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Thrombopoietin (TPO) is a ligand for c-mpl that promotes both proliferation and differentiation of megakaryocytes in vivo and in vitro. We investigated the expression of c-mpl transcripts and the effects of recombinant human TPO (rhTPO) on the proliferation and differentiation of human leukemic cell lines or fresh samples obtained from 32 patients with transient abnormal myelopoiesis (TAM) or acute myeloblastic leukemia (AML). Cells were cultured with TPO alone or combined with rh interleukin-3 (IL-3) or stem cell factor (SCF). Expression of c-mpl was verified in 6 of 13 cases tested. All but one of the cases that showed c-mpl expression responded to TPO. Blasts from all cases of TAM or French-American-British (FAB) subtype M7 showed growth responses to TPO with higher sensitivity than cells of other FAB subtypes and these responses were increased by addition of rhIL-3 or rhSCF in some cases. Responses of cells of other FAB subtypes varied. In addition, increased expression of platelet-specific surface antigens on MO7E cells after incubation with rhTPO was observed. These data suggest that TPO may be involved in the abnormal proliferation and differentiation of human leukemic cells, especially of M7 and TAM cells, considered to be of megakaryocytic lineage.

We investigated the effects of the administration of FLT-3 ligand (FL) on mobilization of primitive and committed progenitor cells in mice. C57bl/6J mice were injected subcutaneously with FL once a day for 5 d at doses of 20, 100 and 200 microg/kg. After the collection of peripheral blood, we determined the number of white blood cells (WBCs) with the differential counts. The formation of colony-forming cells (CFCs) in peripheral blood, bone marrow and spleen was evaluated. Although the administration of FL, 20 microg/kg, did not stimulate leukocytosis, its administration at doses of 100 and 200 microg/kg increased the number of WBC up to 1.7- and 2.4-fold, respectively. Committed progenitor cells were mobilized into the peripheral blood dose-dependently and the number of CFCs was increased up to 5.5-fold by the administration of FL at 200 microg/kg on d 5. The number of CFCs in the bone marrow increased, but not dose-dependently. The number of CFCs in the spleen also increased up to 32-fold at a dose of 200 microg/kg FL. Mobilized peripheral blood mononuclear cells were transplanted into lethally irradiated mice and the number of CFU-S (d 12) was scored. A dose-dependent mobilization of CFU-S (d 12) into peripheral blood was also observed. These observations suggest that FL can mobilize hematopoietic primitive and committed progenitor cells into the peripheral blood of mice and those cells mobilized by FL may be applicable to peripheral blood stem cell transplantation.

We investigated the effect of anti-IL-6 receptor monoclonal antibody (hPM1) on the in vitro proliferation of cloned and freshly isolated myeloma cells from 20 patients with advanced stage multiple myeloma (MM). Humanized PM1 significantly inhibited the growth of a myeloma cell line in a dose-dependent manner and inhibited more than 30% of the proliferation of fresh myeloma cells in 10 of the 19 cases. Flow cytometric analysis using annexin V and 7AAD showed that hPM1 induced apoptosis of myeloma cells. These observations suggest the possibility of using hPM1 for treating some patients with MM whose growth depends on IL-6.

A 17-year-old male with chronic myelogenous leukemia in blast crisis received a non-T cell-depleted (TCD) HLA haplo-identical three-loci mismatched hematopoietic stem cell transplant (HSCT) from his mother. Long-term feto-maternal microchimerism was detected by nested polymerase chain reaction with sequence-specific primer typing. The post-transplantation prophylaxis against graft-versus-host disease (GVHD) was tacrolimus with minidose methotrexate. Sustained engraftment was obtained. Acute GVHD (grade 2) developed, but improved rapidly. Bone marrow aspiration on day 120 showed complete remission. Non-TCD HLA haplo-identical HSCT based on feto-maternal microchimerism might be feasible and has important implications in the selection of alternative family donors in HSCT.

