Process Of Immune Reconstitution Research Articles

Immune characteristics of kidney transplant recipients with acute respiratory distress syndrome induced by COVID-19 at single-cell resolution

BackgroundCOVID-19-induced acute respiratory distress syndrome (ARDS) can result in tissue damage and multiple organ dysfunction, especially in kidney transplant recipients (KTRs) receiving immunosuppressive drugs. Presently, single-cell research on COVID-19-induced ARDS is considerably advanced, yet knowledge about ARDS in KTRs is still constrained.MethodsSingle-cell RNA sequencing (scRNA-seq) analysis was performed to construct a comprehensive single-cell immune landscape of the peripheral blood mononuclear cells (PBMCs) of eight patients with COVID-19-induced ARDS, five KTRs with COVID-19-induced ARDS, and five healthy individuals. Subsequently, we conducted a comprehensive bioinformatics analysis, including cell clustering, enrichment analysis, trajectory analysis, gene regulatory network analysis, and cell–cell interaction analysis, to investigate the heterogeneity of the immune microenvironment in KTRs with ARDS.ResultOur study revealed that KTRs exhibit significant heterogeneity with COVID-19-induced ARDS compared with those of other individuals, with significant reductions in T cells, as well as an abnormal proliferation of B cells and monocytes. In the context of dual influences from immunosuppression and viral infection, KTRs exhibited more specific plasma cells, along with significant enrichment of dysfunctional GZMB and XAF1 double-positive effector T cells and IFI27-positive monocytes. Additionally, robust communication existed among T cells and monocytes in cytokine signaling. These effects impede the process of immune reconstitution in KTR patients.ConclusionOur findings suggest that KTRs with COVID-19-induced ARDS show elevated antibody levels, impaired T cell differentiation, and dysregulation of innate immunity. In summary, this study provides a theoretical foundation for a comprehensive understanding of COVID-19-induced ARDS in KTRs.

Differential analysis of immune reconstitution after allogeneic hematopoietic stem cell transplantation in children with Wiskott-Aldrich syndrome and chronic granulomatous disease.

To investigate similarities and differences in immune reconstitution after allogeneic hematopoietic stem cell transplantation (allo-HSCT) in children with Wiskott-Aldrich syndrome (WAS) and chronic granulomatous disease (CGD). We retrospectively analyzed the lymphocyte subpopulations and the serum level of various immune-related protein or peptide on Days 15, 30, 100, 180 and 360 post-transplantation in 70 children with WAS and 48 children with CGD who underwent allo-HSCT at the Transplantation Center of the Department of Hematology-Oncology, Children's Hospital of Chongqing Medical University from January 2007 to December 2020, and we analyzed the differences in the immune reconstitution process between the two groups. ① The WAS group had higher lymphocyte subpopulation counts than the CGD group. ② Among children aged 1-3 years who underwent transplantation, the WAS group had higher lymphocyte subpopulation counts than the CGD group. ③ Further comparisons were performed between children with non-umbilical cord blood transplantation (non-UCBT) and children with umbilical cord blood transplantation (UCBT) in the WAS group. On Day 15 and 30 post-transplantation, the non-UCBT group had higher B-cell counts than the UCBT group. On the remaining time points post-transplantation, the UCBT group had higher lymphocyte subpopulation counts than the non-UCBT group. ④ Comparisons were performed between children with non-UCBT in the WAS group and in the CGD group, the lymphocyte subpopulation counts were higher in the WAS group compared to the CGD group. ⑤ On Day 100 post-transplantation, the CGD group had higher C3 levels than the WAS group. On Day 360 post-transplantation, the CGD group had higher IgA and C4 levels than the WAS group. ① The rate of immunity recovery was faster in children within the WAS group compared to those children within the CGD group, which may be attributed to the difference of percentage undergoing UCBT and primary diseases. ② In the WAS group, the non-UCBT group had higher B-cell counts than the UCBT group at Day 15 and 30 post-transplantation, however, the UCBT group had higher B-cell counts than the non-UCBT group at Day 100 and 180 post-transplantation, suggesting that cord blood has strong B-cell reconstitution potentiality after transplantation.

