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Biomedical subjects

H Okubo

Publications and source records attributed to H Okubo.

At least 73 records · Page 4Linked to original sources
PubMed1987

Sequence analysis of the putative regulatory region of rat alpha 2-macroglobulin gene.

Rat alpha 2-macroglobulin (alpha 2M) is an acute-phase reactant, concentration of which in serum increases more than 100-fold in the course of inflammation. Glucocorticoid and some protein factors such as interleukin 1 (IL-1) have been known to be involved in the regulation of this plasma protein synthesis. To understand the regulatory mechanism of alpha 2M production at the molecular level, we isolated genomic DNA clones of rat alpha 2M gene and characterized the promoter region of the gene by comparing the nucleotide sequence with those of other acute-phase reactant genes. Several possible regulatory signals were identified. Particularly, a sequence (T/A)T(C/G)TGGGA(A/T) was found about at 170 bp upstream from a putative capping site, which was also found in the 5'flanking region of various acute-phase reactant genes.

Animals↗
PubMed1987

Kupffer cell stimulation of alpha 2-macroglobulin synthesis in rat hepatocytes and the role of glucocorticoid.

When stimulated with lipopolysaccharide (LPS), primary cultures of Kupffer cells from the rat secreted a hepatocyte-stimulating factor (Kupffer factor) which induced the synthesis of alpha 2-macroglobulin (a2M) by hepatocytes. The effect of this Kupffer factor was found only when dexamethasone was present in the hepatocyte culture medium. Dexamethasone alone stimulated the a2M synthesis, to some extent, in a dose-dependent manner. When both Dex and the Kupffer factor were added to the medium, cultured hepatocytes synthesized 100-fold a2M, quantitative evidence that explains the in vivo phenomenon in which the serum a2M concentration increases more than 100-fold in the acute phase. A co-culture study with hepatocytes and Kupffer cells indicated that the latter cells have a major function in the induction of a2M synthesis in vivo. A hybridization study with rat a2M cDNA showed that the concentration of mRNA increased in cultured hepatocytes in the presence of the Kupffer factor, but only when dexamethasone was present. From these results we concluded that Kupffer cells secrete a hepatocyte-stimulating factor when stimulated by LPS and that glucocorticoid is essential for the induction of a2M in rat hepatocytes. The effect of the Kupffer factor and glucocorticoid appears to be regulated in the pretranslational, probably the transcriptional, phase.

Animals↗
PubMed1987

[Clinical and histological studies on vaginal reconstruction after radical hysterectomy].

The extensive resection of vagina has a prime importance to obtain the remission in patients with invasive carcinoma of cervix. Maintenance of optimal length, configuration and physiologic function of vagina has been neglected because of malignant nature of lesions. Thus, sexual frigidity coupled with dyspareunia was usually a serious problem faced by postoperative patients. In order to resolve this problem we devised a new attempt to obtain sufficient regeneration of vagina in postoperative patients. The protection procedure for a shortened vagina consists of enlargement of the vaginal vault and promotion of epithelial regeneration. Vaginal length in 145 patients who had received the protection procedure as well as the radical hysterectomy was 7.0 +/- 0.1 cm, being not different from the one in 187 patients with simple hysterectomy. However, significantly shortened length was observed in 21 patients hysterectomized radically who had not the protection. The influence of preserved ovarian functions into the regeneration process of vagina was investigated. The mean length was 7.3 cm in 75 patients with ovarian conservation and 6.3 cm in 7 patients with bilateral oophorectomy. The latter was significantly shorter than the former. However, administration of synthetic estrogens alone was incomplete for the full reconstruction of remnant vagina. Adjuvant radiotherapy was not an obstacle in obtaining the optimal vaginal length. The mean length of irradiated group was 7.4 cm, being not different from the one of the nonirradiated group. Regeneration process of vagina was investigated histologically. New vaginal lumen consisted originally of the remnant vagina and granulation tissues obliterating dead spaces in the pelvic floor. Then, epithelia stained with iodine followed to cover the granulation. Enlargement of such areas directed into the deep part of vault was evident with the passage of time following the operation. As a result, epithelial covering of the whole reconstructed vagina was completed within 2 months after the operation. Those results indicated the clinical availability of the protection procedure in radically hysterectomized patients. Then, ovaries conserved either by orthotopic manner or transposition plays some role for the reconstruction of remnant vagina.

Adult↗
PubMed1986

Human lymphocytes synthesize C-reactive protein.

