Impact of Acute GVHD on Immune Reconstitution after Cord Blood Transplantation in Adult.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 984-984 ◽  
Author(s):  
Tokiko Nagamura-Inoue ◽  
Satoshi Takahashi ◽  
Jun Ooi ◽  
Akira Tomonari ◽  
Toru Iseki ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Cord Blood ◽  
Nk Cells ◽  
Immune Reconstitution ◽  
Cord Blood Transplantation ◽  
Adult Patients ◽  
Blood Transplantation ◽  
Cd34 Cells ◽  
Grade Ii

Abstract BACKGROUND: Immune reconstitution following unrelated cord blood transplantation (UCBT) in adult patients is of great concern because of immaturity of cord blood immunological cells. STUDY DESIGN AND METHODS: Twenty-six adult patients (15 to 58 year-old) with hematological malignancies, who underwent UCBT and sustained engraftment were enrolled in this study. Infused number of immunological cells in thawed CB units including T cells (CD3+), B cells (CD19+), NK cells (CD3-CD56+), monocytes (CD14+) and also CD34 + cells was analysed using bead-contained TRUCOUNT tube (BD, CA). Dead cells after thawing were excluded by gating out with 7AAD dye. Immune reconstitution was analysed every 30 days by 120 days after CBT. Four-colour FACS Caliber and TRUCOUNT tube were utilized to calculate the absolute number of immune cells concentration in blood after UCBT. We put strict volume of 50μl fresh unmanipulated blood in each TRUCOUNT tube. RESULTS: Thawed-transplanted NC 2.3±x107/kg, CD34 was 0.72±0.3x105/kg (4.1x106 total), T cells; 3.1±1.6x106/kg with CD4/8 ratio of 3.2±2.0, B cells; 1.2±0.5x106/kg, NK cells; 1.0±0.5x106/kg and monocytes; 1.6±0.6x106/kg. There were no correlations between infused CD34+ cells number and T, B, NK and monocytes numbers. Monocytes increased in blood rapidly after CBT at 30 days, then, declined to the normal value. NK cells was recovered in the early after CBT and then did not so change in number from 30 to 120 days after CBT, while T cells increased time dependent manner, and B cells appeared late but influenced by acute GVHD grade. Within 120 days after CBT, T cells showed also CD4+dominant in most cases with relatively high CD25+CD4+ regulatory T (rT) cells compared to normal control. The patients with grade II to IV aGVHD showed significantly higher number of rT cells on 30 days (P<0.05) compared to those with grade 0–I aGVHD. On day 30, the number of rT cells showed 7.7±5.9/μl in grade 0–I aGVHD and 19.4±13.3/ μl in grade II–IV. The patients with grade II to IV aGVHD showed significant delayed recovery of B cells on 90 days after CBT compared to those with 0–I aGVHD (P<0.001). CONCLUSION: aGVHD in adult patients may influence on the number of regulatory T cells in the early period after UCBT and delayed recovery of B cells.

Effects of Cord Blood Cell Subset Populations in the Development of the Dominant Cord Blood Unit in Non-Myeloablative Sequential Double Cord Blood Transplantation (DCBT).

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3148-3148 ◽  
Author(s):  
David T. Ting ◽  
Karen Ballen ◽  
Thomas R. Spitzer ◽  
Richard L. Haspel ◽  
Michelle Dorn ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Cord Blood ◽  
Nk Cells ◽  
Cd8 T Cells ◽  
Cord Blood Transplantation ◽  
Cell Populations ◽  
Blood Transplantation ◽  
Graft Outcome ◽  
The Difference

