Secretion of IFN-γ by specific T cells in HCMV infection.

One major risk for recipients undergoing allogeneic hematopoietic stem cell transplants (allo-HSCTs) is infection with the human cytomegalovirus (HCMV). For HCMV treatment, it is especially crucial to be able to differentiate between recipients who are at high risk of reactivation and those who are not. In this study, HCMV-DNA was collected from 60 HLA-A*02 allo-HSCT recipients before and after transplantation. After transplantation, the release of interferon (IFN)-γ by T cells specific to HCMV was assessed using the enzyme-linked immunospot assay (ELISPOT). The results show that the median viral load (VL) was significantly higher in the HCMV persistent-infection group compared to the non-persistent-infection group (p = 0.002), and that the late-infection rate was considerably higher in the high-VL group compared to the low-VL group (p = 0.014). The uninfected group had a considerably higher median IFN-γ spot-forming cell (SFC) count than the persistent-infection group (p = 0.001), and IFN-γ SFC counts correlated negatively and linearly with VLs (r = -0.397, p = 0.002). The immune-response groups showed significantly difference in median VL (p = 0.018), and the high immune response group had a reduced late-infection rate than the no/low immune response groups (p = 0.049). Our study showed that allo-HSCT recipients with a high VL at an early transplantation stage were at high risk for late HCMV infection. Further HCMV reactivation can be prevented by HCMV-specific T cells secreting enough IFN-γ.

Early immune surveillance to predict cytomegalovirus outcomes after allogeneic hematopoietic stem cell transplantation.

Background: There is no method of predicting human cytomegalovirus (HCMV) outcomes in allogeneic hematopoietic stem cell transplant recipients clinically, leading in some cases to excessive or insufficient antiviral therapy. We evaluated the early immune response of recipients with disparate HCMV outcomes. Methods: The HCMV outcomes of recipients were determined by long-term monitoring of HCMV DNA levels posttransplant. HCMV IgG and IgM concentrations at 1 week before and 1 week after transplantation, absolute lymphocyte counts, and HCMV-specific IFN-γ secreting cells at 1 month posttransplant were evaluated based on HCMV outcome. Results: All recipients were negative for HCMV IgM. Significant differences between recipients with and without HCMV reactivation were observed in pre- and post-transplant HCMV IgG antibody levels, absolute lymphocyte counts, and HCMV-specific IFN-γ secreting cells (P < 0.05). HCMV IgG antibody levels significantly increased after transplantation in recipients with HCMV reactivation (P = 0.032), but not in those without reactivation. Multivariate analysis revealed that except for the absolute lymphocyte count these biomarkers were related to HCMV reactivation, independent of other clinical factors. In time-to-event analyses, lower levels of these biomarkers were associated with an increased 150-day cumulative incidence of HCMV reactivation (log-rank P < 0.05). In recipients with HCMV reactivation, the duration of HCMV DNAemia had negative correlation with HCMV-specific IFN-γ-secreting cells (P = 0.015, r = -0.372). The relationships between the peak HCMV DNA load and absolute lymphocyte count and HCMV-specific IFN-γ-secreting cells followed the same trends (P = 0.026, r = -0.181 and P = 0.010, r = -0.317). Conclusions: HCMV IgG, absolute lymphocyte count, and HCMV-specific IFN-γ secreting cells represent the humoral and cellular immune response. Early monitoring of these immune markers could enable prediction of HCMV outcomes posttransplant and assessment of the severity of HCMV DNAemia.

ELISPOT assay of interferon-γ secretion for evaluating human cytomegalovirus reactivation risk in allo-HSCT recipients.

Human cytomegalovirus (HCMV) is a common cause of significant morbidity and mortality in transplant recipients after allogeneic hematopoietic stem cell transplantation (allo-HSCT). We evaluated interferon-γ (IFN-γ) secretion by HCMV NLV-specific CD8+ T cells in HCMV-reactivated allo-HSCT recipients using an enzyme-linked immunospot (ELISPOT) assay at 3 months post-transplantation. Blood samples from 47 recipients were tested for HCMV DNAemia, HCMV pp65 antigenemia, and anti-HCMV immunoglobulins (IgG/IgM) over 3 months post-transplantation. Of the 47 transplant recipients, 26 were HLA-A*02 positive and 21 were HLA-A*02 negative. The results were essentially consistent between the 47 transplant recipients and the HLA-A*02-positive recipients. HCMV DNAemia was not linearly correlated with IFN-γ spot-forming cells (SFCs) counts; IFN-γ SFCs counts did not differ significantly between the HCMV DNAemia-positive and -negative groups, whereas the HCMV-DNA virus loads were inversely correlated with the IFN-γ SFCs counts. HCMV pp65 antigenemia was not linearly correlated with IFN-γ SFCs counts; IFN-γ SFCs counts in the HCMV pp65 antigenemia-positive and -negative groups were similar. More IFN-γ SFCs counts were detected in transplant recipients with high anti-HCMV-IgG antibody titers than in those with low anti-HCMV-IgG titers pre-transplantation in the 47 recipients. Anti-HCMV-IgG antibody titers were positively linearly correlated with IFN-γ SFCs counts in HLA-A*02-positive recipients. The HCMV infection indicators used to monitor HCMV reactivation had different values in transplant recipients. The use of the IFN-γ SFCs counts measured by ELISPOT to evaluate the risk of HCMV reactivation needs further study.

