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The Ten Eleven Translocation 1 (TET1) gene encodes an epigenetic modifying molecule that is involved in demethylation of 5-methylcytosine. In hematological malignancies, loss-of-function mutations of TET2, which is one of the TET family genes including TET1, are frequently found, while the mutations of TET1 are not. However, clinical studies have revealed that TET1 is highly expressed in some cases of the hematological malignancies including acute myeloid leukemia. Indeed, studies by mouse models using conventional Tet1 knockout mice demonstrated that Tet1 is involved in myeloid leukemogenesis by Mixed Lineage Leukemia (MLL) fusion gene or TET2 mutant. Meanwhile, the other study showed that Tet1 is highly expressed in hematopoietic stem cells (HSCs), and that deletion of Tet1 in HSCs enhances potential self-renewal capacity, which is potentially associated with myeloid leukemogenesis. To examine the role of Tet1 in myeloid leukemogenesis more precisely, we generated novel conditional Tet1-knockout mice, which were used to generate the compound mutant mice by crossing with the inducible MLL-ENL transgenic mice that we developed previously. The leukemic immortalization in vitro was not critically affected by conditional ablation of Tet1 in HSCs with the induced expression of MLL-ENL or in hematopoietic progenitor cells retrovirally transduced with MLL-ENL. In addition, the leukemic phenotypes caused by the induced expression of MLL-ENL in vivo was not also critically affected in the compound mutant mouse model by conditional ablation of Tet1, although we found that the expression of Evi1, which is one of critical target genes of MLL fusion gene, in tumor cells was remarkably low under Tet1-ablated condition. These results revealed that Tet1 was dispensable for the myeloid leukemogenesis by MLL-ENL, suggesting that the therapeutic application of Tet1 inhibition may need careful assessment., Competing Interests: Sanikai is Mie University, Faculty of Medicine, School of Medicine, Alumni Association, a non-profit organization. There are no other relevant declarations relating to employment, consultancy, patents, products in development, or marketed products, etc. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

The aim of this study was to investigate whether a dual endothelin receptor antagonist bosentan modulates the kinetics of bone marrow-derived stem cells in inhibiting the development of pulmonary hypertension. Bone marrow chimeric mice, transplanted with enhanced green fluorescent protein (eGFP)-positive bone marrow mononuclear cells, were exposed to hypobaric hypoxia or kept in the ambient air, and were daily treated with bosentan sodium salt or saline for 21 days. After the treatment period, right ventricular pressure was measured and pulmonary vascular morphometry was conducted. Incorporation of bone marrow-derived cells was analyzed by immunohistochemistry. Gene expression and protein level in the lung tissue were evaluated by quantitative real-time PCR and western blotting, respectively. The results showed that, in hypoxic mice, right ventricular pressure and the percentage of muscularized vessel were increased and pulmonary vascular density was decreased, each of which was reversed by bosentan. Bone marrow-derived endothelial cells and macrophages in lungs were increased by hypoxia. Bosentan promoted bone marrow-derived endothelial cell incorporation but inhibited macrophage infiltration into lungs. Quantitative real-time PCR analysis revealed that interleukin 6, stromal cell-derived factor-1, and monocyte chemoattractant protein-1 were upregulated by hypoxia, in which interleukin 6 and monocyte chemoattractant protein-1 were downregulated and stromal cell-derived factor-1 was upregulated by bosentan. Protein level of endothelial nitric oxide synthase (eNOS) in the whole lung was significantly upregulated by hypoxia, which was further upregulated by bosentan. Bosentan modulated kinetics of bone marrow-derived ECs and macrophages and related gene expression in lungs in ameliorating pulmonary hypertension in mice. Altered kinetics of bone marrow-derived stem cells may be a novel mechanism of the endothelin receptor blockade in vivo and confer a new understanding of the therapeutic basis for pulmonary hypertension., (© The Author(s) 2020.)

A 53-year-old male presented with pancytopenia for 13 months. He had a past history of follicular lymphoma and hypopharyngeal cancer, which was treated via chemotherapy and radiotherapy. Bone marrow aspiration biopsy of the patient revealed a hypocellular marrow with 32% of hypergranular blasts without Auer bodies. There were also erythroid and megakaryocytic dysplasia in the bone marrow. Although the PML/RARA transcript was detected by fluorescence in situ hybridization (FISH) and reverse transcription polymerase chain reaction (RT-PCR), the G-banding karyotype analysis showed a complex karyotype without t (15;17). The PML/RARA fusion signal was identified on chromosome 15 by metaphase FISH. The patient was diagnosed of therapy-related acute promyelocytic leukemia (t-APL) with cryptic PML/RARA. He successfully attained molecular complete remission with all-trans retinoic acid (ATRA) and two courses of arsenic trioxide (ATO). He was subsequently administered nivolumab without ATRA maintenance therapy because of a progressing metastasis of a hypopharyngeal cancer to the lung. The patient had a relapse of t-APL following nine courses of nivolumab, 8 months after ending consolidation therapy with ATO. Reinduction therapy with ATRA was not effective for the relapsed t-APL that was accompanied by del (5q) and monosomy 7. Little has been previously reported on t-APL with cryptic PML/RARA. Therefore, the clinical course of this patient may provide useful insights about the characteristics of t-APL with cryptic PML/RARA.

A 56-year-old man diagnosed with multiple myeloma was treated with CBD (cyclophosphamide, bortezomib, and dexamethasone; DEX), which was discontinued because of bortezomib-associated adverse events. Thereafter, he was treated with Ld (lenalidomide; LEN+DEX) followed by high-dose chemotherapy with autologous stem cell rescue, resulting in a complete response. Ld as maintenance therapy was discontinued because of immune thrombocytopenia, resulting in disease progression. Although treatment was switched to Pd (pomalidomide+DEX), DLd (daratumumab+LEN+DEX), and IRd (ixazomib+LEN+DEX); the patient's M protein level continued to increase and the extramedullary disease expanded despite radiotherapy. He was treated with E-Ld (elotuzumab+LEN+DEX) after 3 cycles of short VAD (vincristine, doxorubicin, and DEX). The extramedullary disease disappeared after 8 cycles of E-Ld. To the best of our knowledge, this is the first report showing the effectiveness of E-Ld treatment for extramedullary disease of a heavily treated patient for multiple myeloma. We believe that the clinical course of this patient provides useful insights about the antimyeloma mechanism of elotuzumab.