Abstract Umbilical cord blood has become an established alternative hematopoietic stem cell source for patients without suitable donors. However, single cord blood transplantation in adults has been associated with high mortality rates due to graft failure, delayed engraftment, and poor immune reconstitution. In order to improve engraftment, sequential cord blood transplantation has been employed. One consistent observation with this strategy has been the eventual dominance of one cord graft over the other. However, the factors that determine the fate of one cord over the other have not been well elucidated. Our previous work demonstrated that the dominant cord tends to have an initial higher CD34+ cell dose. This study attempted to identify other cell populations in the cord units that may determine which unit becomes the ultimate source of hematopoiesis. Forty patients with hematologic malignancies underwent non-myeloablative conditioning with fludarabine, melphalan, and ATG followed by two sequential cord blood infusions. All cord units were at least a 4/6 HLA match with the recipient and each other. GVHD prophylaxis consisted of cyclosporine & mycophenolate mofetil or FK-506 & rapamycin. Chimerism analysis of peripheral blood leukocytes was performed by PCR of short tandem repeat loci. Flow cytometry for T-cells (CD4 and CD8), B-cells (CD19), and NK cells (CD3−CD56+CD16− & CD3−CD56+CD16+) were done using standard immunofluorescence methods. A total of 10 patients were available for this analysis. The Wilcoxon signed rank test was used to evaluate the significance of the difference in cell populations between the transplanted cords on graft outcome. This analysis demonstrated that 80% (8 of 10) of the dominant cords had higher NK cells when compared to the rejected unit with average total NK cells of 4.15 x 106 vs 2.30 x 106 per kg of recipient, respectively. Total NK cells included both CD16− and CD16+ cells, but the difference was much higher for CD16+ NK cells, which represent a more mature NK phenotype. B-cells, CD4+ T-cells, and CD8+ T-cells were similar between the two cords. The mean total cell/kg of recipient for each subpopulation between cord units are summarized in Table 1. A trend towards a difference in NK cells (p=0.1) was observed; the small number of samples, however, limited the interpretation of the analysis. Future studies involving additional DCBT recipients are ongoing to better define the influence of NK cells as well as other cell populations including mesenchymal stem cells and T-reg cells on cord graft outcome. Mean Subpopulation Cell Count/kg of recipient between Cord Units Cord CD3+CD4+ T-Cells CD3+CD8+ T-Cells CD3−CD56+CD16− NK Cells CD3−CD56+CD16+ NK Cells Total NK Cells CD19− B-Cells Dominant Cells/kg 3.92E+06 (1.35–7.30E+06) 1.76E+06 (0.5–3.22E+06) 4.64E+05 (0.06–1.52E+06) 3.68E+06 (0.4–9.65E+06) 4.15E+06 (0.7–9.97E+06) 1.68E+06 (0.3–3.96E+06) Rejected Cells/kg 4.34E+06 (2.04–7.83E+06) 1.71E+06 (0.7–4.86E+06) 3.60E+05 (0.0–0.9E+06) 1.94E+06 (0.3–3.55E+06) 2.30E+06 (0.3–4.10E+06) 1.80E+06 (0.9–3.15E+06)

Cord Blood Transplantation from Unrelated Donors for Children with Acute Lymphoblastic Leukemia in Japan: The Impact of Methotrexate on the Clinical Outcome.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2023-2023
Author(s):  
Koji Kato ◽  
Ayami Yoshimi ◽  
Etsuro Ito ◽  
Kentaro Oki ◽  
Jun Hara ◽  
...  
Keyword(s):  
Cord Blood ◽  
Acute Gvhd ◽  
Lymphoblastic Leukemia ◽  
Cord Blood Transplantation ◽  
Disease Status ◽  
Blood Transplantation ◽  
Gvhd Prophylaxis ◽  
Cd34 Cells ◽  
Advanced Stages ◽  
Grade Ii

