No abstract available.
Neoplasm, Residual*

BACKGROUND: Peripheral blood progenitor cells collected from normal donors after granulocyte- colony stimulating factor (G-CSF) treatment are increasingly used for allogeneic transplantation. Previously, we investigated the mobilization kinetics of CD34+/Thy-1dim progenitor cells during subcutaneous administration of G-CSF in normal donors (Transfusion 1997;37:406-10). Although it is considered to be a relatively safe procedure, there are still uncertainties about the most efficient method of progenitor cell mobilization. Due to the short elimination half-life of G-CSF of about 3-4 hours we considered the subcutaneous administration of G-CSF once or twice daily might be suboptimal. The aim of the present study is to evaluate the mobilization kinetics of CD34+ cells during continuous intravenous (IV) administration of G-CSF in normal donors. METHODS: Fifteen healthy donors were enrolled in this study. The median age was 38 years (range, 20-56). G-CSF (Filgrastim, 10 microgram/kg/day) was administered for 4 consecutive days through continuous IV infusion. Then, we collected PBPC on the day following the 4th dose of G-CSF using a blood cell separator. For measurement of complete blood counts and CD34+ cell levels, peripheral blood sampling was performed immediately before the administration of G-CSF (steady-state) and after 4, 8, 24, 48, 72, 96, 120 hours of continuous IV administration of G-CSF. RESLUTS: After continuous IV administration of G-CSF, the WBC counts increased up to day 5 and reached approximately 8.4-fold above the steady-state level. Changes in the granulocyte count were similar to those in WBC counts. The number of lymphocytes increased up to day 4 (2.7-fold above the steady-state level), but no further increase occurred on day 5. Although there were considerable variations among the healthy donors, the statistical peaks of CD34+ cell levels were consistently observed on day 3 or day 4. Up to the fourth day of G-CSF treatment, the circulating CD34+ cells expanded by 25-fold. The percentage and absolute number of CD34+ cells significantly increased on day 3 (0.55 +/- 0.09%, 51.12 +/- 24.83x103/mL) and day 4 (0.47 +/- 0.09%, 46.66 +/- 24.93x103/mL), compared with steady-state values (0.06 +/- 0.09%, 2.03 +/- 5.69x103/mL). CONCLUSION: Our results showed that continuous IV administration of G-CSF apparently results in more rapid mobilization of CD34+ cells at least 24-48 hours compared with daily subcutaneous administration of G-CSF in normal donors.
Administration, Intravenous* ; Blood Cell Count ; Blood Cells ; Colony-Stimulating Factors ; Granulocyte Colony-Stimulating Factor* ; Granulocytes ; Half-Life ; Humans ; Kinetics* ; Lymphocytes ; Stem Cells ; Tissue Donors* ; Transplantation, Homologous

BACKGROUND: The stromal cells, which one part of the microenvironment, including adipocyte, fibroblasts, endothelial cells, macrophage and smooth muscle cells provide a suitable environment for self-renewal, proliferation and differentiation of hematopoietic stem cells, as well as providing other regulators, cytokines to stimulate and enhance hematopoiesis. Unlike bone marrow, the in vitro development of this stroma cells from the umbilical cord blood (UCB) is not well established and has met very limited success. To see if stromal elements capable of supporting hematopoiesis could be grown from CD34+ enriched umbilical cord blood, we was done stroma-free liquid culture system with thrombopoietin (TPO), flt3-ligand (FL) and the granulocyte- colony stimulating factor (G-CSF) using cord blood CB CD34+ cells. METHODS: Purified umbilical cord blood CD34+ cells were cultured for five weeks in a stroma-free liquid culture system using TPO, FL, and G-CSF in the density of 1x105 cells/mL. When a firmly adherent monolayer of fibroblast like cells were fully formed, we evaluate morphology, immunopheno-type, immunohistochemistry and functional assay, including non-adherent expansion, clonogenic progenitor expansion and apoptosis of established adherent cell layers. RESULTS: By the 4th~5th week, we found that firmly adherent fibroblast-like cells were established. These cells showed characteristics of endothelial cells expressing with VWF, VCAM-1, ICAM-1, CD31 and E-selectin. Furthermore, comparing without established endothelial monolayer, with established endothelial monolayer better expansion of total nucleated cells, CD34+ cells and colony forming units granulocyte-macrophage (CFU-GM) during culture were found. CONCLUSION: These results suggest that the UCB CD34+ cell fraction contains stromal cell precursors and can establish the hematopoietic microenvironment to expand and sustain UCB CD34+ cells through down-regulating apoptosis.
Adipocytes ; Apoptosis ; Bone Marrow ; Colony-Stimulating Factors ; Cytokines ; E-Selectin ; Endothelial Cells ; Fetal Blood* ; Fibroblasts ; Granulocyte Colony-Stimulating Factor* ; Hematopoiesis ; Hematopoietic Stem Cells ; Humans* ; Immunohistochemistry ; Intercellular Adhesion Molecule-1 ; Macrophages ; Myocytes, Smooth Muscle ; Stem Cells ; Stromal Cells ; Thrombopoietin ; Vascular Cell Adhesion Molecule-1

