CMV-specific Immune Response Research Articles

Impact of pretransplant CMV-specific T-cell immune response in the control of CMV infection after solid organ transplantation: a prospective cohort study

IntroductionAlthough solid organ transplant (SOT) recipients with pretransplant serology for cytomegalovirus (CMV-R+) are considered at intermediate risk for CMV infection post transplantation, CMV infection remains a major cause of morbidity in this population. We prospectively characterized whether having pretransplant CMV-specific cellular immunity is independently associated with controlling infection after transplantation in R + SOT recipients. MethodsA prospective cohort of consecutive R + SOT recipients that received pre-emptive treatment for CMV infection was monitored after transplantation and variables were recorded during the follow-up. The cytomegalovirus-specific T-cell immune response was characterized by intracellular cytokine staining and viral loads determined using real-time PCR. ResultsOne hundred and thirty-five R + SOT recipients were included (67 kidney, 64 liver, four liver–kidney). Only one-third of the patients (42; 31.85%) had CMV-specific T-cell immunity (CD8+CD69+INF-γ+ T cells >0.25%) before transplantation. Patients with negative pretransplant immunity had more CMV infection (49, 52.7% vs. 15, 35.7%; p 0.07) and received more antiviral therapy than those with immunity (32, 34.4% vs. 6, 14.3%, p 0.016). Having CMV specific immunity was an independent factor for protection from developing viraemia ≥2000 IU/mL (OR 0.276, 95% CI 0.105–0.725, p < 0.01) and lower administration of treatment (OR 0.398, 95% CI 0.175–0.905, p 0.028). Only patients with no pretransplant CMV-specific T-cell response were diagnosed with CMV-disease (8, 8.6% vs. 0, 0%, p 0.05). Discussion.Our results show that having a pretransplant CMV specific T-cell response may be associated with a lower rate of CMV viraemia and less antiviral treatment after transplantation; however, more prospective studies are needed to confirm these findings.

NKG2C Natural Killer Cells in Bronchoalveolar Lavage Are Associated With Cytomegalovirus Viremia and Poor Outcomes in Lung Allograft Recipients.

Cytomegalovirus (CMV) infection is a risk factor for chronic lung allograft dysfunction (CLAD), which limits survival in lung allograft recipients. Natural killer (NK) cells that express the NKG2C receptor mediate CMV-specific immune responses. We hypothesized that NKG2C NK cells responding to CMV in the lung allograft would reduce CMV-related inflammation and would improve CLAD-free survival. We prospectively followed 130 subjects who underwent lung transplantation from 2012 to 2016. Bronchoalveolar lavage (BAL) NK cells were immunophenotyped for NKG2C, maturation, and proliferation markers. CMV viral load, serologies, serial spirometry, and mortality were recorded from medical records. Natural killer cell subset association with CMV endpoints were made using generalized estimating equation-adjusted linear models. BAL NKG2C NK cell association with CLAD-free survival was assessed by Cox proportional hazards modeling. NKG2C NK cells were more mature and proliferative than NKG2C NK cells and represented a median of 7.8% of BAL NK cells. The NKG2C NK cell proportion increased prior to the first detection of viremia and was nearly tripled in subjects with high level viremia (>1000 copies/mL) compared with no detected viremia. Subjects with increased BAL NKG2C NK cells, relative to the median, had a significantly increased risk for CLAD or death (hazard ratio, 4.2; 95% confidence interval, 1.2-13.3). The BAL NKG2C NK cell proportion may be a relevant biomarker for assessing risk of CMV viremia and quantifying potential CMV-related graft injury that can lead to CLAD or death.

The impact of inflationary cytomegalovirus-specific memory T cells on anti-tumour immune responses in patients with cancer.

Human cytomegalovirus (CMV) is a ubiquitous, persistent beta herpesvirus. CMV infection contributes to the accumulation of functional antigen-specific CD8+ T-cell pools with an effector-memory phenotype and enrichment of these immune cells in peripheral organs. We review here this 'memory T-cell inflation' phenomenon and associated factors including age and sex. 'Collateral damage' due to CMV-directed immune reactivity may occur in later stages of life - arising from CMV-specific immune responses that were beneficial in earlier life. CMV may be considered an age-dependent immunomodulator and a double-edged sword in editing anti-tumour immune responses. Emerging evidence suggests that CMV is highly prevalent in patients with a variety of cancers, particularly glioblastoma. A better understanding of CMV-associated immune responses and its implications for immune senescence, especially in patients with cancer, may aid in the design of more clinically relevant and tailored, personalized treatment regimens. 'Memory T-cell inflation' could be applied in vaccine development strategies to enrich for immune reactivity where long-term immunological memory is needed, e.g. in long-term immune memory formation directed against transformed cells.