Abstract Cord blood transplantation (CBT) became one of the important alternatives in allogeneic stem cell transplantation for children with hematological malignancies. We have analyzed the clinical outcomes of CBT for children with acute lymphoblastic leukemia (ALL) in Japan and identified risk factors of transplant outcome, when they had no prior transplant. From 1997 to 2006, total 332 children with ALL have undergone CBT from unrelated donor and 270 of them had no prior transplant. They are 0–15 yrs (median 5) and 4 to 60kg of body weight (median 18). Serological disparities of HLA for graft-versus-host disease (GVHD) direction were 0(n=54), 1(n=168) and 2(n=47), and disease status at transplant were 1st complete remission (CR) (n=120), 2nd CR (n=71), and more advanced stages (n=75). The median number of nucleated cells in cord blood unit was 4.93×107/kg (1.35–24.9), and that of CD34+ cells was 1.53×105/kg (0.17–15.0). As preconditioning, total body irradiation (TBI) was given in 194 patients and methotrexate (MTX) was given as GVHD prophylaxis in 159 patients. The neutrophil engraftment was achieved in 88.5% (95%CI: 84.1–91.8%) of patients and platelet engraftment (&gt;50k) was obtained in 72.6% (95%CI: 66.8–77.7). The incidence of grade II–IV and III–IV acute GVHD was 45.6% (95%CI: 39.5–51.4) and 20.4% (95%CI: 15.8–25.4) respectively. Non-relapse mortality was observed in 22.6% (95%CI: 17.7–27.8) and 35.2% (95%CI: 29.2–41.3) of patients relapsed after CBT. The five year event free survival (EFS) of all patients was 38.1% (95%CI: 34.9–41.3); 47.4%(95%CI: 42.4–52.4) in 1st CR, 45.5%(95%CI: 38.9–52.1) in 2nd CR and 15.2%(95%CI: 10.8–38.9) in more advanced stages, respectively. The multivariate analysis revealed that the larger number of CD34+ cells (p&lt;0.01) and administration of granulocyte colony stimulating factor after CBT (p&lt;0.01) were associated with earlier neutrophil engraftment. Preconditioning with TBI (p&lt;0.01) and absence of MTX (p=0.037) significantly affected the development of grade II-IV acute GVHD. Advanced disease at transplant was the most predominant factor for leukemic relapse (p&lt;0.01). GVHD prophylaxis with MTX (p=0.042), less allele mismatches (p=0.013) and disease status of 1st and 2nd CR at transplant (p&lt;0.01) were significantly associated with better EFS. Our results showed the favorable effect of MTX for the development of acute GVHD and EFS. In conclusion, GVHD prophylaxis including MTX is important in CBT for children with ALL.

BK Virus Reactivation After Double Umbilical Cord Blood Transplantation in Adults Correlates with Tregs and Delayed Reconstitution of CD4+ and CD8+ T Effector Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4174-4174
Author(s):  
Gowri Satyanarayana ◽  
Sarah Hammond ◽  
Haesook T Kim ◽  
Sean McDonough ◽  
Julia Brown ◽  
...  
Keyword(s):  
T Cells ◽  
Cord Blood ◽  
Immune Reconstitution ◽  
Hemorrhagic Cystitis ◽  
Cord Blood Transplantation ◽  
Bk Virus ◽  
Cellular Mechanisms ◽  
Effector Cells ◽  
T Effector Cells ◽  
Blood Transplantation

