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Biomedical subjects

T Tachikawa

Publications and source records attributed to T Tachikawa.

At least 73 records · Page 4Linked to original sources

[Antitumor effect of carboplatin on in vitro three-dimensional tumor].

We studied the antitumor effect of Carboplatin on the three-dimensional tumor in vitro. First, we observed the three-dimensional growth of cultured tumor cell lines (Hep 2, KB and HT 29 cells), which were developed in the combined culture system of fibrin matrix and agar culture. The tumor cells developed a 3 x 3 mm tumor in diameter in the in vitro 10 day-culture system. This size was large enough to allow the histologic study. When we applied Carboplatin to the three-dimensional tumors, histologic change of the three-dimensional tumor developed from Hep 2 and KB cells were observed in a dose dependent manner. However, no remarkable histologic change was observed in the three-dimensional tumor developed from HT 29 cells. These results suggest the experimental system of three-dimensional tumor can predict histologically the antitumor effect of Carboplatin on various tumors at the solid tumor level.

Carboplatin↗

Elucidation of three-dimensional ultrastructure of Hirano bodies by the quick-freeze, deep-etch and replica method.

To clarify the yet controversial fine structure of Hirano bodies, we made three-dimensional observations of the tissues from the right hippocampus obtained at autopsy of elderly patients by the quick-freeze, deep-etch and replica method. The basic structure of Hirano bodies was a unit lamella, a closely attached pair of sheets composed of parallel-running smooth filaments, 10 to 12 nm in diameter with 12-nm interspaces. In the unit lamella, filaments from each of the overlapping sheets crossed obliquely at acute or obtuse angles to form lattice-like meshworks. The unit lamellae were arranged in a folded, waved or concentric manner, and connected or supported by cross-linking filaments of the same width. The distance between these unit lamellae was about 50 nm. Occasionally the sheets were separated or fused making layers of one to three sheets. At the periphery of the bodies parallel filaments were dispersed into individual filaments of similar size or directly attached to the cytoplasmic membrane.

Aged↗

An experimental study of healing around hydroxylapatite implants installed with autogenous iliac bone grafts for jaw reconstruction.

The purpose of this study was to evaluate the use of hydroxylapatite implants (HA) with autogenous bone grafts for jaw reconstruction. After autogenous iliac bone grafts were transplanted to 36 rabbit mandibles, an HA implant was installed into the graft immediately (IM group), or 90 days (90D group), or 180 days (180D group) later. The animals were killed 7, 14, 30, 60, 90, and 180 days after the HA implant placement. The healing process was examined histologically, and histomorphometric measurements were made with a computer-based image analyzer to quantify the percentage of HA-bone bonding and trabecular bone in the medullary cavity. In the IM group, the HA-bone bonding tended to be deterred by fibrous tissue, and the rate of HA-bone bonding (BBSR) was less than that of the other groups. The trabecular bone around the implants tended to decrease after 30 days in all groups. However, the average of the trabecular bone specific volume (tVsp) in the 90D group was about 10% higher than that in the other groups. From these results, it is concluded that in clinical use of HA-coated dental implants with autogenous bone grafts, the time of installation should not be immediately after the bone graft, but when there is sufficient newly formed trabecular bone to enhance HA-bone bonding.

Animals↗

Novel method for quantifying alpha(1----3)-L-fucosyltransferase activity in serum.

A novel method has been developed to quantify alpha(1----3)-L-fucosyltransferase activity in human sera by applying a sandwich-type immunoradiometric assay. H type 2 trisaccharide (6Fuc alpha 1----2Gal beta 1----4GlcNAc) covalently attached to bovine serum albumin (BSA) was used as an acceptor and incubated with serum samples in the presence of guanosine diphosphate-fucose. The resulting product, Y tetrasaccharide (Fuc alpha 1----2Gal beta 1----4[Fuc alpha 1----3] GlcNAc beta-BSA), was detected by a sequential use of anti-BSA antibody-coated bead and 125I-labeled anti-Y antibody. Inter- and intra-assay CVs for alpha(1----3)-L-fucosyltransferase were both less than 4%, and the results of the dilution linearity and analytical recovery studies were satisfactory. Using the present assay method, we measured alpha(1----3)-L-fucosyltransferase in serum from patients with benign and malignant gastric disorders and in healthy subjects. The detection rate of alpha(1----3)-L-fucosyltransferase for cancer was apparently higher than that of carcinoembryonic antigen measured in the same samples, particularly in the early clinical stage; indeed, no correlation was observed between the concentrations of the two potential markers. The results indicate that the present assay method seems to be excellent for the determination of serum alpha(1----3)-L-fucosyltransferase activity and useful for the detection of cancer-associated increases of the enzyme activity at the early stage of gastric cancer.