Specific Immunity to Cytomegalovirus in Pediatric Cardiac Transplantation.

Cytomegalovirus (CMV) infection is implicated in endothelial dysfunction and graft damage after pediatric heart transplantation. CMV-specific immune responses are thought to be necessary for CMV viral control but there is little data in pediatric heart transplantation. We studied 28 consecutive pediatric heart transplant recipients for 1 year posttransplant. CMV T-cell expressing IFN-γ, TNF-α, and IL-2 in response to ex vivo stimulation with CMV lysates or peptides were measured. Circulating cytokines were measured in plasma. Generalized Additive Models were applied to the data to model changes of cell population dynamics over time. CMV-specific T cell-mediated responses were impaired in the first 8 weeks posttransplant. During this period, 25% of patients had CMV viremia, of which those with VLs of 10 000 or more CMV deoxyribonucleic acid copies/mL were given ganciclovir. In this group, the frequency of CD4+ and CD8+ T cells producing IFN-γ and the CD8+CD57+ granzyme B+ T-cell population increased at 12 to 24 weeks and remained elevated for the duration of the study. We have shown that CMV viremia is associated with CMV-specific immune responses and increased CD8+CD57+ granzyme B+ cells at 1 year posttransplant; however, early responses were not predictive of impending CMV viremia. It remains to be seen if the early CMV immune response detected is associated with endothelial and allograft damage, in light of previous studies demonstrating increased vasculopathy in pediatric patients with CMV viremia.

The Efficacy of CMV Specific Immune Reconstitution is Highly Dependent on Donor/Recipient Serostatus and Type of Solid Organ Transplanted

Introduction The CMV specific immune recovery is known to control viral infection and disease. The CMV specific cell-mediated immunity (CMI) and CMV specific humoral immune response (IgG titer and IgG avidity) was explored as candidate biomarker predictive of CMV infection. Materials 653 observations from 297 SOT including 103 kidney (K), 60 liver (L), 47 heart (H), 87 Lung (Lu) transplants. The follow-up was 0-400 days after transplant. The following parameters were examined: 1) CMV serostatus before transplant 2) CMV IgG titer 3) CMV IgG avidity 4) CMV-ELISPOT 5) primary (D+/R-) or non-primary (D±/R+) CMV infection 6) CMV viremia. Data were statistically analyzed using ANOVA and linear regression analysis and spline model. Results The main findings of the study are: 1) CMV IgG titers and avidity are not predictive biomarker for CMV infection. The CMV IgG titer and IgG avidity levels are comparable either in infected and non-infected patients. 2) CMV viral load is statistically different between infected D±/R+ and D+/R-. The viral load in infected D+/R- is 1 Log higher compared to D±/R+ (10^5 vs 10^4 respectively) 4) The CMV immune reconstitution is more rapid and efficient in D±/R+ compared to D+/R- 5) D±/R+ and D+/R- Kidney and Lung transplants display a statistically significant lower CMI compared to heart and liver transplants; 6) Kidney and Lung transplants display a similar pattern of immune recovery 7) The CMV cell mediated immunity biomarker is predictive of infection in all organs with exception of lung transplants. Conclusion The study shows a marginal role of humoral immunity in controlling CMV infection. The pattern of CMI immune recovery is highly dependent upon pre-transplant CMV serostatus, type of organ transplant and immunosuppression therapy. High levels of CMV specific cell-mediated immunity correlate with a reduced CMV infection risk in heart, kidney and liver transplants. A different scenario emerged in lung transplants: high levels of circulating CMV specific T-cells can be found in lung transplants but cannot prevent CMV infection and clear the virus. In this view the lung may be considered a "sanctuary site" for CMV replication.

