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Biomedical subjects

M Maeda

Publications and source records attributed to M Maeda.

At least 757 records · Page 42Linked to original sources
PubMed1993

Extracorporeal circulation for tissue transplantation (in the case of venous flaps).

During free-tissue transplantation, recipient vessels are often crushed as a result of trauma or radiation therapy, making anastomosis of vessels difficult. In some cases, a second operation is unavoidable because of formation of a postoperative thrombus at sites of the anastomosis. To solve such problems, it should be possible to transplant tissue anywhere in the body without vascular anastomosis by extracorporeal circulation. This study mainly concerned venous flaps that were spindle-shaped and 4 x 3 cm in size. One hundred and forty-six male Japanese White rabbits, weighing 2.0 to 2.5 kg, were used. Stabilized hemoglobin, lactated Ringer's solution, whole blood, and plasma were used as perfusate. Each of these perfusates was injected at a rate of 5 ml per hour continuously for 3 days in good recipient sites and for 7 days in poor recipient sites. Five rabbits were used in each group. The flaps were examined histologically and electron microscopically. Moreover, in the plasma group, microangiography was performed to observe blood circulation between the flap and the surrounding tissue. The flaps survived in all cases in which the plasma group had good recipient beds but did not survive in any of the other cases, in which degeneration of the endothelial cells was detected prior to edema. According to the results of this examination, tissue viability of the venous flap could be maintained for at least 3 days by perfusion of plasma. During this time, in the cases of good recipient sites, blood may begin to recirculate between the flap and the surrounding tissue.

Angiography↗
PubMed1993

[A case of acute hemorrhagic gangrenous acalculous cholecystitis with bile peritonitis during anti-coagulant therapy after coronary-artery bypass grafting].

Acute acalculous cholecystitis is a relatively rare complication occurring after surgery on organs other than the bile duct system. It is often misinterpreted to be a post-operative symptom, and can progress into a very serious condition with high risk of mortality if gangrene and perforation develop. Its occurrence after open heart surgery is relatively rare. We experienced a case of acute hemorrhagic, gangrenous acalculous cholecystitis that developed after coronary-artery bypass grafting. The patient, a 78-year-old man, complained post-operatively of a right upper abdominal pain. The diagnosis of acute gangrenous acalculous cholecystitis was established on the basis of abdominal sonography and CT, and emergency operation performed was successful. Etiological factors in this case may have included post-operative stasis of bile, swelling of the gallbladder, hypotension during cardiopulmonary bypass, and post-operative anti-coagulant therapy administered after open heart surgery. These factors induced intracystic hemorrhage followed by sudden exacerbation, which resulted in gangrenous cholecystitis followed by perforative biliary peritonitis.

Acute Disease↗
PubMed1992

Escherichia coli F0F1-ATPase. Residues involved in catalysis and coupling.

The molecular biological approach has provided important information toward understanding the complexities of the F0F1 ATPase. This article focuses on our recent results on the ATPase catalytic site contained in the beta subunit and the role of the gamma subunit in regulation of proton transport. We used a combination of affinity labeling and mutagenesis to locate several residues of the alpha and beta subunits in the catalytic site. Adenosine triphosphopyridoxal (AP3-PL) labeled beta Lys-155, beta Lys-201 and alpha Lys-201, suggesting that they are near the gamma-phosphate moiety of ATP. Turning to a mutagenesis approach we demonstrated that the two conserved residues, beta Lys-155 and beta Thr-156 in the glycine-rich sequence, are essential for catalysis. Finally, using pseudorevertant analysis, we positioned residue beta Gly-149 (also in the glycine-rich sequence) in proximity to beta Ser-174, beta Glu-192 (binding site for DCCD), and beta Val-198 (only three residues away from the AP3-PL binding site, beta Lys-201). Genetic studies suggested that the gamma subunit plays a role in regulation of catalysis and its coupling with proton conduction. We found that four mutations in the carboxyl-terminal region (gamma Gln-269-->Leu, gamma Gly-275-->Lys, gamma Thr-277-->end, or frameshift) had similar membrane ATPase activities but different ATP-dependent proton pumping and growth by oxidative phosphorylation. These results suggested a perturbation in the coupling between catalysis and proton translocation. We were able to clearly define the "uncoupling" by introducing mutations in the amino-terminal region of the gamma subunit. We were led to gamma Met-23-->Lys and Arg which resulted in an enzyme still regulated by delta microH+, but with profoundly inefficient coupling between ATPase catalytic sites and proton translocation in both ATP-dependent proton pumping and delta microH(+)-driven ATP synthesis. Second-site mutations in the carboxyl-terminal region of the gamma subunit reversed this effect.