Risk Factors Analysis for Human Cytomegalovirus Viremia in Donor+/Recipient+ Hematopoietic Stem Cell Transplantation.

OBJECTIVE: To assess the rate of, and risk factors for, human cytomegalovirus viremia (HCMV) in donor+/recipient+ (HCMV serostatus matched) hematopoietic stem-cell transplantation (HSCT) recipients. METHODS: HCMV DNA from 144 donor+/recipient+ HSCT recipients was examined by quantitative polymerase chain reaction (qPCR). RESULTS: The cumulative incidence of HCMV viremia was 69.4% (100/144) during the 48 weeks after HSCT. In a multivariate analysis, acute graft-versus-host disease (aGVHD) was discovered to be a risk factor for the occurrence of HCMV viremia (P = .006). The cumulative incidence of HCMV viremia and increasing DNA loads were significantly associated with aGVHD occurrence (P = .001 for each). The occurrence of late-term HCMV viremia was associated with aGVHD (P = .001) and a higher DNA load during the first 12 weeks after HSCT (P = .04). CONCLUSIONS: aGVHD is a risk factor for HCMV viremia. Recipients with aGVHD who have a high HCMV DNA load should be strictly monitored to prevent HCMV activation.

Relationship between T-cell receptor beta chain sequences and human cytomegalovirus infection in allogeneic hematopoietic stem cell transplant recipients.

In the present study, clonal amplifications of T-cell receptor ß variable (TCR BV) linked to human cytomegalovirus (HCMV) infection were detected in recipients of allogeneic hematopoietic stem cell transplants (HSCT), and certain relationships between them were identified. Furthermore, the relationship between TCR BV sequences and HCMV infections was investigated. The results indicated that the 3 recipients of HSCT had monoclonal expansion of specific TCR BV clones following HSCT. Among these recipients, 2 suffered from pp65 and immediate early (IE) antigenemia. These patients demonstrated preferential expansion of TCR BV9 (QVRGGTDTQ) and TCR BV11 (VATDFQ). The remaining recipient did not express TCR BV9 and TCR BV11, nor did this individual have pp65 and IE antigenemia. These results suggest that expression of TCR BV9 and TCR BV11 may be associated with HCMV antigenemia, and may be involved in the immune response. The amino acid sequences 'QVRGGTDTQ' and 'VATDFQ' may be involved in HCMV reactivation in patients who have undergone HSCT. Assessment of the TCR BV families may provide valuable insight into HCMV pathogenesis and may aid in the diagnosis and therapy for HSCT recipients infected with HCMV.

Herpesvirus infections in hematopoietic stem cell transplant recipients seropositive for human cytomegalovirus before transplantation.

BACKGROUND: Viral infections are a major cause of morbidity and mortality after hematopoietic stem cell transplantation (HSCT). The effect of herpesvirus infections in human cytomegalovirus (HCMV)-seropositive (IgG-positive/IgM-negative) HSCT recipients remains poorly understood. The risk factors associated with Epstein-Barr virus (EBV), HCMV, and human herpes virus type 6 (HHV-6) infections after HSCT, both alone and in combination, were investigated in this study. METHODS: Peripheral blood specimens were collected from 44 HSCT recipients and examined for viral DNA using quantitative fluorescence PCR assays. Risk factors for EBV, HCMV, and HHV-6 infections were analyzed by binary logistic regression, and relationships between these viruses were analyzed using the Chi-square test. RESULTS: EBV, HCMV, and HHV-6 were detected in 50%, 45.45%, and 25% of HCMV-seropositive (IgG-positive/IgM-negative) HSCT recipients, respectively. Male sex (p=0.007) and conditioning regimens including anti-thymocyte globulin (ATG) (p=0.034) were strongly associated with an increased risk of EBV infection. Graft-versus-host disease (GVHD) prophylaxis with corticosteroids was a risk factor for both EBV (p=0.013) and HCMV (p=0.040) infections, while EBV infection (p=0.029) was found to be an independent risk factor for HHV-6 infection. Pre-existing HHV-6 infection was associated with lower rates of HCMV infection (p=0.002); similarly, pre-existing HCMV infection was protective against HHV-6 infection (p=0.036). CONCLUSIONS: HCMV-seropositive (IgG-positive/IgM-negative) HSCT recipients exhibited a high rate of herpesvirus infections, particularly EBV. ATG and male sex were strongly associated with an increased risk of EBV infection. GVHD prophylaxis with prednisone was found to affect both EBV and HCMV infections. Prior infection with EBV was shown to promote HHV-6 infection. Taken together, these data highlight the need for active monitoring of herpesvirus infections in patients undergoing HSCT.