Biomedical subjects

Wha-Soon Chung

Publications and source records attributed to Wha-Soon Chung.

4 recordsLinked to original sources

PubMed2003

In vitro differentiation of natural killer T cells from human cord blood CD34+ cells.

Natural killer T (NKT) cells are involved in innate immune defence and also in the regulation of adaptive immune responses. However, the development of NKT cells in vitro has not been fully characterized and culture conditions have not been fully optimized. In the present study, we found that an NKT cell fraction developed during the in vitro culture of cord blood (CB) CD34+ cells, and this was subsequently characterized both phenotypically and morphologically. CD34+ cells purified from 10 human CB were cultured in the presence of several cytokines and analysed by flow cytometry, light microscopy and electron microscopy. The NKT cell fraction, defined phenotypically (CD3+CD16+CD56+CD94+) as expressing the invariant T-cell receptor Valpha24 and Vbeta11, appeared in the CD56hi fractions. Intracytoplasmic staining demonstrated that interferon-gamma and interleukin 4 (IL-4) were detected in the CD56hi fractions. IL-15 was essential and, in combination with either flt3-ligand (FL) or stem cell factor (SCF), was sufficient to induce the development of NKT cells. The phenotype of the NKT cell fraction was CD45RO+CD45RA- and CD4+CD8alpha+. Morphologically, they were very large, with either round or oval nuclei, moderately condensed chromatins, voluminous weakly basophilic cytoplasm and various cytoplasmic granules such as dense core granules, multivesicular bodies, and intermediate form granules. When CD34+ cells purified from bone marrow (BM) were compared with those from CB, the latter were consistently more efficient at generating CD56hi NKT cell fractions. In conclusion, IL-15 in combination with FL and/or SCF can induce the differentiation of NKT cells from human CB CD34+ cells.

Antigens, CD↗

PubMed2003

Adherent cells generated during long-term culture of human umbilical cord blood CD34+ cells have characteristics of endothelial cells and beneficial effect on cord blood ex vivo expansion.

Hematopoiesis depends on the association of hematopoietic stem cells with stromal cells that constitute the hematopoietic microenvironment. The in vitro development of the endothelial cell from umbilical cord blood (UCB) is not well established and has met very limited success. In this study, UCB CD34(+) cells were cultured for 5 weeks in a stroma-free liquid culture system using thrombopoietin, flt3 ligand, and granulocyte-colony stimulating factor. By week 4-5, we found that firmly adherent fibroblast-like cells were established. These cells showed characteristics of endothelial cells expressing von Willebrand factor, human vascular cell adhesion molecule-1, human intracellular adhesion molecule-1, human CD31, E-selectin, and human macrophage. Furthermore, when comparing an ex vivo system without an established endothelial monolayer to an ex vivo system with an established endothelial monolayer, better expansion of total nucleated cells, CD34(+) cells, and colony-forming units (CFUs)-granulocyte-macrophage and CFUs-granulocyte-erythroid-megakaryocyte-macrophage were found during culture. This phenomenon was in part due to the fact that a significant reduction of apoptotic fractions was found in the CD34(+) cells, which were cultured on the adherent monolayer for up to 5 weeks. To gather quantitative data on the number of endothelial cells derived from a given number of CD34 cells, we performed limiting dilution assay by using Poisson distribution: the number of tested cells (linear scale) producing a 37% negative culture (logarithmic scale) is the number of cells containing one endothelial cell. By this method, one endothelial cell may be found from 314 CD34(+) cells after 5 weeks of culture. These results suggest that the UCB CD34(+) cell fraction contains endothelial cell precursors, establishing the hematopoietic microenvironment and providing the beneficial effects through downregulating apoptosis on UCB expansion protocols. These observations may provide insight for future cellular therapy or graft engineering.

Antibodies↗

PubMed2002

Decrease in apoptosis and increase in polyploidization of megakaryocytes by stem cell factor during ex vivo expansion of human cord blood CD34+ cells using thrombopoietin.

Thrombopoietin (TPO) is widely used for ex vivo expansion of hematopoietic stem cells. Previously, we have reported that TPO induces a characteristic pattern of apoptosis, and the TPO-induced apoptosis is closely associated with megakaryocyte (MK) differentiation. In the present study, several cytokines, flt3-ligand, stem cell factor (SCF), interleukin-3 (IL-3), IL-6, IL-11, leukemia inhibitory factor, G-CSF, and erythropoietin, which are known to affect megakaryocytopoiesis, have been evaluated to elucidate their effects on the TPO-induced apoptosis. Measurement of apoptosis by flow cytometry revealed that only SCF absolutely reduced the TPO-induced apoptosis in MK fractions, particularly in the late phase of ex vivo expansion. Platelet production was demonstrated by electron microscopy in a later phase when SCF was added. Simultaneous measurement of DNA contents with immunophenotyping demonstrated a significant increase in polyploidization in the CD41+ cell fraction when cultured with SCF. These results suggested that SCF not only inhibited premature senescence but also enhanced maturation of the differentiating cells of MK lineage during ex vivo expansion using TPO.

Antigens, CD34↗

PubMed2002

Functional maturation of myeloid cells during in vitro differentiation from human cord blood CD34+ cells.

CD34+ cells purified from human cord blood were expanded in the presence of several cytokines. The cultured cells were assayed for myeloid effector functions, including phagocytic activity, respiratory burst and microbicidal activity. The results showed that phagocytic activity was observed as early as day 6, irrespective of the type of cytokines used. By contrast, respiratory burst and microbicidal activity peaked on day 15 and were dependent on the type of cytokines used. In particular, granulocyte-colony stimulating factor and interleukin-6 markedly enhanced respiratory burst and microbicidal activity.

Antigens, CD34↗