Abstract Abstract 4174 Umbilical cord blood transplantation (UCBT) in adults is associated with impaired immune function and increased infection-related morbidity and mortality due to lack of antigen experienced cells and delayed immune reconstitution. BK virus (BKV) is a human polyomavirus that remains latent in renal epithelial cells and can be reactivated after hematopoietic stem cell transplantation (HSCT), leading to hemorrhagic cystitis. Data regarding BKV reactivation and its association with immune reconstitution after UCBT is lacking. We evaluated the status, cellular mechanisms, and clinical implications of immune reconstitution on BK viremia in adults with hematologic malignancies undergoing double unit cord blood transplantation. Thirty-two patients with a median age of 50 years with hematopoietic malignancies were treated with reduced intensity conditioning (Flu/Mel/rATG) followed by infusion of two sequential UCB grafts and GvHD prophylaxis with tacrolimus and sirolimus. The grafts were at least a 4/6 match with each other and the recipient. The results are based on 27 evaluable patients. Assessments were done prior to transplant and at 1, 2, 3, 6 and 12 months after UCBT. After UCBT, 15 patients had detectable serum BKV DNA, median 2.6×104 copies/ml (range, 2.5×102–7.9×106) with a median time to viremia of 40 days (range, 26–733). The cumulative probability of developing BK viremia by day 100 was 0.52 (95% CI, 0.33–0.71). In 9 of the 15 patients with detectable serum BKV DNA, urinary BKV PCR was also performed. All 9 tested patients had detectable urinary BKV and developed clinical symptoms ranging from dysuria to hemorrhagic cystitis. To determine whether development of BK viremia was related to the immunological status, we analyzed detection of serum BKV DNA in conjunction with parameters of immune reconstitution. At 6 and 12 months after transplantation development of BK viremia displayed a statistically significant inverse correlation with CD4+ and CD8+ T cells (p<0.05). Development of BK viremia at these time intervals also inversely correlated with CD4+CD45RO+ and CD8+CD45RO+ T cells (p<0.05), consistent with a potentially significant role of these effector populations in preventing and/or clearing BKV. Conversely, simultaneoulsy, there was a significant positive correlation of BK viremia with T regulatory cell numbers (p<0.05) suggesting that cellular mechanisms of Treg-mediated immune suppression were directly involved in regulating this clinical outcome. At 3 months after UCBT there was a significant positive correlation (p<0.05) between BK viremia and T cell receptor excision circles (TRECs), which are expressed in recent thymic emigrant T cells. BK viremia was not dependent on any other immune cell populations including CD20+ B cells, CD16+CD56+ NK cells and CD14+ monocytes. Furthermore, prevention and/or clearance of BK viremia was not dependent on naïve cell numbers as determined by lack of correlation between BK viremia -or absence thereof- with CD4+CD45RA+ T cells and CD8+CD45RA+ T cells. These observations were in complete contrast to our previous findings regarding CMV-specific immunity, which revealed that prevention and/or clearance of CMV viremia depend on reconstitution of thymopoiesis and increase of TRECs and naïve CD4+CD45RA+ cells. In addition, we found no correlation between development of CMV viremia and BK viremia in UCBT recipients. Our results indicate that reactivation of BK virus occurs with high frequency in adult UCBT recipients and is related to the inability of TREC positive cells to control BK viremia, the impaired and delayed reconstitution of CD4+ and CD8+ T effector cells, and the suppressive function of Treg. Furthermore, our results indicate that distinct immunological mechanisms govern CMV-specific and BK-specific anti-viral responses after UCBT. Disclosures: No relevant conflicts of interest to declare.

Expression of CD10 and CD7 Defines Distinct In Vivo Lymphoid Reconstituting Capacity within Human Cord Blood CD34+CD38-Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3184-3184
Author(s):  
Shuro Yoshida ◽  
Fumihiko Ishikawa ◽  
Leonard D. Shultz ◽  
Noriyuki Saito ◽  
Mitsuhiro Fukata ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
B Cells ◽  
Cord Blood ◽  
Nk Cells ◽  
Progenitor Cells ◽  
Human Cord Blood ◽  
Cd34 Cells