Intestinal fibrosis is a serious complication in inflammatory bowel disease (IBD). Despite the remarkable success of recent anti-inflammatory therapies for IBD, incidence of intestinal fibrosis and need for bowel resection have not significantly changed. To clarify the contribution of haematopoietic-derived cells in intestinal fibrosis, we prepared bone marrow (BM) chimeric mice (chimeras), which were reconstituted with BM cells derived from enhanced green fluorescent protein (EGFP)-transgenic mice or CC chemokine receptor 2 (CCR2)-deficient mice. After 2 months of transplantation, BM chimeras were treated with azoxymethane/dextran sodium sulphate. During chronic inflammation, CCR2 + BM-derived monocyte and fibrocyte infiltration into the colon and CC chemokine ligand 2 production increased, leading to colon fibrosis in EGFP BM chimeras. In CCR2-deficient BM chimeras, monocyte and fibrocyte numbers in the colonic lamina propria significantly decreased, and colon fibrosis was attenuated. In colon tissue, mRNA expression of tissue inhibitor of metalloproteinase (TIMP)-1 but not of collagen I, transforming growth factor-β1 or matrix metalloproteinases was significantly different between the two chimeras. CCR2 + monocytes and fibrocytes showed high Timp1 mRNA expression. Our results suggest that infiltrating CCR2 + monocytes and their progenies, fibrocytes, promote colon fibrosis by inhibiting collagen degradation through TIMP-1 production.

Background: Hodgkin lymphoma usually presents with sequential enlargement of peripheral lymph nodes, and bone marrow invasion rarely occurs (approximately 3-5%). However, several cases have been reported as "primary" bone marrow Hodgkin lymphoma, especially among patients with human immunodeficiency virus and the elderly. This type of Hodgkin lymphoma is characterized by no peripheral lymphadenopathies and has been reported to have poorer prognosis., Case Presentation: A 38-year-old Japanese man was admitted to our hospital because of fever of unknown origin and pancytopenia without lymphadenopathies. Bone marrow examination revealed Hodgkin cells mimicking abnormal cells. These were positive for CD30, EBER-1, CD15, PAX-5, and Bob-1 and negative for Oct-2, CD3, CD20, surface immunoglobulin, CD56. On the basis of systemic evaluation and bone marrow examination, he was diagnosed with primary bone marrow Hodgkin lymphoma. We initiated therapy with DeVIC (dexamethasone, etoposide, ifosfamide, and carboplatin) therapy, but remission was not achieved. Then, the patient was treated with brentuximab vedotin combined with systemic chemotherapy (Adriamycin, vinblastine and dacarbazine), which was effective., Conclusions: There is no established treatment strategy for Hodgkin lymphoma, and therapeutic outcomes using ABVD (Adriamycin, bleomycin, vinblastine and dacarbazine)-like or CHOP (cyclophosphamide, Adriamycin, vincristine, and prednisone)-like regimens are reportedly poor. Only a few patients have been reported to achieve long-term remission. Through this case report, we suggest an alternative therapeutic option for primary bone marrow Hodgkin lymphoma.

A 56-year-old female who was diagnosed with acute calculous cholecystitis received intravenous administration of cefmetazole (CMZ) from the day of admission; she underwent laparoscopic cholecystectomy on the 13th hospital day. She was referred to our department because of hematuria that persisted for 3 days and progressive anemia on the day after the surgery. Laboratory data showed the following results: hemoglobin (Hb) level, 6.8 g/dl; reticulocyte count, 3.4%; serum lactate dehydrogenase, 1,505 IU/l; serum creatinine, 1.1 mg/dl; and undetectable haptoglobin. The direct globulin test showed that the patient was positive for IgG. Thus, drug-induced immune hemolytic anemia (DIIHA) was considered. All drugs, including CMZ, were immediately discontinued, and steroid was administered. The signs of hemolysis began to subside 3 days after the initiation of steroid therapy, and the administration of steroid was discontinued on the 5th day of the treatment. The patient's Hb level gradually increased, and the direct globulin test showed that the patient was negative for IgG on the 21st day from the onset of hematuria. Antibodies against CMZ-coated red blood cells were observed in the serum preserved at the onset of hemolysis. DIIHA is a rare but life-threatening disease. Immediate discontinuation of any suspected drugs and the initiation of steroid therapy as necessary are important in cases wherein DIIHA is suspected.

A 23-year-old man from Mie Prefecture, Japan, with past and family history of hematuria was diagnosed with influenza A and admitted to our hospital on the following day because of hemoglobinuria. He was diagnosed with thrombotic microangiopathy and was suspected of having atypical hemolytic uremic syndrome (aHUS). C3 p.I1157T missense mutation, which we had previously reported in eight aHUS patients from six families in Mie Prefecture, was identified. The laboratory findings and symptoms of our patient promptly improved after administering eculizumab. Little information is available on abnormalities of the complement system in aHUS or on mutation-specific outcomes of eculizumab therapy. Eculizumab was effective for treating our aHUS patient with C3 p.I1157T missense mutation.

Wilms' tumor 1 (WT1) is constantly expressed in leukemic cells of acute leukemia and myelodysplastic syndrome (MDS). A T-cell receptor (TCR) that specifically reacts with WT1 peptide in the context of HLA-A*24:02 has been identified. We conducted a first-in-human trial of TCR-gene transduced T-cell (TCR-T-cell) transfer in patients with refractory acute myeloblastic leukemia (AML) and high-risk MDS to investigate the safety and cell kinetics of the T cells. The WT1-specific TCR-gene was transduced to T cells using a retroviral vector encoding small interfering RNAs for endogenous TCR genes. The T cells were transferred twice with a 4-week interval in a dose-escalating design. After the second transfer, sequential WT1 peptide vaccines were given. Eight patients, divided into 2 dose cohorts, received cell transfer. No adverse events of normal tissue were seen. The TCR-T cells were detected in peripheral blood for 8 weeks at levels proportional to the dose administered, and in 5 patients, they persisted throughout the study period. The persisting cells maintained ex vivo peptide-specific immune reactivity. Two patients showed transient decreases in blast counts in bone marrow, which was associated with recovery of hematopoiesis. Four of 5 patients who had persistent T cells at the end of the study survived more than 12 months. These results suggest WT1-specific TCR-T cells manipulated by ex vivo culture of polyclonal peripheral lymphocytes survived in vivo and retained the capacity to mount an immune reaction to WT1. This trial was registered at www.umin.ac.jp as #UMIN000011519., (© 2017 by The American Society of Hematology.)