Reconstitution of T-cell-mediated immunity in patients after allogeneic stem cell transplantation

Background. The timely reconstitution of the donor-derived immune system is a key factor in the prevention of such post-transplant complications as graft versus host disease, relapse or secondary tumours and various infections. These complications affect the long-term survival of patients after allogeneic stem cell transplantation.Aim — to describe the main stages of T Cell–mediated immune recovery in patients after allogeneic stem cell transplantation.General findings. T-cell–mediated immunity is responsible for anti-infective and anti-tumour immune response. The early post-transplant period is characterized by the thymus-independent pathway of T-cell recovery largely involving proliferation of mature donor T cells, which were transplanted to the patient together with hematopoietic stem cells. To a lesser extent, this recovery pathway is realized through the expansion of host naïve and memory T cells, which survived after conditioning. Thymus-dependent reconstitution involves generation of de novo naïve T cells and subsequent formation of a pool of memory T-cells providing the main immunological effects — graft versus tumour and graft versus host reactions. A better understanding of the T-cell immune reconstitution process is important for selecting optimized pre-transplant conditioning regimens and patient-specific immunosuppressive therapy approaches, thus reducing the risks of post-transplant complications and improving the long-term survival of patients after allogeneic stem cell transplantation.

Use of Multivariate Immune Reconstitution Patterns to Describe Immune Reconstitution after Allogeneic Stem Cell Transplantation in Children.

Immune reconstitution after hematopoietic stem cell transplantation (HSCT) is a complex process. Impacts of the reconstitution of different immune cells over time are complex and difficult to understand. New mathematical models are needed to better understand this process. In this study, we used principal component analysis to better analyze the process of immune reconstitution after HSCT. Forty-six consecutive patients receiving HSCT for malignant and nonmalignant disorders were included in the study. All patients were followed for at least 24 months after transplantation with regular blood sampling for analysis of lymphocyte subset numbers and function. Exponentially transformed lymphocyte subset counts and lymphocyte functional markers were analyzed to identify major trends in the reconstitution process. Using our multivariate model for mapping immune reconstitution after HSCT, we showed that dysfunctional reconstitution patterns precede severe complications, such as chronic graft-versus-host disease, relapse, and death.

Posttransplant Cyclophosphamide Promote Naïve-Subset Dominant B Cell Recovery Coordinated with Early Treg Expansion; Implication of Reduced Risk of Pathogenesis into Chronic Gvhd

Posttransplant cyclophosphamide (PTCy) is an effective prophylaxis for both acute and chronic graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (HSCT). We recently studied the immune reconstitution dynamics of each lymphocyte subset after PTCy-based transplant using murine haploidentical BMT model and reported that PTCy strongly promoted Treg-dominant T-cell reconstitution and stem cell-derived mature B-cell generation with broad BCR-diversity. We also found that the early reconstitution of Treg could contribute to promote naïve B cell emergence from bone marrow, indicating the T and B cell recovery might be mutually coordinated after PTCy-based transplant (Iwamoto et al, ASH2017). However, the detailed process of immune reconstitution in patients after haploidentical HSCT with PTCy has not been well studied. To address this issue, we here investigated the early dynamics of donor-lymphocyte subset chimerisms in patient after clinical PTCy-based haploidentical HSCT with comparing those in patients after low-dose ATG-based haploidentical HSCT and patients after cord blood transplantation. Laboratory studies were undertaken in 13 adult patients who received HLA-mismatched allogeneic graft; unrelated cord blood (n=5), and haploidentical related peripheral blood after ATG-based conditioning (n=5) and haploidentical related peripheral blood after PTCy-based conditioning (n=5). Blood samples were obtained before and at 1, 2, 4, 6 and 8 weeks after HSCT. Peripheral blood mononuclear cells (PBMCs) were isolated from blood samples by density gradient centrifugation and cryopreserved before being analyzed. After thawing, to analyze the subset-specific chimerism, PBMCs were stained with anti-HLA monoclonal antibodies and other subset-specific antibodies as follows: Pacific Blue conjugated anti-CD4, eFluor450 conjugated anti-CD3, PE-Cy7 conjugated anti-CD25, anti-CD14, APC conjugated anti-CD127, anti-CD56, and APC-eFluor780 conjugated anti-CD8a, anti-CD19. Gated lymphotes (CD4+Tcons, CD4+Tregs, CD8+T cells, B cells, NK cells, Monocytes) were analyzed their chimerism by flowcytometry. To examine the detailed phenotype of B cells, the expression of CD27, CD24, CD38 and IgD were tested. Flowcytometry-based method enables us to analyze the lymphocyte subset chemerism in the very early phase after HSCT. At 2 weeks after HSCT, our analysis revealed that CD4+Tcons, CD4+Tregs and CD8+T cells had already achieved complete donor chimerisms (>95% in all subsets) in patients after ATG-based SCT and had been approaching complete donor chimerisms (85.8%, 75.4% and 87.2%, respectively) in patients after CBT. In contrast, percentage of donor chimerisms of CD4+Tcons, CD4+Tregs and CD8+T cells after PTCy-based haplo-SCT was 73.5%, 59.6% and 59.2%, respectively, and those remained to be in the lower levels than other 2 groups. However, at 4 weeks after HSCT, all examined patients achieved complete donor chimerism of T cells, NK cells and Monocytes (>90%). At 8 weeks after HSCT, the number of B cells in PTCy-based haplo-group was higher than in ATG-based haplo-group (3494 vs 1901/mm3). Of note, B cell population in PTCy-based haplo-group at 8 weeks contained the significantly higher percentage of CD24+CD27-IgD+CD38+ transitional/naïve subset and the significantly lower percentage of CD24+CD27+IgD-CD38neg/dim activated/switched-memory subset when compared to B cell population in ATG-based haplo-group (59.9% vs 10.2%, 2.6% vs 21.5%, P<0.02 respectively), suggesting PTCy treatment might be associated with the favorable B cell reconstitution with naïve-subset dominant composition. Moreover, in patients after PTCy-based haplo-group, the percentage of activated/switched-memory subsets in B cell population at 8 weeks was inversely correlated with percentage of Treg in CD4 T cells at 4 weeks (P<0.05, r2=0.77). Taken together, consistently with our murine study, the current data from clinical samples again suggest that PTCy-based immune-modulation lead to coordinated T and B cell recovery, especially promoting naïve-subset dominant B cell recovery with help of the early expansion of Treg, which might reduce the risk of subsequent chronic GVHD. These data provide the important information for understanding the immunological reconstitution after PTCy-based haploidentical HSCT. DisclosuresNo relevant conflicts of interest to declare.