Amino Acid Sequence↗
PubMed1992

Effects of mutations of conserved Lys-155 and Thr-156 residues in the phosphate-binding glycine-rich sequence of the F1-ATPase beta subunit of Escherichia coli.

beta Lys-155 in the glycine-rich sequence of the beta subunit of Escherichia coli F1-ATPase has been shown to be near the gamma-phosphate moiety of ATP by affinity labeling (Ida, K., Noumi, T., Maeda, M., Fukui, T., and Futai, M. (1991) J. Biol. Chem. 266, 5424-5429). For examination of the roles of beta Lys-155 and beta Thr-156, mutants (beta Lys-155-->Ala, Ser, or Thr; beta Thr-156-->Ala, Cys, Asp, or Ser; beta Lys-155/beta Thr-156-->beta Thr-155/beta Lys-156; and beta Thr-156/beta Val-157-->beta Ala-156/beta Thr-157) were constructed, and their properties were studied extensively. The beta Ser-156 mutant was active in ATP synthesis and had approximately 1.5-fold higher membrane ATPase activity than the wild type. Other mutants were defective in ATP synthesis, had < 0.1% of the membrane ATPase activity of the wild type, and showed no ATP-dependent formation of an electrochemical proton gradient. The mutants had essentially the same amounts of F1 in their membranes as the wild type. Purified mutant enzymes (beta Ala-155, beta Ser-155, beta Ala-156, and beta Cys-156) showed low rates of multisite (< 0.02% of the wild type) and unisite (< 1.5% of the wild type) catalyses. The k1 values of the mutant enzymes for unisite catalysis were lower than that of the wild type: not detectable with the beta Ala-156 and beta Cys-156 enzymes and 10(2)-fold lower with the beta Ala-155 and beta Ser-155 enzymes. The beta Thr-156-->Ala or Cys enzyme showed an altered response to Mg2+, suggesting that beta Thr-156 may be closely related to Mg2+ binding. These results suggest that beta Lys-155 and beta Thr-156 are essential for catalysis and are possibly located in the catalytic site, although beta Thr-156 could be replaced by a serine residue.

Adenosine Triphosphate↗
PubMed1992

F0F1-ATPase gamma subunit mutations perturb the coupling between catalysis and transport.

We introduced mutations to test the function of the conserved amino-terminal region of the gamma subunit from the Escherichia coli ATP synthase (F0F1-ATPase). Plasmid-borne mutant genes were expressed in an uncG strain which is deficient for the gamma subunit (gamma Gln-14-->end). Most of the changes, which were between gamma Ile-19 and gamma Lys-33, gamma Asp-83 and gamma Cys-87, or at gamma Asp-165, had little effect on growth by oxidative phosphorylation, membrane ATPase activity, or H+ pumping. Notable exceptions were gamma Met-23-->Arg or Lys mutations. Strains carrying these mutations grew only very slowly by oxidative phosphorylation. Membranes prepared from the strains had substantial levels of ATPase activity, 100% compared with wild type for gamma Arg-23 and 65% for gamma Lys-23, but formed only 32 and 17%, respectively, of the electrochemical gradient of protons. In contrast, other mutant enzymes with similar ATPase activities (including gamma Met-23-->Asp or Glu) formed H+ gradients like the wild type. Membranes from the gamma Arg-23 and gamma Lys-23 mutants were not passively leaky to protons and had functional F0 sectors. These results suggested that substitution by positively charged side chains at position 23 perturbed the energy coupling. The catalytic sites of the mutant enzymes were still regulated by the electrochemical H+ gradient but were inefficiently coupled to H+ translocation in both ATP-dependent H+ pumping and delta mu H+ driven ATP synthesis.

Amino Acid Sequence↗
PubMed1992

Conformational feature of neuroactive domoic acid: X-ray structural comparison with isodomoic acid A and alpha-kainic acid.

As an aid for developing a new type of potent insecticide acting on the neuromuscular junction, conformational characteristics of domoic acid and isodomoic acid A, the naturally occurring glutamate agonists, were investigated by X-ray crystal analyses. Conformational comparison with a neuroactive alpha-kainic acid provides information concerning the stereochemical feature responsible for the biological activity.

Insecticides↗
PubMed1992

Spinal cord blood flow decreases following chemical stimulation of the rostral ventrolateral medullary pressor area in anesthetized rats.