Crystal-storing histiocytosis (CSH) is characterized by the accumulation of large histiocytes with intracytoplasmic crystallized immunoglobulin and is typically associated with hematological malignancies. A 69-year-old man, who had a history of left nephrectomy and chemotherapy for renal pelvic cancer six years earlier, had received a CT scan every year thereafter and a small nodule was found in the left lower lobe of his lungs two years prior to the current presentation. Because of progression of this pulmonary nodule, he underwent pulmonary lobectomy on suspicion of lung cancer. He was ultimately diagnosed as having CSH accompanied by mucosa-associated lymphoid tissue lymphoma stage IAE. In the absence of further treatment, he has been well with no recurrence of the disease for 10 months postoperatively. Because CSH could reportedly be an initial presentation of hematological malignancies, careful observation and evaluation for the presence of these blood disorders is essential.

A 36-year-old woman complained of a mass on the sole of her foot in February 200X. She was diagnosed with extranodal NK/T-cell lymphoma, nasal type (ENKL) by skin biopsy. Because the lesion was localized on the subcutaneous tissue of the sole, she was treated with RT/2/3DeVIC, resulting in a complete response (CR). In March of the following year, PET/CT showed significant uptake and mucosal thickening in the right nasal cavity, and a mucosal biopsy confirmed ENKL infiltration. Because the lesion was localized in the nasal cavity, she was re-treated with RT/2/3DeVIC, with a focus on local control, and she achieved a second CR. She subsequently received allogeneic hematopoietic stem cell transplantation in the hope of preventing systemic relapse. She has remained in CR for four years since the transplantation. Our case suggests that allogeneic hematopoietic stem cell transplantation to be a potentially promising approach to curative treatment for recurrent ENKL in younger patients. As nasal lesions may subsequently appear during the course of primary non-nasal ENKL, ongoing meticulous evaluation for nasal lesions is important.

Bing-Neel syndrome is known as Waldenström's macroglobulinemia with central nervous system infiltration by neoplastic lymphoplasmacytoid and plasma cells. A 74-year-old man was admitted because of progressive cognitive impairment. Serum immunoelectrophoresis showed a monoclonal IgM-kappa component. Bone marrow aspiration revealed 59% small lymphocytes showing plasmacytoid differentiation. Bone marrow flow cytometry disclosed a population of kappa light-chain positive lymphoid cells expressing CD19, CD20, CD38, and CD138. Magnetic resonance imaging of the brain demonstrated gadolinium-enhancement in the right temporo-parieto-occipital meninges with sulcal enhancement. Cerebrospinal fluid cytology showed a population of lymphoplasmacytoid cells, positive for CD19, CD20, CD25, and kappa light-chain. Based on these findings, Bing-Neel syndrome was diagnosed. Although combination chemotherapy consisting of intrathecal methotrexate and oral cyclophosphamide was started, his symptoms continued to worsen. Then, we initiated treatment with a regimen consisting of fludarabine/rituximab (FR). After 6 courses of this FR regimen, a complete remission was achieved. Our case suggests the FR regimen to potentially be an effective treatment option for Bing-Neel syndrome of the scattered type.

Aim: The spleen is not believed to contribute to hematopoiesis in healthy adults. However, several reports have demonstrated that the spleen in adults contains a large number of hematopoietic stem/progenitor cells (HSC). Although splenectomy increases platelet and leukocyte counts, the effects of splenectomy on circulating HSC have not been elucidated. In this study, we evaluated the association between the number of circulating HSC and splenectomy in patients with hepatitis C virus (HCV)-associated liver cirrhosis (LC)., Methods: In 48 patients with various stages of HCV-associated chronic liver disease and seven patients with LC who underwent splenectomy, and 10 healthy volunteers, we determined the numbers of circulating CD34 + cells and colony-forming unit culture by flow cytometry and methylcellulose culture, respectively. Plasma stromal cell-derived factor-1α (SDF-1α) concentrations were measured using an enzyme-linked immunosorbent assay., Results: The numbers of circulating CD34 + cells and colony-forming unit culture decreased but the plasma SDF-1α concentration increased with the progression of liver disease. There was an inverse correlation between the number of circulating HSC and the plasma SDF-1α concentration. The numbers of circulating HSC and platelets were determined before and after splenectomy in seven patients with LC. In these patients, the numbers of circulating HSC and platelets increased significantly after splenectomy and the enhancing effect persisted for a long time., Conclusion: Our data suggest that the spleen plays an important role in modulating HSC dynamics in patients with HCV-associated chronic liver disease. Our results also imply that splenectomy may improve liver function in patients with LC., (© 2014 The Japan Society of Hepatology.)

Objectives: Invasive fungal diseases (IFDs) are life-threatening events in patients with haematologic disorders, and the spectrum of the aetiological pathogens continues to expand. This study aimed to evaluate the clinical utility of a panfungal polymerase chain reaction (PCR) assay for the management of IFDs in such patients., Methods: We prospectively analysed 273 consecutive blood samples from 64 risk episodes in 51 patients with haematologic disorders at high risk for IFD who were treated at our hospital between April 2007 and October 2010., Results: PCR-positive results were obtained in 18 of 64 risk episodes (35.3%). IFD was documented in 14 episodes (21.9%, 9 probable IFDs and 5 possible IFDs) according to the revised criteria of the European Organization for Research and Treatment of Cancer/Mycoses Study Group. PCR was positive in all of these 14 episodes, and in 4 of the 50 episodes with no IFD category. Sensitivity, specificity, positive predictive value, and negative predictive value of our assay were 100%, 92%, 78% and 100% respectively. A considerable number of fungi (44.4%) that are less common than Aspergillus and Candida species were positive by PCR. Molecular diagnoses of Cunninghamella species, Aspergillus ustus, Fusarium species, Scedosporium apiospermum, Rhodotorula species and Rhizopus species were beneficial in selecting suitable treatments., Conclusions: Our panfungal PCR approach allows for the highly sensitive and specific detection and identification of a wide spectrum of fungal pathogens, which provides indispensable information for managing IFDs, especially refractory or breakthrough IFDs during antifungal therapy in high-risk patients with haematologic disorders., (© 2013 John Wiley & Sons A/S.)