High Incidence of Transient Monoclonal Gammopathies after Allogeneic Hematopoietic Stem Cell Transplantation in Non-Myeloma Patients

BackgroundThe clinical and biological relevance of a monoclonal gammopathy (MG) that newly arises after allogeneic hematopoietic stem cell transplantation (allo-HSCT) in non-myeloma patients is unknown. In healthy subjects, the incidence of monoclonal gammopathy of undetermined significance (MGUS) ranges from 3-9% and significantly increases with age. In these individuals, the absolute risk of progression to plasma cell myeloma at 20 years is 5-58%, depending on the MGUS subtype. The aim of this study was to investigate the incidence, the course and the clinical relevance of post-allo-HSCT MG.MethodsWe retrospectively analyzed a cohort of 403 non-myeloma patients (median age: 48y, range: 18-69y, 57% men, 43% women) who underwent allo-HSCT at the Division of Hematology of the University Hospital Zurich between January 2004 and December 2014. In these patients, a serum electrophoresis and an immunofixation was performed every 3-6 months post allo-HSCT. Patients with a MGUS before allo-HSCT were excluded from the study. The immunoglobulin subtype and the light chain restriction of the MG were determined by immunofixation. The most frequent indications for allo-HSCT were acute leukemia (AL, n=305), chronic myeloid leukemia (CML, n=30), myelodysplastic syndrome (MDS, n=24), and myeloproliferative neoplasm (MPN, n=20).ResultsThe incidence of a MG after allo-HSCT (56/403 patients, 14%) was higher than the reported incidence of MGUS in age-matched healthy subjects (estimated prevalence of MGUS in 50y old men: 3.2%) and in contrast to healthy subjects did not correlate with age. The majority of patients carried a IgG paraprotein (80%), while IgM paraproteins were detected in 12.5% of patients. Rarely, two types of paraproteins were identified simultaneously in the same patient. In most patients (44/56, 79%), the MG appeared within the first year after allo-HSCT and was transient with a median duration of 5.8 months (range 2.5 - 43.9 months). However, in 7/56 (13%) patients, the MG persisted until the end of the observation period (median duration 33 months, range 12-45.6 months). Only one patient showed a very late (7y post allo-HSCT) but also transient appearance of a MG. Development of plasma cell myeloma was not observed in patients with post allo-HSCT MG. Conditioning regimen, GvHD, immunoglobulin treatment and cessation of immunosuppressive therapy did not predict the appearance of post allo-HSCT MG.ConclusionsOur study reveals a high incidence of a transient MG after allo-HSCT that is not related to age of the recipient and follows the expected immunoglobulin subtype distribution. Importantly, patients with post allo-HSCT were not at increased risk of developing plasma cell myeloma as observed for MGUS in otherwise healthy subjects inferring a distinct pathogenetic mechanism. The transient nature of MG early after allo-HSCT suggests an association with the immune reconstitution process and future studies will determine whether expansion of a specific plasma cell clone can occur during engraftment of donor-derived hematopoiesis. [Display omitted] DisclosuresTheocharides:Novartis: Consultancy, Honoraria.

