Search PubMed⌕ Search

Biomedical subjects

J Watanabe

Publications and source records attributed to J Watanabe.

At least 217 records · Page 12Linked to original sources
PubMed1995

Effects of various natural antioxidants on the Cu(2+)-mediated oxidative modification of low density lipoprotein.

We have reported in our previous paper that several flavan-3-ol derivatives (tea polyphenols) inhibited the Cu(2+)-mediated low density lipoprotein (LDL) oxidation in vitro. (-)-Epigallocatechin gallate (EGCG), in particular, exhibited strong inhibition. In this study, we have compared the antioxidative effects of EGCG with those of other natural antioxidants, such as flavonols, sesaminol, curcuminoid derivatives, tocopherol analogues and theaflavins. The antioxidative effects were monitored by conjugated diene formation in LDL which was carried out at 37 degrees C with 5 microM CuSO4 with or without antioxidants. Dibutyl hydroxytoluene (BHT) was used as a reference compound. The lag-time before the onset of conjugated diene formation was more than 100 min in the presence of 0.5 microM EGCG, theaflavin, myricetin, quercetin, and sesaminol. The ability to prolong the lag-time was in the order of sesaminol > quercetin > EGCG > theaflavin > or = myricetin > BHT > alpha-tocopherol. Among the 4 tocopherol analogues used, alpha-tocopherol showed the strongest antioxidative activity. We have also studied the effects of EGCG, BHT, and alpha-tocopherol on cholesteryl and alpha-tocopherol on cholesteryl ester (CE) degradation and apolipoprotein B 100 (apo B 100) fragmentation in the Cu(2+)-mediated oxidative modification of LDL. EGCG was the most effective inhibitor of CE degradation and apo B 100 fragmentation.

Antioxidants↗
PubMed1995

Uptake of low molecular weight fractionated [3H]heparin by rat hepatocytes in the primary culture.

The uptake of low molecular weight fractionated [3H]heparin (LMWFH: 7000 Da) was examined, for comparison with that of high molecular weight fractionated [3H]heparin (HMWFH:20000 Da), in a primary culture of rat hepatocytes. The uptake of LMWFH increased almost linearly with time up to 60 min (extended uptake), although a faster uptake was observed in the initial 2 min (initial uptake). Both the initial and extended uptake were saturable, and the maximum uptake velocity (Vmax) and the Michaelis constant (Km) were estimated to be 10.7 pmol/min/mg protein and 398 nM, respectively, for the initial uptake and 0.34 pmol/min/mg protein and 116 nM, respectively, for the extended uptake. The Km for the extended uptake was 5 times larger than that of 21 nM for HMWFH, but the other parameters were comparable with those for HMWFH. Thus, an increase in Km, or a decrease in the apparent affinity, with a decrease in molecular weight in the extended uptake may be responsible for the reported lower hepatic uptake of low molecular weight heparin, compared with unfractionated heparin. It was also shown that both the initial and the extended uptake of LMWFH were inhibited by several analogs of heparin, including HMWFH, and anionic compounds such as 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), suggesting that LMWFH and HMWFH, in spite of a large difference in the molecular weight, share the same specialized uptake mechanism, in which an anionic moiety and/or heparin-like structure plays an important role.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1995

Influence of anesthetic regimens on the intestinal absorption of 5-fluorouracil in rats.