The regulation of human early lymphopoiesis remains unclear. B- and T-lineage cells cannot develop simultaneously with conventional stromal cultures. Here we show that telomerized human bone marrow stromal cells supported simultaneous generation of CD19(+) CD34(lo/-) CD10(+) cyCD79a(+) CD20(+/-) VpreB(-) pro-B cells and CD7(+) CD34(+) CD45RA(+) CD56(-) cyCD3(-) early T/Natural Killer (NK) cell precursors from human haematopoietic progenitors, and the generation of both lymphoid precursors was promoted by flt3 ligand (flt3L). On the other hand, stem cell factor or thrombopoietin had little or no effect when used alone. However, both acted synergistically with flt3L to augment the generation of both lymphoid precursors. Characteristics of these lymphoid precursors were evaluated by gene expression profiles, rearrangements of IgH genes, or replating assays. Similar findings were observed with primary human bone marrow stromal cells. Notably, these two lymphoid-lineage precursors were generated without direct contact with stromal cells, indicating that early B and T/NK development can occur, at least in part, by stromal cell-derived humoral factors. In serum-free cultures, flt3L elicited similar effects and appeared particularly important for B cell development. The findings of this study identified the potential of human bone marrow stromal cells to support human early B and T lymphopoiesis and a principal role for flt3L during early lymphopoiesis., (© 2012 Blackwell Publishing Ltd.)

Background and Objective: Interstitial lung diseases (ILD) are characterized by progressive interstitial pulmonary fibrosis and a decline in lung function. Fibrocytes are bone marrow-derived mesenchymal progenitor cells that may play a role in the pathogenesis of pulmonary fibrosis. Circulating fibrocyte numbers have been correlated with the prognosis of patients with idiopathic pulmonary fibrosis. The aim of the present study was to evaluate the relationship between circulating fibrocytes, and parameters of disease activity and progression in several groups of patients with ILD., Methods: The study population comprised 41 patients with ILD and seven healthy control subjects. Circulating CD45(+) collagen-I(+) fibrocytes were evaluated by flow cytometry., Results: The number of circulating fibrocytes was significantly increased in all patients with ILD and particularly in patients with idiopathic interstitial pneumonitis and interstitial pneumonitis associated with collagen vascular disease as compared with healthy control subjects. The numbers of circulating fibrocytes were significantly correlated with pulmonary function test parameters and with serum levels of sialylated carbohydrate antigen, a marker of disease activity. Temporal changes in circulating fibrocyte numbers were evaluated in two patients, and the results suggested that these changes correlated with the activity of ILD., Conclusions: The results from this study provide further evidence for the role of circulating fibrocytes in fibrotic lung diseases., (© 2012 The Authors. Respirology © 2012 Asian Pacific Society of Respirology.)

The BCR-ABL1 induces chronic myelogenous leukemia (CML) and Ph+ acute lymphoblastic leukemia (ALL). Recent studies revealed high ratios of loss of the IKZF1 gene which encodes IKAROS in BCR-ABL1+ ALL and lymphoblastic crisis (LBC) of CML. However, little is known about the cooperativity between the aberrant IKAROS and BCR-ABL1 in primary human hematopoietic cells. We investigated the effects of expression of BCR-ABL1 and/or IK6, a natural dominant negative isoform of IKAROS, on proliferation and differentiation of human CD34+ cord blood cells with or without human bone marrow-derived stromal cells which support early B cell differentiation. Cell proliferation was remarkably enhanced by co-expression of BCR-ABL1 and IK6, with reduced expression of glycophorin A and increased expression of CD41, especially on stromal cells, compared with expression of BCR-ABL1 alone that resulted in expansion of erythroid progenitors. Interestingly, p190BCR-ABL1 showed higher dependency on stromal cells to stimulate cell growth with IK6, than p210BCR-ABL1. Furthermore, the cooperation was found to depend on direct cell adhesive interaction of hematopoietic progenitors with stromal cells. These findings suggest that IK6 and BCR-ABL1 synergistically contribute to leukemogenesis in human bone marrow stromal microenvironment, and may provide a clue to elucidate the mechanisms of leukemogenesis of Ph+ ALL and CML-LBC.

Objective: Although the anticancer activities of histone deacetylase (HDAC) inhibitors have been studied, a role for HDAC in normal hematopoiesis has not been clearly defined. Previous studies have shown that the potent HDAC inhibitor FK228 stimulates interleukin (IL)-3-mediated erythropoiesis. Here, we examined whether the widely used valproic acid (VPA) affects megakaryopoiesis as well as erythropoiesis., Materials and Methods: CD34(+) cells were incubated in serum-free or serum-containing cultures with cytokines, with or without VPA., Results: In the serum-free cultures containing IL-3+stem cell factor (SCF), VPA significantly increased generation of CD61(+)GPA(-) megakaryocytic and a CD61(+)GPA(+) mixture of megakaryocytic and erythroid precursors from CD34(+) hematopoietic precursors at a pharmacological concentration (100 microg/mL). The increase in generation of megakaryocytic and erythroid precursors by VPA was confirmed by replating cultured cells with thrombopoietin+SCF and erythropoietin+SCF, respectively. VPA was as potent as FK228. In cultures with granulocyte-macrophage colony-stimulating factor+SCF, where CD61(-)GPA(+) erythroid precursors were mostly developed, VPA mainly enhanced the generation of CD61(-)GPA(+) erythroid precursors. In serum-containing cultures, only low numbers of CD61(+) or GPA(+) cells were developed with IL-3+SCF. Nevertheless, a substantial number of these cells were generated with VPA. Furthermore, these stimulating effects of VPA were observed by incubating CD34(+) cells from patients with myelodysplastic syndrome. Quantitative reverse transcription polymerase chain reaction showed that VPA enhanced GATA-2, but not GATA-1, messenger RNA expression with IL-3+SCF., Conclusions: These results indicate a novel role for VPA in enhancing the potential of IL-3 to stimulate megakaryopoiesis as well as erythropoiesis and suggest a new therapeutic approach of epigenetic therapy for hematological disease., (2010 ISEH - Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.)