Visfatin concentrations in children with leukemia before and after stem cell transplantation

Visfatin (VF) is an adipocytokine that performs many functions, including enhancing cell proliferation and biosynthesis of nicotinamide mononucleotides and dinucleotides. It also seems to be involved in the development of glucose metabolism disturbances. The goal of the study was the determination of VF concentrations in children with leukemia who are treated with stem cell transplantation. VF concentrations were measured in plasma before and after oral glucose tolerance test (OGTT; 60 and 120 minutes) in 22 children with leukemia treated with hematopoietic stem cell transplantation (HSCT) and healthy control subjects (n = 24). The HSCT group was studied twice: before HSCT (22 children) and approximately 6 months after HSCT (12 of 22 children). After fasting, concentrations of glucose, insulin, triglycerides, total cholesterol, high-density lipoprotein, and high-sensitivity C-reactive protein (hsCRP) were determined. Significantly lower (p < 0.05) median values of VF concentrations at all time points in the OGTT were found in pre- HSCT children compared with control subjects. The median VF concentration was significantly higher after HSCT compared with before HSCT. The decrease in VF in leukemic children in complete remission may be caused by myelosuppression and immunosuppression after prolong chemotherapy and is beneficial because of the decrease in its antiapoptotic activity. VF can serve as an additional biochemical marker for remission in patients with leukemia. Normalization of plasma VF concentration after HSCT might be caused by a process of immune reconstitution and prolonged inflammation (e.g., infections, graft-versus-host disease), injury to organs (e.g., lungs, gut, liver), and endocrinology deficiencies.

Evidence for the cure of HIV infection by CCR5Δ32/Δ32 stem cell transplantation

AbstractHIV entry into CD4+ cells requires interaction with a cellular receptor, generally either CCR5 or CXCR4. We have previously reported the case of an HIV-infected patient in whom viral replication remained absent despite discontinuation of antiretroviral therapy after transplantation with CCR5Δ32/Δ32 stem cells. However, it was expected that the long-lived viral reservoir would lead to HIV rebound and disease progression during the process of immune reconstitution. In the present study, we demonstrate successful reconstitution of CD4+ T cells at the systemic level as well as in the gut mucosal immune system after CCR5Δ32/Δ32 stem cell transplantation, while the patient remains without any sign of HIV infection. This was observed although recovered CD4+ T cells contain a high proportion of activated memory CD4+ T cells, ie, the preferential targets of HIV, and are susceptible to productive infection with CXCR4-tropic HIV. Furthermore, during the process of immune reconstitution, we found evidence for the replacement of long-lived host tissue cells with donor-derived cells, indicating that the size of the viral reservoir has been reduced over time. In conclusion, our results strongly suggest that cure of HIV has been achieved in this patient.

Quality care: a link between clinical and public health approaches to HIV infection in developing countries

As the importance of quality in health care provision is increasingly recognised, it is opportune to consider quality care as a key link between clinical and public health approaches to human immunodeficiency virus (HIV) infection in developing countries, especially in sub-Saharan Africa. This region has the lion's share of the global epidemic and the least resources to respond. Looking at health problems using a 'quality lens' may help bridge the gaps between clinical care and public health, the current and desired standard of care, and prevention and treatment. Quality care, with prompt diagnosis and effective treatment, of people with HIV infection is crucial for good individual health outcomes, public health outcomes (in terms of decreased HIV transmission) and societal outcomes (increased productivity and decreased costs of health provision for HIV-related care). A spotlight on quality care can bring clinicians and public health practitioners together in working towards universal access to quality HIV care and prevention - one of the greatest health challenges faced in developing countries in Africa today.

