Biomedical subjects

M Tsukada

Publications and source records attributed to M Tsukada.

At least 19 recordsLinked to original sources

PubMed1996

Chemical modification of the arginyl residue in silk fibroin: 2. Reaction of 1,2-cyclohexanedione in aqueous alkaline medium.

The arginyl residue of solubilized silk fibroin was chemically modified with 1,2-cyclohexanedione in aqueous alkaline medium to form a stable imidazolidinone ring, and its positive charge was masked. CD spectra of the modified silk fibroin in aqueous solution showed an increase in the fraction of random coil conformation. The increase may be caused by the exposure to alkaline medium in the modification reaction. FT-IR and CD spectra of the silk fibroin films before and after the modification indicated that the conformational change in the modified silk fibroin in the solid state did not occur by the modification of its arginyl residue with 1,2-cyclohexanedione. The chemical stability of the modified silk fibroin film was investigated in vitro with phosphate-buffered saline solution. The modified arginyl residue in the film was stable in the phosphate-buffered solution.

Animals↗

PubMed1996

A novel endogenous mouse mammary tumor virus locus in Asian wild mice and its evolutionary divergency.

The divergency of endogenous mouse mammary tumor virus (MMTV) in European mouse strains indicates acquisition of exogenous MMTV into their genomes during evolution. In the present study, we selected two strains of Asian wild mice, Cas-Hmi (Mus musculus castaneus) and Sub-Kjr (M. m. subspecies) to investigate a possible divergency of endogenous MMTVs among Asian mouse strains. Southern blot analysis and polymerase chain reaction (PCR) demonstrated that Cas-Hmi and Sub-Kjr have the full structure of integrated endogenous MMTVs, suggesting that these mice were diverged during evolution after MMTV endemics.

Animals↗

PubMed1995

Attachment and growth of fibroblast cells on silk fibroin.

The attachment and growth of fibroblast cells (L-929) on matrices of silk fibroin from Bombyx mori domestic silkworm (DSF) and Antheraea pernyi wild silkworm (WSF) were studied by a cell culture method. The performance of the two kinds of silk fibroin was compared to that of collagen. DSF exhibited as high a cell attachment and growth as collagen did. The cells attached to DSF were extensively spread out and their filopodia were visible in the SEM pictures. WSF, which contains the tripeptide sequence Arg-Gly-Asp (believed to be a specific interaction site for cell-attachment), displayed much higher cell attachment and growth compared to DSF. The cells attached on WSF became virtually flat and their filopodia could be seen, indicating that the cells were very strongly held on the surface.

Amino Acid Sequence↗

PubMed1995

Attachment and growth of cultured fibroblast cells on silk protein matrices.

The attachment and growth of L-929 cells on films made of Bombyx mori silk proteins--fibroin and sericin and their mixtures--was studied by a cell culture method. Both cell attachment and growth were dependent on a minimum of around 90% sericin in the mixture. The results from electron micrography as well as from the DSC measurements supported the notion that the mixture of the two proteins fibroin and sericin has a phase-separated structure in the solid state. The observed minimum of sericin in the cell attachment and growth is thought to be a result of this phase-separated structure. Films of pure component proteins (i.e., 100% fibroin or sericin) exhibited as high a cell attachment and growth as collagen, a widely used mammalian cell culture substrate. However, a morphological study of the attached cells revealed that the cells attached to silk fibroin were extended and had a spindle shape, just like the cells attached to collagen, while the cells attached to the silk sericin had a different shape. It is concluded, therefore, that the attachment condition on silk fibroin is ideal for the viability, growth and function of the cells.

Amino Acids↗

PubMed1995

Changes in nitric oxide synthase activities in the cerebellum during development and aging of C57BL/6 mice.