Study Objective: To investigate whether a drug interaction exists between bortezomib and the cytochrome P450 (CYP) 3A4 inhibitor itraconazole and/or the CYP2C19 inhibitor lansoprazole that results in increased severity of bortezomib-induced peripheral neuropathy and thrombocytopenia., Design: Retrospective medical record review., Setting: Hematology-oncology ward of a university-affiliated hospital in Japan., Patients: Six adults with relapsed multiple myeloma who received intravenous bortezomib plus oral dexamethasone as the first course of a 21-day cycle between July 2007 and December 2008. Four of the six patients were treated concomitantly with itraconazole or lansoprazole: two with itraconazole, one with lansoprazole, and one with both itraconazole and lansoprazole., Measurements and Main Results: Using the National Cancer Institute's Common Terminology Criteria for Adverse Events, we identified the presence and graded the severity of peripheral neuropathy and thrombocytopenia before and during each patient's first 21-day course of bortezomib plus dexamethasone therapy. All three patients who received itraconazole experienced new or worsening peripheral neuropathy; they also experienced grade 4 thrombocytopenia. The patient who received lansoprazole alone, as well as the two patients who did not receive itraconazole or lansoprazole, had no changes in either adverse effect. We also evaluated the relationship between peripheral neuropathy and bortezomib plus dexamethasone therapy by using the Naranjo adverse drug reaction probability scale, and a probable relationship was found. We further assessed whether a drug interaction between bortezomib and itraconazole and/or lansoprazole had occurred involving the CYP3A4 and/or the CYP2C19 pathways, respectively-resulting in increased severity of the bortezomib-induced peripheral neuropathy and thrombocytopenia-by using the Horn drug interaction probability scale. We found that the occurrence of this drug interaction was strongly supported., Conclusions: Itraconazole appears to exacerbate peripheral neuropathy and thrombocytopenia induced by bortezomib; however, the mechanism of this drug interaction is unknown. Clinicians should closely monitor for bortezomib-induced adverse effects when itraconazole, or any other potent CYP3A4 inhibitor, is administered concomitantly with bortezomib.

A 30-year-old Brazilian man hospitalized with AIDS developed a high-grade fever. Neither culture studies nor radiological examinations revealed the cause; small yet highly intense signals in the basal ganglia were detected upon gadolinium (Gd)-enhanced T1-weighted magnetic resonance imaging (MRI) of the head. This finding was equivocal at that time but obviously abnormal for his age. A week later, he developed a movement disorder in his right arm, speech apraxia, and a worsening disturbance of consciousness. Repeated Gd-enhanced T1-weighted MRI demonstrated incredible changes in the brain; enhanced lesions in the basal ganglia deteriorated over time, multiple nodular and ring-enhanced lesions were observed in almost the entire brain. A diagnosis of toxoplasma encephalitis (TE) was confirmed by the detection of Toxoplasma gondii DNA in the cerebrospinal fluid. After initiation of intravenous trimethoprim-sulfamethoxazole (TMP-SMX; 10 mg/kg/day of TMP and 50 mg/kg/day of SMX) treatment, his symptoms and radiological findings improved dramatically. Our case suggests that high-intensity signals seen in the basal ganglia of a Gd-enhanced T1-weighted MRI, even at the preclinical stage, is indicative of TE. Because the use of MRI in general has become more widespread, it is predicted that preclinical lesions of TE will be found in various clinical settings more frequently.

A 46-year-old Japanese man was admitted to our hospital because of prolonged fever. Laboratory examination demonstrated leukopenia, thrombocytopenia, marked liver dysfunction, and elevation of serum ferritin. A bone marrow examination showed several hemophagocytic macrophages, and a diagnosis of hemophagocytic syndrome was made. He was treated using HLH-94 protocol, and his clinical symptoms and laboratory data were rapidly improved. After 5 weeks, fever and liver dysfunction reappeared. A repeat bone marrow examination demonstrated that 28.4% of marrow nucleated cells were atypical lymphocytes, which were positive for CD2, CD7, CD16, CD56, and HLA-DR. Clonality of these proliferating NK cells was confirmed by an analysis of EB virus terminal repeat sequence and cytogenetic analysis, and final diagnosis of aggressive NK-cell leukemia was made. After induction chemotherapy consisting of dexamethasone, etoposide, ifosfamide, and L-asparaginase, the patient achieved partial remission. He received allogeneic peripheral blood stem cell transplantation from his one locus mismatched son, and is alive with no evidence of disease 20 months after transplantation.

Thrombotic microangiopathy (TMA) or thrombotic thrombocytopenic purpura (TTP) is a life-threatening syndrome characterized by increased number of fragmented red cells (FRCs) and thrombocytopenia. FRCs can be measured using the recently developed automated hematology analyzer XE-2100. The normal range for FRCs is 0% to 0.205%, as determined by the automated hematology analyzer XE-2100. The FRC count is significantly elevated in patients with TMA associated with liver transplantation, bone marrow transplantation, or TTP. In patients with TMA after liver transplantation, the FRC count is significantly higher than in those without TMA. In receiver operating characteristic analysis for the diagnosis of TMA, the area under the curve is 0.986, suggesting that FRC is a useful marker for the diagnosis of TMA. When the cutoff value of FRC for TMA is 1.2%, the sensitivity is 90% and the specificity is 96%, indicating that FRC is the most useful screening test for the diagnosis of TMA.

Autologous peripheral blood stem cell transplantation (auto-PBSCT) is considered the standard therapy for younger patients with multiple myeloma, however, survival curve doesn't reach plateau. On the other hand, allogeneic reduced-intensity stem cell transplantation (RIST) with graft-versus-myeloma (GVM) effects is expected to be the curable therapy. We had three cases received RIST (one case as a salvage therapy for relapse after tandem auto-PBSCT, and two cases as an intentional RIST after achieving partial response by auto-PBSCT). All of three cases relapsing or progressing after RIST showed disease type of plasmacytoma without plasma cells in the bone marrow. That may be because conditioning regimen doesn't include total body irradiation, or GVM is ineffective in plasmacytoma, but effective in bone marrow. Here, we report clinical course of these three cases with some consideration.