Immune reconstitution during maintenance therapy in children with acute lymphoblastic leukemia, relation to co-existing infection

Immunosuppression is a major side effect of cancer chemotherapy. The process of immune reconstitution can be dissimilar according to the nature of the disease, type and doses of drugs, and age of the patients. Recently, several studies have examined immune reconstitution in children and young adults after intensive chemotherapy for solid tumours or stem cell transplantation.The aim of the present study is to evaluate immune reconstitution (cellular and humoral) in children with acute lymphoblastic leukemia during the maintenance phase of therapy and to correlate between the complicating infections and the abnormalities in immune system during reconstitution. To achieve this goal, 36 children with acute lymphoblastic leukemia (24 females and 12 males) in the maintenance phase of therapy with 12 healthy children of matched age and sex served as a control group were recruited in this study. The patients were taken consecutively from the Hematology/Oncology Outpatient Clinic of Mansoura University Children's Hospital (MUCH). They were subjected to thorough history taking, clinical examination and laboratory investigations in the form of: complete blood count, serum creatinine, liver function tests and evaluation of the immune system by estimation of CD3, CD4, CD8, CD19 and CD56 (cellular immunity) by flow cytometry and immunoglobulins A, M and G (humoral immunity) at the first and the third month of maintenance therapy.The results of the study documented presence and persistence of leucopenia and lymphopenia during maintenance therapy with decreased medians of CD3, CD4 and CD8 from the first to the third month of therapy and in comparison to the control group. The other markers CD19, CD56, IgA, IgM, IgG and CD4/CD8 ratio showed increasing median from the first to the third month of therapy. Also we detect a significant correlation between infection and CD19 and serum IgM at the first month and between infection and CD19, IgM and CD4/CD8 ratio at the third month of therapy. In conclusion, persistent immunosuppression is documented in children with acute lymphoblastic leukemia during maintenance therapy. Reconstitution of B lymphocytes and Natural killer cells occurs early while T cell reconstitution shows delayed recovery of both T helper and T suppressor cells. Immunosupression during maintenance therapy has no major clinical impact in terms of increased incidence or severity of systemic infections.

Immune reconstitution complicated by CMV retinitis in a pediatric patient who underwent haploidentical CD34+-selected hematopoietic stem cell transplant for acute lymphoblastic leukemia

We describe two episodes of CMV retinitis in a pediatric patient who underwent a CD34+ selected graft from his haploidentical father. Both recipient and donor were cytomegalovirus (CMV) seropositive. Both episodes occurred late post-grafting during a phase of complete immunological recovery with sufficient numbers of circulating CMV-specific clones. Antiviral treatment with foscarnet and ganciclovir was successful but prolonged treatment was required to prevent relapses. We hypothesize that this complication was more related to an immune reconstitution process than to an immune-deficient state post-grafting. We conclude that CMV retinitis is a late complication of HSCT that can occur despite satisfactory immune reconstitution. Usually, it is responsive to antiviral therapy. Dilated fundoscopic examination is essential both for examining patients with reduced visual acuity and for screening asymptomatic patients.

Variations in serum IL-7 and 90K/Mac-2 binding protein (Mac-2 BP) levels analysed in cohorts of HIV-1 patients and correlated with clinical changes following antiretroviral therapy.

Serum levels of interleukin-7 (IL-7), a non-redundant cytokine that plays a crucial role in lymphopoiesis, are known to be elevated in HIV-1-infected subjects. To examine further the association between levels of IL-7, CD4+ cell counts and viraemia, we analysed these parameters in a large cohort of HIV-1 patients along with serum levels of 90K, a marker of disease severity but with no established involvement in lymphopoiesis. While IL-7 levels were only found to correlate with CD4+ cell counts, 90K levels presented strong correlations with both CD4+ cell numbers and with plasma viral loads (VLs). These correlations were maintained in patients naive to treatment with antiretrovirals (n = 38) but were abolished when the analysis was restricted to the group receiving highly active antiretroviral therapy (HAART, n = 82). Moreover, although 90K levels were significantly reduced in patients on HAART, IL-7 levels continued to be elevated despite successful treatment. The influence of HAART on the variations in these serum parameters was further assessed in a longitudinal study on 32 subjects. The HAART-induced decrease in VLs and increase in CD4+ counts were found to correlate with a reduced serum level of 90K and IL-7, respectively. Nevertheless, following a median period of 33 months of immunological and virological successful HAART, serum levels of IL-7 continued to be significantly elevated compared with those detected in healthy controls. These findings suggest that immunotherapy with IL-7, aimed to replenish T-cell stock in HAART-treated subjects, may have a limited impact on the process of immune reconstitution.

Low Incidence of Cytomegalovirus Infection Associated with Rapid Cell-Mediated Immune Reconstitution after Cord Blood Transplantation Using Full Conditioning Regimen Containing 12Gy Total Body Irradiation.

