Susanna Prösch

CYTOMEGALOVIRUS INFECTION IN TRANSPLANT RECIPIENTS : THE ROLE OF TUMOR NECROSIS FACTOR

Human cytomegalovirns (CMV) infection is an important cause of morbidity and mortality in transplant recipients. CMV infection commonly results from the reactivation of a latent infection. Using a set of monoclonal anti-CMV antibodies, we found CMV antigen expression in peripheral blood mononuclear cells (PBMNC), particularly in monocytes, in 312 of 816 samples from 190 allograft recipients. The detection of CMV-IE antigens and CMV-IE DNA in PBMNC indicates that positive cells may represent truly infected cells. The relation between increased cytokine plasma levels (particularly following treatment by pan-T cell antibodies) and the appearance of CMV antigens in PBMNC suggests that cytokines may play an important role in the reversal of CMV latency. This hypothesis is supported by our finding that tumor necrosis factor-alpha (TNF) is able to stimulate the activity of the CMV-IE enhancer/promoter region in the human monocytic cell line, HL-60. The interleukins 1,2,3,4,6,8 and 10; transforming growth factor-β: interferongamma; and granulocyte/macrophage colony-stimulating factor did not show any enhancing effect on the CMV promoter activity. Thus, TNF-alpha seems to play a key role in regulating the balance between latency and reactivation of CMV infection. Inhibition of TNF-alpha release or action may be an alternative strategy for preventing CMV-associated morbidity in allograft recipients.

CYCLIC ADENOSINE MONOPHOSPHATE-RESPONSIVE ELEMENTS ARE INVOLVED IN THE TRANSCRIPTIONAL ACTIVATION OF THE HUMAN IL-10 GENE IN MONOCYTIC CELLS

IL‐10 plays an important role in the regulation of immune responses. We and others have demonstrated recently that cyclic adenosine monophosphate (cAMP)‐elevating substances up‐regulate monocytic IL‐10 expression in vitro and in vivo. Computer analysis of the IL‐10 promoter/enhancer region localized four putative cAMP‐responsive elements (CRE1– 4) with homology to the CRE consensus motif. In electrophoretic mobility shift assays CRE1 and CRE4 bound protein complexes consisting of transcription factors CREB‐1 and ATF‐1, while CRE3 bound only marginal amounts of CREB‐1/ATF‐1 in combination with unknown protein(s). CRE2 showed no protein binding activity. In vitro mutation of CRE1 and CRE4 reduced the level of cAMP‐stimulated transactivation in reporter gene assays in comparison to the wild‐type promoter by 20 % and 50 %, respectively, while mutation of CRE3 had no effect. The main action of CRE4 on cAMP‐dependent stimulation is probably based on its adjacent localization to the TATA box and its sequence comprising a perfect half site. Experiments with double and triple mutants and with deleted promoter fragments indicated the participation of additional elements beside the CRE motifs in the cAMP‐dependent stimulation. Our data suggest that intracellular cAMP may directly affect expression of the immunoregulatory cytokine IL‐10 in monocytic cells via activation of the eukaryotic transcription factors CREB‐1 and ATF‐1 and their binding to CRE1 and CRE4 in the upstream enhancer of the IL‐10 promoter.

Mechanisms of human cytomegalovirus (HCMV) (re)activation and its impact on organ transplant patients

Human cytomegalovirus (HCMV) infection plays an important role in transplant patients. Its impact is both direct and indirect. This review focuses on new aspects of HCMV (re)activation and HCMV related pathology, particularly HCMV‐associated renal allograft injury. During the last two years we have learned that HCMV is more frequently (re)activated, even in healthy people, than previously expected. Inflammatory as well as stress mediators and some drugs may (re)activate the virus by using distinct intracellular pathways. Commonly, HCMV (re)activation is accompanied by HCMV antigenemia/DNAemia, suggesting that precursor cells in the bone marrow play an important role as a reservoir of latent virus. However, local HCMV (re)activation (colon, lung) without detection of active HCMV infection in the peripheral blood is possible. In healthy people a sufficient type 1 T‐cell response controls the active HCMV infection, while in patients with severe immune deficiency (AIDS, high‐dose immunosuppression) the virus can spread in an uncontrolled fashion and induce ‘classic’ HCMV disease. In patients with moderate immune deficiency (e.g. long‐term transplant patients on low‐dose immunosuppression) virus spreading is controlled but the elimination of cells harboring the active virus may be insufficient. The resulting persistent HCMV antigenemia may induce chronic inflammatory processes leading to tissue injury, particularly in the allograft. Therefore, antiviral therapy may be useful in patients suffering from graft deterioration with otherwise clinically symptomless HCMV infection. HCMV‐related immune deficiency with an increased risk of developing bacterial/fungal superinfections is frequently seen in patients with symptomatic HCMV disease but not in asymptomatic CMV antigenemia. The risk of developing superinfections can be predicted by flow‐cytometric monitoring of peripheral blood monocytes.

Catecholamines trigger IL-10 release in acute systemic stress reaction by direct stimulation of its promoter/enhancer activity in monocytic cells

Acute stress reactions (e.g. linked with trauma, major surgery, psychic stress and myocardial infarction) are accompanied with temporary systemic release of the anti-inflammatory cytokine IL-10 followed by immunodepression. Since an association between activation of the sympathetic system and IL-10 release has been described, we studied the influence of catecholamines on its promoter activity in vitro. Using reporter gene assays we demonstrated that catecholamines in monocytic cells directly stimulate the IL-10 promoter/enhancer via a cAMP/protein kinase A-dependent pathway. A cAMP responsive element was identified as major target. Thus, catecholamines are directly involved in the regulation of immunoresponsiveness under stressful conditions.

