Biomedical subjects

Y M Bae

Publications and source records attributed to Y M Bae.

At least 19 recordsLinked to original sources

PubMed2001

Ca2+-dependent membrane currents in vascular smooth muscle cells of the rabbit.

The membrane potential in vascular smooth muscle cells contributes to the regulation of cytosolic [Ca2+], which in turn regulates membrane potential by means of Ca2+i-dependent ionic currents. We investigated the characteristics of Ca2+i-dependent currents in rabbit coronary and pulmonary arterial smooth muscle cells. Ca2+i-dependent currents were recorded using the whole-cell patch-clamp technique while cytosolic [Ca2+] was increased by caffeine. The reversal potentials of caffeine-induced currents were between -80 and -10 mV under normal ionic conditions, whereas they were about 0 mV when K+-free NaCl solutions were used both in pipette and bath. The total substitution of extracellular Na+ with membrane-impermeable cation N-Methyl-D-glucamine did not affect caffeine-induced currents, implying no significant contribution of Na+ as a permeant ion to the currents. The substitution of extracellular NaCl with sucrose reduced outward component of the currents and shifted the reversal potentials according to the change in Cl- equilibrium potential. Upon application of the niflumic acid under K+-free conditions, most of the current induced by caffeine was inhibited. Taken together, the results of the present study indicate that K+ and Cl- currents are major components of Ca2+i-dependent currents in vascular smooth muscles isolated from coronary and pulmonary arteries of the rabbit, and the relative contribution of each type of current to total currents are not different between the two arteries.

Animals↗

PubMed2001

CD99 regulates the transport of MHC class I molecules from the Golgi complex to the cell surface.

The down-regulation of surface expression of MHC class I molecules has recently been reported in the CD99-deficient lymphoblastoid B cell line displaying the characteristics of Hodgkin's and Reed-Sternberg phenotype. Here, we demonstrate that the reduction of MHC class I molecules on the cell surface is primarily due to a defect in the transport from the Golgi complex to the plasma membrane. Loss of CD99 did not affect the steady-state expression levels of mRNA and protein of MHC class I molecules. In addition, the assembly of MHC class I molecules and the transport from the endoplasmic reticulum to the cis-Golgi occurred normally in the CD99-deficient cells, and no difference was detected between the CD99-deficient and the control cells in the pattern and degree of endocytosis. Instead, the CD99-deficient cells displayed the delayed transport of newly synthesized MHC class I molecules to the plasma membrane, thus causing accumulation of the molecules within the cells. The accumulated MHC class I molecules in the CD99-deficient cells were colocalized with alpha-mannosidase II and gamma-adaptin in the Golgi compartment. These results suggest that CD99 may be associated with the post-Golgi trafficking machinery by regulating the transport to the plasma membrane rather than the endocytosis of surface MHC class I molecules, providing a novel mechanism of MHC class I down-regulation for immune escape.

12E7 Antigen↗

PubMed2001

Use of transposon Tn5367 mutagenesis and a nitroimidazopyran-based selection system to demonstrate a requirement for fbiA and fbiB in coenzyme F(420) biosynthesis by Mycobacterium bovis BCG.

Three transposon Tn5367 mutagenesis vectors (phAE94, pPR28, and pPR29) were used to create a collection of insertion mutants of Mycobacterium bovis strain BCG. A strategy to select for transposon-generated mutants that cannot make coenzyme F(420) was developed using the nitroimidazopyran-based antituberculosis drug PA-824. One-third of 134 PA-824-resistant mutants were defective in F(420) accumulation. Two mutants that could not make F(420)-5,6 but which made the biosynthesis intermediate FO were examined more closely. These mutants contained transposons inserted in two adjacent homologues of Mycobacterium tuberculosis genes, which we have named fbiA and fbiB for F(420) biosynthesis. Homologues of fbiA were found in all seven microorganisms that have been fully sequenced and annotated and that are known to make F(420). fbiB homologues were found in all but one such organism. Complementation of the fbiA mutant with fbiAB and complementation of the fbiB mutant with fbiB both restored the F(420)-5,6 phenotype. Complementation of the fbiA mutant with fbiA or fbiB alone did not restore the F(420)-5,6 phenotype, but the fbiA mutant complemented with fbiA produced F(420)-2,3,4 at levels similar to F(420)-5,6 made by the wild-type strain, but produced much less F(420)-5. These data demonstrate that both genes are essential for normal F(420)-5,6 production and suggest that the fbiA mutation has a partial polar effect on fbiB. Reverse transcription-PCR data demonstrated that fbiA and fbiB constitute an operon. However, very low levels of fbiB mRNA are produced by the fbiA mutant, suggesting that a low-level alternative start site is located upstream of fbiB. The specific reactions catalyzed by FbiA and FbiB are unknown, but both function between FO and F(420)-5,6, since FO is made by both mutants.