Kinetic of the CMV‐specific T‐cell immune response and CMV infection in CMV‐seropositive kidney transplant recipients receiving rabbit anti‐thymocyte globulin induction therapy: A pilot study

Some studies have suggested that rATG treatment may be associated with an increased incidence of CMV infection and delayed CMV immune response. However, the evidences supporting this matter are scarce. This study aims to characterize the kinetic of the CMV-specific T-cell immune response before and after rATG induction therapy and the relationship with the development of CMV infection in CMV-seropositive kidney transplant recipients. An observational prospective study of CMV-seropositive kidney transplant patients that received rATG induction therapy was performed. A pretransplant sample was obtained before the surgery to determine the CMV-specific immunity. CMV viral load (by PCR) and CMV-specific T-cell immune response (by flow cytometry) were determined during the follow-up at 0.5, 1, 2, 3, 6, and 12months post transplantation. A total of 23 patients were included in the study. CMV prophylaxis was administrated for a media of 90days after transplantation. At the end of follow-up, 18 (78.3%) patients had CMV-specific immunity with a median value of 0.31% CD8+ CD69+ INF-γ+ T cells at a median of 16weeks post transplantation. Five patients never acquired CMV-specific immunity. No statistically significant association between CMV infection and CMV-specific T-cell immune response (P=.086) was observed. However, patients with positive pretransplant CMV-specific immunity developed earlier immunity and achieved higher levels of CD8+ CD69+ INF-γ+ T-cell post-transplantation than patients with negative pretransplant immunity. CMV-specific immune monitoring in addition to CMV-serology may be useful to stratify patient's risk of CMV infection before transplantation.

CMV-infected kidney grafts drive the expansion of blood-borne CMV-specific T cells restricted by shared class I HLA molecules via presentation on donor cells.

We aimed to determine the role of cytomegalovirus (CMV)-infected donor cells in the development of a CMV-specific immune response in kidney transplant recipients. We assessed the CMV pp65-specific immune response by using interferon-ɣ ELISPOT and dextramers in peripheral blood mononuclear cells from 115 recipients (D+R- 31, D+R+44, D-R+40) late after transplantation (mean 59±42months). Receiving a kidney from a D+ donor resulted in a higher number of IFN-ɣ-producing anti-CMV Tcells (P=.004). This effect disappeared with the absence of shared HLA class I specificities between donors and recipients (P=.430). To confirm the role of donor cells in stimulating the expansion of newly developed CMV-specific CD8+ T cells after transplantation, we compared the number of HLA-A2-restricted CMV-specific CD8+ T cells in primo-infected recipients who received an HLA-A2 or non-HLA-A2 graft. The median of anti-CMV pp65 T cells restricted by HLA-A2 was very low for patients who received a non-HLA-A2 graft vs an HLA-A2 graft (300 [0-14638] vs. 17972 [222-85594] anti-CMV pp65 CD8+ T cells/million CD8+ T cells, P=.001). This adds new evidence that CMV-infected kidney donor cells present CMV peptides and drive an inflation of memory CMV-specific CD8+ T cells, likely because of frequent CMV replications within the graft.

Cytomegalovirus-specific T-cells are associated with immune senescence, but not with systemic inflammation, in people living with HIV

In people living with HIV (PLWHIV), coinfection with cytomegalovirus (CMV) has been associated with inflammation, immunological ageing, and increased risk of severe non-AIDS related comorbidity. The effect of CMV-specific immune responses on systemic inflammation, immune activation and T-cell senescence was evaluated in 53 PLWHIV treated with combination antiretroviral therapy (cART). Activated-, terminally differentiated-, naïve-, and senescent T-cells were assessed by flow cytometry, and plasma levels of CMV IgG, interleukin-6, tumor necrosis factor-α, high-sensitivity C-reactive protein and soluble-CD14 were measured. In PLWHIV, expression of interleukin-2, tumor necrosis factor-α and interferon-γ was measured by intracellular-cytokine-staining after stimulation of T-cells with CMV-pp65, CMV-IE1, and CMV-gB. Increased CMV-specific T-cell responses were associated with a higher ratio of terminally differentiated/naïve CD8+ T-cells and with increased proportions of senescent CD8+ T-cells, but not with systemic inflammation or sCD14. Increased CMV-specific CD4+ T-cell responses were associated with increased proportions of activated CD8+ T-cells. In PLWHIV with expansion of CMV-specific T-cells or increased T-cell senescence, CMV-specific polyfunctionality was maintained. That the magnitude of the CMV-specific T-cell response was associated with a senescent immune phenotype, suggests that a dysregulated immune response against CMV may contribute to the immunological ageing often described in PLWHIV despite stable cART.