Background: The objectives of this study were to evaluate the effect of the number of infused CD34+ cells on hematopoietic recovery and on the cost in autologous peripheral blood stem cell transplantation (PBSCT). Methods: Sixty-nine patients who received autologous PBSCT (ABSCT) were divided into three groups defined by the number of infused CD34+ cells. The number of days until 0.5 x 10 9 /l neutrophils and 50 x 10 9 /l platelets, the number of transfused blood products, the febrile days, the duration of parenteral antibiotics and the cost of additional supportive care (transfusions of blood products and parenteral antibiotics) were analyzed. Results: Twenty-three patients received

Summary. This study aimed to identify which graft product subset of cells might be the most predictive of late haematopoietic recovery (three to 12 months) following autologous peripheral blood stem cell transplantation (PBSCT). The relationships between the numbers of reinfused CD34+ cells and their immature subsets such as CD34+/CD90+, CD34+/AC133+, CD34+/CD38– and CD34+/HLA-DR– cells, and haemoglobin, white blood cell (WBC) and platelet counts at 3, 6, 9 and 12 months after PBSCT, were studied in 25 patients with haematological and solid malignancies. The total CD34+ cell number, as well as CD34+/CD90+ and CD34+/AC133+ cell numbers, correlated with platelet counts at 3, 6, 9 and 12 months after PBSCT, but the CD34+/CD90+ cells infused best predicted platelet recovery during the first 12 months after PBSCT (P

Four patients (three males and one female) were diagnosed as myeloid/natural killer (NK) cell precursor acute leukemia in our department. Two patients showed the extramedullary involvement at initial presentation. Leukemic cells expressed CD7, CD33, CD45 and CD56 in all patients. Additionally, CD13, CD34, HLA-DR, cytoplasmic CD3 and myeloperoxidase were expressed in some patients. Trisomy 10 was found in two patients, which has not been reported in this disease. Therefore, myeloid/NK cell precursor acute leukemia might be rather heterogeneous especially in chromosomal abnormality though it seemed to constitute the distinct clinical entity among acute myeloid leukemia of M0 subtype.

We immunophenotyped 128 patients with B-cell non-Hodgkin's lymphoma (B-NHL) of various histological subtypes using two-color flow cytometry (FCM), and found that lymphoma cells obtained from 31 patients (24.2%) coexpressed at least one of the following T-cell associated antigens (T-Ag); CD2 (2.3%), CD5 (18.0%) or CD7 (6.3%). Moreover, 3 patients expressed two kinds of T-Ag (CD2/CD5, CD2/CD7 or CD5/CD7) as reported by other investigators. Though we could not find coexpression of CD3, CD4 or CD8 antigen in any patients analyzed in our study, such T-Ag(+) B-NHL have also been reported in the literature. As clinical features, extranodular involvement and higher International Prognostic Index (high and high intermediate) seemed more frequent in T-Ag(+) B-NHL than T-Ag(-) B-NHL in our study. Such prognostic significance of T-Ag expression is also reported by other investigators especially in CD5(+) diffuse large B-cell lymphoma. In addition, two-color FCM for detecting such aberrant T-Ag expression in B-NHL is useful for monitoring the minimal residual disease in the subgroup with T-Ag(+) B-NHL.

We performed the immunophenotyping of 101 patients with B-cell non-Hodgkin's lymphoma (B-NHL) using two-colour flow cytometry (FCM) and found that lymphoma cells coexpressed at least one kind of T-cell-associated antigen (T-Ag; CD2, CD5, CD7) in 25 patients (24. 8%). Among these three T-Ags, CD5 was the most frequently expressed, in 21 patients (20.8%), followed by CD7, expressed in five patients (5.0%), and CD2, which was expressed in two patients (2.0%). Two kinds of T-Ag were simultaneusly expressed in three patients (CD2/CD5, CD2/CD7, and CD5/CD7, each expressed in one patient). Concerning the expression pattern of T-Ag, there were no significant differences between lymph nodes and extranodal organs in the three patients with T-Ag-positive B-NHL (T-Ag(+) B-NHL) who were analysed. When comparing the clinical features between T-Ag(+) B-NHL and T-Ag-negative B-NHL (T-Ag(-) B-NHL), extranodal involvement and higher International Prognostic Index (H and H.I.) were significantly frequent in the former subgroup (P = 0.0119 and P = 0. 0302 respectively).