The one of crucial questions in cord blood transplantation (CBT) is whether naïvity of cord blood lymphocytes could gain antigen-specific cellular immunity during early phase of transplant. Cytomegalovirus (CMV) infection is serious clinical problem in allogeneic transplant recipients and T cell immunity has known to have an important role in control of virus replication and prevention.During 1998 and 2006, 111 adults has received myeloablative regimens including 12 Gy of total body irradiation followed by CBT and a standard cyclosporine and methotrexate combination as GVHD prophylaxis in our institute. Patients also received intravenous immunoglobulin from day −3 to day 120 if the immunoglobulin level in the serum was less than 500 mg/dl. CMV antigenemia assay was performed twice a week after neutrophil recovery until day 120. Once CMV antigenemia is positive, patients received 5 mg/kg ganciclovir (GCV) once daily for at least 2 weeks as preemptive therapy. Ninety-two patients achieved engraftment with full donor chimerism and survived without disease relapse at the time of 120 days after CBT (82.8%). None of these 92 recipients had CMV disease during first 4 months after CBT.We have investigated the association of CMV reactivation status and their immune reconstitution process for 4 months after CBT in 39 patients who received CBT from 2002 to 2006 in our institute. CMV-specific CD4+ and CD8+ T cell recoveries were assessed by detection of interferon-g (IFN-g) producing cells with CMV antigen stimulation using intracellular cytokine staining. The positive was defined as more than 0.1% IFN-g positive cells among CD4+ or CD8+ T cell population.Six of 39 patients were CMV sero-negative and 33 patients were sero-positive. None of 6 CMV sero-negative receipients and 31 of 33 CMV sero-positive recipients observed CMV reactivation and received GCV therapy within the first 4 months.CMV-specific CD4+ T cells were detected in 30 of 31 recipients with positive CMV antigenemia (% positive: 55% at 1 month and 85% at 2 month), on the other hand, CMV-specific CD8+ T cells were detected in 14 out of 31 cases (% positive: 14% at 1 month and 22% at 2 month), both of which were comparable to post-bone marrow or peripheral blood transplants (CMV-specific CD4+ T cells were detected 18 of 21 recipients with positive CMV antigenemia and CMV-specific CD8+ T cells were detected in 12 out of 21).These data suggest that post-thymic naive T lymphocytes in cord blood might obtain memory and effector function in vivo with antigen-specific manner during early phase of post-transplant.

Recombinant Adenovirus Vaccines Can Successfully Elicit CD8+ T Cell Immunity under Conditions of Extreme Leukopenia

We have examined the efficacy of vaccination with recombinant adenovirus under conditions of extreme leukopenia in lethally irradiated mice reconstituted with autologous bone marrow. The expansion of antigen-specific CD8+ T cells following immunization of lethally irradiated hosts paralleled the recovery of total CD8+ T cells. Surprisingly, the numbers of antigen-specific CD8+ T cells in lethally irradiated mice beyond 6 weeks postimmunization were comparable to the numbers found in nonirradiated controls. CD8+ T cells elicited in the lethally irradiated hosts were functionally indistinguishable from those elicited in normal hosts. Antigen expression and presentation persisted for a longer period of time in the draining lymph nodes of irradiated mice compared to those of nonirradiated animals, suggesting that antigen presentation mechanisms were intact during the reconstitution period. Experiments employing allogeneic bone marrow demonstrated that radioresistant host antigen-presenting cells were responsible for antigen presentation during the process of immune reconstitution. These results demonstrate clear compatibility of adenovirus vaccines and cytotoxic therapy. Furthermore, these observations provide novel insights into the mechanisms of CD8+ T cell activation following adenovirus immunization.

Contribution of T-Cell Reconstitution during the Early Phase after Cord Blood Transplantation to Favorable Clinical Outcomes.