We obtained evidence for the synthesis and secretion of C-reactive protein (CRP) by peripheral mononuclear cells in culture. Human mononuclear cells isolated from peripheral blood, after depletion of platelets, were cultured in glutamine-depleted RPMI 1640 supplemented with [3H]glutamine in the presence of 10-O-tetradecanoyl-phorbol-13-acetate (TPA). Anti-CRP antiserum was added to the culture medium, and the resultant immunoprecipitate was analyzed in SDS-polyacrylamide gel electrophoresis (SDS-PAGE). The immunoprecipitate consisted of CRP, heavy and light chains of IgG, and only the CRP protein band had radioactivity, indicating that CRP was synthesized by mononuclear cells. In the populations of mononuclear cells, T-cell preparations mainly synthesized CRP, under stimulation of a factor derived from activated monocytes. Studies using the inhibitors of phospholipid metabolism suggested that generation of the monocyte factor was relevant to metabolites of an arachidonate cascade.

B-Lymphocytes↗
PubMed1986

Selective partial IgM deficiency: functional assessment of T and B lymphocytes in vitro.

Seven patients with selective IgM deficiency (SIgMD) were studied for cell surface immunoglobulin (SmIg), T-cell subpopulations, and immunoglobulin (Ig) synthesis in vitro by peripheral blood lymphocytes (PBL). Serum IgM levels were less than 25 mg/dl, while IgA, IgG, and IgD were within normal levels. The patients had respiratory or urinary tract infections and two were diagnosed as having systemic lupus erythematosus (SLE). T/B-cell ratios in PBL were within normal ranges. Percentage ratios of B cells bearing SmIg were normal in five patients and decreased in two; however, normal values were seen after 7 days of culture in the presence of PWM. OKT4/OKT8 ratios decreased in five of seven patients, in whom two were due to a decrease in OKT4 and two to an increase in OKT8 cells. One showed a decrease in OKT4 and an increase in OKT8. Analysis of lymphocyte function for Ig synthesis in vitro, using a coculture of counterpart T and B cells from healthy individuals and patients with SIgMD, revealed that the increased function of IgM isospecific suppressor T cells (Ts) was responsible for the IgM deficiency in all seven patients.

Adult↗
PubMed1986

Expression of rat alpha 2-macroglobulin gene during pregnancy.

Rat alpha 2-macroglobulin (alpha 2M) is a typical acute phase protein, the concentration of which in serum increases more than 100-fold after inflammation. It is also known that the protein increases during pregnant (and neonatal) stages. Using a specific cDNA probe, expression of the alpha 2M gene during pregnancy was studied at the mRNA level. During inflammation, the liver is almost the only organ producing alpha 2M, but during pregnancy the placenta and uterus were found to be major organs producing a large amount (70-80% of that of inflamed liver) of alpha 2M mRNA at days 12-15. The yolk sac, maternal liver and fetal (or neonatal) liver also produced a small but significant amount (5-20% of that of inflamed liver) of the mRNA. Southern blotting analysis showed that only one copy of the alpha 2M gene was present in a haploid rat genome. These results indicated that a single alpha 2M gene has the ability to respond to two completely-different physiological states.

Animals↗
PubMed1986

Polymorphonuclear leukocytes and the induction of alpha 2-macroglobulin synthesis.

Peritoneal exudate-polymorphonuclear leukocytes (PMNLs) secreted a factor, PMN factor (PMN-F), which stimulated hepatocytes to synthesize alpha 2-macroglobulin (a2M), in vitro. The synthesis of a2M was enhanced by the factor in the presence of dexamethasone (Dex), and the effect was dose-dependent. After harvesting from the peritoneal exudate, PMNLs released the factor within 1 hr of the incubation. Physicochemical properties of the factor were as follows: nondialyzable, heat-labile, stable in wide pH change, inactivated by a protease inhibitor, aprotinin, and apparent molecular weight being 12,000-21,000, determined by gel filtration. Treatment of PMNLs with zymosan resulted in increases in the activity of lysosomal enzymes but not that of the factor. These results indicate that the factor is a protease other than lysosomal enzymes.

Animals↗
PubMed1986

[Clinical evaluation of imipenem/cilastatin sodium in gynecological infection].

The MK-0787/MK-0791 is a combination of imipenem (a carbapenem antibiotic) and cilastatin sodium (a dehydro peptidase-I inhibitor) in a 1 to 1 ratio, which produces a higher urinary recovery of imipenem than imipenem alone. The MK-0787/MK-0791 was used in 8 female patients with intrapelvic infections. Clinical efficacies were very good in all the patients. There were neither subjective nor objective side effects nor abnormal laboratory findings.

Adult↗
PubMed1985

Molecular cloning of DNA complementary to rat alpha 2-macroglobulin mRNA.

