Biomedical subjects

H M Kim

Publications and source records attributed to H M Kim.

At least 271 records · Page 15Linked to original sources

PubMed1995

Isolation of calcium-phosphate crystals of bone by non-aqueous methods at low temperature.

We have developed low temperature nonaqueous solution methods as well as low power plasma ashing for the degradation of the organic matrix of bone power which have permitted us to obtain bone crystals essentially free of organic matrix constituents without any significant change in their composition, overall structure, or internal short-range order. We have also been able to disperse the crystals, which has made it possible to examine the isolated crystals by X-ray diffraction and resolution-enhanced Fourier transform infrared (FTIR) spectroscopy and isolated single crystals by high resolution transmission electron microscopy (TEM) and electron diffraction. TEM of isolated single crystals of fish, chicken, mouse and bovine bone have clearly demonstrated that the crystals are very thin plates. No rod or needle-like crystals were observed in any of the bone samples in the four species studied including the earliest crystals deposited. Although there were variations in the size distribution of the crystals in the different species studied, in general the average crystal dimensions were very similar.

Animals↗

PubMed1995

Effect of qigong training on proportions of T lymphocyte subsets in human peripheral blood.

The effect of Qigong training on proportions of T lymphocyte subsets was investigated in human peripheral blood. We observed that the ratio of CD4+/CD8+ T lymphocytes was increased as much as 50% in a trainee group who practiced Qigong training more than 5 months compared to a normal healthy group who did not practice. The absolute number of CD4+ T lymphocytes was also elevated in trainee group with 100 cells/mm3 more than in normal healthy group. The positive correlation between the ratio of CD4+/CD8+ T lymphocytes and the ratio of CD4+45RA-/CD4+CD45RA+ T lymphocytes was shown in the trainee group. In contrast, there was a negative correlation between the ratio of CD4+/CD8+ T lymphocytes and the ratio of CD8+CD57+/CD8+CD57- T lymphocytes in the trainee group. The data indicate that Qigong training affects the profile of lymphocyte subsets in human peripheral blood, especially the proportion of CD4+ T lymphocytes.

Adult↗

PubMed1995

Delayed cutaneous hypersensitivity reactions in Qigong (chun do sun bup) trainees by multitest cell mediated immunity.

To determine the difference of cellular immunity between a Qigong trainee group and a normal healthy group, skin tests for delayed cutaneous hypersensitivity (DCH) were carried out with ubiquitous seven antigens. The maximal antigen response time was faster in Qigong trainee group (24 hr) and the response antigen number was also higher in the Qigong trainee group (6 antigens) than in normal healthy person (48 hr and 4 antigens). Qigong trainee also had a larger induration diameter (5.14 mm) than normal healthy person (3.79 mm) at 24 hr. Our results represent the difference in cell mediated immunity (CMI) between Qigong trainees and normal healthy subjects.

Adult↗

PubMed1995

B-lymphocyte-stimulating polysaccharide from mushroom Phellinus linteus.

Hot water extract prepared from the mycelial culture of mushroom Phellinus linteus stimulated polyclonal antibody production in an in vitro culture system. The active fraction PLP was purified from the extract ca. 1030-fold by ethanol precipitation followed by DEAE-cellulose and gel permeation chromatography. PLP contained 13.2% (w/w) peptide and 82.5% (w/w) carbohydrate. About 6.8% (w/w) of the total carbohydrate was uronic acid. The molecular weight distribution of PLP was found to be nearly homogeneous (153 kDa) in gel permeation HPLC analysis. Neutral sugar composition analysis revealed Ara (7.5%), Xyl (3.7%), Glc (21.1%), Gal (24.1%) and Man (44.2%). Uronic acid was identified as a glucuronic acid by gas chromatography. Ten amino acids were detected and Asp and Glu were the major components. In our assay system, the half-maximal concentration of PLP for B-lymphocyte stimulation was ca. 3 micrograms/ml. Partial acid hydrolysis as well as sodium periodate treatment of PLP decreased the activity significantly, suggesting that both the full molecular size and the sugar moiety were essential. However, proteinase K treatment for up to 48 h did not affect the activity.

Acetylation↗

PubMed1995

Functional human hepatocytes: isolation from small liver biopsy samples and primary cultivation with liver-specific functions.