Assessing the risk of CMV reactivation and reconstitution of antiviral immune response post bone marrow transplantation by the QuantiFERON-CMV-assay and real time PCR

BackgroundCMV reactivation is a major cause of severe complications in allogeneic hematopoietic stem cell transplant (HSCT) recipients. The risk of CMV reactivation depends on the serostatus (+/−) of the donor (D) and recipient (R). The reconstitution of CMV-specific T-cell responses after transplantation is crucial for the control of CMV reactivation. ObjectivesThe study aimed to determine the cellular immune status correlating with protection from high-level CMV viremia (>5000 copies/ml) and disease. Study designWe monitored CMV-specific cellular immune responses in 9 high-risk (D−/R+), 14 intermediate risk (D+/R+) and 3 low risk individuals (D+/R−), and 8 CMV negative controls (D−/R−). Interferon- γ (IFN-γ) levels as a marker for the CD8+ T-cell response were determined by the QuantiFERON-CMV-assay and compared to viral loads determined by PCR. ResultsEarly CMV reactivation was detected in all high-risk and 13/14 intermediate risk individuals. High-level viremia was detected in 5/7 high and 7/14 intermediate risk patients. Reconstitution of the CMV-specific cellular immune response started from 3 months after transplantation and resulted in protection against CMV reactivation. Re-establishing of CMV-specific T-cell immune responses with IFN- γ levels >8.9 IU/ml is crucial for protection from high-level CMV viremia. ConclusionsMonitoring of HSCT-recipients with the QuantiFERON-CMV-assay might be of great benefit to optimize antiviral treatment.

Phase 1 Clinical Trial of a Replication-Defective Human Cytomegalovirus (CMV) Vaccine

BackgroundCongenital CMV remains an unmet medical need worldwide. Naturally acquired CMV immunity in women prior to pregnancy has been shown effective in reducing maternal-fetal transmission. V160 is engineered as a replication-defective CMV, and its replication in culture is controlled by a synthetic chemical. V160 can’t replicate in humans but it maintains all virological properties for presentation of viral antigens, including gH/gL/pUL128-131 pentameric complex, important for potent neutralizing antibodies (NABs).MethodsApproximately 190 CMV seronegative and seropositive adults at study entry received 3 doses of V160 or placebo administered via intramuscular (IM) or intradermal (ID) route on Day 1, Month 1, and Month 6. Four antigen levels (10, 30, 100, and 250 units per dose) formulated with or without aluminum phosphate adjuvant were evaluated. In each vaccination group, approximately 10 and 4 subjects received study vaccine and placebo, respectively. Injection site and systemic adverse events (AEs) were collected for 14 days after each vaccination. Serious AEs (SAEs) were assessed up to Month 18. Viral shedding (urine and saliva) were monitored up to Month 12. CMV-specific NABs and cell-mediated immune responses (CMI) were measured prior and 1 month after each vaccination, and at Months 12 and 18.ResultsDuring the study, no serious AEs were reported and only one CMV seropositive subject had non-vaccine type viral shedding. In both seronegative and seropositive cohorts, proportion of subjects who reported injection site AEs was higher in V160 recipients than placebo controls. Proportion of subjects who reported systemic AEs was comparable across V160 doses/formulations and placebo. In the CMV seronegative cohort, immune responses increased with incremental dosing. More importantly, recipients of V160 from several dose levels mounted NAB and CMI responses at 1 month post dose 3 (PD3) that were comparable to baseline levels measured in seropositive subjects.ConclusionV160 had acceptable safety profile across all dose levels and formulations studied; Vaccine was immunogenic and elicited NAB and CMI responses at 1 month PD3 that were comparable to natural CMV infection.Disclosures S. Adler, Merck: Investigator, Research grant. N. Lewis, Merck: Employee, Salary. A. Conlon, Merck: Employee, Salary. M. Christiansen, Merck: Investigator, Research grant. M. S. Al-Ibrahim, Merck: Investigator, Research grant R. Rupp, Merck: Investigator, Research grant. T. M. Fu, Merck: Employee, Salary. O. Bautista, Merck: Employee, Salary. H. Tang, Merck: Employee, Salary.T. Culp, Merck: Employee, Salary. R. Das, Merck: Employee, Salary. K. Beck, Merck: Employee, Salary. G. Tamms, Merck: Employee, Salary. L. Musey, Meck: Employee, Salary.

