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Biomedical subjects

N Harada

Publications and source records attributed to N Harada.

At least 199 records · Page 11Linked to original sources
PubMed1999

Gabexate mesilate, a synthetic protease inhibitor, reduces ischemia/reperfusion injury of rat liver by inhibiting leukocyte activation.

OBJECTIVE: To investigate whether gabexate mesilate, a synthetic protease inhibitor with anticoagulant properties, prevents hepatic damage by inhibiting leukocyte activation, we examined its effect on ischemia/reperfusion injury of rat liver in which activated leukocytes play a critical role. DESIGN: Prospective, randomized, controlled study. SETTING: Research laboratory at a university medical center. SUBJECTS: Male Wistar rats weighing 220 to 280 g. INTERVENTIONS: Hepatic damage was evaluated by changes in bile flow and serum transaminase concentrations after ischemia/ reperfusion. Rats received continuous intravenous infusions of gabexate mesilate (10 mg/kg/hr) or intravenous administration of an inactive derivative of activated factor X (Xa), a selective inhibitor of thrombin generation (3 mg/kg), immediately before the induction of ischemia in the median and left lobes of the liver. To determine whether gabexate mesilate inhibits leukocyte activation, we examined the effects of gabexate mesilate on hepatic concentrations of tumor necrosis factor-alpha and rat interleukin-8 and on hepatic myeloperoxidase activity after ischemia/reperfusion. MEASUREMENTS AND MAIN RESULTS: Hepatic dysfunction, observed after 60 mins of ischemia/reperfusion, showed a reduction in bile flow. The ischemia/reperfusion-induced decrease in bile flow was prevented by administration of gabexate mesilate. Serum transaminase concentrations increased after hepatic ischemia/reperfusion, peaking 12 hrs after reperfusion. Gabexate mesilate significantly inhibited the ischemia/reperfusion-induced increase in serum transaminase levels seen 12 hrs after reperfusion. Although an inactive derivative of factor Xa inhibited the increases in serum levels of fibrin and fibrinogen degradation products 6 hrs after reperfusion, it did not prevent ischemia/ reperfusion-induced liver injury. Hepatic levels of tumor necrosis factor-alpha, rat interleukin-8, and myeloperoxidase were significantly increased after ischemia/reperfusion. These increases were significantly inhibited by gabexate mesilate but unaffected by an inactive derivative of factor Xa. CONCLUSION: Gabexate mesilate reduced ischemia/reperfusion-induced hepatic injury not by inhibiting coagulation, but by inhibiting leukocyte activation.

Analysis of Variance↗
PubMed1999

Identification of amino acid residues of the T-cell epitope of Mycobacterium tuberculosis alpha antigen critical for Vbeta11(+) Th1 cells.

Stimulation of Mycobacterium tuberculosis-primed lymph node cells from C57BL/6 mice with alpha antigen (also known as antigen 85B and MPT59) induced cell proliferation, production of interleukin 2 and gamma interferon, and expansion of Vbeta11(+) CD4(+) T cells in conjunction with antigen-presenting cells in an I-A(b)-restricted manner. Using a series of 15-amino-acid peptides that overlapped each other by 5 amino acids and spanned the mature alpha antigen, we identified the antigenic epitope for alpha antigen-specific Vbeta11(+) Th1 cells. That peptide (peptide-25), which corresponds to amino acid residues 240 to 254 of alpha antigen, contains a motif that is conserved in I-A(b) and requires processing by antigen-presenting cells. Using peptide-25-reactive Vbeta11(+) T-cell clones and substituted peptide-25 mutants, we determined which amino acid residues within peptide-25 were critical for T-cell receptor (TCR) recognition. Our results showed that the amino acid residues at positions 245, 246, 248, 250, and 251 are important for recognition of TCRVbeta11 and that residues at positions 244, 247, 249, and 252 are I-A(b) contact residues. We also observed that active immunization of C57BL/6 mice with peptide-25 can lead to decreased bacterial load in the lungs of M. tuberculosis H37Rv-infected mice. These results should provide us with a useful tool for delineating the regulation of Vbeta11(+) Th1-cell development during M. tuberculosis infection and for developing a vaccine inducing a Th1-dominant immune response.

Acyltransferases↗
PubMed1999

Establishment and characterization of a CD95 (Fas/Apo-1)-negative myeloma cell line.