Rat alpha 2-macroglobulin (alpha 2M) is an acute-phase protein synthesized in the liver. Using an in vitro translation system coupled with solid-phase radioimmunoassay, alpha 2M mRNA activity was found to rise to a maximum level in 16-24 h after turpentine injection. Poly(A)+ RNA from turpentine-injected rat liver was converted to cDNA by the method of Okayama-Berg, and about 50,000 transformants were obtained. From these transformants, clones containing alpha 2M cDNA were selected using the following criteria: 1) alpha 2M cDNA should hybridize with synthetic oligonucleotides encoding portions of the alpha 2M amino acid sequence, 2) alpha 2M cDNA should hybridize preferentially with RNA which increases during inflammation, 3) mRNA which hybridizes with alpha 2M cDNA should encode a polypeptide which specifically reacts with antibody against alpha 2M, and 4) the cDNA should contain the nucleotide sequences encoding the amino acid sequences of alpha 2M. We found clones which fulfilled these criteria. Using the cDNA clone as a probe, we demonstrated that the level of alpha 2M mRNA in the liver of inflamed animal markedly increased up to 1000-fold. The size of the alpha 2M mRNA was about 4800 nucleotides in length by Northern analysis.

Animals↗
PubMed1985

Alpha 2-macroglobulin secretion enhanced in rat hepatocytes by partially characterized factor from Kupffer cells.

Isolated rat Kupffer cells produced a factor which stimulated the synthesis of alpha 2-macroglobulin (alpha 2M) in primary cultured rat hepatocytes. Although Kupffer cells placed in culture produced the factor without stimulation by lipopolysaccharide (LPS), the LPS-stimulated cells produced larger amounts of the factor. On the other hand, the production of the factor was inhibited by addition of actinomycin D. The induction of alpha 2M synthesis by cultured hepatocytes was enhanced in the presence of dexamethasone (Dex), in that hepatic synthesis of alpha 2M increased by addition of the factor alone and with Dex 1.5 and three- to four-fold, respectively. The factor was nondialyzable and stable at 60 degrees C for 30 min. When the factor was fractionated using the molecular sieve method, the activity recovered in the fraction had a molecular weight of over 30,000.

Animals↗
PubMed1985

Influence of acute-phase proteins on the activity of natural killer cells.

The effects of alpha 1-antitrypsin (alpha 1-AT), alpha 1-acid glycoprotein (alpha 1-AGP), and haptoglobin (Hp), the main constituents of alpha-globulin and which belong to acute-phase proteins, on NK activity were examined using K562 cells as the NK target cells. Among the three proteins, alpha 1-AT and alpha 1-AGP had inhibitory effects on NK activity for "fast target" K562 cells. The alpha 1-AT preparations having the same protein concentration and a different trypsin inhibitory capacity (TIC) had an equal effect. Although alpha 1-AT and alpha 1-AGP equally reduced the NK activity, the mechanism involved in the reduction differed, in that the effect of alpha 1-AT directed toward NK cells reduced their binding capacity with the target cells. alpha 1-AGP probably interacts with a cytotoxic factor secreted from NK cells following effector-target interaction. These studies suggest that each of the acute-phase proteins, which increase following inflammation, inhibits NK cell function by two distinct mechanisms.

Albumins↗
PubMed1985

[Effect of iron concentrations on the DNA-degradation reaction by bleomycin].

DNA-degradation reaction by bleomycin was conducted with form I of SV40 DNA as the substrate under "iron-limited conditions" and "iron-sufficient conditions". For "iron-limited conditions", the iron ions in reaction mixtures were derived from contaminant iron in the bleomycin preparation, the molar ratio of bleomycin/iron being 1/0.05. For "iron-sufficient conditions", Fe(II) was added to reaction mixtures to attain the molar ratio of bleomycin/iron at 1/1 to 1/29. DNA-degradation reaction by Fe(II) without bleomycin, or "Fe(II) alone", was also run for comparison. In any case, the reaction was allowed to proceed at such low rates that other products than form II and form III, due to further fragmentation, were negligible. The mass of DNA in each form was quantitatively determined by spectrofluorometry after agarose gel electrophoresis and staining with ethidium bromide. "Iron-limited conditions" and "iron-sufficient conditions" showed similar reaction characteristics as follows. As form I decreased with incubation time, form II increased faster than form III, hit a plateau (ca. 65% of total DNA) and decreased gradually thereafter. Form III kept increasing throughout the incubation. The reaction was dependent on incubation time and on doses of both bleomycin and Fe(II) but rather independent of temperature. The reaction was inhibited by acidic pH, Tiron (a specific chelator of iron) and Cu(II) but not EDTA. Octanucleotides with specific base sequence inhibited the reaction. DNA-degradation reaction by "Fe(II) alone" showed different characteristics as follows. The reaction proceeded rapidly at first, but ceased within 2 minutes. The extent of reaction was dependent on dose of Fe(II). The reaction product was form II accompanied by little amount of form III. The reaction was highly dependent on temperature, inhibited by EDTA, and not inhibited by Cu(II) and acidic pH. Octanucleotides, irrespective of their base sequences, inhibited the reaction.

Base Sequence↗