Morphologically and functionally intact human hepatocytes were isolated from small liver biopsy samples weighing about 1-2 g by initial digestion with collagenase followed by repeated digestions with trypsin. The usual yield of hepatocytes was greater than 1 x 10(7) cells per g of liver sample and cell viability, as judged by dye exclusion test, was routinely over 90%. The isolated human hepatocytes showed intact morphology under scanning electron microscope. Formation of membrane protrusions upon phalloidin addition demonstrated that the actin in isolated hepatocytes was maintained with its structural integrity. The cultured human hepatocytes retained a variety of liver-specific functions which were similarly exhibited by rat hepatocytes isolated using the same procedure. The cultured human hepatocytes exhibited a specific cytochrome P-450 related enzyme activity, and active amino acid uptake that increased upon addition of hormones like glucagon and dexamethasone. Additionally, the cultured human hepatocytes synthesized DNA actively and, human serum albumin, and was found to be responsive to modulation by growth modulating hormones, cytokines and hepatotoxic agents. Based on the profile of activity described above, the presently established conditions for isolation and culturing of human hepatocytes demonstrate that functional liver cells can be obtained from small biopsied liver samples.

Albumins↗

PubMed1995

Administration of dehydroepiandrosterone reverses the immune suppression induced by high dose antigen in mice.

Several factors including antigen concentration, the route of antigen administration, hormones and cytokines have shown to affect T cells to produce the distinct patterns of lymphokines which exert regulatory and effector functions of immune response. In this study, we asked whether administration of dehydroepiandrosterone (DHEA) to mice which were tolerized by high dose of antigen could modulate T cell functions to restore the suppressed cellular immune response and to produce the distinct lymphokines. An intravenous injection of high dose of sheep red blood cells induced suppression of delayed type hypersensitivity (DTH) and a single subcutaneous injection of the tolerant mice with DHEA restored the suppressed DTH response. Furthermore, in vitro treatment of spleen cells from tolerant mice with DHEA abolished the transfer of tolerance to naive recipients. Lymphocytes from the DHEA-treated tolerant mice produced more IFN-gamma and less IL-4 and IL-6 than the cells from tolerant animals without DHEA treatment. These findings indicate that DHEA could recover antigen-specific immune suppression by differentially affecting T cells to produce the distinct lymphokines.

Animals↗

PubMed1995

Intracellular Ca2+ pool depletion is linked to the induction of nitric oxide synthesis in murine peritoneal macrophages.

The ability of putative Ca(2+)-ATPase inhibitor of endoplasmic reticulum (ER), thapsigargin (TG), to induce nitric oxide (NO) synthesis in murine peritoneal macrophages was examined. TG alone had small effect on NO synthesis, whereas TG in combination with LPS markedly increased NO synthesis in a dose dependent manner. This increase in NO synthesis was reflected as increased amount of inducible NO synthase (iNOS) mRNA by Northern blotting. In addition, the ability of TG on NO synthesis could be mimicked by another chemically unrelated inhibitor of Ca(2+)-ATPase, 2,5-DI-(t-butyl)-1, 4-benzohydroquinone (tBuBHQ). Adding EGTA, a calcium chelator, to the incubation medium significantly reduced the ability of macrophages to induce NO synthesis in response to the optimal stimulation of TG or TG plus LPS. These results therefore demonstrate that intracellular Ca2+ pool depletion is linked to the induction of NO synthesis in murine peritoneal macrophages and further suggest that it is also related with interferon-gamma (IFN-gamma)-induced signaling.

Animals↗

PubMed1995

Down-regulation of transforming growth factor-beta gene expression by antisense oligodeoxynucleotides increases recombinant interferon-gamma-induced nitric oxide synthesis in murine peritoneal macrophages.