The immaturity of T cells in cord blood is well known in functional assays and phenotypic analyses. During the first several months after cord blood transplantation (CBT), the T cell compartment is recovered by peripheral expansion from those mature and naïve T cells in cord blood grafts and plays an important role in acute graft-versus-host disease (GVHD) and graft-versus-leukemia reaction. Recently, we have reported that adult patients with hematological malignancies receiving CBT from HLA-partially-mismatched unrelated donors (n=68) had a lower risk of severe acute GVHD (> grade II, 7% versus 26%) and transplant-related mortality (9% versus 29% at 1 year) and a higher probability of disease-free survival (74% versus 44% at 2 years) than HLA-matched unrelated bone marrow transplant (BMT) recipients (n=45) in our multivariate analysis (Takahashi et al., Blood, in press). We speculated that the immune reconstitution process over a period of several months after CBT might have contributed to these promising clinical results. Using four-color analysis with CD4, CD8, CD45RA, and CD62L, more than 90% of cord blood CD4+ and CD8+ T cells in the grafts belonged to the naïve fraction. Cytokine expression in cord blood T cells was also suppressed to 0.1% in CD4+ and to 0.9% in CD8+ with positive interferon-γ by intracellular staining, which were significantly lower than those in adult T cells (16.2% in CD4+ and 37.8% in CD8+). Circulating T cell counts normalized after 3 months for CD8+ and 4 months for CD4+ in our CBT recipients, both of which were significantly faster than in previously published studies, which were 9 months for CD8+ and 12 months for CD4+. After T cell recovery, peripheral blood T cells moved from the naïve to the central memory fraction immediately, and then moved to the effector memory fraction. A naïve subset of CD4+ T cells remained (median: 38 cells/μl on day 90, n=12) during the first 3 months, which was significantly higher than in the BMT control (median: 9 cells/μl on day 90, n=5, p=0.015), but showed a low level of CD8+ T cells (median: 14 cells/μl on day 90, n=12), almost the same as in BMT recipients (median: 13 cells/μl on day 90, n=5). Intracellular interferon-γ-producing T cells were detected at 3.4% (0.1–34.2%) in CD4+ and 32.3% (1.1–86.9%) in CD8+ at 1 month post-CBT (n=16), both of which were comparable to post-BMT. To investigate whether these T cells with memory phenotype are functional, we analyzed antigen-specific T-cell recovery using cytomegalovirus (CMV) as a specific antigen. CMV-responsive CD4+ T cells were detected within the first 4 months in all recipients with positive CMV antigenemia (n=13), but CD8+ T cells were detected only in 5 out of 13 cases, probably because of pre-emptive Gancyclovir administration in most antigenemia-positive patients. To conclude, naïve cord blood T cells rapidly increased in number and adopted a memory phenotype showing cytokine-production and antigen-recognition capacity in the early phase after CBT. These data suggest that mature T lymphocytes in cord blood have unique properties and contribute to the favorable clinical outcome of CBT.

Hematopoietic stem cell transplantation in multiple sclerosis: experimental evidence to rethink the procedures

Acute immunosuppression with lymphocytic agents given at maximally tolerated doses, followed by hematopoietic stem cell rescue achieved by autologous bone marrow or peripheral blood stem cell transplantation (BMT), has proved effective in various experimental models of autoimmunity. The rationale for such an approach in autoimmune diseases is based on the concept of lymphoablation of self-reactive lymphocytes followed by de novo immune system reconstitution, which, in the presence of the autoantigens in the thymus, may reinduce self-tolerance. Our previous work shows that in experimental autoimmune encephalomyelitis (EAE), autologous/syngeneic BMT not only prevents the appearance of paralytic signs, but can also partially reverse chronic disease and induce long-term, antigen-specific tolerance. However, there are serious reservations to be considered when interpreting these data and before applying similar protocols in patients with multiple sclerosis. (1) The model of EAE is not a completely reliable model of multiple sclerosis. (2) In animals with chronic EAE, although further relapses were prevented, the established paralysis was usually not reversible. According to recent data, in chronic multiple sclerosis (MS) lesions, damage caused by axonal loss/transection and cortical/spinal cord atrophy is irreversible and probably amenable to immunotherapy. (3) Long-term, antigen-specific tolerance may be induced with BMT, but not in all cases; in passively induced CR-EAE, many of the mice relapsed upon challenge with myelin antigens, which may indicate that the presence of the immunizing, myelin antigens (on the site of immunization) during the process of immune reconstitution is critical for induction of tolerance. Finally, one should weigh the procedure-related risks (including mortality of up to 5%) of bone marrow or peripheral stem cell transplantation (SCT). A more radical solution for autoimmunity may involve the use of non-myeloablative allogeneic transplantation.

Immune reconstitution in pediatric stem-cell transplantation

Hematopoietic stem-cell transplantation (HSCT) has successfully been used to cure many pediatric disorders. However, the immunologic alterations associated with transplantation result in profound immunodeficiencies in the transplant recipient, resulting in significant infectious morbidity and mortality. The precarious process of immune reconstitution in the transplant recipient is neither instantaneous nor complete, but influenced by multiple factors such as graft-versus-host disease (GVHD), conditioning regimen, patient age and underlying disease. Studies in pediatric HSCT have revealed unique attributes of immune recovery in pediatric transplant recipients. Future studies addressing these findings are needed to complement novel immunotherapies emerging from the field of transplant immunology.