Abstract Human cord blood (CB) CD34+ cells are known to contain both long-term hematopoietic stem cells (LT-HSCs) and lineage-restricted progenitor cells. In the past, in vitro studies suggested that CD10, CD7 or CD127 (IL7Ra) could be candidate surface markers that could enrich lymphoid-restricted progenitor cells in human CB CD34+ cells (Galy A, 1995, Immunity; Hao QL, 2001, Blood; Haddad R, 2004, Blood). However, in vivo repopulating capacity of these lymphoid progenitors has not been identified due to the lack of optimal xenogeneic transplantation system supporting development of human T cells in mice. We aim to identify progenitor activity of human CB CD34+ cells expressing CD10/CD7 by using newborn NOD-scid/IL2rgKO transplant assay that can fully support the development of human B, T, and NK cells in vivo (Ishikawa F, 2005, Blood). Although LT-HSCs exist exclusively in Lin-CD34+CD38- cells, not in Lin-CD34+CD38+ cells, CD10 and CD7 expressing cells are present in Lin-CD34+CD38- cells as well as in Lin-CD34+CD38+ cells (CD10+CD7+ cells, CD10+CD7- cells, CD10-CD7+ cells, CD10-CD7- cells accounted for 4.7+/−2.7%, 10.5+/−1.9%, 7.6+/−4.4%, and 77.1+/−5.2% in Lin-CD34+CD38- CB cells, respectively). We transplanted 500–6000 purified cells from each fraction into newborn NOD-scid/IL2rgKO mice, and analyzed the differentiative capacity. CD34+CD38-CD10-CD7- cells engrafted long-term (4–6 months) in recipient mice efficiently (%hCD45+ cells in PB: 30–70%, n=5), and gave rise to all types of human lymphoid and myeloid progeny that included granulocytes, platelets, erythroid cells, B cells, T cells, and NK cells. Successful secondary reconstitution by human CD34+ cells recovered from primary recipient bone marrow suggested that self-renewing HSCs are highly enriched in CD34+CD38–CD10–CD7- cells. CD10–CD7+ cells were present more frequently in CD34+CD38+ cells rather than in CD34+CD38- cells. Transplantation of more than 5000 CD34+CD38+CD10–CD7+ cells, however, resulted in less than 0.5% human cell engraftment in the recipients. Within CD34+CD38–CD10+ cells, the expression of CD7 clearly distinguished the distinct progenitor capacity. At 8 weeks post-transplantation, more than 70% of total human CD45+ cells were T cells in the CD10+CD7+ recipients, whereas less than 30% of engrafted human CD45+ cells were T cells in the CD10+CD7– recipients. In the CD10+CD7- recipients, instead, more CD19+ B cells and HLA–DR+CD33+ cells were present in the peripheral blood, the bone marrow and the spleen. Both CD34+CD38–CD10+CD7+ and CD34+CD38–CD10+CD7- cells highly repopulate recipient thymus, suggesting that these progenitors are possible thymic immigrants. Taken together, human stem and progenitor activity can be distinguished by the expressions of CD7 and CD10 within Lin-CD34+CD38- human CB cells. Xenotransplant model using NOD-scid/IL2rgKO newborns enable us to clarify the heterogeneity of Lin-CD34+CD38- cells in CB by analyzing the in vivo lymphoid reconstitution capacity.

In Vitro Assessment of Tolerance and Rejection Occurring After Co-Transplantation of Allogeneic Umbilical Cord Blood and Haploidentical CD34+ Cells as Treatment for Severe Aplastic Anemia

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2352-2352
Author(s):  
Nicole J. Gormley ◽  
Aleah Smith ◽  
Maria Berg ◽  
Lisa Cook ◽  
Catalina Ramos ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cord Blood ◽  
Cord Blood Transplantation ◽  
Neutrophil Recovery ◽  
Post Transplant ◽  
Blood Transplantation ◽  
Cd34 Cells ◽  
Cord Blood Cells