Carbohydrate Sequence↗

Three-dimensional structure of Alzheimer's neurofibrillary tangles of the aged human brain revealed by the quick-freeze, deep-etch and replica method.

The three-dimensional structure of Alzheimer's neurofibrillary tangles in the pyramidal cells of the hippocampus and in the nerve cells of the parahippocampal gyrus was examined by the quick-freeze, deep-etch and replica method. The tangles consisted of either parallel bundles of or randomly arranged paired helical filaments (PHF), occupying the perikaryotic cytoplasm and extending to the dendritic processes. On the stereophotographs the PHF, measuring 28 to 36 nm in width, had two component filaments of 14 to 18 nm in diameter which were coiled anti-clockwise (left-handed) around each other with periodicity of 70 to 90 nm. The PHF in compact parallel bundles were cross-linked to each other with thin filaments, of about 6 nm in diameter, at relatively regular intervals. Randomly arranged PHF had no cross-bridges or side arms. Straight-type tangles of about 24 nm in diameter were rarely found in the dendritic processes. There were no discernible differences between the PHF of the patients with senile dementia of Alzheimer type and those of nondemented brains.

Aged↗

Monoclonal antibody to 2-amino-3-methylimidazo(4,5-f)quinoline, a dietary carcinogen.

In order to investigate relationships between human carcinogenesis and dietary carcinogens, one hybridoma cell line secreting a monoclonal antibody against 2-amino-3-methylimidazo(4,5-f)quinoline (IQ), a dietary carcinogen, was produced by fusing splenocytes from Balb/c mice immunized with IQ-Lysine(Lys)-Ascaris protein conjugate. The subclass of monoclonal anti-IQ antibody was determined by double immunodiffusion using culture medium and identified as IgG1. Monoclonal anti-IQ antibody was purified from ascites fluids of Balb/c mice with affinity chromatography on Protein A-Sepharose CL4B and analyzed concerning its cross-reactivity and sensitivity with RIA. Finally, we showed that our monoclonal antibody recognized IQ, 2-amino-3,4-dimethylimidazo(4,5-f)quinoline (MeIQ) and several beta-carbolines more intensely and that the sensitivity to IQ was 23 nmol in 50% displacement.

Animals↗

Abnormally high expression of proteasomes in human leukemic cells.

Proteasomes are eukaryotic ring-shaped or cylindrical particles with multicatalytic protease activities. To clarify the involvement of proteasomes in tumorigenesis of human blood cells, we compared their expression in human hematopoietic malignant tumor cells with that in normal peripheral blood mononuclear cells. Immunohistochemical staining showed considerably increased concentrations of proteasomes in leukemic cells from the bone marrow of patients with various types of leukemia and the predominant localization of these proteasomes in the nuclei. Moreover, enzyme immunoassay and Northern blot analysis indicated that the concentrations of proteasomes and their mRNA levels were consistently much higher in a variety of malignant human hematopoietic cell lines than in resting peripheral lymphocytes and monocytes from healthy adults. Proteasome expression was also greatly increased in normal blood mononuclear cells during blastogenic transformation induced by phytohemagglutinin; their expression increased in parallel with induction of DNA synthesis and returned to the basal level with progress of the cell cycle. Thus, abnormally high expression of proteasomes may play an important role in transformation and proliferation of blood cells and in specific functions of hematopoietic tumor cells.

Antibodies, Monoclonal↗

Morphological changes of rabbit skin by application of dentin primer.

The side effects of dentin primer applications on soft tissue were examined pathologically. Of the 4 experimental dentin primers tested, dimethylaminoethyl methacrylate (DMAEM) and glutaraldehyde (GLU) caused irritation to the skin of rabbits whereas hydroxyethyl methacrylate (HEMA) and glyceryl methacrylate (GM) caused only insignificant changes in the substrate. It was concluded that an aqueous solution of GM can be used clinically because of its low irritation to skin and high bonding efficacy, as previously reported, though possible irritation to pulp tissues should perhaps still be investigated. The mechanisms of irritation to operator fingertips caused by repeated direct contact with dentin primers containing HEMA remain unknown.

