Oxidative and Inflammatory Markers Are Higher in Full-Term Newborns Suffering Funisitis, and Higher Oxidative Markers Are Associated with Admission.

The aim of this study was to assess whether oxidative and inflammatory mediators in the cord blood of newborns with funisitis and chorioamnionitis can serve as indicators of their inflammatory status, and whether there is a positive association between higher mediator levels and an increased risk of admission to the neonatal intensive care unit (NICU). This study was conducted prospectively in a neonatology department of a university hospital. In total, 52 full-term newborns were evaluated, including 17 funisitis cases, 13 chorioamnionitis cases, and 22 control newborns without funisitis or chorioamnionitis. Cord blood samples were measured for oxidative stress and inflammatory status markers. The oxidative stress markers included the total nitric oxide (NO), total hydroperoxide (TH), biological antioxidant potential (BAP), and TH/BAP ratio, comprising the oxidative stress index (OSI). Inflammatory markers included interleukin (IL)-1b, IL-6, IL-8, IL-10, tumor necrosis factor alpha (TNFα), interferon γ (IFNγ), and complement component C5a. TH, OSI, IL-1b, IL-6, and IL-8 concentrations were higher in the funisitis group than in the chorioamnionitis and control groups. C5a was higher in the funisitis and chorioamnionitis groups than in the control group. Among all enrolled newborns, 14 were admitted to the NICU. Multiple logistic regression analysis showed that elevated umbilical cord blood levels of OSI and TH were associated with a higher risk of admission to the NICU (OSI: R = 2.3, 95% CI 1.26-4.29, p = 0.007 and TH: R = 1.02, 95%CI = 1.004-1.040, p = 0.015). In conclusion, OSI and TH in cord blood from full-term newborns can provide an index of inflammatory status, and higher levels are associated with the risk of admission to the NICU and, therefore, could serve as an early indicator of inflammatory conditions in newborns.

Inferior outcomes of stage III T lymphoblastic lymphoma relative to stage IV lymphoma and T-acute lymphoblastic leukemia: long-term comparison of outcomes in the JACLS NHL T-98 and ALL T-97 protocols.

T cell lymphoblastic lymphoma (T-LBL) accounts for 30 % of all childhood non-Hodgkin's lymphomas (NHL) in Japan. Twenty-nine patients with T-LBL in stages III and IV were eligible for and enrolled in the JACLS NHL-T98 trial (1998-2002), and 72 patients with T-ALL were enrolled in the JACLS ALL-T97 trial (1997-2001). The 10-year overall survival (OS) (61.1 ± 11.5 %) and the 10-year event-free survival (EFS) (44.4 ± 11.7 %) of stage III LBL were lower than those of other diseases, and the OS and EFS were nearly the same when comparing stage IV LBL and ALL (OS: stage IV LBL, 80.0 ± 12.7 % vs. ALL, 80.2 ± 4.9 %; EFS: stage IV, LBL 70.0 ± 14.5 % vs. ALL, 70.7 ± 5.5 %). Outcomes were worse for stage III LBL than for stage IV LBL or T-ALL. Given that the treatment results of T-ALL and LBL stage IV did not differ when compared with previous reports, LBL stage III in Japanese children may differ from LBL stage III in children in other countries.

A novel mutation of ribosomal protein S10 gene in a Japanese patient with diamond-Blackfan anemia.

Diamond-Blackfan anemia (DBA) is an inherited bone marrow disease. The condition is characterized by anemia that usually presents during infancy or early childhood and congenital malformation. Several reports show that DBA is associated with mutations in the ribosomal protein (RP) genes, RPS19, RPS24, RPS17, RPL35A, RPL5, RPL11, and RPS7. Recently, 5 and 12 patients with mutations in RPS10 and RPS26, respectively, were identified in a cohort of 117 DBA probands. Therefore, we screened the DBA patients who were negative for mutations in these DBA genes for mutations in RPS10 and RPS26. The present case report describes the identification of the first Japanese DBA patient with a novel mutation in RPS10.