We investigated the influence of anesthetic regimens on the intestinal absorption of 5-fluorouracil (5-FU), which is known to be absorbed by concurrent Na(+)-dependent, carrier-mediated transport and passive transport, in single-pass perfusion experiments in rats. Compared with the absorption in unanesthetized rats, the regular dose of urethane (1.13g/kg) reduced the maximum transport rate (Jmax), the Michaelis constant (Km) and the membrane permeability coefficient of passive transport (P m,d); a low dose of urethane (0.7g/kg) reduced Jmax and Kmax, but did not affect Pm,d; pentobarbital sodium (50 mg/kg) increased Jmax without affecting Km, and reduced Pm,d. The reductions in Jmax and Km were comparable for the regular and low doses of urethane. Thus, urethane and pentobarbital, which have been most commonly used in laboratory animal experiments, exerted qualitatively different effects on the carrier-mediated transport of 5-FU, although they similarly inhibited the passive transport. For urethane, the effect on the passive transport was avoided by reducing the dose, but the effect on the carrier-mediated transport was not. This influence of anesthetic regimens on intestinal drug absorption may not be easily scaled for normalizing absorption data. When compiling them for such purposes as establishing in situ-in vivo quantitative correlation, the absorption data in perfusion (in situ) should be categorized on the basis of anesthetic regimens, to avoid ending up with poor outcomes. We also examined the effect of urethane on the exsorption of Na+ in the intestinal loop where Na+-free buffer was introduced, and found a minimal effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Anesthesia↗
PubMed1995

Ras amplification in BHK-21 cells produces a host cell line for further rapid establishment of recombinant protein hyper-producing cell lines.

To rapidly establish recombinant protein hyper-producing cell lines we introduced a reporter plasmid into BHK-21 cells that had been 'primed' by transfection and amplification of the ras oncogene. The reporter plasmid used carries the human interleukin-6 (hIL-6) gene which is under control of the human cytomegalovirus immediate early promoter. The primed BHK cell lines were shown to produce many stable hIL-6 hyper-producing cells achieving about 15 times higher productivity than the control BHK-21 cells.

Animals↗
PubMed1995

E1A and ras oncogenes synergistically enhance recombinant protein production under control of the cytomegalovirus promoter in BHK-21 cells.

The amplified ras oncogene greatly enhanced the production of recombinant human interleukin-6 (hIL-6) under control of the cytomegalovirus immediate early promoter (CMV promoter) in BHK-21 cells. When the adenovirus E1A oncogene was further transfected into the above mentioned ras-amplified hIL-6 hyperproducing BHK cells, the transfectants had about 10 times higher productivity than non-transfectants. However, the E1A gene alone did not enhance productivity. These results implicate a ras and E1A synergistic ability that acts to enhance hIL-6 production.

Adenoviridae↗
PubMed1995

Glycation accelerates the oxidation of low density lipoprotein by copper ions.

We investigated the in vitro effect of glycation on LDL oxidation. Native LDL (nLDL) was glycated in 0, 5, 10, or 20 mM glucose. This glycated LDL (gLDL) was oxidized by 1 microM copper ion. Compared to nLDL and gLDL, oxidized gLDL (ogLDL) has a greater negative charge. The thiobarbituric acid reactive substance (TBARS) value of ogLDL increased with the glucose concentration tested during glycation in a dose-dependent manner. OgLDL glycated in 20 mM glucose had a significantly higher TBARS level than did oxidized LDL incubated without glucose. In conclusion, LDL glycated in vitro is prone to oxidation. Thus, glycated LDL, which increases in the diabetic state, may contribute to the pathogenesis of atherosclerosis in diabetic patients.

Copper↗
PubMed1995

[Combined immunotherapy using interferon-alpha, interleukin-2 and lymphokine-activated killer cells--improvement of quality of life in patients with advanced renal cell carcinoma].