Toll-like receptor 3 (TLR3) is a pattern-recognizing receptor that is involved in immune signaling and plays a crucial role in survival by being able to recognize various viral components including double-stranded RNA (dsRNA). TLR3 expression and function in cancer cells are not well understood. We investigated the expression of TLR3 in hepatocellular carcinoma (HCC) cells and the function of TLR3 signaling by stimulation and transfection with polyinosinic-polycytidylic acid (Poly I:C), a synthetic form of dsRNA. TLR3 mRNA was expressed in HCC tissues as well as in non-tumor tissues. Positive immunohistochemical staining for TLR3 was observed in 52.7% of HCC tissues, and in HCC cells we found both membranous and cytoplasmic expression of TLR3. While cell surface stimulation of TLR3 with Poly I:C did not affect cell viability, it did activate NF-kappaB levels. In contrast, cytoplasmic stimulation with transfected Poly I:C significantly induced apoptosis accompanied by the down-regulation of anti-apoptotic protein. Transfected Poly I:C also synergistically augmented TRAIL-induced apoptosis, but only with low levels of transfected Poly I:C was IFN-beta production not observed. In conclusion, our results indicate that TLR3 expression in HCC plays an important role with regard to cell survival and proapoptotic activity. Endogenously expressed TLR3 may provide new clinical prospects for TLR3 agonists as cytotoxic agents in HCC.

Hepatic stellate cells are believed to play a key role in the development of liver fibrosis. Several studies have reported that bone marrow cells can give rise to hepatic stellate cells. We hypothesized that hepatic stellate cells are derived from hematopoietic stem cells. To test this hypothesis, we generated chimeric mice by transplantation of clonal populations of cells derived from single enhanced green fluorescent protein (EGFP)-marked Lin(-)Sca-1(+)c-kit(+)CD34(-) cells and examined the histology of liver tissues obtained from the chimeric mice with carbon tetrachloride (CCl(4))-induced injury. After 12 weeks of CCl(4) treatment, we detected EGFP(+) cells in the liver, and some cells contained intracytoplasmic lipid droplets. Immunofluorescence analysis demonstrated that 50% to 60% of the EGFP(+) cells were negative for CD45 and positive for vimentin, glial fibrillary acidic protein, ADAMTS13, and alpha-smooth muscle actin. Moreover, EGFP(+) cells isolated from the liver synthesized collagen I in culture. These phenotypes were consistent with those of hepatic stellate cells. The hematopoietic stem cell-derived hepatic stellate cells seen in male-to-male transplants revealed only one Y chromosome. Our findings suggest that hematopoietic stem cells contribute to the generation of hepatic stellate cells after liver injury and that the process does not involve cell fusion.

Interleukin (IL)-6 plays pleiotropic roles in human hematopoiesis and immune responses by acting on not only the IL-6 receptor-alpha subunit (IL-6Ralpha)(+) but also IL-6Ralpha(-) hematopoietic progenitors via soluble IL-6R. The Notch ligand Delta-1 has been identified as an important modulator of the differentiation and proliferation of human hematopoietic progenitors. Here, it was investigated whether these actions of IL-6 are influenced by Delta-1. When CD34(+)CD38(-) hematopoietic progenitors were cultured with stem cell factor, flt3 ligand, thrombopoietin and IL-3, Delta-1, in combination with the IL-6R/IL-6 fusion protein FP6, increased the generation of glycophorin A(+) erythroid cells but counteracted the effects of IL-6 and FP6 on the generation of CD14(+) monocytic and CD15(+) granulocytic cells. Although freshly isolated CD34(+)CD38(-) cells expressed no or only low levels of IL-6Ralpha, its expression was increased in myeloid progenitors after culture but remained negative in erythroid progenitors. It was found that Delta-1 acted in synergy with FP6 to enhance the generation of erythroid cells from the IL-6Ralpha(-) erythroid progenitors. In contrast, Delta-1 antagonized the effects of IL-6 and FP6 on the development of monocytic and granulocytic cells, as well as CD14(-)CD1a(+) dendritic cells, from the IL-6Ralpha(+) myeloid progenitors. These results indicate that Delta-1 interacts differentially with gp130 activation in IL-6Ralpha(-) erythroid and IL-6Ralpha(+) myeloid progenitors. The present data suggest a divergent interaction between Delta-1 and gp130 activation in human hematopoiesis.

The management of elderly patients with acute myeloid leukemia (AML) and a poor performance status is challenging. An 89-year-old man with AML secondary to myelodysplastic syndrome (MDS) and a poor performance status (4) underwent treatment with methylprednisolone (mPSL) (125 mg/body), which resulted in a remarkable reduction of blast cells in the peripheral blood. Neutrophil counts were maintained or increased. Although the suppression was of short duration, mPSL was useful for disease control because it selectively reduced blast counts while maintaining the patient's performance status. In vivo and in vitro findings suggested that mPSL had direct inhibitory actions on the survival of blast cells. On the basis of this experience, we gave the same mPSL dose to other elderly patients with MDS/AML (n=5) or AML-M4 (n=1) who had a poor performance status (3 or higher) and appeared unable to tolerate standard cytotoxic chemotherapies. Selective and significant blast cell reduction was observed in 4 of the 5 patients with MDS/AML, whereas no effects were seen in the AML patient. Although our experience is limited, these findings may provide a clue to understanding the mechanisms regulating the survival of blast cells of MDS/AML and indicate that mPSL may provide a benefit to a subset of these patients.

The appearance of monocytes before neutrophils in the blood during haematopoietic recovery in myelosuppressive patients is commonly observed, thus suggesting a difference in the cell division history between these two lineages in the differentiation from granulocyte-macrophage (GM) progenitors. We investigated the cell division histories of murine GM progenitors. When analysed by the dye dilution method, GM progenitors gave rise to Gr-1+Fms+ and Gr-1+Fms- cells that passed through similar rounds of cell division during initial 5 d of culture. The Gr-1+Fms+ cells showed morphological features of monocytes, while Gr-1+Fms- cells exhibited an immature morphology of neutrophils. In the subsequent culture, a decline in the number of Gr-1+Fms+ cells was observed, while Gr-1+Fms- cells increased. The proliferation of Gr-1+Fms- cells and no cell division of Gr-1+Fms+ cells were confirmed by DNA staining, Ki-67 expression, membrane dye staining and bromodeoxyuridine incorporation. These Gr-1+Fms- cells acquired mature neutrophil morphology, whereas Gr-1+Fms+ cells became macrophages. These results demonstrate that GM progenitors generate postmitotic monocytes earlier than mature neutrophils. Our data may also offer one explanation for the rapid recovery of monocytes in comparison with neutrophils in the early phase of haematopoietic regeneration.