Only a small number of reports have described the cytogenetic analysis of minimally differentiated acute myeloid leukemia (AML, M0). We performed a cytogenetic analysis on a patient with AML (M0) with a normal platelet count. It revealed a chromosomal translocation between chromosome bands 3q21 and 12q24. 3q. Abnormalities in AML are known to be associated with normal or elevated platelet counts. 3q21 and 12q24 are common translocation sites in AML patients, but this is the first report of translocation t(3;12)(q21;q24) in an AML patient.

We have previously shown that FLT-3 ligand (FL) mobilizes murine hematopoietic primitive and committed progenitor cells into blood dose-dependently. Whether FL also acts synergistically with granulocyte colony-stimulating factor (G-CSF ) to induce such mobilization has now been investigated. Five- to 6-week-old C57BL/6J mice were injected subcutaneously with recombinant human G-CSF (250 μg/kg), Chinese hamster ovarian cell-derived FL (20 μg/kg), or both cytokines daily for 5 days. The number of colony-forming cells (CFCs) in peripheral blood increased approximately 2-, 21-, or 480-fold after administration of FL, G-CSF, or the two cytokines together, respectively, for 5 days. The number of CFCs in bone marrow decreased after 3 days but was increased approximately twofold after 5 days of treatment with G-CSF. The number of CFCs in the bone marrow of mice treated with both FL and G-CSF showed a 3.4-fold increase after 3 days and subsequently decreased to below control values. The number of CFCs in spleen was increased 24.2- and 93.7-fold after 5 days of treatment with G-CSF alone or in combination with FL, respectively. The number of colony-forming unit-spleen (CFU-S) (day 12) in peripheral blood was increased 13.2-fold by G-CSF alone and 182-fold by G-CSF and FL used together after 5 days of treatment. Finally, the number of preCFU-S mobilized into peripheral blood was also increased by the administration of FL and G-CSF. These observations show that FL synergistically enhances the G-CSF–induced mobilization of hematopoietic stem cells and progenitor cells into blood in mice, and that this combination of growth factors may prove useful for obtaining such cells in humans for transplantation.

Recently, the ligand for c-mpl has been cloned and initial studies have shown it to be the platelet regulatory factor, thrombopoietin (TPO). To elucidate the role of TPO in the reconstitution of megakaryopoiesis and platelet production after stem cell transplantation, we measured serum TPO levels in nine patients undergoing autologous peripheral blood stem cell transplantation (PBSCT) and in healthy volunteers. Serum TPO levels significantly correlated with the degree of peripheral thrombocytopenia and a strong inverse correlation between serum TPO level and platelet count was observed (r = −0.700, P < 0.001). serum tpo levels began to rise as the platelet count decreased after chemotherapy. tpo levels peaked at over 25.00 fmoles/ml between days 0 and 10; tpo levels then decreased gradually as the platelet count began to rise. one patient with multiple myeloma received purified cd34+ peripheral blood stem cells. No difference was observed in the kinetics of serum TPO levels between unfractionated and purified PBSCT. These observations suggest that TPO plays a critical role in the reconstitution of megakaryopoiesis and platelet production after PBSCT.

症例は73歳,男性.小脳出血,多発性脳梗塞にて臥床状態にあり近医で在宅中心静脈栄養(IVH)管理を受けていた.徐々に貧血,白血球減少が進行するため当科入院.骨髓穿刺で血球貪食像認めたためステロイドパルス療法施行するも無効.銅欠乏状態が判明し微量元素製剤を補充したところ骨髓所見も含め速やかな改善を得た.長期間の人工栄養療法施行時には微量元素欠乏症の可能性に充分留意すべきであると考え報告した.

There have been some reports on the efficacy and tolerability of an oral itraconazole (ITCZ) solution as prophylaxis for fungal infection in patients with hematological malignancies. However, there are some cases where the bitter taste of oral ITCZ solution leads to an interruption of administration because the patient refuses to take this medicine. Therefore, we prospectively investigated the pharmacokinetics and promotion of treatment adherence in patients taking oral ITCZ solution mixed with a beverage. Compared with the responses of patients taking oral ITCZ solution with water, the taste of the agent was improved significantly when mixed with orange juice, although the plasma concentration of the agent did not differ between the two groups. Using this method, we can expect an improvement in treatment adherence and this method can easily be applied in clinical practice. This method is highly useful and should become common knowledge.