Although expression of CD95 (Fas/Apo-1) on myeloma cells has been reported, its significance is not clearly understood. We established a myeloma cell line, KHM-11ad (11ad), from a parental cell line, KHM-11, by collecting cells adhered to a plastic dish. KHM-11 cells have been reported to be positive for CD45 and CD95 (Fas/Apo1), and negative for a myelomonocytic antigen, CD13. Interestingly, CD95 was not detected in 11ad. Expression of CD45 was also significantly decreased in 11ad cells while expression of CD13 was detected in these cells. The growth rate of 11ad cells was 1.7 times lower than that of KHM-11 cells. Analysis of adhesion molecules showed that expression of VLA4 and CD44 was significantly suppressed in 11ad. The IC50 of melphalan (L-PAM) for 11ad cells was 50 times higher than that for KHM-11, indicating that 11ad is significantly refractory to L-PAM than KHM-11 cells. Induction of apoptosis by doxorubicin and cycloheximide was suppressed in 11ad cells compared with those in KHM-11 cells. Western blot analysis for Bcl-2 family of proteins showed that Bax was expressed at a 2.2 times lower level in 11ad cells than in KHM-11 cells while there was no difference in expression of Bcl-2, Bcl-Xs nor Bcl-XL. These results suggest that CD95-negative myeloma cells may have characteristics as follows: (1) slow proliferation; (2) low sensitivity to apoptosis; (3) low expression of VLA4, CD44 and Bax. Although these intraclonal variations were based on the findings of cell lines, these may reflect similar variations in vivo. The 11ad line may be a suitable model for analyzing intraclonal variation of myeloma cells.

Antigens, Surface↗
PubMed1999

A plasma cell leukemia patient showing bialleic 14q translocations: t(2;14) and t(11;14).

We report here an IgG/lambda-type plasma cell leukemia patient showing bialleic 14q32 translocations. All immunoglobulins were suppressed in this patient, but a small amount of monoclonal IgG was detected by immunoelectrophoresis. Two cells of six peripheral blood mononuclear cells showed 46,XY,t(2;14)(q11;q32), i(8)(q10), t(11;14)(q13;q32), del(12)(q13.1) by karyotypic analysis. We confirmed the juxtaposition of IgH and PRAD1/Cyclin D1 genes by fluorescent in situ hybridization and overexpression of the PRAD1/Cyclin D1 gene, but Southern analysis showed no bcl-1 rearrangement. We analyzed the t(2;14)(q11;q32) using DNA fragments derived from childhood B-chronic lymphocytic leukemia cases bearing t(2;14)(p13;q32). Southern and Northern analyses demonstrated no alteration of these genes, indicating that this t(2;14) was different from that of childhood B-chronic lymphocytic leukemia. At the IgH loci, Southern analysis showed two rearranged bands and one germ-line band of JH. Cmicro was deleted on one rearranged allele but remained on the other, suggesting that the chromosome translocation occurred after productive class switch recombination on the Cmicro deleted allele.

Alleles↗
PubMed1999

Aromatase and intracrinology of estrogen in hormone-dependent tumors.

Aromatase (estrogen synthetase) has been shown to occur in various extragonadal tissues as well as gonadal tissues, to be tissue-specifically regulated by various factors, and to play important roles in physiological functions in various tissues. The human gene is revealed to contain multiple variants of exon 1 which are tissue-specifically selected. Each exon 1 is flanked with a unique promoter region, which may explain tissue-specific transcription. All results strongly support an idea of intracrinology of estrogen. The validity of this concept is verified in connection with plasma levels of various steroid hormones and association constants of aromatase and estrogen receptor. 17beta-Estradiol is locally produced through several metabolic pathways. In this context, aromatase, steroid sulfatase, estrogen sulfotransferase, and 17beta-hydroxysteroid dehydrogenases I and II are considered to be important factors in the development of hormone-dependent tumors.

17-Hydroxysteroid Dehydrogenases↗
PubMed1999

Aromatase deficiency caused by a novel P450arom gene mutation: impact of absent estrogen production on serum gonadotropin concentration in a boy.