Animals↗

Long-term morphology of spastic or flaccid muscles in spinal cord-transected rabbits.

Despite difficulty in long-term maintenance of spinalized rabbits, muscular pathologic changes in chronic spinalized rabbits could be observed for a period of four weeks. Rabbits were prepared by spinal cord transection at T10 (spastic paralysis) or by spinal cord removal below L7 (flaccid paralysis). Spastic preparations showed hind-limb spasticity and reflex incontinence one to two days after operation. Hypertrophic fibers began to appear in spastic muscles after two weeks. This hypertrophy, thought to be caused by phasic repetitive contraction, was verified by electron microscopy to be different from normal exercise hypertrophy. Flaccid preparations maintained hind-limb flaccidity and overflow incontinence. In flaccid muscle, marked muscle fiber necrosis indicated rapid atrophy. Spinal deformity and joint contracture inactivate spinalized rabbits, and cause pressure sores. However, feeding assistance and avoidance of complications make long-term maintenance possible.

Animals↗

[Study of the penetration of extrinsic tracers into exposed cementum in vitro].

The purpose of the experiment was to study the possible penetration of extrinsic tracers with different molecular weights into exposed cementum in vitro and pathway of penetration. 75 human extracted teeth-55 periodontally diseased teeth and 20 embedded 3rd molars-were used. Each tooth was maintained in 0.005% fluorecein isothiocyanate (FITC) solution (M.W. 400), 0.05% FITC conjugated peroxidase solution (M.W. 40,000), or 0.01% FITC conjugated human IgG solution (M.W. 160,000) for 10 days. Morphological observations were made by means of fluorescence microscopy. Other extracted teeth were maintained in 0.05% microperoxidase solution (M.W. 1,900) for 5 days. Observations were then made using the TEM, and micrographs were taken and analyzed with an image analyzer. As a result, solutions of higher molecular weight showed lower penetration, and unexposed cementum showed a tendency toward greater penetration. Furthermore, the penetration of tracers on the surface of the cementum was along collagen fibers, but in the inner cementum, was into cementocyte lacunae and canaliculi.

Dental Cementum↗

Quantitative and qualitative characterization of human cancer-associated serum glycoprotein antigens expressing epitopes consisting of sialyl or sialyl-fucosyl type 1 chain.

The levels of carbohydrate antigens having epitopes consisting of type 1 chain (R----Gal beta 1----GlcNAc beta 1----3Gal beta 1----R) in the sera of patients with various malignant and nonmalignant disorders have been investigated with the use of three monoclonal antibodies, N-19-9, FH-7, and FH-9. Serum levels of 2----3 sialylated Lea antigen and 2----6 sialylated Lea antigen, defined respectively by antibodies N-19-9 and FH-7, were found to be frequently high in patients with cancer of the digestive system, particularly pancreatic cancer. High levels of 2----3,2----6 disialylated Lc4 antigen, defined by antibody FH-9, were less frequent in cancer patients when compared with the other two antigens. In patients with nonmalignant disorders, especially renal and autoimmune diseases, serum levels of the two type 1 chain antigens defined by FH-7 and FH-9 were more frequently high than that defined by N-19-9. Molecular weights and other general biochemical characteristics of serum mucin carrying the type 1 chain determinants were not significantly different in cancer patients as compared with patients with nonmalignant disorders. However, the degree of glycosylation of the antigen, as assessed by its solubility in perchloric acid, showed significant differences; i.e., the mucin antigen carrying 2----6 sialylated Lea determinant in the sera of patients with nonmalignant disorders had the highest carbohydrate/protein ratio, followed by the mucin carrying the same determinant in the sera of cancer patients. Mucin antigen carrying 2----3 sialylated Lea antigen or 2----3, 2----6 disialylated Lc4 antigen in cancer patients had the lowest carbohydrate/protein ratio among the four groups tested. Thus, the carbohydrate/protein ratio in the type 1 chain mucin antigens in sera of normal subjects is higher than that in sera of cancer patients (P less than 0.05). This finding is in contrast to previous findings on the mucin antigens carrying the type 2 chain determinant (R. Kannagi et al., Cancer Res., 46: 2619-2626, 1986), in which the mucin antigen in cancer patients was found to have a much higher carbohydrate/protein ratio than that carrying the same antigenic determinants in patients with nonmalignant disorders.