Enzyme-Linked Immunospot Assay as a Complementary Method to Assess and Monitor Cytomegalovirus Infection in Kidney Transplant Recipients on Pre-emptive Antiviral Therapy: A Single-Center Experience

BackgroundCytomegalovirus (CMV) disease represents a major cause of post-transplantation morbidity and mortality. To estimate the risk of infection and monitor response to antiviral therapy, current guidelines suggest combination of viral load monitoring with direct assessment of CMV-specific immune response. We used enzyme-linked immunospot (ELISpot) for the evaluation of CMV-specific T-cell response in kidney transplant recipients with CMV viremia and investigated how information gained could help manage CMV infection. MethodsSeventeen patients on pre-emptive antiviral therapy and CMV quantitative polymerase chain reaction (qPCR) ≥500 copies/mL (first episode after transplantation) were assessed using ELISpot and divided into Weak (9 patients with baseline ELISpot <25 spot-forming colonies [SFCs]/200,000 peripheral blood mononuclear cells [PBMCs]) and Strong Responders (8 patients with baseline ELISpot ≥25 SFCs/200,000 PBMCs). CMV-specific T-cell response, infection severity, viral load, and antiviral therapy were prospectively recorded and compared between groups at 1, 2, and 24 months of follow-up. ResultsDemographic and transplant characteristics of Weak and Strong Responders were similar. No episodes of CMV disease were observed. Weak Responders were more likely to experience CMV syndrome (56% vs 36.5%) and late virus reactivation (56% vs 25%) than Strong Responders. Weak Responders showed higher baseline median viral loads (19,700 vs 9265 copies/mL) and needed antiviral therapy for longer (179 vs 59.5 days). T-cell response showed 2 main patterns: early and delayed. ConclusionsELISpot provides prognostic information about infection severity, risk of late reactivation, and response to therapy. Randomized trials, evaluating the need for antiviral therapy in kidney transplant recipients with asymptomatic infection and effective virus-specific T-cell immune response, are warranted.

Preserved immune functionality and high CMV-specific T-cell responses in HIV-infected individuals with poor CD4+ T-cell immune recovery

Poor CD4+ T-cell recovery after cART has been associated with skewed T-cell maturation, inflammation and immunosenescence; however, T-cell functionality in those individuals has not been fully characterized. In the present study, we assessed T-cell function by assessing cytokine production after polyclonal, CMV and HIV stimulations of T-cells from ART-suppressed HIV-infected individuals with CD4+ T-cell counts >350 cells/μL (immunoconcordants) or <350 cells/μL (immunodiscordants). A group of HIV-uninfected individuals were also included as controls. Since CMV co-infection significantly affected T-cell maturation and polyfunctionality, only CMV+ individuals were analyzed. Despite their reduced and skewed CD4+ T-cell compartment, immunodiscordant individuals showed preserved polyclonal and HIV-specific responses. However, CMV response in immunodiscordant participants was significantly different from immunoconcordant or HIV-seronegative individuals. In immunodiscordant subjects, the magnitude of IFN-γ+ CD8+ and IL-2+ CD4+ T-cells in response to CMV was higher and differently associated with the CD4+ T-cell maturation profile., showing an increased frequency of naïve, central memory and EMRA CMV-specific CD4+ T-cells. In conclusion, CD4+ and CD8+ T-cell polyfunctionality was not reduced in immunodiscordant individuals, although heightened CMV-specific immune responses, likely related to subclinical CMV reactivations, may be contributing to the skewed T-cell maturation and the higher risk of clinical progression observed in those individuals.

CMV-specific T-cell immunity in solid organ transplant recipients at low risk of CMV infection. Chronology and applicability in preemptive therapy

To characterize whether the CMV-specific cellular immune response can be used as a predictor of the control of CMV infection and disease and determine thresholds in solid organ transplant (SOT) recipients seropositive for CMV (R+). The CMV-specific T-cell response was characterized using intracellular cytokine staining and the evolution of clinical and virological parameters were recorded during the first year after transplantation. Besides having positive CMV serology, only 28.4% patients had positive immunity (CD8+CD69+IFN-γ+ ≥0.25%) at 2 weeks after transplantation. These patients had less indication of preemptive treatment (p=0.025) and developed less high grade (≥2000IU/ml) CMV replication episodes (p=0.006) than patients with no immunity. Of the 49 patients with a pretransplant sample, only 22.4% had positive immunity, and had a detectable immune response early after transplantation (median of 3.7 weeks). However, only 50% of patients with negative pretransplant immunity acquired a positive immune response and it was significantly later, at a median of 11 weeks (p<0.001). Patients that developed CMV disease had no CMV-specific immunity. Having CMV-specific CD8+IFN-γ+ cells ≥0.25% before transplant; 0.15% at two weeks or 0.25% at four weeks after transplantation, identifies patients that may spontaneously control CMV infection and may require less monitoring.