Antitubercular Agents↗

PubMed2000

Viral latent membrane protein 1 (LMP-1)-induced CD99 down-regulation in B cells leads to the generation of cells with Hodgkin's and Reed-Sternberg phenotype.

Recently we reported that the down-regulation of CD99 (Mic2) is a primary requirement for the generation of Hodgkin's and Reed-Sternberg (H-RS) cells seen in Hodgkin's disease. In this study, we provide evidence that the down-regulation of CD99 is induced by high expression of Epstein-Barr virus (EBV) latent membrane protein 1 (LMP-1), which is highly expressed in H-RS cells of EBV-associated Hodgkin's disease. To investigate the effect of LMP-1 on the expression of CD99 in vitro, we established a stable cell line by transfecting an SV40-early promoter driven-LMP-1 expression construct into a neoplastic lymphoblastoid B cell line, IM9, in which the level of endogenous LMP-1 expression is almost negligible. In this cell line, the overexpression of LMP-1 led to the down-regulation of CD99 and the acquisition of morphological and functional characteristics of H-RS cells indistinguishable from those in lymph nodes of Hodgkin's disease patients and in CD99-deficient B cells. In addition, induced LMP-1 expression in an EBV-negative B cell clone, BJAB, directly caused the down-regulation of surface CD99 expression. Northern and Western analysis data, showing that overexpression of LMP-1 negatively influenced the expression of CD99, were supported by experiments in which a CD99 promoter-driven luciferase promoter reporter construct transfected into 293T cells was down-regulated when LMP-1 was coexpressed. Therefore, our data strongly suggest that the EBV LMP-1 protein plays a pivotal role in the down-regulation of CD99 via transcriptional regulation, which leads to the generation of the H-RS cells. (Blood. 2000;95:294-300)

12E7 Antigen↗

PubMed1999

Development of new adherent mutant from human myeloma-derived cell line: in vitro model of anaplastic transformation of myeloma.

Anaplastic myeloma is a rare but distinct, biologically aggressive variant of myeloma which usually results from dedifferentiation or anaplastic transformation of the myeloma cells. The molecular mechanisms that determine the biologic behavior of anaplastic myeloma and effective treatment modalities have not been well known due to lack of in vitro models. In the present study, we have developed an anaplastically transformed mutant from a human myeloma-derived cell line. In the process of long-term culture of the myeloma-derived IM-9 cell line in low serum and nutrient conditions, an adherent mutant line was developed and named IM-9/AD. This mutant cell line displayed several characteristics resembling anaplastic myeloma such as: 1, large cells with large vesicular nucleus and prominent nucleolus, multinuclearity and high mitotic figures; 2, loss of leukocyte-associated antigens; and 3, higher tumorigenecity in scid mice than its parental cell line. This newly developed mutant cell line may serve as a readily available in vitro model to investigate the biology of anaplastic myeloma.

Animals↗

PubMed1999

LFA-1- and ICAM-1-dependent homotypic aggregation of human thymocytes induced by JL1 engagement.