Nitric oxide (NO) is one of the neurotransmitters in the cerebellum. Activities of NO synthase and densities of granule cells were determined in the cerebellum in an attempt to elucidate a mechanism of biochemical changes on the anatomical basis during aging. Activities of Ca2+/calmoduline dependent NO synthase were measured in the cerebellum in male and female mice, from 18 gestation days to 28 months of age. There were significant differences in NO synthase activities among the groups of prenatal (18 gestation days), middle aged (5-18 months) and old aged (21-28 months) mice. Between groups of old and middle ages, a significant decrease with aging was found in the mean density of granular cells, which express NO synthase. Changes in NO synthase activities and granule cell densities occurred to a similar degree between old and middle ages. The present data suggest that an age-associated rise and fall of its activities likely occur in parallel with those of granule cell densities, and thus may represent impairment of cerebellar function during aging.

Aging↗

PubMed1995

Detection of genomic variability among isolates of Campylobacter jejuni from chickens by crossed-field gel electrophoresis.

Digestion with Sal I facilitated the subclassification of 41 strains of Campylobacter jejuni into seven types, and digestion with Sma I enabled subclassification into twelve types. Sma I was potentially more useful for the detection of variability among the 41 strains, but both restriction enzymes seemed to be potentially useful for detecting variability among crossed-field gel electrophoresis profiles of the strains. The results clearly demonstrated that C. jejuni strains from different sources and with different routes of transmission had invaded the three farms investigated. At farms Sa and Ai, approximately 70% (23 strains) of isolates of C. jejuni (33 strains) were subclassified into the two major genotypes (I and II) on the basis of cleavage profiles with both Sal I and Sma I. These two major genotypes appeared to have invaded, expanded in and occupied the two chicken farms. All nine strains from farm Ai belonged genotype I.

Animals↗

PubMed1994

Mercury, not sulphur dioxide, poisoning as cause of smelter disease in industrial plants producing sulphuric acid.

Several episodes of "smelter disease", previously assumed to be caused by sulphur dioxide (SO2) poisoning, have been reported in workers replacing pipes in sulphuric acid manufacturing plants. One such incident, affecting 20 men, was recorded in Akita, Japan, in July, 1993, but the protection these workers used suggested that some cause other than SO2 needed to be looked for. 10 workers were affected despite wearing respirators with SO2 cartridges, the symptoms including dyspnoea, diarrhoea, colicky pain, muscle pain and eczema with erythema. Subsequently 10 other workers using face masks with supplied air were affected, though without respiratory symptoms. Sludge in the piping contained mercuric sulphate, and mercury fumes resulted when pipes were cut with gas burners. Blood and urine measurements confirmed heavy exposure to the metal, and simulation experiments in rats showed that skin absorption was likely too. The masks with supplied air ought to have excluded both SO2 and mercury fumes. The only way to avoid smelter disease reliably is to wear an encapsulated suit that prevents inhalation and skin absorption of industrial toxins.

Acute Disease↗

PubMed1994

Temporal pattern sensitivity of long-term potentiation in hippocampal CA1 neurons.

High-frequency electrical stimulation in the hippocampus leads to an increase in synaptic efficacy that lasts for many hours. This long-term potentiation (LTP) of synaptic transmission is presumed to play a crucial role in learning and memory in the brain. However, the frequency of stimulation generally used to obtain LTP is beyond the normal physiological range of activity of hippocampal neurons. We found that LTP can be induced by an electrical stimulation whose frequency is comparable to that of the naturally occurring firing activity of hippocampal neurons if the stimulating pulse-interval train has a special time structure. In the present experiment, we compared the magnitude of LTP induced by the four types of stimuli which have the same pulse number and the same mean frequency but different time structure in interstimulus intervals. One type of stimuli has regular intervals, and this served as a control stimulus. In the other three types of stimuli, the adjacent interstimulus interval had the following statistical properties: in type 1, their correlations are positive; in type 2, negative; and in type 3, independent. The magnitude of LTP induced by these four types of stimuli showed clear order relationships: type 3/type 1 >> control > type 2. Detailed analysis of the evoked potential during a period of temporal pattern stimulation revealed that the amplitude of the population spikes of repetitive firing, especially of the second and third population spikes, had the same order relationship as the LTP.(ABSTRACT TRUNCATED AT 250 WORDS)

2-Amino-5-phosphonovalerate↗

PubMed1994

Transcription of multiple exogenous mouse mammary tumor viruses in a single mammary tumor of an SHN mouse.