Clinical Usefulness of Monitoring Cytomegalovirus-Specific Immunity by Quantiferon-CMV in Pediatric Allogeneic Hematopoietic Stem Cell Transplantation Recipients.

Cytomegalovirus (CMV) is a well-established cause of morbidity and mortality in pediatric recipients of allogeneic hematopoietic stem cell transplantation (allo-HSCT). CD8+ T-cells are important for controlling CMV infection. We conducted a prospective pilot study to investigate the clinical utility of measuring the CMV-specific T-cell immune response using the QuantiFERON-CMV assay (QF-CMV) in pediatric allo-HSCT recipients. Overall, 16 of 25 (64%) patients developed CMV infection. QF-CMV was evaluated in these 16 patients during the early and late phases of the first CMV infection post allo-HSCT. Whereas the initial QF-CMV results during the early phase of CMV infection did not correlate with the course of the corresponding infection, the QF-CMV results post resolution of the first CMV infection correlated with the recurrence of CMV infection until 12 months post allo-HSCT; no recurrent infections occurred in the four QF-CMV-positive patients, while recurrent infections manifested in five of eight QF-CMV-negative (62.5%) and all three QF-CMV-indeterminate patients (P=0.019). In spite of the small number of patients examined, this study supports the potential application of monitoring CMV-specific T-cell immunity using the QF-CMV assay to predict the recurrence of CMV infection in pediatric allo-HSCT recipients.

Immunological Prediction of Cytomegalovirus (CMV) Replication Risk in Solid Organ Transplantation Recipients: Approaches for Regulating the Targeted Anti-CMV Prevention Strategies.

The current cytomegalovirus (CMV) prevention strategies in solid organ transplantation (SOT) recipients have contributed towards overcoming the detrimental effects caused by CMV lytic infection, and improving the long-term success rate of graft survival. Although the quantification of CMV in peripheral blood is the standard method, and an excellent end-point for diagnosing CMV replication and modulating the anti-CMV prevention strategies in SOT recipients, a novel biomarker mimicking the CMV control mechanism is required. CMV-specific immune monitoring can be employed as a basic tool predicting CMV infection or disease after SOT, since uncontrolled CMV replication mostly originates from the impairment of immune responses against CMV under immunosuppressive conditions in SOT recipients. Several studies conducted during the past few decades have indicated the possibility of measuring the CMV-specific cell-mediated immune response in clinical situations. Among several analytical assays, the most advancing standardized tool is the QuantiFERON®-CMV assay. The T-Track® CMV kit that uses the standardized enzyme-linked immunospot assay is also widely employed. In addition to these assays, immunophenotyping and intracellular cytokine analysis using flow cytometry (with fluorescence-labeled monoclonal antibodies or peptide-major histocompatibility complex multimers) needs to be adequately standardized and validated for potential clinical applications.

T-Cell Activation Independently Associates With Immune Senescence in HIV-Infected Recipients of Long-term Antiretroviral Treatment.

Aging-associated noncommunicable comorbidities are more prevalent among human immunodeficiency virus type 1 (HIV)-infected individuals than among HIV-uninfected individuals. Residual HIV-related chronic immune activation and senescence may increase the risk of developing comorbidities. Immune phenotyping, thymic output, and telomere length were assessed in 94 HIV-infected individuals who were aged >45 years and receiving antiretroviral therapy (ART; cases) and 95 age-matched uninfected controls. Cases had lower CD4(+) T-cell counts, higher CD8(+) T-cell counts, and increased levels of immune activation (ie, increased soluble CD14 [sCD14] level and increased percentages of CD38(+)HLA-DR(+) cells among both CD4(+) and CD8(+) T cells), regulatory T cells, and percentage of programmed cell death 1 (PD-1)-expressing cells among CD4(+) T cells. Immune senescence levels (ie, percentages of CD27(-)CD28(-) cells or CD57(+) cells) were comparable between cases and controls. Peripheral blood mononuclear cells from cases had shorter telomeres but increased single-joint T-cell receptor excision circle content and CD31(+) naive CD4(+) T cells. Although cytomegalovirus (CMV) antibody titers were higher in cases, CMV-specific T-cell responses were comparable between cases and controls. T-cell senescence in cases was independently associated with T-cell activation but not with CMV-specific immune responses. Despite long-term receipt of ART, HIV-infected adults had higher levels of immune activation, regulatory T cells, and PD-1-expressing CD4(+) cells and shorter telomeres. The increased soluble CD14 levels and percentage of CD38(+)HLA-DR(+) cells among CD4(+) T cells correlated with shorter telomeres and increased regulatory T-cell levels. This suggests that HIV influences immune function irreversibly, with several pathways that are persistently abnormal during effective ART. Therapies aimed at improving immune health during ART are needed.

