Yoshinari Kawabata

A clinical analysis of 52 adult patients with hemophagocytic syndrome: the prognostic significance of the underlying diseases.

We retrospectively analyzed 52 adult patients with hemophagocytic syndrome (HPS). The underlying diseases were heterogeneous, including malignant lymphoma (lymphoma-associated hemophagocytic syndrome [LAHS]) in 26 patients, systemic lupus erythematosus in 3 patients, viral infections in 7 patients, and bacterial or fungal infections in 6 patients. More than 83% of patients received prednisolone as an initial treatment. Multiple-agent chemotherapies (cyclophosphamide, doxoru-bicin, and vincristine) were administered to 96% of LAHS patients after a histopathological diagnosis of lymphoma. HPSs were controllable and remissions were achieved except for those patients with LAHS, fulminant Epstein-Barr virus-ssociated HPS, and an immunosuppressive state. Twenty-one (81%) of the LAHS patients had uncontrollable HPS and died of multiple organ failure and disseminated intravascular coagulation.The median survival time of LAHS patients was 83 days. In contrast, 3 (12%) of the other HPS patients died of multiple organ failure within 44 days.The clinical manifestations and the laboratory findings of LAHS and the other HPSs were too variable to establish the prognosis based only on the findings at the onset of HPS. The prognostic factors of adult HPS were found to be the underlying diseases, notably malignant lymphoma and infections, accompanied by the immunosuppressive state.

Allogeneic haematopoietic stem cell transplantation as a promising treatment for natural killer-cell neoplasms

The efficacy of allogeneic haematopoietic stem‐cell transplantation (allo‐HSCT) for natural killer (NK)‐cell neoplasms is unknown. We investigated the results of allo‐HSCT for NK‐cell neoplasms between 1990 and 2003 through questionnaires. After reclassification by a haematopathologist, of 345 patients who underwent allo‐HSCT for malignant lymphoma, 28 had NK‐cell neoplasms (World Health Organization classification): extranodal NK/T‐cell lymphoma (n = 22), blastic NK‐cell lymphoma (n = 3), and aggressive NK‐cell leukaemia (n = 3). Twelve were chemosensitive and 16 chemorefractory. Twenty‐two had matched‐related donors. Stem‐cell source was bone marrow in eight and mobilised peripheral blood in 20. Conditioning regimens were myeloablative (n = 23) and non‐myeloablative (n = 5). Grade 2–4 acute graft‐versus‐host disease (GVHD) and chronic GVHD developed in 12 and 8 respectively. Eight died of disease progression, three of infection, two of acute GVHD, one of veno‐occlusive disease, one of interstitial pneumonitis, and one of thrombotic microangiopathy. Two‐year progression‐free and overall survivals were 34% and 40% respectively (median follow‐up, 34 months). All patients who did not relapse/progress within 10 months achieved progression‐free survival (PFS) during the follow‐up. In multivariate analysis, stem cell source (BM versus peripheral blood; relative risk 3·03), age (≥40 years vs. <40 years; relative risk 2·85), and diagnoses (extranodal NK/T‐cell lymphoma versus others; relative risk 3·94) significantly affected PFS. Allo‐HSCT is a promising treatment for NK‐cell neoplasms.

Distinct TCRAV and TCRBV repertoire and CDR3 sequence of T lymphocytes clonally expanded in blood and GVHD lesions after human allogeneic bone marrow transplantation

Acute graft-versus-host disease (GVHD) is a disorder involving the skin, gut and liver that is caused by mismatches of major and/or minor histocompatibility antigens between the HLA-identical donor and recipient. If T lymphocytes infiltrating GVHD lesions recognize antigens expressed in these organs, T cell clones should expand in inflammatory tissues. We previously reported that recipients of allogeneic bone marrow grafts have clonally expanded TCRαβ+ T lymphocytes soon after transplantation, which leads to a skew of TCR repertoires. To establish whether or not the same antigens cause clonal expansion of T lymphocytes in both blood and GVHD tissues, we examined the usage of TCR α and β chain variable regions (TCRAV and TCRBV) and determined the complementarity-determining region 3 (CDR3) of T lymphocytes clonally expanded in circulating blood and GVHD lesions. We found that the repertoires and CDR3 diversity of TCRAV and TCRBV differed between the GVHD lesions and circulating blood, suggesting the selective recruitment of antigen-specific T cells into GVHD tissues. We also found that the usage of TCRAV and TCRBV by the clonally expanded T lymphocytes and their CDR3 sequences differed between the GVHD tissues and blood. These results suggest that the antigen specificity of TCRαβ+ T lymphocytes clonally expanded in blood and GVHD lesions is different.

