Assessment of human herpesvirus-6 infection in mesenchymal stromal cells ex vivo expanded for clinical use.

Infection or reactivation of human herpesvirus (HHV)-6 represents a potentially serious complication (often involving the central nervous system) in patients receiving either solid organ or hematopoietic stem cell transplantation. The objective of this study was to assess the risk of HHV-6 infection/reactivation in mesenchymal stromal cells (MSCs). MSCs are multipotent cells displaying immunomodulatory properties that have been already successfully used in the clinical setting to enhance hematopoietic stem cell engraftment and to treat steroid-refractory acute graft-versus-host disease. We analyzed 20 samples of ex vivo expanded MSCs, at different passages of culture, isolated both from bone marrow and from umbilical cord blood. Through Western blotting and immunocytochemistry techniques, we investigated the presence of the HHV-6 receptor (CD46) on cell surface, whereas the presence of HHV-6 DNA was evaluated by nested polymerase chain reaction assay. All of the MSC samples tested were positive for the virus receptor (CD46), suggesting their potential susceptibility to HHV-6. However, none of the MSC samples derived from cultures, performed in the perspective of clinical use, was found to harbor HHV-6. This preliminary observation on a consistent number of MSC samples, some of them tested at late in vitro passages, indicates a good safety profile of the product in terms of HHV-6 contamination. Nevertheless, it remains important to set up in vitro experimental models to study MSCs' susceptibility to HHV-6 (and HHV-7) infection, to verify their capacity to integrate the virus into cellular DNA, and to investigate which experimental conditions are able to induce virus reactivation.

An assessment of herpesvirus co-infections in patients with CMV disease: correlation with clinical and virologic outcomes.

The effect of herpesvirus co-infections (HHV-6, HHV-7) on cytomegalovirus (CMV) disease and its response to therapy is unknown. We prospectively analyzed herpesvirus co-infections in transplant recipients with CMV disease. All patients received 3 weeks of antiviral therapy. Samples were collected at baseline (day 0) and then day 3, 7, 14 and 21 poststart of therapy. Viral load testing for CMV, HHV-6 and HHV-7 was done using quantitative PCR assays in 302 patients of whom 256 had documented symptomatic CMV viremia. In this subset, day 0 HHV-6 co-infection was present in 23/253 (9.1%) and HHV-7 in 17/253 (6.7%). Including those positive at any time point raised the prevalence to 79/256 (30.9%) for HHV-6 and 75/256 (29.3%) for HHV-7. Viral co-infection did not influence the response of CMV disease to antiviral therapy. Baseline CMV viral loads, time to eradication and risk of recurrence were similar in patients with and without HHV-6 or HHV-7 co-infection. Ganciclovir and valganciclovir had no clear effect on HHV-6 and HHV-7 viremia. In conclusion, herpesvirus co-infections are common in patients with CMV disease but with standard antiviral therapy, no clear clinical effects are discernable. Routine monitoring for viral co-infection in patients with CMV disease is not indicated.

Cardiovascular magnetic resonance assessment of human myocarditis: a comparison to histology and molecular pathology.

BACKGROUND: Myocarditis can occasionally lead to sudden death and may progress to dilated cardiomyopathy in up to 10% of patients. Because the initial onset is difficult to recognize clinically and the diagnostic tools available are unsatisfactory, new strategies to diagnose myocarditis are needed. METHODS AND RESULTS: Cardiovascular MR imaging (CMR) was performed in 32 patients who were diagnosed with myocarditis by clinical criteria. To determine whether CMR visualizes areas of active myocarditis, endomyocardial biopsy was taken from the region of contrast enhancement and submitted to histopathologic analysis. Follow-up was performed 3 month later. Contrast enhancement was present in 28 patients (88%) and was usually seen with one or several foci in the myocardium. Foci were most frequently located in the lateral free wall. In the 21 patients in whom biopsy was obtained from the region of contrast enhancement, histopathologic analysis revealed active myocarditis in 19 patients (parvovirus B19, n=12; human herpes virus type 6 [HHV 6], n=5). Conversely, in the remaining 11 patients, in whom biopsy could not be taken from the region of contrast enhancement, active myocarditis was found in one case only (HHV6). At follow-up, the area of contrast enhancement decreased from 9+/-11% to 3+/-4% of left ventricular mass as the left ventricular ejection fraction improved from 47+/-19% to 60+/-10%. CONCLUSIONS: Contrast enhancement is a frequent finding in the clinical setting of suspected myocarditis and is associated with active inflammation defined by histopathology. Myocarditis occurs predominantly in the lateral free wall. Contrast CMR is a valuable tool for the evaluation and monitoring of inflammatory heart disease.

HHV-6 A- or B-specific P41 antigens do not reveal virus variant-specific IgG or IgM responses in human serum.

The etiology of multiple sclerosis (MS) remains unknown, but there are indications of a role of human herpesvirus 6 (HHV-6), especially variant A, in the pathogenesis. Higher serum antibody reactivity against an HHV-6 early protein, p41, has been found in MS cases than in controls. The antigen, however, was purified from infected cells with a monoclonal antibody also reactive with a protein (p38) likely to be of cellular origin. To avoid serological crossreactivity with the cellular protein, recombinant p41 proteins from HHV-6A strain GS and HHV-6B strain Z29 were expressed as glutathione-S-transferase fusion proteins (p41-GST), and used as antigens in an enzyme-linked immunosorbent assay (ELISA). p41 variant specific monoclonal antibodies reacted strongly with the respective recombinant proteins. Serum IgM and IgG reactivities with the recombinant p41 antigens were analysed in patients with manifest MS, patients with optic neuritis, patients with other neurological diseases, and in one group of healthy controls. All sera were HHV-6 IgG seropositive by immunofluorescence. The serum IgM or IgG reactivities against the recombinant p41 antigens did not differ significantly between the groups, and the reactivities against the variant A and B antigens were identical. In many samples, the reactivity was very low. The results indicate that p41 is not an optimal target for HHV-6 serology studies, and that the data obtained with the p41 antigen prepared from infected cells (possibly including also p38) should be interpreted with caution.