Abstract Abstract 2352 Introduction/Methods: The administration of highly purified haploidentical peripheral blood CD34+ cells combined with an unrelated cord blood transplant results in earlier neutrophil engraftment than is typically seen with a cord blood transplant alone. Chimerism data from pilot trials evaluating this strategy have reported 3 phases of engraftment: 1) early myeloid engraftment from transplanted haplo-CD34+ cells followed by 2) cord blood engraftment resulting in dual chimerism and 3) the subsequent disappearance of haploidentical donor cells with resultant full donor cord chimerism. The mechanism accounting for the disappearance of haploidentical cells has not been defined. Here the clinical results and an in vitro assessment of alloreactivity in three patients that underwent combined haploidentical CD34+ cell and cord blood transplantation for severe aplastic anemia (SAA) are described. The conditioning regimen consisted of cyclophosphamide (60mg/kg/day on days -7 and -6), fludarabine (25mg/m2/day on days -5 to -1), horse ATG (40mg/kg/day on days -5 to -2), and total body irradiation (200cGy on day -1). GVHD prophylaxis consisted of tacrolimus and mycophenolate mofetil. PCR of STRs was used to assess chimerism in T-cell and myeloid lineages and mixed lymphocyte reaction assays(MLR) were performed on peripheral blood samples collected at different time-points post-transplant to assess for alloreactivity against the recipient, the haploidentical donor, or the cord unit. Stimulator cord blood cells for the MLR were obtained from residual cord blood cells remaining in the infusion bag after patient administration and expanded in vitro using anti-CD28/CD3 Dynabeads. Results: Prior to transplantation, all three pts had transfusion dependent SAA associated with severe neutropenia that was refractory to conventional immunosuppressive therapy. Pt 1 had an early transient myeloid recovery (ANC 400 on day+11) from the haploidentical donor followed by engraftment of the cord unit (Cord ANC > 500) on day 21. The patient is currently 2 years post transplant and has 100% cord blood chimerism and is transfusion independent. An MLR assay performed when donor T-cell chimerism was 100% cord, showed evidence for rejection of the haploid cells by cord blood T-cells, with the MLR response to haploidentical donor cells being seven fold higher than the response to fully HLA-mismatched 3rd party cells. In pt 2, neutrophil recovery from the transplanted haploidentical donor occurred on day +10, with chimerism studies showing no evidence for cord engraftment in either myeloid or T-cell lineages at any point post-transplant. The patient is currently 15 months post transplant and is transfusion independent with normal blood counts and sustained “split” chimerism (T-cells recipient in origin with myeloid cells being 100% haploidentical donor). MLR assays showed that the recipient was tolerant to the haploid donor, with no statistically significant difference in the alloreactive response to the haploid donor compared to self. In pt 3, neutrophil recovery from the transplanted haploidentical donor occurred on day +10, with chimerism studies showing split chimerism (T-cell chimerism >90% cord and myeloid chimerism 88–100% haploid donor in origin). MLR assays again showed evidence of rejection of the haploid cells by cord blood T-cells, with a trend towards greater alloreactivity against the haploid donor compared to an HLA mismatched 3rd party on post-transplant day +63. Conclusions: Combined haploidentical CD34+ cell and unrelated cord blood transplantation following highly immunosuppressive conditioning represents a viable treatment option for patients with SAA who lack an HLA-matched donor. Using this approach, 2 of 3 pts had cord blood engraftment associated with early neutrophil recovery from the haploidentical donor. In one pt, the cord unit failed to engraft. Remarkably, sustained engraftment from the haploidentical donor in this pt resulted in transfusion independence. MLR appears to be a useful approach to assess the in vitro alloreactivity of this unique stem cell graft source. In the two pts who had cord engraftment, in vitro MLR assessments established that the disappearance of haploid cells occurred as a consequence of rejection of the haploidentical cells by engrafting cord blood T-cells, rather than from non-immunological haploidentical cell graft failure. Disclosures: No relevant conflicts of interest to declare.

Development of CMV-SPECIFIC Immunity after Cord Blood Transplantation in Adults Depends on Reconstitution of Thymopoiesis and Regeneration of NAÏVE CD8+ T Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1167-1167
Author(s):  
Julia Brown ◽  
Sean McDonough ◽  
Kristin Stevenson ◽  
Carol Reynolds ◽  
Haesook Kim ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Immune Reconstitution ◽  
Cord Blood Transplantation ◽  
Adult Patients ◽  
Normal Range ◽  
Cell Numbers ◽  
Post Transplant ◽  
Blood Transplantation