Identification of the T cell clones expanding within both CD8+CD28+ and CD8+CD28- T cell subsets in recipients of allogeneic hematopoietic cell grafts and its implication in post-transplant skewing of T cell receptor repertoire

We have previously reported that skewed repertoires of T cell receptor-β chain variable region (TCRBV) and TCR-α chain variable region (TCRAV) are observed at an early period after allogeneic hematopoietic cell transplantation. Furthermore, we found that T lymphocytes using TCRBV24S1 were increased in 28% of the recipients of allogeneic grafts and an increase of TCRBV24S1 usage was shown to result from clonal expansions. Interestingly, the arginine residue was frequently present at the 3′ terminal of BV24S1 segment and was followed by an acidic amino acid residue within the CDR3 region. These results suggest that these clonally expanded T cells are not randomly selected, but are expanded by stimulation with specific antigens. This study was undertaken to elucidate the mechanisms of the post-transplant skewing of TCR repertoires. Since the CD8+CD28−CD57+ T cell subset has been reported to expand in the peripheral blood of patients receiving allogeneic hematopoietic cell grafts, we examined the TCRAV and TCRBV repertoires of the CD8+CD28− T cell and CD8+CD28+ T cell subsets, and also determined the clonality of both T cell populations. In all three recipients examined, the CD8+CD28− T cell subset appeared to define the post-transplant TCR repertoire of circulating blood T cells. Moreover, the CDR3 length of TCRBV imposed constraints in both CD8+CD28− T cell and CD8+CD28+ T cell subsets. The DNA sequences of the CDR3 region were determined, and the same clones were identified within both CD8+CD28− and CD8+CD28+ T cell subsets in the same individuals. These results suggest that the clonally expanded CD8+CD28− T cells after allogeneic hematopoietic cell transplantation derive from the CD8+CD28+ T cell subset, possibly by an antigen-driven mechanism, resulting in the skewed TCR repertoire. Bone Marrow Transplantation (2001) 27, 731–739.

Reconstitution of γδ T cell repertoire diversity after human allogeneic hematopoietic cell transplantation and the role of peripheral expansion of mature T cell population in the graft

We have examined the reconstitution of γδ T cell repertoire diversity after human allogeneic hematopoietic cell transplantation using a polymerase chain reaction (PCR)-based complementarity-determining region (CDR) 3 size spectratyping and DNA sequencing. The CDR3 complexity in the variable region of the T cell receptor (TCR)-δ chain was different amongst the individuals studied. Furthermore, CDR3 size distribution patterns of allogeneic hematopoietic cell transplant recipients were almost completely recovered by a few months after transplantation. In some patients, clonal predominance of the TCRDV1+ T cells became evident during the period after transplantation. In one particular donor/recipient pair, clonal predominance of TCRDV1+ T cells was already present in blood lymphocytes of the donor, and was also observed in the recipient after transplantation. Using this donor/recipient pair, we have questioned whether γδ T cell regeneration occurs via the peripheral expansion of mature T cells in the graft. In the donor lymphocytes, two expanding γδ T cell clones, which were demonstrated by CDR3 sequences of the TCR-δ chain, were recognized. These two clones were identified in the T cells from the recipient post transplant, but not before transplantation. One of the two clones was still detectable 1½ years after the transplant procedure. These results strongly suggest that peripheral expansion of mature T cells in the graft is the principal pathway of γδ T cell regeneration after allogeneic hematopoietic cell transplantation in adults. Bone Marrow Transplantation (2000) 26, 177–185.

Extensive clonal expansion of T lymphocytes causes contracted diversity of complementarity-determining region 3 and skewed T cell receptor repertoires after allogeneic hematopoietic cell transplantation

We previously described skewed repertoires of the T cell receptor-β chain variable region (TCRBV) and the TCR-α chain variable region (TCRAV) soon after allogeneic hematopoietic cell transplantation. To determine the characteristics of skewed TCRBV after transplantation, we examined the clonality of T lymphocytes carrying skewed TCRBV subfamilies and determined the CDR3 sequences of expanded T cell clones. In all 11 recipients examined, TCR repertoires were skewed, with an increase of certain TCRBV subfamilies that differed among individuals. In nine of 11 patients, clonal/oligoclonal T cell expansion was observed, although the expanded T cells were not necessarily oligoclonal. The extent of expansion after transplantation appeared to predict clonality. The arginine (R)-X-X-glycine (G) sequence was identified in clonally expanded T cells from four of five recipients examined, and glutamic acid (E), aspartic acid (D) and alanine (A) were frequently inserted between R and G. These results suggest that T lymphocyte expansion may result from the response to antigens widely existing in humans, and that the extensive clonal expansion of a limited number of T cells leads to contracted CDR3 diversity and post-transplant skewed TCR repertoires. Bone Marrow Transplantation (2001) 27, 607–614.