Cell-cell adhesion is essential for the appropriate immune response, differentiation, and migration of lymphocytes. This important physiological event is reflected in vitro by homotypic cell aggregation. We have previously reported that a 120 kDa cell surface glycoprotein, JL1, is a unique protein specifically expressed by immature double positive (DP) human thymocytes which are in the process of positive and negative selections through the interaction between thymocyte and antigen-presenting cells (APCs). The function of the JL1 molecule, however, is yet to be identified. We show here that anti-JL1 monoclonal antibody (mAb) induced the homotypic aggregation of human thymocytes in a temperature- and Mg2+-dependent manner. It required an intact cytoskeleton and the interaction between leucocyte function associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) since it was blocked by cytochalasin B and D, and mAb against LFA-1 and ICAM-1 which are known to be involved in the aggregation of thymocytes. Translocation of phosphatidylserine (PtdSer) through the cell membrane was not detected, implying that the molecular mechanism of JL-1-induced homotypic aggregation is different from that of CD99-induced homotypic aggregation. In summary, JL1 is a cell surface molecule that induces homotypic adhesion mediated by the LFA-1 and ICAM-1 interaction and cytoskeletal reorganization. These findings suggest that JL1 may be an important regulator of thymocyte development and thymocyte-APC interaction.

Antibodies, Monoclonal↗

PubMed1999

Contribution of Ca2+-activated K+ channels and non-selective cation channels to membrane potential of pulmonary arterial smooth muscle cells of the rabbit.

1. Using the perforated patch-clamp or whole-cell clamp technique, we investigated the contribution of Ca2+-activated K+ current (IK(Ca)) and non-selective cation currents (INSC) to the membrane potential in small pulmonary arterial smooth muscle cells of the rabbit. 2. The resting membrane potential (Vm) was -39.2 +/- 0.9 mV (n = 72). It did not stay at a constant level, but hyperpolarized irregularly, showing spontaneous transient hyperpolarizations (STHPs). The mean frequency and amplitude of the STHPs was 5.6 +/- 1. 1 Hz and -7.7 +/- 0.7 mV (n = 12), respectively. In the voltage-clamp mode, spontaneous transient outward currents (STOCs) were recorded with similar frequency and irregularity. 3. Intracellular application of BAPTA or extracellular application of TEA or charybdotoxin suppressed both the STHPs and STOCs. The depletion of intracellular Ca2+ stores by caffeine or ryanodine, and the removal of extracellular Ca2+ also abolished STHPs and STOCs. 4. Replacement of extracellular Na+ with NMDG+ caused hyperpolarization Vm of without affecting STHPs. Removal of extracellular Ca2+ induced a marked depolarization of Vm along with the disappearance of STHPs. 5. The ionic nature of the background inward current was identified. The permeability ratio of K+ : Cs+ : Na+ : Li+ was 1.7 : 1.3 : 1 : 0. 9, indicating that it is a non-selective cation current (INSC). The reversal potential of this current in control conditions was calculated to be -13.9 mV. The current was blocked by millimolar concentrations of extracellular Ca2+ and Mg2+. 6. From these results, it was concluded that (i) hyperpolarizing currents are mainly contributed by Ca2+-activated K+ (KCa) channels, and thus STOCs result in transient membrane hyperpolarization, and (ii) depolarizing currents are carried through NSC channels.

Animals↗

PubMed1998

Development of a new anti-CD4 monoclonal antibody (YG23) which inhibits the formation of colonies of human bone marrow progenitor cells.

We have previously reported CD4 expression in CD34+ hematopoietic progenitor cells and suggested a role of CD4 in normal hematopoiesis and its possible relationship with the pathogenesis of acquired immunodeficiency syndrome (AIDS). To investigate whether CD4 expression in bone marrow progenitor cells can explain bone marrow suppression in AIDS, monoclonal antibodies (mAbs) against human CD4 were developed by immunizing Balb/c mice with human thymocytes. Three mAbs completely blocked the binding of Leu3a antibody, a well-known anti-CD4 mAb, to thymocytes, which indicates overlap between the epitopes recognized by these and Leu3a antibodies. Interestingly, one of these mAbs, YG23, significantly inhibited colony formation of human bone marrow progenitor cells treated with GM-CSF. This is the first demonstration that ligation of CD4 by an anti-CD4 mAb suppresses GM-CSF mediated proliferation and differentiation of human hematopoietic progenitor cells by modifying the intracellular signaling pathway through CD4 molecules. Based on these findings, we propose that alteration of CD4 signaling by either cross-linked gp120 or antibodies directed against a certain epitope shared with the YG23 binding site of the CD4 molecule may play a role in bone marrow dysfunction in AIDS patients.

Animals↗

PubMed1998

A cell surface molecule, JL1; a specific target for diagnosis and treatment of leukemias.