Brain atrophy caused by vitamin B12-deficient anemia in an infant.

Vitamin B12 deficiency in infants often presents with nonspecific hematological, gastrointestinal, and neurological manifestations. It is usually caused by inadequate intake, abnormal absorption, or congenital disorders of vitamin B12 metabolism, including transport disorders. We describe a vitamin B12-deficient infant with severe anemia who was breastfed. His mother had undiagnosed vitamin B12 deficiency having undergone total gastrectomy 18 years earlier. The infant developed normally after taking vitamin B12. It is important to suspect vitamin B12 deficiency in mothers who have undergone gastrectomy. Early diagnosis and treatment of vitamin B12 deficiency in infants is important and will help improve long-term prognosis.

Incidence and risk factors of early bacterial infections after unrelated cord blood transplantation.

Incidence and characteristics of early bacterial infection within 100 days after unrelated cord blood transplantation (UCBT) were assessed for 664 pediatric and 1208 adult recipients in Japan. Cumulative incidence of early bacterial infection at day 100 post-UCBT was 11% (95% confidence interval [CI], 8%-13%) for children and 21% (CI, 19%-24%) for adults (P < .0001). Early bacterial infection in adults had a significant impact on mortality (hazard ratio [HR] = 2.1, CI, 1.7-2.6; P < .0001), although no significant risk factors were identified. Multivariate analysis identified older age group (6-10, and 11-15 years versus 0-5 years of age) at transplant (HR = 2.0 and 2.7, CI, 1.1-3.5 and 1.4-4.9; P = .020 and .002, respectively) as an independent risk factor of early bacterial infection for children. Early bacterial infection in children did not have a significant impact on mortality when adjusted. Of 315 bacteremia, 74% were caused by Gram-positive microorganisms. Pneumonia occurred in 39 patients including 13 cases of Stenotrophomonas maltophilia pneumonia. Early bacterial infection had a negative effect on survival for adults and the median day of development was 10 days after transplant, suggesting that the prevention of bacterial infection in the very early post-UCBT phase is important.

HapMap scanning of novel human minor histocompatibility antigens.

Minor histocompatibility antigens (mHags) are molecular targets of allo-immunity associated with hematopoietic stem cell transplantation (HSCT) and involved in graft-versus-host disease, but they also have beneficial antitumor activity. mHags are typically defined by host SNPs that are not shared by the donor and are immunologically recognized by cytotoxic T cells isolated from post-HSCT patients. However, the number of molecularly identified mHags is still too small to allow prospective studies of their clinical importance in transplantation medicine, mostly due to the lack of an efficient method for isolation. Here we show that when combined with conventional immunologic assays, the large data set from the International HapMap Project can be directly used for genetic mapping of novel mHags. Based on the immunologically determined mHag status in HapMap panels, a target mHag locus can be uniquely mapped through whole genome association scanning taking advantage of the unprecedented resolution and power obtained with more than 3 000 000 markers. The feasibility of our approach could be supported by extensive simulations and further confirmed by actually isolating 2 novel mHags as well as 1 previously identified example. The HapMap data set represents an invaluable resource for investigating human variation, with obvious applications in genetic mapping of clinically relevant human traits.

Hematopoietic engraftment in recipients of unrelated donor umbilical cord blood is affected by the CD34+ and CD8+ cell doses.

Umbilical cord blood (UCB) transplantation is limited by the low number of hematopoietic stem cells in UCB units, which results in a low engraftment rate in transplant recipients. Here, we measured the total nucleated cell count and CD34(+), CD3(+), CD4(+), CD8(+), CD14(+), and CD16(+)/56(+) cell doses in each UCB unit and evaluated their influence on engraftment and other outcomes in 146 recipients. Multivariate analysis showed a significant association between a higher incidence of successful engraftment and a dose of CD34(+) and CD8(+) cells above the median (1.4 x 10(5) and 15.7 x 10(5) cells/kg, respectively). Engraftment occurred 4 days earlier in patients who received UCB with more than the median dose of CD34(+) cells than those receiving UCB at or below the median. Stratification of the group according to CD34(+) cell dose revealed a significant influence of the CD8(+) cell dose on the time to achieve neutrophil engraftment in patients receiving a lower CD34(+) cell dose, whereas there was no significant influence in the patients receiving a higher CD34(+) cell dose. These results suggest that consideration of CD34(+) and CD8(+) cell doses in UCB units may improve the engraftment in recipients of UCB transplantation.