In urethane-anesthetized, paralyzed and artificially ventilated rats, the neurons in the rostral ventrolateral medullary pressore area (VLPA) were chemically stimulated by microinjections of L-glutamate (1.7-5.0 nmol in 100 nl of 0.9% sodium chloride solution) and the spinal cord blood flow (SCBF) was determined using a combination of labeled microspheres (57Co, 113Sn and 46Sc). In order to measure SCBF at normotension, moderate hypotension within the lower limit of spinal cord autoregulation was induced by controlled hemorrhage (n = 12). Unilateral chemical stimulation of the VLPA in these rats increased arterial blood pressure (ABP) but it remained within normotensive range. The SCBFs of cervical, thoracic and lumbar cord decreased significantly from 27 +/- 3 (mean +/- S.E.M.) to 20 +/- 2 (P less than 0.01), from 22 +/- 1 to 17 +/- 2 (P less than 0.05), and from 41 +/- 5 to 26 +/- 3 (P less than 0.05) ml.min-1.(100 g)-1, respectively. The spinal cord vascular resistances (SCVRs) of cervical, thoracic and lumbar cord increased significantly from 3.7 +/- 0.4 to 5.0 +/- 0.6 (P less than 0.05), from 4.2 +/- 0.2 to 5.9 +/- 0.7 (P less than 0.05), and from 2.5 +/- 0.2 to 3.8 +/- 0.4 (P less than 0.05) mmHg per [ml.min-1.(100 g)-1], respectively. During the chemical stimulation of the VLPA, SCBF increased in response to the changes in arterial PaCO2 indicating that the reactivity of spinal cord vasculature was intact (n = 5).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Determination of conjugated bile acids in human urine by high-performance liquid chromatography with chemiluminescence detection.

A qualitative and quantitative analysis of the conjugated 1 beta- and 6 alpha-hydroxy bile acids, including common bile acids, in human urine using high-performance liquid chromatography with chemiluminescence detection is described. After extraction of urine with C18 silica cartridges, the bile acids were separated into non-conjugated, glycine, taurine and sulphate fractions by ion-exchange chromatography on a lipophilic gel. Solvolysis of the sulphate was carried out by treatment with trifluoroethanol in acetone containing hydrochloric acid, and the liberated amino acid conjugates were fractionated again. The individual bile acids were separated on a reversed-phase C18 column (Bile Pak II), with detection by an immobilized 3 alpha-hydroxysteroid dehydrogenase enzyme reactor and chemiluminescence reaction of the generated NADH using 1-methoxy-5-methylphenazinium methylsulphate-isoluminol-microperoxidase system. The assay method showed the detection limits ranging from 8 to 250 pmol for the bile acids tested. Analysis of urine samples obtained from newborns, non-pregnant women and women in late pregnancy showed a large difference in bile acid composition and conjugation mode, suggesting that bile acid metabolism is different during fetal and neonatal periods.

3-Hydroxysteroid Dehydrogenases↗
PubMed1992

Molecular abnormalities of a phosphoglycerate kinase variant generated by spontaneous mutation.

A new case of X chromosome-linked phosphoglycerate kinase (PGK) abnormality is described. The male proband was mentally retarded, had behavior disorders, and displayed episodes of hemolytic anemia. The enzyme activity of red blood cells from the patient was about 10% of normal, and that of the cultured fibroblasts was about 50% of normal cells. The variant PGK was characterized by a lower affinity for the substrates, reduced thermostability, and increased anodal electrophoretic mobility. The pH activity profile of the variant enzyme was different from that of normal. The amount of messenger RNA (mRNA) in the variant fibroblasts was comparable to that of normal fibroblasts. The mRNA coding for PGK was subjected to coupled reverse transcription followed by amplification by the polymerase chain reaction. Nucleotide sequence of the variant cDNA showed a point mutation, T/A----C/G transition, in exon 9 of the variant gene. No other mutation was found in all coding regions of the variant. The mutation should cause Cys----Arg substitution at the 315th position from the NH2-terminal Ser of PGK, and it created an additional Ava II (or isoschimatic) cleavage site in the variant gene. Because the variant gene was not detected in the proband's mother and siblings, it must have been generated by spontaneous mutation during oogenesis.

Adult↗
PubMed1992

Involvement of a non-proton pump factor (possibly Donnan-type equilibrium) in maintenance of an acidic pH in lysosomes.