The SHN mouse strain, established in Japan independently from other mammary tumor-prone strains, is known to have a very high incidence of early-onset mammary tumors. In this study, we demonstrate that exogenous mouse mammary tumor virus (MMTV) plays a crucial role in early-onset mammary tumorigenesis in this strain. We subsequently isolated MMTV RNA from a mammary tumor of an SHN female mouse, reverse transcribed it, and amplified a 1.2-kb coding region of the long terminal repeat (LTR) by use of the polymerase chain reaction. The products were then subcloned into a pUC18 vector. Six clones were obtained, and their nucleotide sequences were determined. Nucleotide sequences of the six cloned LTRs indicated that two MMTVs are transcribed in the tumor. Both of them had point mutations and were different from known LTR sequences. Both LTRs had a potential open reading frame encoding 320 amino acids. The amino acid variabilities were predominantly between positions 280 and 320 codons when compared with deduced open reading frames. In contrast, only a few point mutations were detected in the regulatory area that contains five glucocorticoid elements, the nuclear factor-1 binding sites, the octamer consensus sequence, and the TATA box. Because point mutations occur more predominantly between positions 280 and 320, it is possible that mutations in this area endow MMTV with beneficial functions for propagation through mutational selection during tumorigenesis.

Animals↗

PubMed1994

Macro-fingerprinting analysis at the chromosomal genomic DNA level of isolates of thermophilic Campylobacter coli and C. jejuni, by pulsed-field gel electrophoresis.

Chromosomal genomic DNA from 25 strains of thermophilic Campylobacter coli and C. jejuni were analysed after pulsed-field gel electrophoresis (PFGE) of restriction fragments generated by SalI and SmaI to evaluate the utility of macro-fingerprinting analysis and to assess the possible subtyping of the isolates. Two rare-cutting restriction enzymes, SalI and SmaI, allowed the subclassification of eight strains of C. coli into six types. SalI facilitated the subclassification of seventeen strains of C. jejuni into thirteen types and SmaI the subclassification of seventeen strains into twelve types. The diversity in the PFGE profiles was demonstrated and, therefore, the two enzymes were found to be potentially useful for detecting variability of PFGE profiles among strains of C. coli and C. jejuni. The two enzymes and PFGE may be useful in molecular epidemiology.

Animals↗

PubMed1993

Isolation and characterization of autophagy-defective mutants of Saccharomyces cerevisiae.

Autophagy in the yeast is similar to that in mammalian cells. A mutant designated as apg1 (autophagy) defective in accumulation of autophagic bodies in the vacuoles was isolated by selection using a light microscope from a mutagenized proteinase-deficient strain. In the apg1 strain, which has normal vacuolar proteinases, nitrogen starvation did not induce protein degradation. The apg1 mutant lost its viability faster than wild-type cells during nitrogen starvation. By using the loss of viability as a first screening test, 75 other apg mutants were selected. These apg mutants including apg1 fell into 15 complementation groups. Genetic analyses of representative apg mutants revealed that they all had single recessive chromosomal mutations. Strains with each apg mutation were defective in protein degradation in the vacuoles induced by nitrogen starvation and homozygous diploids for each apg mutation did not sporulate. These results on the apg mutants suggest that autophagy via autophagic bodies is indispensable for protein degradation in the vacuoles under starvation conditions, and that at least 15 APG genes are involved in autophagy in yeast.

Autophagy↗

PubMed1993

Increase in housing temperature can alleviate decrease in white blood cell counts after energy restriction in C57BL/6 female mice.