Increasing evidence indicates that the production of nitric oxide (NO) by inducible NO synthase (iNOS) is tightly regulated. Transforming growth factor-beta (TGF-beta) is a family of multifunctional peptides secreted during macrophage activation, but several lines of evidence suggest that TGF-beta is selectively suppressive for macrophage NO production. We therefore reasoned that a strategy employing oligodeoxynucleotides (ODN) complementary to TGF-beta mRNA (antisense ODN) might increase NO production in interferon-gamma (IFN-gamma) treated murine peritoneal macrophages. To evaluate this concept, we tested the effects of antisense ODN targeted to TGF-beta mRNA (25-mer ODN complementary to TGF-beta mRNA sequences) by introducing them into the medium of cultured macrophages. Phosphorothioation of ODN was employed to retard their degradation. Antisense ODN had no effect on NO production by itself, whereas IFN-gamma alone had a modest effect. When antisense ODN were used in combination with IFN-gamma, there was a marked cooperative induction of NO production. These effects of antisense ODN were associated with decreased TGF-beta expression in activated macrophages. However, sense ODN had no effect. Adding anti-TGF-beta antibodies to the IFN-gamma-treated macrophages mimicked the positive effect of antisense ODN on NO production. In addition, the effects of either antisense ODN or anti-TGF-beta antibodies were blocked by adding exogenous TGF-beta in cultured macrophages. These results indicate that the generation of TGF-beta by activated macrophages provides a self-regulating mechanism by which the temporal and perhaps spatial production of NO, a reactive and potentially toxic mediator, can be finely regulated.

Animals↗

PubMed1995

Expression of protein kinase C delta gene in germ cells.

PURPOSE: To investigate the biological functions of protein kinase C delta (PKC delta) in spermatogenesis. MATERIALS AND METHODS: We examined PKC delta transcript in mouse testis by means of in situ hybridization and Northern blotting. RESULTS: In testes of normal mice, signals of PKC delta gene expression were detected specifically at the spermatid stage. The PKC delta gene was weakly expressed in 8-week-old mice and highly expressed by 12 weeks. However, the expression was not detected in testes of germ cell-deficient W/Wv mice even at 12 weeks. CONCLUSIONS: Protein kinase C delta gene expression may be controlled by specific developmental processes and PKC delta may play a role in spermatogenesis.

Animals↗

PubMed1994

Synergistic cooperation between phorbol ester and IFN-gamma for induction of nitric oxide synthesis in murine peritoneal macrophages.

The role of protein kinase C (PKC) in the induction of nitric oxide (NO) synthesis in murine peritoneal macrophages was examined. Phorbol ester, a PKC activator, had no effect on NO synthesis by itself, whereas IFN-gamma alone had modest activity. When phorbol ester was used in combination with IFN-gamma, there was a marked cooperative induction of NO synthesis in a dose-dependent manner. This increase in NO synthesis was reflected as increased amount of inducible NO synthase (iNOS) mRNA, as determined by Northern blotting. The optimal effect of phorbol ester was shown at 6 h after treatment with IFN-gamma. Phorbol ester also induced the release of NO to the incubation medium by bacillus Calmette-Guerin-infected peritoneal macrophages. Prolonged incubation of cells with phorbol ester, which down-regulates PKC activity, abolished the synergistic cooperative effect on NO production with IFN-gamma. In addition, such PKC inhibitors as staurosporin or polymyxin B reduced NO production induced by IFN-gamma plus phorbol ester. When the cells were treated with both actinomycin D and phorbol ester after IFN-gamma stimulation, more NO was produced and more iNOS mRNA was expressed than in the cells treated with actinomycin D alone. On the basis of these observations, we conclude that PKC might not be directly involved in the expression of NO synthase, but, instead, might be involved in the stabilization of the iNOS mRNA already expressed by the treatment of IFN-gamma.

Alkaloids↗

PubMed1994

Nitric oxide inhibits the expression of protein kinase C delta gene in the murine peritoneal macrophages.

Since there is increasing evidence that protein kinase C (PKC) has a crucial role in the production of nitric oxide (NO) from activated macrophages, this study was undertaken to address whether NO could regulate the expression of the gene of this enzyme. Stimulation of the cells with lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA) after treatment with recombinant interferon-gamma (rIFN-gamma) resulted in the increased production of NO in the medium. rIFN-gamma in combination with either LPS or PMA showed marked inhibition of the expression of PKC delta gene, whereas rIFN-gamma alone showed modest inhibition. The inhibition of gene expression was correlated with the amount of NO produced by activated macrophages. The inhibitory effect of NO on the expression of PKC delta gene is mimicked by the treatment of NO generating agent, sodium nitroprusside (SNP). On the other hand, a specific inhibitor for NO synthase, NG-monomethyl-L-arginine (NGMMA), blocked the inhibition of the expression of PKC delta gene by blocking the NO production in the rIFN-gamma and LPS-stimulated cells. However, production of NO did not affect the expression of both TNF-alpha and TGF-beta gene which were induced by the stimulation of macrophages, as well as beta-actin gene, which was constitutively expressed in the macrophages. In conclusion, these findings show that NO has a regulatory role for the expression of the gene of PKC delta which is crucially involved in the process of NO synthesis.

