Expression of beta-hexosaminidase isozymes in peripheral lymphocytes from patients with aplastic anemia.
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Biomedical subjects
Publications and source records attributed to T Kaneda.
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One hybridoma cell line that produces an antibody directed against 10S DNA polymerase alpha purified from calf thymus was obtained. The monoclonality of the antibody was tested by sodium dodecyl sulfate polyacrylamide gel electrophoresis, isoelectrofocusing and antibody subclass determination. The antibody specifically recognized the 10S DNA polymerase alpha and 6.5S DNA polymerase alpha-2 from calf thymus, but not 6.5S DNA polymerase alpha-1. The antibody precipitated both polypeptides of 140-150,000 and 46-50,000 dalton of 10S DNA polymerase alpha. The antibody also recognized the DNA polymerase alpha purified from human cells, but did pig DNA polymerase alpha only partially. The antibody did not crossreact with rat DNA polymerase alpha, calf DNA polymerase beta, virus DNA polymerase and E. coli DNA polymerase I. This antibody will be a useful tool for studying the mechanism of DNA replication in eukaryotic cells.
Terminal deoxynucleotidyl transferase (TdT, EC 2.7.7.31) has been purified 4365-fold from pig thymus. It was further separated into two molecular forms of 57,000 and 45,000 dalton by Sephadex G-100 gel-filtration. The former sedimented at 4.2s through a sucrose gradient, while the latter, at 3.6s. By a SDS-polyacrylamide gel-electrophoresis, their molecular weights were estimated as 57,000 and 42,000 dalton, respectively. Thus each of the large and small pig TdT consists of a single polypeptide of 57,000 or 42,000 dalton and has no subunit structure. These two forms were indistinguishable in antigenicity by a neutralization assay of 42,000 dalton-TdT antibody. The enzymological properties of 42,000 dalton-TdT from pig thymus were very similar to those of calf TdT which has a two-subunits structure.
Vascular changes in rat skin after irradiation were examined microangiographically. Revascularization of the skin transplanted during the chronic stage after irradiation was also studied. The results obtained through these examinations revealed higher vascular densities at the acute and the subacute stages, and low values at the chronic stages compared with those of the control. Furthermore, when the skin grafts were transplanted to the irradiated beds in the chronic stage, primary revascularization was scant, and the inhibited capillary proliferation in the recipient sites prevented new vessel penetration. This explains to why grafts transplanted to previously irradiated beds fail to survive.
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1. The conversion mechanism for 14CO2 from [14C]Leu, [14C]Ile and [14C]Val and the incorporation of 14C-amino acids into lipids of subcutaneous tissue (melon and blubber), liver and muscle of the adult whale, Stenella caeruleo-alba were studied using a tissue culture method. 2. Iso-acids such as -5:0, -11:0; -13:0 and -15:0 in melon triglycerides were biosynthesized from [14C]Leu. On the other hand, anteiso-acids such as -5:0, -11:0, -13:0, -15:0 and n-11:0 - n-16:0 acids were derived from [14C]Ile. 3. Iso-acids such as -4:0, -12:0, -14:0 and 16:0 were biosynthesized from [14C]Val. 4. Analytical data indicate that the compositions of these acids biosynthesized were similar to that of fatty acids in whale oil. 5. The branched-chain fatty acid biosynthesized in blubber was iso-5:0 acid from [14C]Leu, with no long-chain branched acids, but n-12:0, -14:0 and -16.0 acids were also biosynthesized. 6. Radioactivities derived from 14C-amino acids incorporated into lipids in liver and muscle were extremely low compared with those in subcutaneous tissue. 7. The ratios of 14C-lipids/14CO2 in every 14C-amino acid incorporation were higher in subcutaneous tissue than in liver and muscle, and especially in the case of [14C]Leu, the ratio in melon was quite high. 8. The incorporation of the 14C-amino acids into short- and long-chain acids and alcohols of wax esters was recognized in all experiments.