Twenty years' experience in allogeneic hematopoietic stem cell transplantation for Philadelphia chromosome-positive acute lymphoblastic leukemia in the Nagoya Blood and Marrow Transplantation Group.

Between October 1981 and December 2000, 46 patients with Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL) underwent allogeneic hematopoietic stem cell transplantation (HSCT) in the Nagoya Blood and Marrow Transplantation Group. The median age was 28.5 years (range, 4-51 years). All but one patient achieved engraftment. Grade II-to-IV acute graft-versus-host disease (GVHD) developed in 32.5% of patients, and chronic GVHD developed in 40.5%. The incidences of relapse and treatment-related mortality (TRM) at 5 years were 65% and 26%, respectively. The estimated overall survival rate at 5 years was 23%. Univariate analysis showed that improved disease-free survival (DFS) was independently associated with complete remission (CR) at transplantation (39%), compared with non-CR (8%) (P = .023). Non-CR at transplantation was associated with a higher risk of relapse. Donor type, acute GVHD, and time from diagnosis to HSCT all had a significant effect on TRM. In a multivariate analysis, 9 months or more from diagnosis to HSCT was the only variable statistically significant for DFS (relative risk, 3.22; P = .01). This study demonstrates that allogeneic HSCT cures a significant population of patients with Ph+ ALL. Relapse is the major obstacle limiting the success of HSCT. Early transplantation during CR from donors, including unrelated persons or mismatched relatives, may offer improved long-term DFS.

In vitro efficacy of l-asparaginase in childhood acute myeloid leukaemia.

To explore the potential efficacy of l-asparaginase treatment in acute myeloid leukaemia (AML) patients, we studied the in vitro resistance of French-American-British (FAB) subtypes of childhood AML to l-asparaginase using a methyl-thiazol-tetrazolium assay. We tested leukaemic cells obtained from 177 common acute lymphoblastic leukaemia (cALL) and 228 AML children at diagnosis. The median 70% lethal dose of l-asparaginase (LD70asp) (U/ml) was 0.46 in the cALL and 6.70 in the AML samples. The median LD70asp among each FAB subtype of AML was 0.76 (M0), 0.46 (M1), 10.00 (M2), 10.00 (M3), 1.18 (M4), 1.35 (M5) and 10.00 (M7). Type M3 samples had the highest LD70asp. The LD70asp of the M2 samples was significantly higher than that of the M1, M4 and M5 samples. When the LD70asp values were classified as low (0.016-0.159), intermediate (0.16-1.59) or high (1.6-10.00), the frequency of low, intermediate or high LD70asp among the M1 samples were similar to those among the cALL samples. In conclusion, cells from AML types M1, M4 and M5 were relatively sensitive to l-asparaginase, and M1 cells were as sensitive as those of cALL, suggesting that l-asparaginase treatment may be effective for these subtypes of AML.

Identification of a polymorphic gene, BCL2A1, encoding two novel hematopoietic lineage-specific minor histocompatibility antigens.