Abstract The treatment of hematological malignancies with umbilical cord blood transplantation (UCBT) is rapidly increasing for adult patients. Disadvantages of UCBT include insufficient cell numbers for adult patient reconstitution, a lack of antigen experienced cells, and deficits in T cell signal transduction mechanisms. Consequently, UCBT is frequently associated with impaired immune function and high infection-related mortality. To counter these difficulties, transplantation with two UCB units has been employed to improve immune reconstitution in adult patients. We evaluated both the quantitative and functional reconstitution of cellular immunity in a group of adult patients undergoing UCBT. Thirty-two patients with a median age of 50 years with hematopoietic malignancies were treated with reduced intensity conditioning (Flu/Mel/rATG) followed by infusion with two sequential UCB grafts and GvHD prophylaxis with tacrolimus and sirolimus. The grafts were at least a 4/6 match with each other and the recipient. Here we report the results of 27 patients who have completed at least one year of follow up. Assessments were done prior to transplant and at various time intervals until 12 months post UCBT. Neutrophil and platelet engraftment occurred at a median of 21 days and 42 days, respectively. CD3+ populations remained severely depressed until 8 wks post-transplant when they gradually began to re-emerge. However the CD4+ and CD8+ populations demonstrated distinctly different reconstitution kinetics. At 6 months the median value of absolute numbers of CD4+ lymphocytes was 35% of pre-transplant levels increasing to 42% at 1 yr post-transplant, a median value far below the normal range for adults. In contrast, at 6 months post-transplant CD8+ lymphocytes remained severely depressed to 12% of pre-transplant levels, but dramatically increased and reached normal levels by 1 yr after UCBT. Interestingly, both the CD14+ monocyte and the CD16+CD56+ NK cell populations expanded dramatically at 4 wks post-transplant and reached pre-transplant levels and were within the normal range by 6 months. CD20+ B cell repopulation began at 8 wks post-transplant, displayed a striking expansion leading to a 17-fold increase in the median value for B cell numbers over pre-transplant values at 1 year and resulting in a median value of absolute numbers near the top of the normal range. To evaluate functional T cell immune reconstitution in vivo, we performed IFN-γ ELISpot analysis on CMV stimulated PBLs and compared the results to a PCR-based assay for CMV viremia. Additionally, we assessed the reconstitution of thymopoiesis with the T cell receptor excision circle (TREC) assay and real-time PCR. 16/27 patients and 26/52 UCB products were CMV seropositive prior to transplant. In the post-UCB period, development of CMV-specific effectors as determined by ELISpot did not always correlate with clearance of CMV viremia. Specifically, prior to 8 weeks post-UCBT, 8 out of 12 (67%) patients with CMV-positive ELISpot displayed CMV viremia, between 8 weeks and 100 days post-UCBT 4 out of 11 (36%) patients with CMV-positive ELISpot displayed CMV viremia and after 100 days post-UCBT only 3 out of 10 patients (30%) who developed CMV-positive ELISpot remained positive for CMV viremia. Identification of functional CMV effectors was only associated with the numbers of CD8+CD45RA+ cells (p=0.01) and the development of high TREC concentrations (p=0.01) that were detected after 6 months of UCBT, and was independent of GvHD or mixed chimerism. Taken together these results indicate that reconstitution of T cell immunity after UCBT is characterized by delayed recovery of CD4+ and CD8+ T cells and correlates with reconstitution of thymopoiesis and increase of naïve CD8+CD45RA+ T cells that can develop into efficient pathogen-specific effectors.

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.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2926-2926
Author(s):  
Satoshi Takahashi ◽  
Jun Ooi ◽  
Akira Tomonari ◽  
Nobuhiro Tsukada ◽  
Takaaki Konuma ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cord Blood ◽  
Cd8 T Cells ◽  
Immune Reconstitution ◽  
Cord Blood Transplantation ◽  
Blood Transplantation ◽  
Total Body ◽  
Cmv Reactivation ◽  
Cmv Antigenemia

Abstract 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.