We previously reported a novel differentiation antigen, which is specifically expressed in stage II double positive (CD4+CD8+) human cortical thymocytes (Park et al, J Exp Med 1993; 178: 1447-1451). This study was designed to investigate the expression pattern of JL1 in various types of leukemic cells from patients and normal hematopoietic cells to evaluate the possibility as a tool for diagnosis and treatment of leukemia. The expression of JL1 antigen was observed in 75.6% of leukemic cases (117 out of 154 leukemic patients tested) on flow cytometric analysis. The percentage of JL1-positive cases of T lineage acute lymphoblastic leukemia (T-ALL) (92.6%) was higher than that of other types of leukemias (75%). The presence of JL1 antigen was also confirmed by immunoblotting and immunoprecipitation. Since the JL1 antigen is selectively expressed on the surface of human leukemic cells but not on the mature human peripheral blood cells, normal bone marrow cells and various types of normal tissues, JL1 could be an excellent candidate for an immunodiagnostic and immunotherapeutic tool for hematopoietic malignancies such as leukemia.

Antibodies, Monoclonal↗

PubMed1997

Thymocytes positively select thymocytes in human system.

We previously demonstrated the expression of MHC class II molecules in a significant percentage of human fetal and postnatal thymocytes. These results, at that time, raised the question as to whether the MHC class II molecules on immature thymocytes could actively be involved in the selection of immature T cells. We have developed a human reaggregate culture system to address this issue. Surprisingly, despite the fact that thymic epithelial cells (TECs) have been shown to be a major selecting cell type of positive selection, we were clearly able to see the involvement of MHC class II+ thymocytes during selection process through T-T interaction. In addition, maturation to single positive (SP) cells occurred only in the presence of MHC class II molecules and immature thymocytes were found to be arrested at the double positive (DP) stage of differentiation by blocking of TCR recognition of MHC class II molecules. All these results strongly suggest that human MHC class II+ thymocytes actively participate in the selection of the TCR repertoire, for which TCR recognition of peptide/MHC class II may be an initial determining step.

Cell Aggregation↗

PubMed1997

Modulation of voltage-dependent K+ channel by redox potential in pulmonary and ear arterial smooth muscle cells of the rabbit.

It has been suggested that hypoxic pulmonary vasoconstriction (HPV) results from the depolarization that is induced by the suppression of K+ current in pulmonary arterial smooth muscle cells (PASMC). We tested the hypothesis that the effect of the cellular redox potential on voltage-sensitive K+ (Kv) current is involved in HPV as a primary sensing mechanism. Kv current in PASMC and ear arterial smooth muscle cells (EASMC) of the rabbit was recorded using the whole-cell patch-clamp technique, and the effect of redox agents [dithiothreitol, DTT and 2,2'-dithio-bis(5-nitropyridine), DTBNP] was tested. Kv current was decreased by DTT, but increased by DTBNP. DTT accelerated the inactivation kinetics, but did not affect steady-state activation and inactivation, whereas DTBNP accelerated activation kinetics. Kv current has a non-inactivating window in the range of from -40 mV to +10 mV. The resting membrane potential measured using the nystatin-perforated-patch method, however, lay between -50 mV and -30 mV and was not depolarized by 5 mM 4-aminopyridine. The membrane-impermeable oxidizing agent DTNB has no effect on Kv current, suggesting that redox modulation sites are intracellular sulphydryl groups. In EASMC, Kv current was decreased by DTT, but increased by DTBNP, indicating that the redox-potential-induced modulation of Kv current in EASMC and in PASMC is the same. It is therefore concluded that Kv current is modulated by the cellular redox potential, but that this modulation is not involved in HPV as a primary sensing mechanism.

Animals↗

PubMed1996

DN4: a novel cell surface molecule on cortical epithelial cells of the human thymus.