We identified a new point mutation in the CYP19 gene responsible for aromatase (P450arom) deficiency in a 46,XY male infant with unremarkable clinical findings at birth. This boy is homozygote for a 1-bp (C) deletion in exon 5 of the aromatase gene causing a frame-shift mutation. The frame-shift results in a prematurely terminated protein that is inactive due to the absence of the functional regions of the enzyme. Aromatase deficiency was suspected prenatally because of the severe virilization of the mother during the early pregnancy, and the diagnosis was confirmed shortly after birth. Four weeks after birth, the baby boy showed extremely low levels of serum estrogens, but had a normal level of serum free testosterone; in comparison with the high serum concentration of androstenedione at birth, a striking decrease occurred by 4 weeks postnatally. We previously reported elevated basal and stimulated FSH levels in a female infant with aromatase deficiency in the first year of life. In contrast, in the male infant, basal FSH and peak FSH levels after standard GnRH stimulation tests were normal. This finding suggests that the contribution of estrogen to the hypothalamic-pituitary gonadotropin-gonadal feedback mechanism is different in boys and girls during infancy and early childhood. In normal girls, serum estradiol concentrations strongly correlate with circulating inhibin levels, and thus, low inhibin levels may contribute to the striking elevation of FSH in young girls with aromatase deficiency. In contrast, estradiol levels are physiologically about a 7-fold lower in boys than in girls, and serum inhibin levels remain elevated even though levels of FSH, LH, and testosterone are decreased.

Androstenedione↗
PubMed1999

[Adenosine reduces ischemia/reperfusion-induced liver injury by inhibiting leukocyte activation].

To examine whether adenosine reduces ischemia/reperfusion (I/R)-induced liver injury by inhibiting leukocyte activation via A2 receptor (A2R), we investigated the effects of adenosine and YT-146, selective A2A receptor (A2AR) agonist, on I/R-induced liver injury in rats. Adenosine and YT-146, in the range of concentrations of 10(-7)-10(-5) M, significantly inhibited the formyl-methionyl-leucyl-phenylalanine (fMLP)-induced neutrophil elastase release from isolated neutrophils by about 35% in vitro. Adenosine and YT-146, in the range of concentrations of 10(-7)-10(-5) M, significantly inhibited the endotoxin-stimulated TNF-alpha production by monocytes to less than 50% of the control. Although ZM241385, a selective A2AR antagonist, significantly enhanced the fMLP-induced neutrophil elastase release in isolated neutrophils in vitro, this agent did not affect the endotoxin-stimulated TNF-alpha production by monocytes. Male Wistar rats were subjected to complete ischemia of median and left lobes of liver for 60 min and the subsequent reperfusion. Serum levels of transaminases increased over time after hepatic I/R, peaking at 12 hrs after reperfusion. Intravenous infusion of Adenosine (1 and 10 mg/kg/hr) and YT-146 (0.1 and 1 mg/kg/hr) significantly inhibited the I/R-induced increases in serum transaminase levels 12 hrs after reperfusion. The I/R-induced decrease in hepatic tissue blood flow was significantly inhibited by adenosine and YT-146. Hepatic levels of TNF-alpha, cytokine-induced neutrophil chemoattractant (a member of interleukin-8), and myeloperoxidase were significantly increased after I/R. These increases were significantly inhibited by administration of adenosine and YT-146. However, ZM241385 did not reduce the I/R-induced liver injury and it inhibited neither the decrease in hepatic tissue blood flow, nor the indicators of leukocyte activation. These findings suggest that adenosine may reduce I/R-induced liver injury mainly by inhibiting hepatic TNF-alpha production via A2AR, thereby reducing neutrophil activation.

Adenosine↗
PubMed1999

Aromatase and sex steroid receptors in human vena cava.

Among sex steroids, especially estrogen metabolism has been considered to play a role in the function and pathology of human veins. We investigated the expression and activity of the estrogen-producing enzyme aromatase and estrogen receptor (ER) in human vena cava to assess possible in situ biosynthesis of estrogens and their modes of action. We first examined aromatase expression by immunohistochemistry in human inferior vena cava obtained from 29 autopsy cases (11 males, 18 females, 63.6 +/- 3.0 years old). We then semiquantitated the level of aromatase mRNA by reverse transcriptase-polymerase chain reaction in 24 cases and aromatase activity by 3H-water assay in 15 cases to examine whether or not and in which cell types aromatase was expressed. We also studied alternative use of multiple exon 1s of its gene and immunolocalization of 17beta-hydroxysteroid dehydrogenase type I (17beta-HSD I), which converts estrone produced by aromatase to estradiol, a biologically active estrogen and ER. Aromatase and 17beta-HSD I immunoreactivity were both detected in smooth muscle cells (SMC) of the media in all the cases and in endothelial cells (EC) in 20 and 22 cases, respectively. ER immunoreactivity was detected in SMC of vena cava in 21 cases. The amount of aromatase mRNA was significantly greater in the cases utilizing 1c (I.3) or 1d (P.II) of exon 1 (9 cases, 191.1 +/- 26.3 attomol/ng total RNA) than those utilizing 1b (I.4) as the promoter (14 cases, 50.6 +/- 13.0 attomol/ng total RNA) (p < 0.01). Significant correlation (p < 0.05) was observed between the amount of aromatase mRNA and aromatase activity in 15 cases examined. No significant correlation was detected between the amount of aromatase mRNA or aromatase labeling index and the ER status. These results suggest that estrone and estradiol are produced in the human vena cava and that their production is mediated by aromatase and 17beta-HSD I, respectively but not all of these locally synthesized estrogens may not work directly in situ.