Animals↗

PubMed1994

Involvement of protein kinase C in the inhibition of nitric oxide production from murine microglial cells by glucocorticoid.

The effects of glucocorticoid on the production of nitric oxide (NO) by murine microglial cells were investigated. Stimulation of the cells with lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA) after the treatment of recombinant interferon-gamma (rIFN-gamma) resulted in the increased accumulation of nitrite in the medium. Concomitant incubation of the cells with dexamethasone (DEX) markedly inhibited the production of NO in a dose dependent manner. DEX also suppressed both rIFN-gamma and rIFN-gamma plus LPS-induced activity of the enzyme protein kinase C (PKC), a putative regulator of NO synthesis, but had only a modest inhibitory effect on basal activity. In addition, the inhibitory effect of DEX on NO generation was mimicked by the treatment of PKC inhibitors such as staurosporine (STSN) and polymyxin B. Our findings show that glucocorticoids have the potential to modulate central nervous system (CNS) NO production via the inhibition of PKC activity particularly under the conditions of stimulated production of NO, such as inflammatory and demyelinating CNS disorders.

Alkaloids↗

PubMed1994

Comparative studies of adriamycin and 28-deacetyl sendanin on in vitro growth inhibition of human cancer cell lines.

The limonoid compound (28-deacetyl sendanin) isolated from the fruit of Melia toosendan SIEB. et ZUCC. was evaluated on anticancer activity. According to a standard in vitro cytotoxicity assay, eight human cancer cell lines and SRB assay were introduced for present evaluation. As a positive standard, adriamycin was tested in parallel. The cell lines were originated from six different organs. In view of dose-response profiles to 28-deacetyl sendanin, the most sensitive cells were SF-539 and PC-3 which were derived from CNS and prostate, respectively. In contrast, all the cell lines responded similarly to adriamycin to give rise to nearly identical dose-response profiles. By comparison of Gl50 between 28-deacetyl sendanin and adriamycin, six cell lines were more sensitive to 28-deacetyl sendanin and two were more resistant. As a result, 28-deacetyl sendanin had more sensitive and selective inhibitory effects on in vitro growth of human cancer cell lines in a comparison with adriamycin.

Antineoplastic Agents↗

PubMed1994

Direct exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) increases infectivity of human erythrocytes to a malarial parasite.

Direct exposure to 10 nM 2,3,7,8-TCDD caused a 75% increase and a 2-fold increase in the infectivity of isolated human erythrocytes to P. falciparum after 48 hours when the parasites were in an unsynchronized or synchronized state of growth, respectively. Treatment of human erythrocytes with 10 microM sodium orthovanadate (NaOV), an inhibitor of plasma membrane Ca-ATPase and phosphotyrosine phosphatase, decreased parasitemia by 30%. Co-treatment of RBCs with TCDD and NaOV completely blocked the TCDD-induced increase in parasitemia. Because erythrocytes are anucleated, these results are discussed as evidence for biochemical changes by TCDD without requiring the activation of gene products.

Animals↗

PubMed1994

Differential expression of protein kinase C genes in cultured mast cells derived from normal and mast-cell-deficient mice and mast cell lines.

We investigated the expression of mRNA of protein kinase C (PKC) isozymes (alpha, beta, gamma, delta, epsilon, xi, and eta) in cultured mast cells (CMC) derived from normal (+/+) mice, CMC derived from genetically mast-cell-deficient (W/W, Wv/Wv, and mi/mi) mice, and murine mast cell lines (IC2, MC9, and P-815) by Northern blotting. In +/+ CMC, abundant expression of PKC delta and moderate expression of PKC alpha and beta was observed, while other PKCs (types gamma, epsilon, xi, and eta) were not detected. In vivo expression of PKC delta was demonstrated in the skin by in situ hybridization. In mast cell lines, the expression pattern of PKC isozymes was similar to that of +/+ CMC, except that the expression of PKC eta was detected in the IC2 cell line. The expression levels of PKC delta in CMC derived from c-kit-deficient mutants, W/W, Wv/Wv, and mi/mi, were lower than that of +/+ mice. These results indicate that PKC delta is the main isozyme in various types of murine mast cells and also suggest that the reduced level of PKC delta expression in mutant mice may be caused by a deficit in the signal transduction system through c-kit receptor.