Clinical Applications of CD34+ Cell‐selected Peripheral Blood Stem Cells

Abstract: Peripheral blood stem cells (PBSC) are increasingly used for stem cell transplantation after high dose chemotherapy. CD34+ cell selection has also been done for use in autologous transplantation studies. Bone marrow (BM) may contain tumor cells at the time of harvesting, and on re‐infusion, these cells could contribute to a subsequent relapse. Similarly, tumor cell contamination of PBSC collections has been found in a number of studies. Therefore, purging contaminating tumor cells may prevent cases of relapse. As most tumor cell types do not express CD34 antigen, one of the most widespread applications of CD34+ cell selection is likely to be in tumor cell purging. Similarly, CD34+ cell selection has aided allogeneic transplantation studies. Acute graft‐versus‐host disease (aGVHD) is a major cause of morbidity and mortality in cases of allogeneic transplantation. As aGVHD is mediated by donor T cells, removal of T cells from the graft by CD34+ cell selection may ensure prophylaxis against aGVHD. Further, high‐dose immunosuppression followed by CD34+ cell‐selected stem cell rescue is theoretically reasonable as a therapeutic tool for patients with autoimmune disease resistant to conventional therapy. However, patients given T cell‐depleted transplantation have an increased risk of opportunistic infection as well as malignancies related to immunosuppression; therefore, close monitoring is warranted. We describe here clinical applications of CD34+ cell‐selected PBSC for a variety of diseases, with special emphasis on the efficacy as well as drawbacks of this novel technique.

Oligoclonal expansion of CD4(+)CD28(-) T lymphocytes in recipients of allogeneic hematopoietic cell grafts and identification of the same T cell clones within both CD4(+)CD28(+) and CD4(+)CD28(-) T cell subsets.

Recipients of allogeneic bone marrow grafts have clonally expanded CD8+CD28− T lymphocytes during the early period after transplantation, which leads to skewing of T cell receptor (TCR) repertoires. Here, we have addressed the question of whether clonal expansion of CD28− T cells is also observed in CD4+ T lymphocytes after human allogeneic hematopoietic cell transplantation. We found that the fraction of T cells lacking CD28 expression in the CD4+ subset was increased after transplantation, and expanded CD4+CD28− T lymphocytes carrying certain TCRBV subfamilies showed limited TCR diversity. In order to further study the ontogeny of CD4+CD28− T cells, we analyzed the complementarity-determining region 3 (CDR3) of the TCR-β chain of CD4+CD28+ and CD4+CD28− cells. We identified the same T cell clones within both CD4+CD28− and CD4+CD28+ T cell subsets. These results suggest that both subsets are phenotypic variants of the same T cell lineage. Bone Marrow Transplantation (2001) 27, 1095–1100.

Gene Expression Profiling of Human Erythroid Progenitors by Micro-Serial analysis of Gene Expression

We compared the expression profiles of highly purified human CD34+ cells and erythroid progenitor cells by micro-serial analysis of gene expression (microSAGE). Human CD34+ cells were purified from granulocyte colony-stimulating factor-mobilized blood stem cells, and erythroid progenitors were obtained by cultivating these cells in the presence of stem cell factor, interleukin 3, and erythropoietin. Our 10,202 SAGE tags allowed us to identify 1354 different transcripts appearing more than once. Erythroid progenitor cells showed increased expression of LRBA, EEF1A1, HSPCA, PILRB, RANBP1, NACA, and SMURF. Overexpression of HSPCA was confirmed by real-time polymerase chain reaction analysis.MicroSAGE revealed an unexpected preferential expression of several genes in erythroid progenitor cells in addition to the known functional genes, including hemoglobins. Our results provide reference data for future studies of gene expression in various hematopoietic disorders, including myelodysplastic syndrome and leukemia.

Late-onset herpes simplex virus-associated interstitial pneumonia after allogeneic bone marrow transplantation

Late-onset herpes simplex virus-associated interstitial pneumonia after allogeneic bone marrow transplantation