This study aimed to identify which graft product subset of cells might be the most predictive of late haematopoietic recovery (three to 12 months) following autologous peripheral blood stem cell transplantation (PBSCT). The relationships between the numbers of reinfused CD34+ cells and their immature subsets such as CD34+/CD90+, CD34+/AC133+, CD34+/CD38- and CD34+/HLA-DR- cells, and haemoglobin, white blood cell (WBC) and platelet counts at 3, 6, 9 and 12 months after PBSCT, were studied in 25 patients with haematological and solid malignancies. The total CD34+ cell number, as well as CD34+/CD90+ and CD34+/AC133+ cell numbers, correlated with platelet counts at 3, 6, 9 and 12 months after PBSCT, but the CD34+/CD90+ cells infused best predicted platelet recovery during the first 12 months after PBSCT (P0.0238 at any time-point). The CD34+/AC133+ cell dose also correlated with WBC counts at 3 months post PBSCT. In addition, all patients receiving more than 80 x 10(4) CD34+/CD90+ cells/kg showed platelet counts greater than 100 x 10(9)/l at all points after PBSCT, suggesting that this value of the CD34+/CD90+ cells infused was a threshold dose for durable haematopoietic engraftment after PBSCT.

Objective One of the mechanisms for mobilization of hematopoietic stem cells and progenitor cells is alternation of adhesion molecules. We investigated the mobilization of hematopoietic progenitor cells in blood by administration of anti-vascular cell adhesion molecule (VCAM)-1 antibody (Ab) in mice. Materials and Methods Twelve- to 14-week-old C57BL/6J mice were injected intravenously with anti-VCAM-1 Ab and anti-very late antigen (VLA)-4 Ab at a dose of 5 mg/kg for 2 days. Results The number of colony-forming cells (CFCs) in blood was increased 11.4-fold after anti-VCAM-1 Ab treatment, but the number of CFCs was not increased after treatment with anti-VLA-4 Ab. The number of colony-forming unit spleen (CFU-S) also was increased 21.6-fold in the peripheral blood by administration of anti-VCAM-1 Ab. The number of CFCs and CFU-S in the bone marrow of mice treated with anti-VCAM-1 Ab was decreased and that in the spleen also was decreased. On administration of recombinant human granulocyte colony-stimulating factor (125 μg/kg twice daily) with anti-VCAM-1 Ab, the numbers of CFCs and CFU-S were increased 141.8-fold and 439-fold, respectively. Conclusion These observations demonstrated that administration of anti-VCAM-1 Ab induced mobilization of hematopoietic progenitor cells into blood from bone marrow and spleen and that granulocyte colony-stimulating factor has synergistic effects on anti-VCAM-1 Ab-induced mobilization.

Wilms' tumor (WT1) gene expression is increased in patients with leukemia as well as myelodysplastic syndrome (MDS) and is useful for detection of minimal residual disease (MRD). A 47-year-old man given a diagnosis of refractory anemia with excess of blasts in transformation (RAEB-T) received myeloablative therapy followed by autologous peripheral blood stem cell transplantation (PBSCT). MRD by WT1 expression was not detected in the graft. The patient has been in CR for 25 months after PBSCT. These observations suggest that PBSCT is feasible for patients with RAEB-T and analysis of WT1 expression can be applied for patients with high risk MDS.

Reliable markers for megakaryocytic reconstitution after peripheral blood stem cell transplantation (PBSCT) have not been established. To determine a convenient and reliable predictor, we measured the number of megakaryocyte progenitor cells in PBSC grafts by clonogenic and flow cytometric assays. Seventeen patients with hematological and solid malignancies were included in this study. For the clonogenic assay, we used thrombopoietin (TPO) as a growth factor to evaluate the maximum number of megakaryocyte progenitor cells. Using a flow cytometric assay, we examined the expression of platelet glycoproteins on CD34+ cells to count the number of megakaryocyte progenitor cells. We used buffer containing EDTA to prevent platelet adhesion to CD34+ cells and selected CD34+ cells by immunomagnetic beads. The best correlation was observed between the number of CD34+/CD41a+ cells and the time to platelet recovery (P = 0.0205), rather than the total number of CD34+ cells. In addition, a close correlation was observed between the number of CD34+/CD41a+ cells and colony-forming unit megakaryocyte (CFU-MK) (P = 0.0018). These observations suggest that the number of CD34+/CD41a+ cells is the best predictor for platelet reconstitution after PBSCT.

