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Biomedical subjects

K Inagaki

Publications and source records attributed to K Inagaki.

At least 145 records · Page 8Linked to original sources
PubMed1987

Ischemic damage prevention by coenzyme Q10 treatment of the donor before orthotopic liver transplantation: biochemical and histologic findings.

This study was undertaken to determine whether pretreatment of the donor rat with coenzyme Q10 (CoQ10) would protect against hepatic ischemia induced for 30 minutes at normothermic body temperature. Fresh liver transplants were used as controls (minus warm ischemia of 30 minutes) and gave a 1-week survival rate of 84.6%. CoQ10 was administered intravenously (10 mg/kg body weight) to the donor rat 1 hour before induction of warm ischemia (group A). In another group (B), the same dose was given intravenously not only to the donor rat but also to the recipient rat 1 hour before grafting. None of the placebo group survived more than 2 days. The 1-week survival rates of the groups pretreated with CoQ10 were 45.5% for group A and 50% for group B. There was no significant difference between groups A and B. A statistically significant difference was demonstrated between the placebo group and both CoQ10-treated groups (p less than 0.05). It was therefore assumed that CoQ10, accumulated in the donor liver, was a primary factor in improving survival. Serum glutamic oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), serum alkaline phosphatase (SALP), total bilirubin, and total protein were measured by means of light and electron microscopic examination of the liver 6 months after transplantation. Long-term-surviving rats with transplanted, ischemically damaged liver that was pretreated with CoQ10 showed a decrease in the activity of SGOT and SGPT and an increase in levels of total protein to the normal range (as well as to those levels exhibited by fresh-liver-transplanted rats) with practically no change in levels of SALP, total bilirubin, or in histologic findings. These results indicate that donor pretreatment with CoQ10 is useful for increasing survival after warm ischemic damage of rat liver grafts.

Alanine Transaminase↗
PubMed1986

Thermostable alanine racemase from Bacillus stearothermophilus: molecular cloning of the gene, enzyme purification, and characterization.

The alanine racemase (EC 5.1.1.1) gene of a thermophilic bacterium, Bacillus stearothermophilus, was cloned and expressed in Escherichia coli C600 with vector plasmid pICR301, which was constructed from pBR322 and the L-alanine dehydrogenase gene derived from B. stearothermophilus. A coupled assay method with L-alanine dehydrogenase and tetrazolium salts was used to detect visually the alanine racemase activity in the clones. Alanine racemase overproduced in a clone carrying the plasmid pICR4, 12 kilobases of DNA, was purified from cell extracts about 340-fold to homogeneity by five steps including heat treatment. The overproduced enzyme was confirmed to originate from B. stearothermophilus by an immunochemical cross-reaction with the enzyme of B. stearothermophilus. The purified enzyme has a molecular weight of about 78 000 and consists of two identical subunits of Mr of 39 000. At the optimum temperature (50 degrees C), the enzyme has a specific activity of 1800 units/mg (Vmax, D- to L-alanine). Resolution and reconstitution experiments together with the absorption spectrum of the enzyme clearly indicate that alanine racemase of B. stearothermophilus is a pyridoxal 5'-phosphate enzyme.

Alanine Racemase↗
PubMed1986

Time-dependent inhibition of Bacillus stearothermophilus alanine racemase by (1-aminoethyl)phosphonate isomers by isomerization to noncovalent slowly dissociating enzyme-(1-aminoethyl)phosphonate complexes.

An alanine racemase encoded by a gene from the thermophilic Gram-positive bacterium Bacillus stearothermophilus is overproduced to 0.3% of the soluble protein when carried on plasmid pICR4 in Escherichia coli [Inagaki, K., Tanizawa, K., Badet, B., Walsh, C. T., Tanaka, H., & Soda, K. (1986) Biochemistry (third paper of four in this issue)]. Purification of large quantities (50 mg) of racemase permits study of time-dependent inactivation by D and L isomers of the antibacterial (1-aminoethyl)phosphonate (Ala-P), the phosphonate analogue of alanine. The time-dependent activity loss by this compound now appears general to Gram-positive but not to Gram-negative racemases [Badet, B., & Walsh, C. (1985) Biochemistry 24, 1333] and is shown to occur by extremely slow dissociation of a noncovalent E X Ala-P complex. Ala-P binds initially in a weak, reversible (KI = 1 mM) competitive manner but is slowly isomerized (kinact = 6-9 min-1) to a stoichiometric enzyme complex, which in turn dissociates extremely slowly, with a half-time about 25 days. Thus, Ala-P is a slow but not a tight-binding inhibitor. The E X Ala-P complex is not reducible by borohydride but does perturb the fluorescence of bound pyridoxal 5'-phosphate coenzyme. Determination of the sequence of an active site octapeptide of the B. stearothermophilus alanine racemase shows homology with the sequence of a Gram-negative Salmonella typhimurium alanine racemase that is not susceptible to time-dependent inhibition by Ala-P. Studies with Ala-P analogues suggest the phosphonate dianion is crucial for stable formation of an isomerized long-lived E X Ala-P-inhibited complex.

Alanine Racemase↗
PubMed1985

A rapid isolation technique of unmodified human T cells on a polystyrene resin column.

An easy and rapid isolation technique of human T cells on a polystyrene resin particle column has been developed. The cells of the effluent fraction contained more than 90% sheep erythrocyte (SRBC) rosette-forming cells and less than 1% of cells bearing surface immunoglobulin (Ig) or peroxidase. The T cell (SRBC rosette-forming cells) recovery rate was 80%. The distribution of OKT antigen T cell subsets was essentially the same as that of T cells separated by rosette sedimentation. Cell functions such as tritiated thymidine uptake by T cells and helper activity in Ig production were also the same as that of T cells separated by SRBC rosette sedimentation. Natural killer-like activity of the T cells isolated by the present method increased more than that of T cells obtained by the conventional method. Moreover, it was free from functional modification which tends to result from stimulation such as by the SRBC antigen in the SRBC centrifugation method. The combination of a T cell population offered by the present method and B cells depleted of SRBC-binding B cells minimized background plaque formation and enabled us to quantify the plaque-forming cell number in an antigen-specific plaque-forming assay. Furthermore, these populations produced relatively pure interleukin 2 (IL 2) by stimulation of an autologous mixed lymphocyte reaction without any absorption of IL 2 produced in the same culture. It seemed to be useful to evaluate the ability of lymphocytes from normal individuals and patients to produce IL 2.

Antigens, Surface↗
PubMed1985

Hydrophobic interaction in Candida albicans and Candida tropicalis adherence to various denture base resin materials.

The effects of hydrophobicities of substrate surfaces on microbial adherence were examined by using Candida albicans and Candida tropicalis and 21 denture base resin materials. With increasing surface free energy of resin plates, increasing adherence of C. albicans and decreasing adherence of C. tropicalis were observed. The surface free energy of C. albicans is higher than that of all resin material surfaces, and C. tropicalis has surface free energy lower than that of all materials used. In calculation of the changes of free energy accompanying the adherence, the higher adherence tendency was accompanied by a lower value for the free energy change in both species. From a different standpoint, the closer the surface free energy of the substrate surface and the microorganism, the higher was the probability of adherence.

Candida↗