Antibodies, Monoclonal↗

Glycolipid-directed FH6 monoclonal antibody recognizes high molecular weight glycoprotein antigen carrying sialyl Lex-i determinant in the culture supernatant of PC-9 cells.

The properties of the antigen recognized by monoclonal antibody FH6 have been analyzed. FH6 was originally generated against a glycolipid, i.e. a difucoganglioside isolated from human colonic adenocarcinoma, and specifically reacts with sialyl Lex-i determinant. Several culture supernatants of human carcinoma cell line cells were found to have high levels of FH6-reactive antigen, and PC-9, a human lung carcinoma cell line was used for the analysis. A solid-phase sandwich radioimmunoassay was performed to detect the antigen. The antigenic activity was extractable in 0.6 M PCA or 7% TCA, and was sensitive to mild alkaline treatment and to Pronase digestion. Most of the antigen was eluted in the void volume of a Sepharose CL-2B column, which indicates that its molecular weight is greater than several million. It was eluted from a DEAE-cellulose column at a NaCl concentration in the range of 0.2-0.25 M. The immunoaffinity-purified antigen has a high carbohydrate content of more than 80%. These data indicate that the antigen recognized by FH6 in the culture supernatant of PC-9 is not a glycolipid, but a high molecular weight glycoprotein which could be referred to as a mucin, or a proteoglycan, which contains keratan-sulfate like glycosaminoglycan chains, as judged from the results of the glycosidase treatments.

Antibodies, Monoclonal↗

Inhibition of membrane Ca2+-ATPase in vitro by mating pheromone in Rhodosporidium toruloides, a heterobasidiomycetous yeast.

Direct addition of physiological concentrations of rhodotorucine A, a lipopeptide mating pheromone of Rhodosporidium toruloides, to the particulate fraction of the target cell strongly inhibited Ca2+-ATPase activity. The pheromone effect was mating-type specific. Membrane Ca2+-ATPase solubilized by a nonionic detergent and further purified by calmodulin-affinity chromatography was also inhibited by the pheromone. Rhodotorucine A S-oxide, a biologically inactive analogue, had no effect on Ca2+-ATPase. The results suggested that the inhibition of membrane Ca2+-ATPase is a critical event in the signaling of mating pheromone and the inhibition of membrane Ca2+-pump could be responsible for the pheromone-induced rapid raise of intracellular Ca2+ concentration reported.

Basidiomycota↗

In vitro early changes in intercellular junctions by treatment with a chemical carcinogen.

To examine early intercellular junction changes caused by treatment with 9,10-dimethyl-1,2-benzanthracene (DMBA), rat lingual epithelium was cultivated in isolation and observed by electrophysiological, freeze-fracture and whole-mount electron microscopy. Electrophysiological measurements showed a transient decrease in membrane potential of -10.2 mV 6 h after the treatment. It returned to almost the same level as that of the control group 1 day later. Six hours after treatment, input resistance decreased rapidly to 5.3 M omega but increased to 18.0 M omega 12 h after treatment. Transient reduction of input resistance and membrane potential occurred prior to the decrease in the coupling ratio 6 h after treatment with DMBA. In freeze-fracture replicas, the number of gap junctions decreased by approximately 45% of the control value 6 h after treatment with DMBA. At 12 h and thereafter, the number and area of gap junctions subsequently decreased by 60-80% of the control value. Alterations in the number and area of desmosomes were similar to those of the gap junctions. The formation of epithelial cytoskeletons, partially devoid of the 2-4 and 5-8 nm filaments was also observed. A decrease in the density of filament networks beneath the plasma membranes was especially apparent. Treatment with a carcinogen brought about morphological cellular changes as early as 6 h after treatment, and such early changes might trigger metabolic cellular abnormalities. Affected cells appear to move away from normal cells in a process of repeated destruction and revision of intercellular junctions, and cytoskeletons.

9,10-Dimethyl-1,2-benzanthracene↗