Böbrek Nakil Hastalarında Sitomegalovirusa Özgül CD4+ ve CD8+ T Hücre Yanıtının Sitokin Akım Sitometri Yöntemiyle İzlemi

In spite of the improvements in the clinical management of solid organ transplant (SOT) recipients provided by immunosuppresion and universal prophylaxis, human cytomegalovirus (CMV) infections continue to be one of the most leading causes of morbidity and mortality. Cell-mediated immunity specific to CMV (CMV-CMI) plays an important role in the control of CMV replication. Therefore, monitoring of CMV-specific T-cell response can be used to predict individuals at increased risk of CMV disease. The aim of this study was to investigate the levels of CMV-specific interferon (IFN)-γ producing CD4(+) and CD8(+) T cells in kidney transplant recipients before and after the transplantation, by cytokine flow cytometry. A total of 21 kidney transplant recipients (14 male, 7 female; age range: 18-66 years, mean age: 34.5 ± 9.9) who were all CMV seropositive have been evaluated in the study. Blood samples from the patients were obtained before and at the 1(st), 3(rd) and 6(th) months after transplantation. CMV seropositive healthy kidney donors (n= 20) constituted the control group. The main stages of our procedure were as follows; isolation of peripheral blood mononuclear cells from whole blood, freezing and storing of the samples, later on thawing the samples, ex vivo stimulation of lymphocytes with pooled CMV peptides and counting CMV-specific IFN- producing CD4(+) and CD8(+) T cells by flow cytometry following surface and intracellular cytokine staining. Monitoring of the viral load (CMV-DNA) was performed in 10 days intervals in the first 3 months followed by 3 week intervals until 6 months using COBAS AmpliPrep/COBAS TaqMan CMV test system (Roche Diagnostics, USA). The frequencies of pretransplant CMV-specific IFN-γ producing CD8(+) T cells in patient (3.53 ± 4.35/µl) and control (4.52 ± 5.17/µl) groups were not statistically different (p= 0.266). The difference between the number of virus-specific CD4(+) T cells in patients (8.84 ± 9.56/µl) and those in the control group (8.23 ± 11.98/µl) was at the borderline of significance (p= 0.057). The age and gender of the patients and type of antiviral prophylaxis protocols [valgancyclovir (n= 4); valacyclovir (n= 17)] did not have any significant effect on CMV-CMI (p> 0.05). Similarly, induction therapy administered to four patients did not show any effect on CMV-CMI (p> 0.05). CMV-specific immune responses of patients who received different immunosuppression protocols [tacrolimus + mycophenolate mofetil (MMF) + steroid (n= 17); cyclosporine + MMF + steroid (n= 2); mTOR inhibitor + MMF + steroid (n= 2)] were not different (p> 0.05). The number of CMV-specific CD4(+) T cells in all patients were significantly decreased in the 3rd month compared to the 1st month after the transplantation (p=0.003), indicating a relationship with the period of immunosuppressive therapy. In one of the patients who did not have CMV-specific CD4+ T-cell response but had cytotoxic T-cells (CD8(+) T= 0.6%) before transplantation, CD4(+) T-cell response have developed during monitorization (1.4%, 1.5% and 0.5% in 1st, 3rd and 6th months, respectively), and no viral reactivation was detected. Out of the two patients who had no CD4(+) and CD8(+) T cell response in the 3rd month, one of them developed low level viremia (150 copies/ml) in the 6th month. In this patient the level of CMV-CMI in the 6th month (CD4(+)T + CD8(+)T= 0.9%), have reached higher values than the values obtained before the transplantation (CD4(+) T + CD8(+) T= 0.5%). The viremia was cleared spontaneously in this patient and no antiviral therapy was required. In conclusion, our results suggested that pretransplant and posttransplant monitoring of CMV-specific T-cell responses might be helpful as well as viral load in the clinical management of CMV infection in SOT patients.