Early immune reconstitution after potent antiretroviral therapy in HIV-infected children correlates with the increase in thymus volume.

Despite significant rises in total CD4 T cells, the process of immune reconstitution in adults with HIV infection treated with potent antiretroviral treatment results in a rather slow increase in phenotypically naive lymphocytes. In children more than in adults, thymic function may be at least partly restored when disease-induced immunosuppression is attenuated by pharmacological means. Twenty-five vertically infected and antiretroviral-experienced [zidovudine (ZDV)/ZDV plus didanosine (ddl)] children were prospectively followed during 12 months of treatment with lamivudine (3TC), stavudine (d4T) and indinavir (IDV). The plasma HIV viral load and phenotypic and functional cellular immunity-defining parameters were examined. The relationship between the degree of immune reconstitution and thymus volume assessed by nuclear magnetic resonance was also examined. An early and steep increase in CD45RA+62L+ T cells was observed in parallel with a sustained decrease in plasma HIV RNA levels and a significant rise in total CD4 T cells. This increase was significantly greater than that observed in CD4+CD45RO+ T cells. Analysis of the CD4 T cell receptor (TCR) beta repertoire and T helper function showed the ability to reconstitute families almost completely absent at baseline, and a substantial improvement of antigen-specific responses by peripheral blood lymphocytes. The rise in CD4 cells and in CD4+CD45RA+62L+ T cells was statistically associated with changes in thymus size observed over time. These data suggest a relevant contribution of the thymus to reconstitution of the peripheral pool of T cells in vertically HIV-infected children treated with potent antiretroviral regimens.

In vitro regulation of immunoglobulin synthesis after marrow transplantation. I. T-cell and B-cell deficiencies in patients with and without chronic graft-versus-host disease

Twenty-four patients with aplastic anemia or acute leukemia were treated by marrow grafts from HLA-identical donors after conditioning with high doses of cyclophosphamide and/or today body irradiation. They were studied between 4 and 63 mo (median 14.2) after transplantation. Seventeen patients had chronic graft-versus-host disease (C-GVHD) and 7 were healthy. They were studied for defects in their T- and B-cell function using and indirect hemolytic plaque assay for Ig production after 6 days of culture in the presence of pokeweek mitogen. T or B cells from the patients with or without C-GVHD were cocultured with T or B cells from their HLA-identical marrow donors or unrelated normal controls. Intrinsic B-cell defects, lack of helper T-cell activity, and suppressor T-cell activity were more frequently found in patients with C-GVHD than in healthy patients. Fifteen of the 17 patients with C-GVHD showed on or more defects in their T-and B-cell function compared to only 3 of the 7 patients without C-GVHD. None of the healthy controls, including the marrow donors, showed defects in their T- and B-cell functions. These in vitro findings may be helpful in assessing the process of immune reconstitution and the immunologic aberration found after human marrow transplantation.

In Vitro Regulation of Immunoglobulin Synthesis After Marrow Transplantation. I. T-Cell and B-Cell Deficiencies in Patients With and Without Chronic Graft-Versus-Host Disease

AbstractTwenty-four patients with aplastic anemia or acute leukemia were treated by marrow grafts from HLA-identical donors after conditioning with high doses of cyclophosphamide and/or total body irradiation. They were studied between 4 and 63 mo (median 14.2) after transplantation. Seventeen patients had chronic graft-versus-host disease (C-GVHD) and 7 were healthy. They were studied for defects in their T- and B-cell function using an indirect hemolytic plaque assay for Ig production after 6 days of culture in the presence of pokeweed mitogen. T or B cells from the patients with or without C-GVHD were cocultured with T or B cells from their HLA-identical marrow donors or unrelated normal controls. Intrinsic B-cell defects, lack of helper T-cell activity, and suppressor T-cell activity were more frequently found in patients with C-GVHD than in healthy patients. Fifteen of the 17 patients with C-GVHD showed one or more defects in their T- and B-cell function compared to only 3 of the 7 patients without C-GVHD. None of the healthy controls, including the marrow donors, showed defects in their T- and B-cell functions. These in vitro findings may be helpful in assessing the process of immune reconstitution and the immunologic aberrations found after human marrow transplantation.