White blood cell (WBC) counts decrease after energy restriction (ER) in both rodents and humans. ER reduces cellular proliferation rates in various organs of rodents. Increases in housing temperatures can cancel this inhibitory effect of ER. We investigated whether a high housing temperature can alleviate decreases in WBC counts after ER, since peripheral WBC counts at basal levels are considered to represent cellular proliferation rates in the bone marrow. C57BL/6 female mice were allocated to three groups: the control (397 kJ/week), the ER (176 kJ/week) and the ER housed-in-incubator group (ERI: 134 kJ/week). The former two groups were housed at 21-23 degrees C and the third group was housed at 30 degrees C. At 4 and 13 months of age, WBC counts were significantly lower in ER or ERI mice than in control mice. WBC counts, however, were significantly higher in ERI mice (194%) than in ER mice at 13 months of age. A high housing temperature was proved to antagonize the effect of ER on WBC counts, as it antagonizes suppressive effects of ER on cellular proliferation rates in various organs.

Animals↗

PubMed1993

Retroviral gene expression as a possible biomarker of aging.

We examined effects of aging on endogenous retrovirus gene expression of mouse lymphocytes with a hypothesis that it may be a useful biomarker of aging. Mice have endogenous murine leukemia viruses (MuLVs) in their chromosomes. We detected the gene expression of long terminal repeats (LTRs) of MuLVs. Brains, livers and spleens were taken from young (3 months old) and old (27 months old) male C57BL/6 mice. In addition to these control (C) mice, we also determined gene expression in dietary restricted (DR) mice, in which rates of aging are known to be slowed. RNA was extracted from the tissues and converted into cDNA. The MuLV-LTR portion of cDNA was amplified by polymerase chain reaction (PCR). The PCR products were analyzed by agarose gel electrophoresis. Gene expressions of young mice were found to be tissue-specific. Expressed LTRs from brains, livers and spleens were that of 370 base-pairs (bp), those of 370 and 620 bp, and those of 370, 400 and 620 bp, respectively. Old mice of C group, however, decreased tissue specificity: expressed LTRs became those of 370-400 bp in any tissues. In contrast the tissue specific gene expression was conserved in old DR mice which had to get prolonged life span and decreased lymphoma incidence. Thereby, gene expression of endogenous retroviruses appears to change during aging and to be modifiable by life-prolonging DR. It may be therefore used as a biomarker of aging in mice. Humans are known to have similar gene elements like MuLV. The present findings demonstrate a possibility of application of endogenous gene expressions to the epidemiology of aging.

Aging↗

PubMed1993

O,O,S-Trimethyl phosphorothioate induces hypothermia in Fischer 344 rats in a manner dependent on both doses and housing temperatures.

We explored the effects of O,O,S-trimethyl phosphorothioate (OOS-TMP) on body temperatures in Fischer 344 female rats. The 7-day LD50 p.o. for Fischer 344 female rats was found to be 11.8 mg/kg. OOS-TMP induced long-lasting (more than 48 h) and extensive hypothermia at doses > or = 14 mg/kg at a typical laboratory temperature (22 degrees C) while it produced typical symptoms at 10 mg/kg without hypothermia. In contrast, pair-fed (to 20 mg/kg rats) rats (n = 4) did not become hypothermic, negating any role of hypophagia in OOS-TMP associated hypothermia. We next investigated the effects of housing temperatures on toxicities at a LD50 dose (12 mg/kg). At 30 degrees C (n = 11) and 22 degrees C (n = 13), rats did not have hypothermic bouts but at 15 degrees C, eight out of ten rats had. Evidence that changes of housing temperatures neither modified clinical symptoms nor changed mortality rates discards a possibility of hypothermia being involved in delayed toxicity. A novel result of the present study suggests that thermoregulation may be heavily impaired by a special class of organophosphorus compounds.

Animals↗

PubMed1993

Energy restriction suppresses microsomal Ca(2+)-ATPase activities in various organs in C57BL/6 female mice in both euthermic and torpor states.