BACKGROUND: The aim of the present study is to evaluate the modulation of CD44 and CD31 expression and apoptotic status on mobilized CD34+ cells during continuous intravenous (i.v.) administration of granulocyte-colony stimulating factor (G-CSF), elucidating the mechanism of peripheral blood progenitor cells (PBPC) mobilization in normal donors. METHODS: Fifteen healthy donors were enrolled in this study. G-CSF (10 microgram/kg/day) was administered for 4 consecutive days through continuous i.v. infusion. Measurement of CD34+ cell levels and their expression of CD44, CD31 and 7-aminoactinomycin D (7-AAD) was performed. PB sampling was drawn immediately before the administration of G-CSF (steady-state) and after 4, 8, 24, 48, 72, 96 and 120 hours of G- CSF administration. RESULTS: Although there were considerable variations among the healthy donors, the absolute number of CD34+ cells significantly increased at day 3 (51.12+/-24.83x103/mL) and day 4 (46.66+/-24.93x103/mL),compared to the steady-state level (2.03+/-5.69x103/mL). The expression of CD44 on CD34+ cells revealed a significant reduction from the steady-state level (579.59+/-133.69) after day 3 (281.02+/-105.15, P=0.0059) and day 4 (164.76+/-107.44, P=0.0002) of G-CSF administration. The expression of CD31 on CD34+ cells also significantly decreased from the steady-state level (145.86+/-30.52) after day 4 (34.47+/-38.87, P=0.0055) and day 5 (17.33+/-50.68, P=0.0134) of G-CSF administration. The proportions of apoptotic (7-AADdIm) CD34+ cells were also significantly decreased after day 3 of G-CSF administration. CONCLUSION: Down-regulation of CD44 and CD31 on CD34+ cells is likely to be involved in the mobilization of PBPC and G- CSF may acts as a survival factor for mobilized CD34+ cells by the suppression of apoptosis during continuous i.v. administration of G-CSF in normal donors.
Administration, Intravenous* ; Apoptosis ; Down-Regulation ; Granulocyte Colony-Stimulating Factor ; Humans ; Stem Cells ; Tissue Donors*

No abstract available.
Stem Cells*

Emergence of new clonal chromosomal abnormality (CCA) has been reported in Philadelphia-negative cells in patients with chronic myeloid leukemia (CML) undergoing the tyrosine kinase inhibitor (TKI) treatment. However, the time of emergence and clinical significance of CCA remains to be elucidated. In this study, we report a CML patient undergoing TKI treatment who developed myelodysplastic syndrome (MDS) after 206 months since the diagnosis of CML. Results of droplet digital PCR performed with serial bone marrow samples revealed that monosomy 7 in Philadelphia-negative cells appeared at the time of MDS development that did not exist initially at the time of CML diagnosis.

Emergence of new clonal chromosomal abnormality (CCA) has been reported in Philadelphia-negative cells in patients with chronic myeloid leukemia (CML) undergoing the tyrosine kinase inhibitor (TKI) treatment. However, the time of emergence and clinical significance of CCA remains to be elucidated. In this study, we report a CML patient undergoing TKI treatment who developed myelodysplastic syndrome (MDS) after 206 months since the diagnosis of CML. Results of droplet digital PCR performed with serial bone marrow samples revealed that monosomy 7 in Philadelphia-negative cells appeared at the time of MDS development that did not exist initially at the time of CML diagnosis.