The goal of any treatment strategy for cancer is to improve not only patient survival but also quality of that survival. Between March 1990 and February 1993, we treated 10 patients with advanced RCC (9 men and 1 woman) by combined immunotherapy using natural interferon-alpha (IFN-alpha), recombinant interleukin-2 (rIL-2) and lymphokine-activated killer (LAK) cells, and resulting the quality of life (QOL) issues examined. The ages of the patients ranged from 36 to 78 years (mean: 60.2) and the performance status (PS) ranged from 30 to 100% (mean: 77%). There were 8 lung, 3 bone, 2 brain and 1 neck and para-aortic lymph node metastases. We could evaluate 8 patients, 2 patients dropped out because of bone fracture and acute pneumonia. The protocol was as follows; 1 x 10(6) IU of rIL-2 as an intravenous infusion and 6 x 10(6) IU of IFN-alpha intramuscularly on days 1-7 and 15-21. In additions LAK cells obtained from the patients were given on days 14, 21, 28, and 35 intravenously. This protocol was repeated for more than three cycles (mean: 4.13 cycles) in each patient. The maintenance therapy on outpatient basis were performed in 4 patients after confirmation of the safety of the combined immunotherapy. This outpatient regimen was composed of 1 x 10(6) IU of rIL-2 intravenously, 6 x 10(6) IU of IFN-alpha intramuscularly on days, 1, 8, 15, 22, and 29, plus LAK cells on days 15 and 29. We repeated this protocol for 3-5 cycles (mean: 4.25 cycles).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Monamidocin, a novel fibrinogen receptor antagonist. I. Production isolation, characterization and structural elucidation.

Monamidocin, a novel fibrinogen receptor antagonist, has been isolated from the culture broth of Streptomyces sp. NR 0637 BY carbon adsorption, n-BuOH extraction, SP-Toyopearl, silica gel 60 silanised and Sephadex LH-20 column chromatographies and preparative HPLC. The molecular formula of monamidocin has been determined to be C15H22N4O4 from HRFAB-MS and 13C NMR spectral data. The structure of monamidocin has been determined to be N-[(S)-5-guanidino-2-hydroxypentanoyl]-L-phenylalanine by 2D NMR experiments.

Chemical Phenomena↗
PubMed1995

[Collagen synthetic activity following filtration surgery in rabbits].

We evaluated collagen synthetic activity, which plays an important role in wound healing, following experimental filtration surgery in rabbits. Collagen synthetic activity was measured by immunohistochemistry for prolyl 4-hydroxylase beta-subunit and type I procollagen. Trabeculectomy was performed on albino rabbit eyes, with the filtering site harvested on postoperative days 1, 4, 7, 14, and 28. Samples were fixed with 10% buffered formalin for 12 hours and prepared for paraffin section, and each antigen was detected in filtering site tissue using avidin-biotinylated peroxidase complex. Immunoreaction of prolyl 4-hydroxylase beta-subunit or type I procollagen increased from day 4 to 14 and markedly decreased at day 28. These findings show that prolyl 4-hydroxylase and type I procollagen are markedly produced almost simultaneously, and collagen synthetic activity following filtration surgery continues for over 14 days in the process of wound healing.

Animals↗
PubMed1995

[Usefulness of the 24-hour delayed film of upper gastrointestinal series for the diagnosis of sigmoidovesical fistula: report of 3 cases].

Recently, sigmoidovesical fistula is not a rare disease as a result of the change of our food style and increase in the age ratio. However, in general, the preoperative image diagnosis is difficult. We have experienced three cases of sigmoidovesical fistula, examined by barium enema, cystography, upper gastrointestinal series, cystoscopy, colonic fiberscopy and computed tomography. The fistulas were identified preoperatively by the 24-hour delayed film of upper gastrointestinal series. By this method, the regions of sigmoidovesical fistula were identified, and the patients were operated. We concluded that the 24-hour delayed film in upper gastrointestinal series might be useful to diagnose the sigmoidovesical fistula.

Adult↗
PubMed1995

Endoplasmic reticulum proliferates without an increase in cytochrome P-450 in hepatocytes of mice treated with phenobarbital and cobalt chloride.