The effect of granulocyte colony-stimulating factor (G-CSF) on accelerating the rate of neutrophil recovery after peripheral blood stem cell transplantation (PBSCT) remains controversial.We retrospectively analyzed 37 patients who received high-dose chemotherapy followed by PBSCT. Patients were divided into three groups: those who received no G-CSF (control), a low dose (50 mg/m2), or a high dose (150 mg/m2), subcutaneously. Seven patients who received increased numbers of colony forming units-granulocyte macrophage (CFU-GM) (>100x104/kg) and either low-or high-dose G-CSF were separately analyzed. Neutrophil recovery (0.5 x109/1) was accelerated in the G-CSF-treated patients. It occurred at a median of 10 and 9 days in the low-and high-dose G-CSF groups compared with 14 days in the control group (p< .0001). In patients receiving increased numbers of progenitor cells, the time until recovery was 9 days. No statistically significant differences were observed for platelet and reticulocyte recovery, transfusions, days febrile and parenteral antibiotic requirement. Thus, the administration of low-dose G-CSF is recommended in PBSCT.

A 48-year-old Japanese man was admitted to our hospital because of general fatigue, nasal bleeding, and petechiae on his extremities. He was diagnosed with acute myelomonocytic leukemia with trilineage myelodysplasia (T-MDS). Chromosomal analysis of bone marrow cells revealed t(7;11)(p15;p15), which has been rarely reported but known to be characteristic of Japanese patients. Although t(7;11)(p15;p15) has been reported mainly in acute myelogenous leukemia (AML), it can be occasionally found in so-called stem cell diseases such as chronic myelogenous leukemia or chronic myeloproliferative disorders. Therefore, t(7;11)(p15;p15) might affect trilineage progenitors or stem cells as well as myeloid lineage cells, subsequently resulting in AML with T-MDS, as in our case reported here.

We analyzed the expression of Thy-1 (CD90) antigen on CD34+ bone marrow mononuclear cells (BMMNC) obtained from 25 patients with myelodysplastic syndrome (MDS) by two-color flow cytometry. Five of nine patients (55.6%) with refractory anemia with excess of blasts (RAEB) and two of 16 (12.5%) with RAEB in transformation (RAEB-t) showed more than 20% of Thy-1 expression of their CD34+ BMMNC. Regarding chromosomal abnormalities, -5/5q- or -18 might be correlated with the expression of Thy-1 on CD34+ BMMNC in MDS. Nine patients were analyzed twice, once before and once after leukemic transformation and showed no significant change in Thy-1 expression. These results show that Thy-1 was expressed on CD34+ BMMNC in certain patients with MDS before leukemic transformation and that it was maintained during the disease progression. In contrast, expression of Thy-1 did not seem essential to the leukemic transformation in MDS though the patients with high Thy-1 expression might have poorer prognosis compared with those with low Thy-1 expression.

We analyzed the expression of the Thy-1 antigen (CD90) in CD34+ acute leukemia using two-color flow cytometry. Leukemic cells were obtained from the bone marrow (BM) and/or the peripheral blood (PB) of 57 patients: 37 with acute myelogenous leukemia (AML) including nine with secondary AML following myelodysplastic syndrome (MDS/AML), and 20 with acute lymphoblastic leukemia (ALL) including three with chronic myelogenous leukemia in blast crisis (CML-BC) of the lymphoid type. Among these patients, one (3.6%) with de novo AML, two (22.2%) with MDS/AML, three (17.6%) with de novo ALL, and two (66.7%) with CML-BC coexpressed CD34 and Thy-1 (CD34+ Thy-1+) on more than 20% of the mononuclear cells within 'lymph' plus 'blast' window. Thy-1 was rarely expressed in de novo acute leukemia especially in AML. Interestingly, in 1 patient with CML-BC, the leukemic cells in BM were divided into two subpopulations (CD34+ Thy-1low and CD34+ Thy-1high), whereas most of the CD34+ leukemic cells in PB were Thy-1high. However, the mechanism for the mobilization of CD34+ Thy-1high leukemic cells into the PB is unknown.