Change of the internal pH of isolated lysosomes was measured with fluorescein isothiocyanate-dextran. In buffer of pH 7.0, isolated lysosomes had an acidic pH of about 5.5, which decreased to pH 5.2 on addition of ATP. Addition of bafilomycin inhibited the acidification by H(+)-ATPase and resulted in an increase of the internal pH to 5.5 due to passive diffusion of protons across the lysosomal membrane. However, no further alkalization was observed. The acidic pH (pH 5.5) of isolated lysosomes could be maintained for at least 48 h in the absence of ATP, but increased gradually to pH 5.9-6.4 upon incubation with monovalent cations (K+ or Na+), amines, or ionophores. These results suggest that a non-proton pump factor (possibly Donnan equilibrium) is involved in maintaining the acidic pH of isolated lysosomes.

Animals↗
PubMed1992

Molecular design of a eukaryotic messenger RNA and its chemical synthesis.

A designed mRNA consisting of 42 ribonucleotides having the cap structure was synthesized. The capped leader sequence of the brome mosaic virus (BMV) mRNA 4, m7G5'pppGUAUUAAUA (F-1), was synthesized by the phosphotriester method and followed by the capping reaction. A 32-mer consisting of an initiation codon (AUG), the coding region corresponding to a bacterial pheromone cAD1 and two stop codons, was constructed by the 18-mer (F-2) and 14-mer (F-3), which were synthesized by the phosphoramidite method. 2'-,3'-O-Methoxymethylene-guanosine 5'-phosphate was condensed with F-3 using P1-2',3'-O-methoxymethyleneguanosine-5'-yl P2-adenosine-5'-yl pyrophosphate (9) with T4 RNA ligase. The chemically synthesized RNA fragments were ligated successively with T4 RNa ligase to afford the whole RNA molecule.

Amino Acid Sequence↗
PubMed1992

Vacuolar type H(+)-ATPase genes: presence of four genes including pseudogenes for the 16-kDa proteolipid subunit in the human genome.

Genes for the human vacuolar type H(+)-ATPase proteolipid (16-kDa) subunit were cloned and their nucleotide sequences were determined. Comparison of the deduced sequences indicated that at least four genes including pseudogenes are present in the human genome. One of them corresponded to that for the 16-kDa subunit expressed in HeLa cells. The coding sequence was separated by two introns. The second intron was located in the DNA segment giving a loop between the second and third transmembrane helices, supporting the idea that the 16-kDa subunit was evolved by gene duplication. The primary sequence determined from the second clone had a termination codon behind the third transmembrane helix. Possible translation products from the other two clones had no putative acidic residues essential for proton transport function of the 16-kDa subunit. Thus, it is interesting to know whether these genes are transcribed, since they may have unique cellular functions.

Base Sequence↗
PubMed1992

Sequence motif in control regions of the H+/K+ ATPase alpha and beta subunit genes recognized by gastric specific nuclear protein(s).

A nuclear protein(s) from rat or pig stomach recognized a conserved sequence in the 5'-upstream regions of the rat and human H+/K(+)-ATPase alpha subunit genes. A gel retardation assay suggested that part of the binding site was located in the TAATCAGCTG sequence. No nuclear proteins capable of the binding could be detected in other tissues of rat (liver, brain, kidney, spleen and lung) or pig liver. The sequence motif (GATAGC) located 5'-upstream of the beta-subunit gene also seemed to be recognized by the same protein, because the binding of nuclear protein to the sequence motifs in the alpha and beta subunits was mutually competitive. Considering the sense-strand sequence of the binding motif in the alpha-subunit gene, we conclude that (G/C)PuPu(G/C)NGAT(A/T)PuPy is a core sequence motif for the gastric specific DNA binding protein (PCSF, parietal cell specific factor).

Adenosine Triphosphatases↗
PubMed1992

Induction of synthesis of manganous superoxide dismutase in L-M(pNTnF) cells carrying an inducible TNF gene.

Based on findings that the cytotoxic effects of tumor necrosis factor (TNF) are closely related to levels of intracellular oxygen radicals, and on the results of TNF gene transfection studies, the hypothesis was made that endogenous TNF (enTNF) acts as a protective factor against exogenous TNF by inducing inhibitors or scavengers of oxygen radicals. In order to test this hypothesis, we investigated the intracellular levels of manganous superoxide dismutase (MnSOD) and glutathione (GSH) in L-M(pNTnF) cells carrying a TNF gene induced by dexamethasone (DM). When L-M(pNTnF) cells were treated with DM they expressed enTNF, and acquired resistance to exogenous TNF. There was no change in the GSH concentration after enTNF induction, but a 1.9- to 3.9-fold increase in MnSOD levels was noted. Our findings suggest that enTNF exerts its protective function against the cytocidal effect of exogenous TNF by inducing MnSOD production.

Animals↗