We describe a patient with low-titer cold agglutinin disease (CAD) who developed mixed-type autoimmune hemolytic anemia (AIHA) and idiopathic thrombocytopenia following chicken pox infection. At least 1 year before admission to hospital, the patient had mild hemolytic anemia associated with low-titer cold agglutinins. A severe hemolytic crisis and thrombocytopenia (Evans' syndrome) occurred several days after infection with chicken pox, and the patient was referred to our hospital. Serological findings revealed the presence of both cold agglutinins and warm-reactive autoantibodies against erythrocytes, and the diagnosis was mixed-type AIHA. Following steroid therapy, the hemoglobin (Hb) level and platelet count improved. The patient was closely followed over a 10-year period with recurrent documented hemolysis after viral or bacterial infections. Warm-reactive autoantibodies have not been detected in the last 2 years, and only the immunoglobulin M anti-I cold agglutinins with a low titer and wide thermal amplitude have remained unchanged. Therefore, the patient has received at least 10 mg prednisolone daily to maintain a Hb level of 10 g/dL. To the best of our knowledge, no adult case of low-titer CAD that has evolved into mixed-type AIHA and Evans' syndrome after chicken pox infection has been previously reported in the literature.

Histone acetylation controls the expression of specific genes in eukaryotic cells. We investigated the role of histone deacetylases (HDACs) in the differentiation of human erythroid cells, using pharmacological approaches. When CD36+ erythroid precursor cells, generated from CD34+ cells with stem cell factor, flt-3 ligand, thrombopoietin, interleukin-3, interleukin-6, and erythropoietin, were cultured with an HDAC inhibitor FK228 (depsipeptide) at a specified dose in the presence of erythropoietin, their differentiation was inhibited, as determined by the expression of CD45 and glycophorin A. Addition of the same dose of FK228 to cultures did not affect the growth of CD36+ cells. Regardless of the presence or absence of FK228, cultured CD36+ cells displayed similar proliferation kinetics. Analysis of acetylated histones revealed that FK228 upregulated the acetylation status of histones H3 and H4 in CD36+ cells. The inhibition of CD36+ cell differentiation was restored by removal of FK228 from the culture, indicating that the modification of CD36+ cell differentiation by FK228 is reversible. Furthermore, interference with histone deacetylation by FK228 inhibited the generation of CD36+ erythroid cells from CD34+ hematopoietic progenitor cells. Our results indicate the possible involvement of HDACs in human erythropoiesis, especially the regulation of erythroid cell differentiation.

A 62-year-old Japanese woman was diagnosed as having follicular lymphoma (FL, grade 3, CS IIIA, IPI high-intermediate risk) in May 1998. After eight courses of CHOP therapy, she achieved a complete remission (CR). In November 1998, her FL relapsed, and she achieved a second CR after two courses of MINE therapy. High-dose etoposide was used for autologous peripheral stem cell mobilization. In May 1999, she underwent high-dose chemotherapy with autologous peripheral blood stem cell transplantation (auto-PBSCT). Four months after the auto-PBSCT, bilateral cervical lymphadenopathy developed. Histopathological findings from a biopsied cervical lymph node showed angioimmunoblastic T-cell lymphoma (AILT). The patient was treated with modified CVP therapy, and she is alive with no evidence of lymphoma five years after auto-PBSCT. Clinical and histopathological findings showed that the FL and AILT in this case were not concomitant. It is thought that in this case, the AILT developed as a post-transplant lymphoproliferative disorder after auto-PBSCT for the FL.

We previously demonstrated the ability of hematopoietic stem cells (HSCs) to generate glomerular mesangial cells by trans-planting clonal populations of cells derived from a single enhanced green fluorescent protein (EGFP)-positive HSC into lethally irradiated mice. To define more precisely the hematopoietic differentiation pathway through which mesangial cells are derived, we studied the relationship between mesangial cell expression and individual hematopoietic lineages by means of a transplantation strategy. In a series of clonal HSC transplantation experiments, we generated 3 mice engrafted predominantly by granulocytes and macrophages (GMs) and 4 mice engrafted with B-cells or with B-cells and T-cells. When the kidneys of these mice were analyzed, the mice exhibiting high GM lineage engraftment revealed much higher levels of EGFP-positive mesangial cells than those with predominantly lymphocyte engraftment. Fluorescence in situ hybridization analysis of the kidneys from a male recipient of an EGFP-positive female donor excluded cell fusion as the cause for the observed differentiation. These results support the notion that glomerular mesangial cells share their origin with GMs.

Targeted disruption of the Friend leukemia integration 1 (Fli-1) proto-oncogene results in severe dysmegakaryopoiesis and embryonic lethality. We used morula-stage aggregation as a strategy to further clarify the hematopoietic defects of the Fli-1 gene-targeted mice. Analyses of lineage expression of Fli-1(+/-) and Fli-1(-/-) cells in the peripheral blood and bone marrow of chimeric mice consistently demonstrated reduced numbers of neutrophilic granulocytes and monocytes and increased numbers of natural killer (NK) cells. Transplantation studies using sorted Fli-1 mutant cells produced similar findings. Clonal culture studies of bone marrow cells revealed increased numbers of granulocytic and early erythroid progenitors in the Fli-1(+/-) cells. The sorted Fli-1(-/-) bone marrow cells revealed specific down-regulation of CCAAT/enhancer binding protein-alpha (C/EBPalpha) and C/EBPepsilon, and the receptors for granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage CSF (GM-CSF), consistent with their critical roles in granulopoiesis. Collectively, these observations suggest previously unknown physiologic roles for Fli-1 in granulocytic, erythroid, and NK cell proliferation and differentiation. Production of chimeras by morula-stage embryo aggregation is an effective way to unravel cell-autonomous hematopoietic defects in gene-targeted mice.