We report the identification of two novel minor histocompatibility antigens (mHAgs), encoded by two separate single nucleotide polymorphisms on a single gene, BCL2A1, and restricted by human histocompatibility leukocyte antigen (HLA)-A*2402 (the most common HLA-A allele in Japanese) and B*4403, respectively. Two cytotoxic T lymphocyte (CTL) clones specific for these mHAgs were first isolated from two distinct recipients after hematopoietic cell transplantation. Both clones lyse only normal and malignant cells within the hematopoietic lineage. To localize the gene encoding the mHAgs, two-point linkage analysis was performed on the CTL lytic patterns of restricting HLA-transfected B lymphoblastoid cell lines obtained from Centre d'Etude du Polymorphisme Humain. Both CTL clones showed a completely identical lytic pattern for 4 pedigrees and the gene was localized within a 3.6-cM interval of 15q24.3-25.1 region that encodes at least 46 genes. Of those, only BCL2A1 has been reported to be expressed in hematopoietic cells and possess three nonsynonymous nucleotide changes. Minigene transfection and epitope reconstitution assays with synthetic peptides identified both HLA-A*2402- and B*4403-restricted mHAg epitopes to be encoded by distinct polymorphisms within BCL2A1.

Targeted cloning of cytotoxic T cells specific for minor histocompatibility antigens restricted by HLA class I molecules of interest.

BACKGROUND: T cells specific for minor histocompatibility antigens (mHAgs) play a major role in both graft-versus-host disease and the graft-versus-tumor effect after HLA-identical bone marrow transplantation (BMT). However, characterization of individual T-cell responses to mHAgs is difficult and has generally involved extensive screening of T-cell clones isolated from bulk T-cell cultures generated from BMT recipients. In this report, we describe a new approach that permits both direct visualization of CD8 T-cell responses to mHAgs and cloning of T cells reacting with mHAgs presented by individual HLA alleles of interest. METHODS AND RESULTS: Panels of Epstein-Barr virus-transformed B-cell lines (B-LCL) expressing retrovirally transduced HLA cDNA were used as stimulator cells in an enzyme-linked immunospot (ELISPOT) assay to identify CD8 T cells reacting with mHAgs in cultures generated from postBMT recipient peripheral blood. T cells specific for mHAgs presented by selected HLA alleles could then be captured and cloned using an interferon-gamma secretion assay and magnetic bead selection. A majority of T-cell clones thus isolated exhibited cytolytic activity against the same HLA-transfected B-cell lines used for the ELISPOT assay. CONCLUSION: The ELISPOT assay was useful for identification of the HLA alleles presenting mHAgs recognized by individual T-cell lines. This approach for isolating mHAgs-specific CD8 T-cell clones should assist in characterizing responses restricted by HLA alleles of interest, which are common in a certain ethnic group.

Lineage conversion from acute lymphoblastic leukemia to acute myeloid leukemia on rearrangement of the IgH gene in a patient with Down syndrome.

A patient with Down syndrome (DS) at the time of diagnosis of acute lymphoblastic leukemia (ALL) had a relapse with acute myeloid leukemia (AML) after 4 years of complete remission. Although the diagnosis was AML, the leukemic blasts at relapse showed an immunoglobulin H rearrangement that turned out to be identical to that of the initial ALL blasts. It is thought that the leukemic precursor cells of this patient had the potential to differentiate into both lymphoid and myeloid lineages. This case is important for investigating target cells for leukemogenesis in DS.

Human leucocyte antigen-Cw-specific cytotoxic T lymphocytes generated from naive cord blood used for cord blood stem cell transplantation.

Human leucocyte antigen (HLA)-Cw-reactive cytotoxic T lymphocytes (CTL) were generated from cord blood (CB) lymphocytes of two cases used for cord blood stem cell transplantation (CBSCT). In both cases, the CTL were cytotoxic against the patient's leukaemic cells, as well as the patient's Epstein-Barr virus (EBV)-lymphoblastoid cell line (EBV-LCL) and phytohaemagglutinin blasts, and the cytotoxicity was blocked by anti-HLA-class I monoclonal antibodies. In the first case, the CTL recognized Cw 3 (Cw 9 and Cw 10)-positive EBV-LCL, while in the second case, the CTL recognized Cw1 and/or Cw7. These cases suggest that CB T cells may be competent enough for generating CTL to induce a graft-versus-leukaemia effect and/or graft-versus-host disease in patients with CBSCT and that the mismatching of Cw antigens between patient and CB may be related to the outcome of CBSCT.