17-Hydroxysteroid Dehydrogenases↗
PubMed1999

Aromatase inhibitors and enzyme stability.

The effects of two steroidal (4-hydroxyandrostenedione and atamestane) and three non-steroidal (fadrozole, vorozole, and pentrozole) aromatase inhibitors on the levels of aromatase mRNA and protein were examined in vitro and in vivo. Immunocytochemistry revealed increased quantities of immunoreactive aromatase in human choriocarcinoma-derived JEG-3 cells in response to pretreatment with the non-steroidal inhibitors. To elucidate this effect in detail, aromatase protein in JEG-3 cells after treatment with various inhibitors was quantified using an enzyme-linked immunosorbent assay (ELISA). A time-dependent increase in aromatase protein in the cells was observed with all the aromatase inhibitors except 4-hydroxyandrostenedione, whereas aromatase mRNA levels in the cells remained unchanged during the inhibitor treatment. The three non-steroidal agents caused an approximately fourfold increase in aromatase protein in the cells 24 h after the treatment, as compared with untreated controls. The increase in aromatase protein in the cells was not blocked by treatment with cycloheximide, an inhibitor of protein synthesis. The inhibitors also appeared to block the rapid degradation observed in JEG-3 cells after induction by forskolin. In vivo, daily injection of the inhibitors into adult female mice caused increases in levels of both aromatase mRNA and protein in the ovary. The increase in aromatase mRNA in this in vivo study could be explained by an increase in gonadotropin concentrations in response to decreased plasma concentrations of estrogens. In conclusion, we suggest that aromatase inhibitors increase aromatase protein through stabilization and reduced protein turnover.

Animals↗
PubMed1999

The present state and future prospects of occupational health in Bangladesh.

Bangladesh is a relatively young and developing country. At the present time, like in most developing countries, a clear demarcation between occupational health care and general medical care is difficult to be recognized in Bangladesh. Occupational health is a fairly new field, as the country is undergoing industrialization and occupational health activities are operated by several ministries, such as Labour, Health, Industry and Transport. Legal foundations of the occupational health-care system based on British India and Pakistani era, were adopted and amended by the Government of Bangladesh after the liberation of the country in 1971. Most of the Labour laws have been rectified by the Government of Bangladesh according to the ILO Conventions. Reconsideration of the occupational health service system avoiding duplication for the 'occupational health' component in several ministries might be helpful to achieve the successful provision of an occupational health service in the developing Bangladesh.

Bangladesh↗
PubMed1999

Heart rate variability in response to psychological test in hand-arm vibration syndrome patients assessed by frequency domain analysis.

To investigate heart rate variability in response to psychological tests (Japanese version of Stroop color word test and mirror drawing test) in 29 hand-arm vibration syndrome (HAVS) patients, 16 of them with vibration-induced white finger (VWF) and 13 without VWF, and 10 healthy controls of similar age, heart rate variability during spontaneous and deep (6 cycles a minute) breathing in supine position before and after exposure to the psychological tests was examined calculating frequency domain components such as low frequency (LF) power-index of both the sympathetic and parasympathetic activity, high frequency (HF) power-index of the parasympathetic activity and LF/HF-index of the sympathovagal balance. The group of all patients and the group without VWF indicated significant increase in LF/HF in the deep breathing measurement after exposure to the psychological tests. The result suggests that the sympathetic tone in the sympathovagal balance predominated in the HAVS patients which means that they had larger sensitivity of the sympathetic nervous system to the psychological tests.

Aged↗
PubMed1999

Aromatase and estrogen receptor immunoreactivity in the coronary arteries of monkeys and human subjects.