The Enhancement of Short-Term Engraftment in NOD/Scid Mice after Double Cord Blood Transplantation Is Mediated by CD34+ Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4596-4596
Author(s):  
Yvette van Hensbergen ◽  
Jan J. Cornelissen ◽  
Willem E. Fibbe ◽  
Anneke Brand
Keyword(s):  
Stem Cells ◽  
Cord Blood ◽  
Cord Blood Transplantation ◽  
Scid Mice ◽  
Adult Patients ◽  
Short Term ◽  
Enhancing Effect ◽  
Cell Numbers ◽  
Blood Transplantation ◽  
Cd34 Cells

Abstract Introduction: Cord blood (CB) is an attractive source of hematopoietic stem cells, however, the low number of cells available hampers its use in adult patients. Transplantation of two unrelated CB units seems to overcome the cell dose limitations for adult patients with at least similar engraftment levels compared to single CB transplantation with sufficient cell numbers. Pre-clinical studies show an important role for total cell numbers in the graft, but also CD34+ stem cells or CD34− cells, including NK cells and/or Lin+, CD3+ T cells may play a facilitating role, possibly via a graft-versus-graft effect. In this study we explored the role of the CD34+ and CD34− cells in facilitating engraftment in NOD/Scid mice after double CB transplantation. Methods: Cohorts (ch.) of irradiated (3.5Gy) NOD/Scid mice (n=9–18 mice per cohort; 3 independent experiments) were transplanted with a combination of 1.105 CD34+ cells (MACS purified; &gt;94% pure) and 1.107 CD34- cells derived from one CB unit. In addition, mice received a combination of CD34+ and CD34− cells, or CD34+, or CD34− cells only, derived from a second CB unit (Table 1). BM engraftment was assessed by the percentage of human CD45+ cells at 6 weeks after transplantation. In addition, the percentage of human lymphoid (CD19/CD3) and myeloid (CD33) cells in the BM were evaluated at 6 weeks. Human platelet (hPLT) and human CD45+ cell recovery in blood was measured from 3 to 6 weeks after transplantation. Results: At 6 weeks, 67% of the mice in the single CB cohort (ch.1), 75% in the cohort co-transplanted with CD34− cells (ch.4), 83% in the cohort co-transplanted with CD34+ cells (ch.3) and 89% in the cohort co-transplanted with the combination of CD34+ and CD34− cells of the second donor, showed engraftment (&gt;10% human CD45+ cells) in the BM. Mice that were co-transplanted with CD34+ cells only (ch.3) had a significantly higher level of engraftment in the BM than mice receiving no second CB unit (ch.1) (54% vs. 22%; p=0.011). A similar engraftment enhancing effect was observed in mice co-transplanted with a combination of CD34+ and CD34− cells (ch.2) (50% p=0.016 vs. ch.1). In contrast, no significant enhancing effect was observed after co-transplantation of CD34− cells only (ch.4) (36%). The distribution between human lymphoid and myeloid cells in BM was similar between al cohorts. All engrafted mice had sustained mixed chimerism in the BM without pre-dominance of a single CB unit during the time of evaluation. hPLT recovery was significantly increased in mice co-transplanted with CD34+ cells, either alone (ch.3; 607plt/μl) or in combination with CD34− cells (ch.2; 1076 plt/μl) compared to mice receiving no second CB (ch.1; 183 plt/μl) (p=0.04 and p&lt;0.001 resp.). The observed mean hPTL concentration in cohort-2 and -3 was greater than expected by the sum of the single CBs (305 plt/μl), indicating a synergistic effect. In mice that were cotransplantated with CD34− cells from a second donor (ch.4) no significant effect on hPLT recovery (253 plt/μl) was observed. Similar results were obtained for hCD45 recovery in peripheral blood. Conclusion: These results show that the engraftment facilitating effect of co-transplantation of CD34+ cells only are similar compared to co-stransplantation of both CD34+ and CD34− cells. Therefore, it is suggested that enhancement of short-term engraftment in NOD/Scid mice by double cord blood transplantation is mainly mediated by the CD34+ cells in the second CB. Table 1: Composition of the transplants per cohort. Cohort-1 received a single CB transplantation of donor-1 or donor-2. Cohort-2 received a normal double CB transplantation. Cohort Transplanted cells of donor-1 Co-transplanted cells of donor 2 1 CD34+ + CD34− OR CD34+ + CD34− 2 CD34+ + CD34− AND CD34+ + CD34− 3 CD34+ + CD34− AND CD34+ 4 CD34+ + CD34− AND CD34−

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