A novel cell surface molecule, DN4, defined by an mAb raised against human thymic epithelial cells, showed a specificity for epithelial cells of the thymic cortex. This antigen was not expressed at detectable levels on any other types of tissues in the human body except for the thymus and bone marrow. Immunohistochemical analysis revealed that the reactivity of anti-DN4 mAb was restricted to the thymic cortex, and the antigen-expressing cells were arranged in a reticular network with long processes between thymocytes. The cellular nature of DN4-positive cells was identified as cortical epithelial cells, as DN4 was expressed in a subpopulation of freshly prepared thymic stromal cells which contain a large amount of keratin and expression of DN4 was strictly confined to the cortical area within the thymus on immunohistochemical analysis of frozen tissues. Immunofluorescence and flow cytometric analysis revealed that a subpopulation of bone marrow cells was also positive for DN4 (20%). The large blasts of normal bone marrow cells were clearly labeled with anti-DN4 mAb, in contrast to small-sized bone marrow cells. This finding suggests that DN4 seems to be transiently expressed in certain blastic stages during the differentiation of bone marrow cells. Immunoprecipitation of 125I-labeled cell lysates from THP-1 and U937 cell lines with anti-DN4 mAb yielded a single chain glycoprotein with an approximate size of 80-85 kd. There was a reduction in apparent molecular weight of approximately 40 kd in the immunoprecipitation of cell lysates after endoglycosidase F treatment. Thus, DN4 seemed to have a considerable amount of carbohydrate group. DN4 appears to be a novel cortical epithelial cell antigen of the human thymus, and although the role of this molecule has not been well established experimentally, the possibility can be suggested that the DN4 molecule might be involved in the positive selection of thymocytes which occurs predominantly in the thymic cortical area.

Antibodies, Monoclonal↗

PubMed1995

A novel T-cell differentiation antigen expressed in immature human thymocytes and neuroglial cells.

We report a novel human thymocyte differentiation antigen ICT-1 with a molecular weight of 49 kDa that is noncovalently associated with another 12-kDa protein. The ICT-1 antigen is expressed in 50-70% of total thymocytes, but not in resting or PHA-activated peripheral blood T-cells and bone marrow cells. The thymocytes expressing ICT-1 antigen appear after the 18th week of gestation during fetal development. Since the distribution pattern of the ICT-1 antigen within thymus partly overlaps with that of the CD1 antigens, we investigated whether ICT-1 was one of the CD1 antigen family. However, the failure of anti-ICT-1 antibody to react with mouse L cells transfected with cDNA of CD1a, -b, and -c and the different histologic distribution patterns from that of CD1d strongly suggest that the anti-ICT-1 antibody recognizes an antigen distinct from CD1. Furthermore, ICT-1 is also expressed in human neuroglial cells such as oligodendroglioma, glioblastoma multiforme, Ewing's sarcoma, and cerebellar astrocyte. Hence we believe that the ICT-1 antigen may be a novel thymus-leukemia (TL) antigen or a nonclassical MHC class I antigen.

Animals↗

PubMed1994

Cloning, sequencing and expression of dinoflagellate luciferase DNA from a marine alga, Gonyaulax polyedra.

The marine dinoflagellate, Gonyaulax polyedra emits light in a reaction involving the enzymatic oxidation of its tetrapyrrole luciferin by molecular oxygen; its luciferase (LCF) single chain has an estimated molecular mass of 130 kDa, and exhibits a circadian rhythm in its activity. A cDNA expression library in the lambda ZAPII vector was constructed from the polyadenylated RNA isolated from the Gonyaulax cells during the early night phase, the time at which LCF synthesis is believed to be greatest. Of the approx. 1.2 . 10(5) phages from the library screened with antibody against Gonyaulax LCF, 13 positive plaques were obtained. The nucleotide sequences of two of the larger inserts (2.4 kb and 1.6 kb in length), both carrying the poly(A) tail, were determined and found to be identical in the overlapping region. When expressed in Escherichia coli, both cDNA clones produced active luciferase. A Northern hybridization using the cDNA as a probe showed that the length of the lcf mRNA is approx. 4.1 kb, sufficiently long to encode the 130 kDa LCF. Analyses of polymerase chain reaction products, prepared using both the cloned cDNA and Gonyaulax chromosomal DNA as templates, indicated that the cloned region of the luciferase gene does not carry any introns. This represents the first dinoflagellate luciferase to be cloned and sequenced; its deduced amino acid sequence bears no significant homologies with that of any other luciferase, or any other sequence in the data base.

Amino Acid Sequence↗

PubMed1994

A monoclonal antibody to cell surface antigen of human thymic epithelial cell.