In vitro studies of the impact of maribavir on CMV-specific cellular immune responses

BackgroundGanciclovir has demonstrated immunosuppressive effects in vitro which may lead to delayed cytomegalovirus (CMV)-specific immune reconstitution when the drug is given prophylactically. Maribavir is a new and more potent anti-CMV drug that is under evaluation for therapeutic use in transplant recipients. ObjectivesThe objective of this study was to evaluate the potential effect of maribavir on CMV-specific T cell function in comparison to ganciclovir. Study designIn ten immunocompetent CMV seropositive donors, maribavir and ganciclovir were compared over a broad range of concentrations (0.2–500μM) regarding their effects on lymphoproliferation, CMV-specific CD4+ and CD8+ cytokine expression, T cell multifunctionality, degranulation and apoptosis. ResultsMaribavir inhibited lymphocyte proliferation at concentrations of 50μM and above, however, cytokine expression, cellular degranulation and multifunctionality of CD4+ and CD8+ T cells in response to CMV lysate and pp65 peptide mix were not impaired except at the highest concentration of 500μM. Ganciclovir inhibited lymphoproliferative responses starting at 10μM. As with maribavir, other cellular responses following stimulation with CMV lysate and pp65 peptide mix were only impaired at the highest concentration of 500μM of ganciclovir. Neither maribavir nor ganciclovir showed induction of lymphocyte apoptosis. ConclusionsMaribavir exhibits a low potential to suppress CMV-specific T cell function. This finding supports the use of higher doses in the prophylactic setting than originally proposed.

Persistent CMV infection after allogeneic hematopoietic stem cell transplantation in a CMV-seronegative donor-to-positive recipient constellation: Development of multidrug resistance in the absence of anti-viral cellular immunity

We describe a case of persistent cytomegalovirus (CMV) infection after allogeneic hematopoietic stem cell transplantation (allo-HSCT) with discordant and high-risk (D−/R+) constellation of CMV serostatus. Despite the use of different and innovative antiviral strategies, viral replication could not be suppressed successfully promoting a protracted CMV colitis associated with severe gastrointestinal graft-versus-host disease (GI GVHD). We illustrate that the development of multidrug viral resistance, the failure to mount a CMV-specific cellular immune response, as confirmed by QuantiFERON®-CMV (Qiagen) assay, and the refractory GVHD requiring prolonged immunosuppression were the main factors contributing to persistent viral replication and the fulminant unfavorable course.

Physical function impairment of older, HIV-infected adults is associated with cytomegalovirus immunoglobulin response.

Cytomegalovirus (CMV) is associated with poor outcomes, including physical function impairment, in older HIV-uninfected adults. Whether CMV is associated with physical functional impairment in HIV-infected adults is unknown. The primary objective of this study was to determine the relationship between CMV-specific humoral and cell-mediated immune responses with functional impairment in well-controlled HIV infection. In a case-control study, low-function cases were matched by age, gender, and time from HIV diagnosis to high-function controls. Quantitative CMV IgG and %CMV-specific CD8(+) and CD4(+) T cells (interferon-γ expression following CMV pp65 stimulation) were used to estimate physical function. Among 30 low-function cases and 48 high-function matched controls, CMV IgG ranged from <10 to 8,830 EU/ml, including four controls with results <10 EU/ml. Each log10 increase in CMV IgG was associated with 5-fold greater odds of low function (p=0.01); these findings were robust to adjustment for concomitant CD4(+) count, tobacco use, and age; to exclusion of subjects with CMV IgG <10 EU/ml; and to adjustment for hepatitis C viremia. %CMV-specific CD4(+) or CD8(+) T cells were not associated with low function. In bivariable models, the relationship between CMV IgG and physical function was attenuated and was no longer significant when including IL-6, CD4/CD8 ratio, or the Veterans Aging Cohort Study Index score. High levels of CMV-specific IgG were associated with impaired physical function. Attenuation of the strength of this association in bivariable models suggests an indirect relationship mediated by systemic inflammation and immune suppression.