Calf thymus DNA polymerases alpha and beta [EC 2.7.7.7] and terminal deoxynucleotidyl transferase [EC 2.7.7.31] were analyzed on two-dimensional gel slabs. DNA polymerase beta appeared as a single spot on two-dimensional gel at the position of 40,000 daltons and pI 8.0 using non-equilibrium pH gradient gel electrophoresis for the first-dimensional run. By overlapping gel slabs, it was possible to identify the distinct spot of DNA polymerase beta among many polypeptide spots of a crude enzyme fraction. 10S DNA polymerase alpha showed two clusters of polypeptide spots on two-dimensional gel slab. One cluster was composed of three large polypeptides of 140,000-150,000 daltons and another was composed of four smaller polypeptides of 46,000-50,000 daltons. All these spots were arranged in a narrow pI range (6.5-6.8) although each spot showed a distinct pI value. Purified terminal deoxynucleotidyl transferase showed three polypeptides of 57,000, 42,000, and 33,000 daltons at similar pI values (7.0-7.2). Each polypeptide consisted of plural spots which differed slightly in pI but were the same in molecular weight. These results suggest a microheterogeneity of polypeptides of terminal deoxynucleotidyl transferase as well as those of 10S DNA polymerase alpha.
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Deficient coagulation factor replacement therapy is often indispensable for dental extractions and oral mucosal bleeding in hemophiliac patients. High concentrates are often infused in large amounts of volume, until complete hemostasis is achieved. Considering the various side effects of replacement therapy the volume infused should be reduced to a minimum. Several aspects associated with replacement therapy for the control of oral bleeding are studied. Our conclusion is that local hemostatic procedures are even more important than replacement therapy. The use of surgical packs and band retainers proved to be of benefit in this study.
Terminal deoxynucleotidyl transferase [EC 2.7.7.31] has been purified 4,365-fold from pig thymus. It was further separated into two molecular forms of 57,000 and 45,000 daltons by Sephadex G-100 gel-filtration. The former sedimented at 4.2S through a sucrose gradient, while the latter sedimented at 3.6S. By sodium dodecyl sulfate-polyacrylamide gel-electrophoresis, their molecular weights were estimated at 57,000 and 42,000 daltons, respectively. Thus, the large and small pig terminal deoxynucleotidyl transferases both consist of a single polypeptide of 57,000 and 42,000 daltons and have no subunit structure. These two forms were indistinguishable in antigenicity as examined by a neutralization assay with an anti-calf terminal deoxynucleotidyl transferase antibody. The enzymatic properties of 42,000-dalton terminal deoxynucleotidyl transferase from pig thymus were very similar to those of the calf enzyme, which has a two-subunit structure.
Extra-weak chemiluminescence was detected in the organ homogenate and blood of tocopherol-deficient rats by use of a newly devised single photon counting apparatus. The spectrum distribution showed that the chemiluminescence had emission peaks at wavelengths between 500 and 650 nm corresponding to the simultaneous transition of singlet molecular oxygen (1O2), 2[1 delta g] leads to 2[3 epsilon g-]. This chemiluminescence was quenched by the presence of free radical scavengers, butyl hydroxytoluene and d-alpha-tocopherol. It was stimulated by D2O and 1O2-emission enhancer, 1,4-diazabicyclo[2,2,2]octane. The chemiluminescent intensities in tissue of rats fed a tocopherol-free diet for seven months were higher than those of rats fed a normal diet which contained 15 mg of tocopherol per 100 g of diet. The considerable increase of light emission was observed especially in liver, kidney, heart, lung and brain homogenates. The intensity of tocopherol-deficient liver chemiluminescence corresponded to 22 x 10(3) photons per sec . cm2. The results indicated that the chemiluminescence was directly related to the generation of 1O2 involving free radical reactions in the tocopherol-deficient rat tissues.
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Terminal deoxynucleotidyl transferase (TdT) activity was assayed in leukemic cells of 34 patients in blastic phase of Philadelphia-positive chronic myelogenous leukemia (CML). The TdT levels were correlated with the response to vincristine and prednisone therapy. None of 20 Tdt-negative patients, but 10 of 14 TdT-positive patients responded to vincristine and prednisone. There was poor correlation between blast cell morphology and response to this therapy. This study confirms that TdT levels in the majority of patients predict effectiveness of vincristine and prednisone therapy in blastic phase of CML.