To determine whether endoplasmic reticulum (ER) proliferation in hepatocytes after phenobarbital (PB) administration relates closely to cytochrome P-450 (P-450) increase, we have measured the amount of total P-450 per unit cytoplasmic volume (P-450 content) by microphotometry and estimated the area of ER per unit cytoplasmic volume (ER area) by morphometry in periportal, midzonal, and perivenular hepatocytes of mice injected daily with PB (100 mg/kg), or with PB (100 mg/kg) plus cobalt chloride (50 mg/kg) for three days. After injection of PB, the P-450 content and ER area increased in hepatocytes of the three sublobular zones. In mice treated with PB plus cobalt chloride, however, the ER area increased, but the P-450 content decreased or remained unchanged in hepatocytes of the three zones. We conclude that cobalt chloride inhibits the increase in total P-450 but has no effect on the proliferation of ER of hepatocytes in mice treated with PB, indicating a dissociation of ER proliferation and P-450 increase after administration of PB.

Animals↗
PubMed1994

Transglycosylation activity of Mucor hiemalis endo-beta-N-acetyl-glucosaminidase which transfers complex oligosaccharides to the N-acetylglucosamine moieties of peptides.

A novel endo-beta-N-acetylglucosaminidase in the culture fluid of Mucor hiemalis isolated from soil was found to have transglycosylation activity. This endo-beta-N-acetylglucosaminidase, Endo-M, could liberate the complex type of asparagine-linked oligosaccharides by hydrolysis of diacetylchitobiose linkage from glycoproteins. The treatment of Endo-M with N-acetyl-glucosamine and asialotransferrin glycopeptide having the complex type of oligosaccharides resulted in the transfer of the released oligosaccharide from the glycopeptide to N-acetyl-glucosamine. The structure of the product after transfer was deduced to be (GlcNAc)2-Man-(Gal-GlcNAc-Man)2 by a combination method of pyridylamination and high performance liquid chromatography, and mass-spectrometry. The enzyme could transfer the complex type of oligosaccharide from asialotransferrin glycopeptide to bovine ribonuclease with the high-mannose type of oligosaccharide. This will lead to the construction of neoglycoproteins containing different types of oligosaccharides.

Acetylglucosamine↗
PubMed1994

Molecular cloning of the human AH receptor gene promoter.

A lambda phage clone containing a promoter region of the human Ah receptor gene was isolated. This clone spanned 13.8 kb and contained the 1st exon, the sequence of which completely matched the reported Ah receptor cDNA. Using RNase protection assay and primer extension analysis, the transcription initiation sites were determined to be 643 and 615 bp upstream of the translational initiation codon ATG. This promoter did not contain a TATA box, while multiple GC boxes were present close to the determined transcription initiation sites. Comparison of the 5' flank sequence of the human Ah receptor with its murine equivalent showed several well conserved regions, containing binding sites for known transcription factors, such as Sp1. The promoter activity was confirmed by transient transfection of chimeric constructs of the Ah receptor gene and reporter gene luciferase into hepatoma HepG2 cells.

Animals↗
PubMed1994

Ras oncogene enhances the production of a recombinant protein regulated by the cytomegalovirus promoter in BHK-21 cells.

In order to enhance recombinant protein productivity in animal cells, we developed the oncogene activated production (OAP) system. The OAP system is based on the premise that oncogenes are able to enhance promoter activity. To this end, we constructed reported plasmids by fusing various promoters to the human interleukin-6 (hIL-6) cDNA, and the effector plasmids by inserting individual oncogenes, for example c-myc, c-fos, v-jun, v-myb and c-Ha-ras, downstream from the human cytomegalovirus immediate early (CMV) promoter. Results of transient expression experiments with BHK-21 cells suggest that the CMV promoter is the most potent promoter examined and that the ras product is able to transactivate the beta-actin, CMV and SR alpha promoters. Recombinant BHK-21 cells producing hIL-6 under the control of the CMV promoter were contransfected with the ras oncogene and dihydrofolate reductase gene, then selected with 50 nM methotrexate to coamplify the ras oncogene. We were able to rapidly establish a stable and highly productive clone which exhibited a 35-times higher production rate as compared to the control value.

Actins↗