Difference in Cell Characteristics among the Monoclonal Cell Populations Obtained from the Human Umbilical Cord Blood Derived Mesenchymal Stem Cell Population / 대한정형외과학회잡지

PURPOSE: The aim of this study was to obtain single cell-derived clones from human umbilical cord blood (hUCB) derived mesenchymal stem cell (MSC) population, to compare the gene expression patterns and differentiation characteristics among the hUCB derived MSC population and its monoclonal cell populations, and to determine if the MSC population is homogenous. MATERIALS AND METHODS: The single cells were isolated from a hUCB derived MSC population and cultured on each well of a culture plate. The gene expression pattern of each monoclonal cell population expanded from the single cells was detected by RT-PCR for osteogenic, chondrogenic, and adipogenic specific genes. The monoclonal cell populations were differentiated into osteogenic, chondrogenic, and adipogenic lineages and were confirmed by specific staining. RESULTS: Fifteen monoclonal cell populations were obtained from the total seeding of 864 single cells. The cell morphology and gene expression patterns among the hUCB derived MSCs and its monoclonal cell population were different. Tri-lineage differentiation potency was different among the monoclonal cell populations. CONCLUSION: The difference in the cell morphology, gene expression patterns, and differentiation characteristics among the monoclonal cell populations suggest heterogeneity of the MSC population isolated using the currently available method.

Difference in HLA-DR Expression of Human Umbilical Cord Blood Derived Mesenchymal Stem Cells after Tri-lineage Differentiation / 대한정형외과연구학회지

PURPOSE: The aim of this study was to examine the expression of HLA-DR surface antigen in undifferentiated human umbilical cord blood (hUCB) derived mesenchymal stem cells (MSCs) and after osteogenic, chondrogenic, and adipogenic differentiation. MATERIALS AND METHODS: hUCB-derived MSCs were differentiated into osteogenic, chondrogenic, and adipogenic lineages. Differentiation was assessed by immunohistochemical staining and RT-PCR. The expression of HLA-DR was assessed with antihuman HLA-DR antibody in undifferentiated hUCB-derived MSCs and after tri-lineage differentiation. RESULTS: HLA-DR expression was negative in undifferentiated hUCB-derived MSCs and after osteogenic and adipogenic differentiation. However, HLA-DR surface antigen was expressed after chondrogenic differentiation. CONCLUSION: The immunologic properties of hUCB-derived MSCs differ from known reports on bone marrow derived MSCs from the results of this study. Careful immunological survey seems to be needed in case of considering the transplantation of hUCB-derived MSCs differentiated into chondrocytes or cartilaginous tissue.

Establishment of Embryonic Stem Cell Line from Isolated Blastomeres from Mouse Preimplantation Embryos / 대한불임학회지

OBJECTIVE: The aim of this study was to investigate whether embryonic stem (ES) cells can be established from isolated blastomeres of mouse embryos. METHODS: Blastomeres were separated from mouse (C57Bl/6J) 2- or 4-cell embryos. Isolated blastomeres or whole 4-cell embryos were co-cultured with mitosis-arrested STO feeder cells in DMEM supplemented with recombinant murine leukemia inhibitory factor and ES-qualified fetal bovine serum. After the tentative ES cell lines were maintained from isolated blastomeres or whole embryos, some of them were frozen and the others were sub-cultured continually. Characteristics of tentative ES cell lines as were evaluated for specific gene expressions with immunocytochemistry and RT-PCR. RESULTS: One ES cell line (3.0%) was established from isolated blastomere of 2-cell embryo and one cell line (4.0%) from isolated two blastomeres of 4-cell embryo. And five cell lines (16.7%) were established from whole 4-cell embryos. Both cell lines from isolated blastomere and whole embryo expressed mouse ES cells specific markers such as SSEA-1, Oct-4 and alkaline phosphatase. Marker genes of three germ layers were expressed from embryoid bodies of both cell lines. CONCLUSION: This study suggests that mouse ES cells could be established from isolated blastomeres, although the efficiency is lower than whole embryos. This animal model could be applied to establishment of autologous human ES cells from biopsied blastomeres of preimplantation embryos in human IVF-ET program.

Induced Differentiation of Embryonic Stem Cells to Insulin Secreting Cells / 대한불임학회지

OBJECTIVE: Embryonic stem (ES) cells could be differentiated into the specific cell types by alternation of culture condition and modification of gene expression. This study was performed to evaluate the differentiation protocol for mouse and human ES cells to insulin secreting cells. METHODS: Undifferentiated mouse (JH-1) and human (Miz-hES1) ES cells were cultured on STO feeder layer, and embryoid bodies (EBs) were formed by suspension culture. For the differentiation, EBs were cultured by sequential system with three stage protocol. The differentiating ES cells were collected and marker gene expressions were analyzed by semi-quantitative RT-PCR in each stage. Amount of secreted insulin levels in culture media of human ES cells were measured by human insulin specific RIA kit. RESULTS: During the differentiation process of human ES cells, GATA-4, alpha-fetoprotein, glucose transporter-2 and Ngn-3 expression were increased whereas Oct-4 was decreased progressively. Insulin and albumin mRNAs were expressed from stage II in mouse ES cells and from stage III in human ES cells. We detected 3.0~7.9 microU/ml secretion of insulin from differentiated human ES cells by in vitro culture for 36 days. CONCLUSION: The sequential culture system could induce the differentiation of mouse and human ES cells into insulin secreting cells. This is the first report of differentiation of human ES cells into insulin secreting cells by in vitro culture with serum and insulin free medium.

Isolation and Culture of Human Embryonic Stem-like Cells from Abnormal Blastocysts / 대한불임학회지

No abstract available.

Interaction of HIV-1 core p24 antigen with human monocytic cell line THP1 results in TNF-alpha dependent secretion of matrix metalloproteinase-9

Immunological mechanisms involving the release of inflammatory factors by HIV-1 infected microglia in the brain have been implicated in the pathogenesis of HIV dementia (HIVD). Since the regulation of matrix metalloproteinases (MMPs) activity can be influenced by variety of inflammatory mediators, this study was undertaken to look for a correlation between the MMP-9 release and the production of TNF-alpha in response to HIV-1 p24 in the human monocyte cell line THP-1 as a model for microglia. First, it was shown that HIV-1 core p24 antigen induced THP-1 to secrete MMP-9 in a dose response manner while it elicited a little effect on MMP-2 release in human astroglial cell line T98G. Next, it was found that p24 induced THP-1 to secrete TNF-alpha without prior differentiation into macrophages by phorbol myristate acetate (PMA) treatment. Furthermore, anti-TNF-alpha neutralizing antibodies significantly blocked p24-induced MMP-9 release in a dose dependent manner. Our data indicate that p24 antigen induces monocytic MMP-9 release by triggering up-regulation of TNF-alpha secretion.