Hemostatic parameters were examined in 39 patients who underwent allogeneic bone marrow transplantation (BMT). Twenty-six patients survived and 13 patients died within 6 months after BMT. The main causes of death were acute graft-versus-host disease (GVHD: n=6), veno-occlusive disease (VOD: n=2), and thrombotic microangiopathy (TMA: n=2). Plasma levels of D-dimer and thrombomodulin (TM) were significantly elevated in the non-survivor group. Plasma levels of soluble fibrin (SF) and Fas were significantly elevated in the non-survivor group at 1 to 4 weeks after BMT. Plasma levels of thrombin-antithrombin complex (TAT), D-dimer, and tissue plasminogen activator-plasminogen activator inhibitor-1 complex (tPA-PAI-1 complex) were significantly elevated in patients with complications after BMT. Plasma levels of TAT, D-dimer, and tPA-PAI-1 complex were significantly elevated in patients with GVHD. These results suggest that abnormalities of hemostatic parameters might predict poor outcomes or complications in patients with BMT.

It is generally recognized that postmitotic neutrophils give rise to polymorphonuclear neutrophils alone. We obtained evidence for a lineage switch of human postmitotic neutrophils into macrophages in culture. When the CD15+CD14- cell population, which predominantly consists of band neutrophils, was cultured with granulocyte macrophage-colony-stimulating factor, tumor necrosis factor-alpha, interferon-gamma, and interleukin-4, and subsequently with macrophage colony-stimulating factor alone, the resultant cells had morphologic, cytochemical, and phenotypic features of macrophages. In contrast to the starting population, they were negative for myeloperoxidase, specific esterase, and lactoferrin, and they up-regulated nonspecific esterase activity and the expression of macrophage colony-stimulating factor receptor, mannose receptor, and HLA-DR. CD15+CD14- cells proceeded to macrophages through the CD15-CD14- cell population. Microarray analysis of gene expression also disclosed the lineage conversion from neutrophils to macrophages. Macrophages derived from CD15+CD14- neutrophils had phagocytic function. Data obtained using 3 different techniques, including Ki-67 staining, bromodeoxyuridine incorporation, and cytoplasmic dye labeling, together with the yield of cells, indicated that the generation of macrophages from CD15+CD14- neutrophils did not result from a contamination of progenitors for macrophages. Our data show that in response to cytokines, postmitotic neutrophils can become macrophages. This may represent another differentiation pathway toward macrophages in human postnatal hematopoiesis.

Hemostatic abnormalities in 26 patients following bone marrow transplantation (BMT) were examined. In the event-free survival group, the plasma levels of antithrombin (AT) and protein C (PC) were significantly decreased 1 and 2 weeks after BMT, and the plasma levels of thrombomodulin (TM) and tissue plasminogen activator-plasminogen activator inhibitor-1 complex (tPA-PAI-I complex) were significantly increased from 4 weeks to 13 weeks after BMT. Excepting AT, there was no significant difference in hemostatic parameters before BMT among the event-free survival, 6-month survival, and death within 6 months groups. On day 0 following BMT, only plasma AT levels were significantly lower in the 6-month survival group than in the death within 6 months group. From 1 to 3 weeks after BMT, plasma levels of AT or PC were significantly lower in the death within 6 months group than in the 6-month survival group. From 1 to 5 weeks after BMT, the plasma levels of TM and tissue type plasminogen activator-plasminogen activator inhibitor-I complex (tPA-PAI-I complex) were significantly higher in the 6-month survival group than in the death within 6 months group. From 1 to 13 weeks after BMT, the plasma levels of D-dimer or soluble fibrin monomer (SFM) were significantly higher in the death within 6 months group than in the 6-month survival group. There was no remarkable difference in plasma levels of thrombin-antithrombin comlex or plasmin-plasmin inhibitor complex following BMT between these groups of patients. These findings suggest that the decrease in the plasma AT or PC level reflects early occurrence of complications of prognostic significance and that the increase in vascular endothelial cell markers such as plasma levels of TM or tPA-PAI-I complex reflects occurrence of complications during the middle course of BMT. Plasma levels of D-dimer and SFM may be useful markers for predicting complications associated with poor prognosis after BMT.

The Notch/Notch ligand system controls diverse cellular processes. The proteolytic cleavage generates transmembrane and soluble forms of Notch ligands. We examined the effect of a soluble Notch ligand, human Jagged-1, on human cord blood (CB) CD34+ cells, under serum-deprived conditions, using soluble human Jagged-1-immunoglobulin G1 chimera protein (hJagged-1). Soluble hJagged-1 inhibited myeloid colony formation but not erythroid-mix or erythroid colony formation, in the presence of stem cell factor (SCF), interleukin-3, granulocyte-macrophage colony-stimulating factor (GM-CSF), G-CSF, thrombopoietin, and erythropoietin. Cytological analysis revealed that the decrease in myeloid colonies resulted mainly from the inhibition of macrophage colony formation. Furthermore, soluble hJagged-1 led to the inhibition of macrophage colony formation supported by M-CSF plus SCF and GM-CSF plus SCF. Delayed-addition experiments and the analysis of colony sizes demonstrated that soluble hJagged-l inhibited the growth of macrophage progenitors by acting in the early stage of macrophage development. The direct action of hJagged-1 was confirmed by the enhanced expression of the HES-1 (hairy enhancer of the split-1) gene. These results suggest that soluble hJagged-1 may regulate human hematopoiesis in the monocyte/macrophage lineage.

We describe a 55-year-old Japanese woman with therapy-related myelodysplastic syndrome (t-MDS) with 2 independent clones, t(1;2)(p36;p21) and t(11;12)(pl5;ql3). She was diagnosed with acute myeloid leukemia (AML) with cytological features of the bone marrow and peripheral blood. Cytogenetic evaluation revealed a 46,XX karyotype. She received chemotherapy and achieved complete remission (CR). Despite maintenance chemotherapy, she suffered a relapse. Chromosomal analysis showed t(1;2)(p36;p21) in 2 of 20 metaphases. At second CR, this clone transiently disappeared. Nine months later, t(1;2) (p36;p21) was detected again in 3 of 20 metaphases while the patient remained in CR. Six months later, bone marrow examination disclosed trilineage dysplasia without an excess of blasts, suggesting MDS. t(1;2)(p36;p21) was observed in 16 of 20 metaphases. The clinical course and serial cytogenetic findings were diagnostic of t-MDS. The duration of t-MDS was 6 years. During this period, persistent t(1;2)(p36;p21) and transient t(11;12)(p15;q13) were found. When t-MDS evolved toAML, cytogenetic evaluation revealed 46,XX,t(1;2)(p36;p21),del(7)(q22),add(19)(p13).