Human Cytomegalovirus Reactivation during Lactation and Mother-to-Child Transmission in Preterm Infants

ABSTRACT In a clinical trial, the incidence of cytomegalovirus reactivation in breastfeeding mothers and transmission to their preterm infants were studied. Breast milk from 73 mothers as well as urine and tracheal and pharyngeal aspirates from their 89 infants were screened weekly for human cytomegalovirus (HCMV) DNA during the first 2 months after delivery. Of the 73 mothers, 48 (66%) were positive for HCMV DNA in the lactating breast. HCMV reactivation could be confirmed for 19 of 20 (95%) immunoglobulin G-positive mothers. Of the eight immunoglobulin G-negative mothers one was positive for HCMV DNA in breast milk. In only 2 out of 13 seropositive mothers with HCMV DNA in breast milk could viral DNA be detected in the peripheral blood. HCMV mother-to-child transmission was concluded for 20 of the 48 (42%) mothers positive for DNA or 7 of 19 (37%) seropositive for HCMV and positive for HCMV DNA in breast milk and one of one mother seronegative for HCMV but positive for HCMV DNA in breast milk. One mother transmitted the virus to her twins. In addition, one infant acquired postnatal HCMV infection despite the mothers being negative for HCMV DNA in breast milk; altogether, we found 22 infants with HCMV infection. In 13 of these 22 infants, virus infection occurred definitively postnatally; two of them developed severe symptomatic HCMV infection. HCMV-infected infants demonstrated higher incidences of amniotic infection, respiratory distress syndrome, bronchopulmonary dysplasia, and retinopathia praenatalis than noninfected infants, however, the differences were not statistically significant. In summary, our study confirmed a very high incidence of HCMV reactivation in mothers during lactation and a significant risk of transmission to preterm infants with the possibility of severe disease in these babies.

Human Cytomegalovirus Reactivation in Bone-Marrow-Derived Granulocyte/Monocyte Progenitor Cells and Mature Monocytes

Monocyte/granulocyte progenitor cells of the bone marrow are a major site of human cytomegalovirus (HCMV) latency. The mechanisms of establishment and maintenance of HCMV latency are still unknown. Reactivation of the latent virus in bone-marrow-derived progenitor cells has been demonstrated in vitro and suggested to occur also in vivo. Clinical studies have shown that reactivation is a rather frequent event not only in immunosuppressed but also in nonimmunosuppressed patients and in healthy blood donors. At least three independent mechanisms of virus reactivation are discussed: systemic inflammation connected with strong tumor necrosis factor alpha release; application of cAMP-elevating drugs, and highly stressful events associated with increased plasma catecholamine levels.

Human adenovirus and human cytomegalovirus infections in preterm newborns: no association with bronchopulmonary dysplasia.

Connatal infection with human adenovirus (HAdV) has been recently proposed as a cofactor for the development of bronchopulmonary dysplasia (BPD) in preterm infants [Couroucli et al. 2000 Pediatr Res 47:225–232]. In another study, BPD was associated with an increased incidence of human cytomegalovirus (HCMV) infection [Sawyer et al. 1987 Am J Dis Child 141:303–305]. During a 18-mo study period, we investigated tracheal aspirates or pharyngeal aspirates and urine samples collected during the first month of life from 66 preterm newborns with very low birth weight (≤1.500 g) for replication-potent HAdV as well as for adenoviral and HCMV DNA by virus culture and qualitative DNA PCR. Thus, our study included not only prenatal but also peri- and postnatal infections. Thirty-seven percent (24/66) of infants developed BPD1, as defined by persistent oxygen dependency at day 28 of life. Replication-potent HAdV and/or adenoviral DNA could be detected repeatedly in tracheal aspirates/pharyngeal aspirates and/or urine from 20% (13/66) of preterm infants. Seventeen percent (4/24) of infants in the BPD1 group and 21% (9/42) of infants in the non-BPD group had an HAdV infection, indicating that in our study the very recently proposed association between HAdV infection of the lung and BPD could not be confirmed. For comparison, active HCMV infection was diagnosed in 18% (12/66) of infants, 3 of which developed HCMV disease. 29% (7/24) in the BPD1 group and 12% (5/42) in the non-BPD group were positive for HCMV. Again, there was no statistically significant association between HCMV infection and BPD. In summary, our findings indicate that HAdV and HCMV infection are frequent in preterm newborns with very low birth weight; however, a causal association with the development of BPD seems unlikely.

Persistent CMV infection correlates with disease activity and dominates the phenotype of peripheral CD8+ T cells in psoriasis

Background:  Previously, we have reported a frequent association of active plaque psoriasis with inflammation‐mediated cytomegalovirus (CMV) reactivation.

Mutations in the hemagglutinin gene associated with influenza virus resistance to norakin

SummaryThe nucleotide sequences of the hemagglutinin genes of four norakinresistant mutants of Influenza A/FPV/Weybridge were determined and compared to the wild-type hemagglutinin. All mutants show one or two amino acid substitutions which are discussed to destabilise the pH 7 conformation of hemagglutinin.

Frequent detection of viral nucleic acids in heart valve tissue.

ABSTRACT Due to a paucity of published data concerning the prevalence of viral nucleic acid in homografts, we analyzed tissue from 30 donor hearts for the presence of viral genome sequences of enteroviruses, adenoviruses, human cytomegalovirus, and influenza virus using different PCR techniques. Viral DNA was amplified in 64 and 52% of the subvalvular myocardial tissue and non-coronary valve samples, respectively. These findings, compared with clinical history and histologic and serologic analysis, demonstrate the importance of viral safety measures in heart valve banking.