Translocations leading to fusions between the immunoglobulin heavy chain gene (IGH) and various partner genes have been reported in B-cell precursor acute lymphoblastic leukemia (B-ALL). However, submicroscopic deletions within IGH in B-ALL have not been rigorously assessed. In this study, we investigated characteristics of IGH submicroscopic deletions, by FISH, in B-ALL with IGH rearrangements. FISH was performed by using commercially available IGH dual-color break-apart rearrangement probes (Abbott/Vysis, Downers Grove, IL, USA; Kreatech, Amsterdam, Netherlands). The study group included seven B-ALL patients with IGH rearrangements, observed by FISH. Among them, two exhibited deletion of the 5' variable region of IGH by FISH. The B-ALL in these two patients included two kinds of abnormal cells; one had an IGH rearrangement without any IGH submicroscopic deletion, while the other had an IGH submicroscopic deletion, which showed that one normal fusion signal and one 3' IGH signal were detected. Thus, submicroscopic deletion of the IGH 5' variable region may have occurred in either the native or rearranged chromosome 14. These findings indicate that B-ALL with IGH rearrangements may be accompanied by submicroscopic deletions of the IGH 5' variable region, which can be detected by FISH. The clinical significance of such deletions is unclear, but the loss of part of the IGH gene in B-ALL warrants further study.
Adult ; Child ; Female ; *Gene Deletion ; *Gene Rearrangement ; Humans ; Immunoglobulin Heavy Chains/*genetics ; In Situ Hybridization, Fluorescence ; Infant ; Male ; Middle Aged ; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/*genetics/pathology ; Young Adult

A genomic instability called chromothripsis occurs as a single catastrophic event, generating massive complex genomic rearrangement with a possible characteristic pattern of copy number oscillations. Here, we report a case of secondary plasma cell leukaemia (PCL) showing chromothripsis identified by single nucleotide polymorphism array (SNP-A)-based karyotyping. A 53-year-old male patient was diagnosed as having secondary PCL four years after he was diagnosed with multiple myeloma, and he died four days later due to intracerebral haemorrhage. Chromosomal analysis and fluorescence in situ hybridization (FISH) revealed the deletions of 13q and 17p and an insertion of 1q. Further, genomic aberrations that were not detected by chromosomal analysis and FISH were identified by SNP-A. In particular, SNP-A revealed numerous alternating copy number state switches involving one, two, or three copy number states on chromosome 7q, suggesting the presence of chromothripsis. The present case suggests that chromothripsis may occur in secondary PCL and can be inferred from genomic copy number profiles identified by SNP-A.
Fluorescence ; Genomic Instability ; Humans ; In Situ Hybridization ; Karyotyping ; Male ; Middle Aged ; Multiple Myeloma ; Plasma Cells* ; Polymorphism, Single Nucleotide

A variant Philadelphia chromosome (Ph) is generated from translocation of one or more partner chromosomes in addition to chromosomes 9 and 22. We have described the cases of 2 patients bearing variant Ph detected by interphase FISH but not detected by G-banded karyotyping and metaphase FISH. FISH was performed using BCR/ABL dual color dual fusion translocation probes (Abbott Molecular, USA). A 52-year-old man was diagnosed with acute leukemia of mixed phenotype. G-banded karyotyping showed 46,XY,t(9;22)(q34;q11.2)[12]/47,idem,+der(22)t(9;22)[5]/46,XY[3]. Interphase FISH revealed nuc ish(ABL1,BCR)x3(ABL1 con BCRx2)[329/450]/(ABL1,BCR)x4(ABL1 con BCRx3)[5/450]/(AL1,BCR)x3(ABL1 con BCRx1)[44/450]. Metaphase FISH showed ish (9;22)(ABL1+,BCR1+;BCR+,ABL+)[22]/der(22)(BCR+,ABL1+)[3]. The other case was that of a 31-yr-old male patient diagnosed with CML in the blastic phase. G-banded karyotyping of all 20 metaphase cells showed 47,XYYc,dup(1)(q21q32),del(7)(p11.2),t(9;22)(q34;q11.2). Interphase FISH revealed nuc ish(ABL1,BCR)x3(ABL1 con BCRx2)[254/600]/(ABL1,BCR)x3(ABL1 con BCRx1)[191/600]. Metaphase FISH showed ish t(9;22)(ABL1+,BCR+;BCR+,ABL1+)[16]. These results suggest that typical t(9;22) and variant Ph may coexist in the same patient, and interphase FISH may facilitate the detection of the variant Ph that cannot be detected by G-banded karyotyping alone.
Adult ; Chromosomes, Human, Pair 22 ; Chromosomes, Human, Pair 9 ; Humans ; In Situ Hybridization, Fluorescence/*methods ; Interphase ; Karyotyping ; Leukemia/diagnosis/genetics ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive/diagnosis/genetics ; Male ; Metaphase ; Middle Aged ; Phenotype ; *Philadelphia Chromosome ; Translocation, Genetic