The cell surface molecule identified by a monoclonal antibody(TE-1) to human thymic epithelial cell showed the specificity for thymic epithelial cells of both the cortex and medulla. TE-1 reacted with the epithelial cells of normal thymus and thymoma in fresh frozen tissues. The antigen recognized by TE-1 was mostly confined to the cell surface membrane and arranged in reticular network with long processes between thymocytes. On immunohistochemical analysis, TE-1 did not recognize normal epithelial cells of the uterine cervix, skin and stomach, and neoplastic cells of squamous cell carcinoma and gastric adenocarcinoma, all of which were stained with anti-cytokeratin monoclonal antibody. Among the tumor cell lines tested with flow cytometry, most of epithelial and all of hematopoietic cell origin were not labeled with TE-1. In summary, TE-1 appears to be a monoclonal antibody against a surface antigen of human thymic epithelial cell that is immunohistologically different from known epithelial cell surface antigens reported so far.

Animals↗

PubMed1993

JL1, a novel differentiation antigen of human cortical thymocyte.

Expression of a novel thymocyte differentiation antigen, JL1, defined by a monoclonal antibody (mAb) developed against human thymocytes showed a specificity for stage II double positive (CD4+CD8+) human cortical thymocytes. This antigen was not expressed at detectable levels on medullary thymocytes, mature peripheral leukocytes, bone marrow cells or on other types of tissues elsewhere in the human body. Immunohistologic analysis revealed that JL1 had a clear pattern of distribution on cortical thymocytes. Immunoprecipitation of 125I-labeled cell lysates from human thymocytes and Molt-4 leukemic cell line with anti-JL1 mAb yielded a 120-130-kD single chain glycoprotein. When immunoprecipitation of cell lysate was done after endoglycosidase F treatment, JL1 antigen was still detected by antibody but the band showed a reduction in apparent molecular mass of approximately 5 kD. This suggests that, although JL1 molecule contains carbohydrate group, this does not form a critical part of the antigenic determinant for anti-JL1 antibody. JL1 antigen appears to be the first double positive, stage-specific differentiation antigen of human thymocyte reported so far. This antigen would be a useful marker for lymphoblastic malignancy of stage II thymocyte origin and it may be involved in the thymocyte education process.

Antigens, Differentiation, T-Lymphocyte↗

PubMed1992

HLA-DR expression in human fetal thymocytes.

We analyzed the expression of MHC class I (W6/32) and class II (HLA-DR) antigens on human fetal and postnatal thymocytes by fluorescence-activated cell sorting. Less than 5% of prenatal thymocytes expressed HLA-DR before week 12 of gestation. However, the number of HLA-DR-positive cells significantly increased during the late second and third trimester of gestation, when greater than 50% of prenatal thymocytes expressed HLA-DR. Such high-level expressions of HLA-DR in fetal thymocytes were also demonstrated by Northern-blot analysis and immunohistochemistry. After birth, the percentage of HLA-DR-positive cells in thymocytes decreased gradually. A high-level expression of class I antigen was also observed in thymocytes from the early stages of gestation, but, in contrast to MHC class II, a majority of postnatal thymocytes maintained high levels of class I antigen after birth.

Cell Separation↗

PubMed1992

A monoclonal antibody to common acute lymphoblastic leukemia antigen (CALLA) and its expression on several human tumor cell lines.

We describe a newly-made murine monoclonal antibody to the common acute lymphoblastic leukemia antigen (CALLA), named SHB-10. The antigen detected by SHB-10 has a molecular weight of about 105 kDa. This antibody is very similar to that of conventional anti-CD10 Ab on indirect flowcytometric analysis using lymphoid malignant cell lines and peripheral lymphocytes of acute lymphoblastic leukemia (ALL) patients. The binding of anti-CD10 to Daudi cell and peripheral lymphocytes of ALL patients is blocked by SHB-10. Thus this monoclonal antibody is thought to detect the CALLA. The distribution of antigen detected by SHB-10 on several cell lines of neuroectodermal tumor and lymphoid malignancy was analysed and a slight difference in their cell surface expression is observed when compared with that by conventional anti-CD10. Further biochemical analysis is now under way for a better characterization of this antigen.

Animals↗