Animals↗

PubMed1994

Generation of nitric oxide inhibits formation of superoxide in macrophages during activation.

We have studied the production of nitric oxide (NO) and superoxide by murine peritoneal macrophages during activation. The production of NO was induced by activation of cells with recombinant interferon-gamma (rIFN-gamma) and lipopolysaccharide (LPS). Phorbol 12-myristate 13-acetate (PMA)-induced formation of superoxide also increased during activation. However, NO released by the activated macrophages exerted the inhibitory effect on the superoxide formation in the same cells. This fact is supported by the increased production of superoxide when the cells were treated with NG-monomethyl-L-arginine (NGMMA) in addition to stimulation with rIFN-gamma and LPS. The production of superoxide was also inhibited by treatment with sodium nitroprusside (SPN), which spontaneously released nitric oxide in vitro, and at the same time there was increased adenosine diphosphate (ADP)-ribosylation of 37 kDa proteins of the cytoplasm. The 3-aminobenzamide (3-AB) treatment, which decreased ADP-ribosylation, partially reversed SNP-induced inhibition of superoxide generation in macrophages. The above data provide evidence that NO decreases superoxide formation possibly via ADP-ribosylation.

Adenosine Diphosphate↗

PubMed1993

PKC gamma gene expression is delayed in postnatal central nervous system of mi/mi mice.

In the central nervous system (CNS), the expression of protein kinase C (PKC) genes is strictly controlled by the developmental stage. We have examined the expression of PKC genes (cPKC alpha, beta, gamma, and nPKC delta, epsilon) in the process of the postnatal development in normal (+/+) C57BL/6 and microphthalmic (mi/mi) C57BL/6 mouse brains by Northern blotting and in situ hybridization. By Northern blotting, the expression level of cPKC gamma mRNA in mi/mi mice was significantly lower than that in +/+ littermates at d 9, 13, and 17. By in situ hybridization analysis, cPKC gamma mRNA-positive cells were detected in hippocampal and Purkinje cells in +/+ and mi/mi mice, but the magnitude of the signals in mi/mi mice was lower than that of +/+ mice, and the number of positive cells was smaller, whereas other isozymes (cPKC alpha, beta, and nPKC delta, epsilon) showed no significant difference between normal and mi/mi mice. The neuronal morphometric analysis by anti-P400 antibody revealed the same number and expression level of P400 protein in cerebellar Purkinje cells compared with +/+ mice. These results indicate that the deficiency of mi gene product causes the delayed expression of the cPKC gamma gene.

Animals↗

PubMed1993

Absence of immature mast cells in the skin of Ws/Ws rats with a small deletion at tyrosine kinase domain of the c-kit gene.

Ws/Ws rats have a small deletion at the tyrosine kinase domain of the c-kit gene, and practically no mast cells were detectable when the tissues were stained with alcian blue. Because alcian blue stains proteoglycans, there is a possibility that immature mast cells that do not contain a sufficient amount of proteoglycans are not detectable by this method. We examined this possibility by using other markers of mast cells. The histamine content in the skin of Ws/Ws rats was 0.3% that of control normal (+/+) rats. Because the number of alcian blue-positive mast cells in the skin of Ws/Ws rats was also 0.3% that of +/+ rats, histamine in the skin seemed to be concentrated to alcian blue-positive mast cells. Mast cells in the skin of +/+ rats express messenger RNA of Fc epsilon RI beta-subunit and c-kit protein. Because c-kit messenger RNA was normally expressed at least in the brain of Ws/Ws rats despite the small deletion, we examined the expression of Fc epsilon RI beta-subunit and c-kit messenger RNA in the skin and stomach of Ws/Ws rats by reverse transcriptase modification of polymerase chain reaction. Expression of either Fc epsilon RI beta-subunit or c-kit messenger RNA in the skin and stomach of Ws/Ws rats was estimated to be less than 1% that of +/+ rats. Moreover no Fc epsilon RI beta-subunit-expressing and no c-kit-expressing cells were detectable in the skin of Ws/Ws rats by in situ hybridization histochemistry. The present result suggests the absence of immature mast cells in tissues of Ws/Ws rats.

Animals↗