a further risk factor (Einsele et al., 1998). In a major Author for correspondence : Matthias Reddehase.Fax j49 6131 395604. e-mail Matthias.Reddehase!uni-mainz.de histocompatibility complex (MHC) class II-identical but MHC class I-disparate setting, selective depletion of class I-restricted CD8 T cells might be discussed as a regime for GvHD prophylaxis. However, if CD8 T cells were the only effectors Fig. 1. (A) Lethality of CMV infection after selective T-cell subset depletion. Syngeneic BMT was performed on day 0 with BALB/c mice as BM cell donors and recipients. Recipients were infected with 1i10 5 p.f.u. of purified murine CMV ca. 2 h after BMT. In vivo depletion of T-cell subsets was performed twice, namely on days 7 and 14, by intravenous administration of 1 mg of the respective antibody. Kaplan-Meyer plots show the survival rates for 20 recipients per experimental group as a function of time. (B) Efficacy and specificity of the T-cell subset depletions. Pulmonary infiltrate leukocytes were isolated on day 21 and analysed by three-colour cytofluorometry for the expression of TCR α/β (phycoerythrin fluorescence, FL-2), CD4 (RED613 fluorescence, FL-3) and CD8 (FITC fluorescence, FL-1). Gates were set on lymphocytes and on positive FL-2 in ...

Cytomegaloviruses express glycoproteins that interfere with antigen presentation to CD8 T cells. Although the molecular modes of action of these "immunoevasins" differ between cytomegalovirus species, the convergent biological outcome is an inhibition of the recognition of infected cells. In murine cytomegalovirus, m152/gp40 retains peptide-loaded major histocompatibility complex class I molecules in a cis-Golgi compartment, m06/gp48 mediates their vesicular sorting for lysosomal degradation, and m04/gp34, although not an immunoevasin in its own right, appears to assist in the concerted action of all three molecules. Using the L d -restricted IE1 epitope YPHFMPTNL in the BALB/c mouse model as a paradigm, we provide here an explanation for the paradox that immunoevasins enhance CD8 T-cell priming although they inhibit peptide presentation in infected cells. Adaptive immune responses are initiated in the regional lymph node (RLN) draining the site of pathogen exposure. In particular for antigens that are not virion components, the magnitude of viral gene expression providing the antigens is likely a critical parameter in priming efficacy. We have therefore focused on the events in the RLN and have related priming to intranodal viral gene expression. We show that immunoevasins enhance priming by downmodulating an early CD8 T-cell-mediated "negative feedback" control of the infection in the cortical region of the RLN, thus supporting the model that immunoevasins improve antigen supply for indirect priming by uninfected antigen-presenting cells. As an important consequence, these findings predict that deletion of immunoevasin genes in a replicative vaccine virus is not a favorable option but may, rather, be counterproductive.

Bone marrow (BM) failure associated with cytomegalovirus (CMV) infection is a feared complication after clinical BM transplantation. Experiments in long-term BM cultures have indicated that BM stromal cells (BMSC) are targets of productive CMV infection, but an in situ infection of BM stroma remained to be documented, and the pathomechanism is open to question. Here we describe a murine in vivo model of lethal CMV aplastic anemia (CMV-AA). The reconstitution of hematopoietic progenitor cells expressing stem cell factor (SCF) receptor was found to be defective in CMV-AA. While murine CMV replication in permissive parenchymal tissues is cytolytic, the hematopoietic cord was found to be a site of very limited virus production with foci of reticular BMSC expressing the intranuclear viral IE1 protein, but with only a few BMSC positive for viral genome in the in situ hybridization. XX-XY BM chimeras were established in order to quantitate Y-chromosome-tagged BMSC by a PCR specific for the male-sex-determining gene Tdy. This approach revealed that murine CMV infection is not associated with a significant loss of BMSC. Despite the physical integrity of the stromal network, the functional integrity of the stroma was impaired. While housekeeping genes were expressed normally in BMSC of infected mice, the expression of genes encoding the essential hemopoietins SCF, granulocyte colony-stimulating factor, and interleukin-6 was markedly reduced. In conclusion, the mechanism of BM failure is not a stromal lesion but an insufficient stromal function. These findings explain CMV-AA as a manifestation of multiple hemopoietin deficiency.

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During immunodeficiency after sublethal haematoablative treatment, cytomegalovirus (CMV) infection interferes with haematopoietic reconstitution and can cause lethal bone marrow (BM) aplasia. The in vivo model of murine CMV infection has identified the BM stroma as the principal target site of CMV in the haematopoietic cord. The infected cell type is the reticular stromal cell which forms the stromal network and produces essential haemopoietins, such as stem-cell factor (SCF). The expression of SCF was found to be reduced in the infected stroma, but the stromal network was not disrupted and the number of infected stromal cells was too low to explain the functional deficiency. These facts call for a negatively regulating cytokine that is induced by the infection and that potentiates the direct effect of infection by down-regulating haemopoietins in un-