OBJECTIVE: The objective of this study was to determine whether estrogen could be formed locally in the coronary arteries. DESIGN: Coronary arteries were examined from monkeys (Macaca fascicularis, one male and one female) and human subjects (one premenopausal woman, one postmenopausal woman, and one man) by immunocytochemistry, using purified antisera against human placental estrogen synthetase (aromatase) and ER alpha. The arteries were graded for the amount of atherosclerosis. RESULTS: There was clear immunopositivity for both aromatase and estrogen receptors in all arteries studied. Although all endothelial cells (CD31 positive) stained for both antigens, the staining in macrophages, fibroblasts, and smooth muscle cells was irregular. CONCLUSION: The present results provide the first evidence for the local formation of estrogen in the coronary arteries. In addition to complementing the evidence of a cardioprotective effect of estrogen on the coronary circulation, our results highlight the potential importance of local regulation of estrogen formation and the role of available precursor androgens in maintaining the cardiovascular system.

Animals↗
PubMed1999

[Myelodysplastic syndrome after autologous peripheral blood stem cell transplantation for adult acute myelogenous leukemia].

A 65-year-old woman was admitted to our hospital with arthritis and agranulocytosis. She had given a diagnosis of acute myelogenous leukemia (FAB classification M1) a year ago and treated with 3 cycles of cytarabine and anthracycline or etoposide for 4 months, achieving complete remission state. Her bone marrow aspirate revealed normocellularity with normal karyotype (46, XX [20]) without apparent dysplastic feature at this time. She received autologous peripheral blood stem cell transplantation subsequently after conditioning regimen consisted of granulocyte colony-stimulating factor, busulfan, Ara-C and etoposide. Three months later, she started to manifest low grade fever, polyarthralgia and agranulocytosis and she admitted to our hospital after nine months. Bone marrow aspirate revealed marked hypocellularity with dysplastic features in three series of hematopoietic cells. Arthritis was dramatically improved after administration of prednisolone, but low granulocyte count continued. Bone marrow aspirate revealed karyotypic abnormality with monosomy 7 and we diagnosed her as myelodysplastic syndrome. Chemotherapy-induced myelodysplasia has been reported so far. This case would represent secondary myelodysplastic syndrome after chemotherapy. We could not clarify the etiology for polyarthritis but could be one of the paraneoplastic syndrome. We should note subsequent occurrence of myelodysplasia when planning treatment schedule.

Aged↗
PubMed1998

Identification by site-directed mutagenesis of amino acid residues in ribosomal protein L2 that are essential for binding to 23S ribosomal RNA.

The ribosomal protein L2 (BstL2) from Bacillus stearothermophilus is a primary 23S rRNA binding protein. We made use of site-directed mutagenesis to identify essential basic and aromatic amino acid residues for 23S rRNA binding. Four mutants, R68Q, K70Q, R86Q, and R155Q, in which Arg-68, Lys-70, Arg-86, and Arg-155, respectively, are replaced by the Gln residue. showed reduced binding affinities as compared with that of the wild type BstL2 (a binding constant K=8.93 microM(-1)): K values of these mutants range between 0.24 and 1.86 microM(-1). As for aromatic amino acids, replacements of Phe-66, Tyr-95 or Tyr-102 by alanine significantly abolished the binding affinities. CD analysis of the mutant proteins indicated that the mutations of four basic residues (Arg-68, Lys-70, Arg-86 and Arg-155) did not affect protein structure, whereas those of aromatic residues (Phe-66, Tyr-95, and Tyr-102) appeared to cause slight structural perturbations. These results, together with sequence comparison of L2 family proteins, suggest that Arg-86 and Arg-155 in BstL2 may act as positively charged recognition groups for negatively charged phosphate backbone of the 23S rRNA, and that Phe-66, Tyr-95, and Tyr-102 may be candidate residues which stabilize the BstL2-23S rRNA interaction through intramolecular interactions.

Alanine↗
PubMed1998

Disruption of sexual behavior in male aromatase-deficient mice lacking exons 1 and 2 of the cyp19 gene.

Aromatase is known to be involved in the control of morphological, neuroendocrine, and behavioral sex differences in the brain. To study the detailed functions of aromatase in mouse brain, we generated aromatase-deficient mice that lack exons 1 and 2 and the proximal promoter region of the cyp19 gene by homologous recombination. The genotypes of offspring from heterozygous matings conformed to the Mendelian rule. The aromatase knockout (ArKO) male and female mice displayed no significant external morphological abnormalities, although the uteri of adult ArKO female mice were remarkably underdeveloped. Male-typical sexual behavior directed toward females was, however, strongly modified in the ArKO male mice. The ArKO male mice showed significantly prolonged latencies to mount and decreased numbers of mounts in response to receptive stimulus females. These results suggest an important participation of the aromatase gene in the reproductive behaviors.

Animals↗