C57BL/6 female mice were fed a daily control diet (n = 5, 3.9 g/day, 95 kcal/week) or ER diet (n = 5, 2.3 g/day, 48 kcal/week) at 1800 h from 6 weeks of age. Telemetry, conducted at 6 months of age, confirmed that all ER mice entered daily torpor (core body temperature less than 31 degrees C) for 6.63 +/- 2.34 h/day while control mice were euthermic (> 35 degrees C). In vitro activities of microsomal Ca(2+)-ATPase were determined in the brain, liver, salivary gland and kidney from these mice at 6 months of age. Assays were performed at three incubation temperatures of 37 degrees C, 31 degrees C and 25 degrees C. In assays at 37 degrees C, the activities of Ca(2+)-ATPase in the brain and salivary gland from ER mice were lower than those in corresponding organs from control mice. The suppression became profound as the incubation temperature decreased. On the other hand, at 37 degrees C Ca(2+)-ATPase activities in the liver and kidney from ER mice were not lower than those in corresponding organs from control mice, but decreased significantly at low temperatures. Microsomal Ca(2+)-ATPase activities thus appeared to be reduced in ER mice, although it remains unknown whether the present results represent reduced in vivo capacities to regulate cytosolic Ca2+ concentrations.

Animals↗

PubMed1993

Treatment of standard risk acute lymphoblastic leukemia in children with the Tokyo Children Cancer Study Group (TCCSG) L84-11 protocol in Japan.

During the period June 1984 through February 1989, 207 children, newly diagnosed with standard-risk acute lymphoblastic leukemia (ALL), were treated with the Tokyo Children Cancer Study Group L84-11 protocol. The patients were randomized into two regimen groups (S1 and S2). All patients received identical induction therapy with vincristine, prednisolone, and L-asparaginase. For central nervous system (CNS) prophylaxis, the patients in regimen S1 received cranial irradiation (18 Gy) and intrathecal chemotherapy starting at 5 weeks, while those in regimen S2 received this therapy starting at 24 weeks. Consolidation consisted of cyclic therapy with vincristine and dexamethasone, and then, after 128 weeks, with medium-dose methotrexate, CNS chemoprophylaxis, and dexamethasone. Regimen S2 employed early consolidation with three doses of medium-dose methotrexate and CNS chemoprophylaxis. The treatment duration was 3.5 years. Complete remission was achieved in 96.0% and 99.0% of the patients in regimens S1 and S2, respectively. Event-free survival (EFS) rates were 80.0 +/- 3.5% in total, 74.5 +/- 5.7% in regimen S1, and 85.1 +/- 5.7% in regimen S2 at a median follow-up of 42 months (< 85 months). There was no significant difference in the EFS between the two regimens. As compared with recently reported results of treatment for this disease, our protocol can yield similar EFS rates with mild toxicity.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1993

Energy restriction that inhibits cellular proliferation by torpor can decrease susceptibility to spontaneous and asbestos-induced lung tumors in A/J mice.

BACKGROUND: Energy restriction (ER) inhibits various tumors in mice. A/J mice have a very high incidence of lung tumors, that are correlated with elevated levels of pulmonary cell proliferation in this strain. Using A/J mice, we studied the effects of ER on spontaneous and asbestos-induced lung tumors and on labeling indices in the lung as a proximal marker of susceptibility to lung tumors to obtain better understanding of the mechanism of ER in reducing tumorigenesis. EXPERIMENTAL DESIGN: Experiment 1: A/J female mice were instilled intratracheally with asbestos or titanium dioxide or simple saline at 20 weeks of age. At 21 weeks of age, mice of the ER diet group were switched from the control diet (350 kJ/week) to the ER diet (175 kJ/wk) until sacrifice at 105 weeks of age, whereas mice of the control diet group were continued on the control diet. Experiment 2: A/J female mice were begun on ER or control diets at 6 weeks of age. The control and ER mice were kept at 20 to 22 degrees C, whereas mice of another ER group (ER+I) were kept at 30 degrees C until 24 weeks of age. Body temperatures of these mice were monitored by telemetery. RESULTS: The present ER was shown to suppress the development of both asbestos-induced and spontaneous lung tumors. ER mice were confirmed to become torporfic, whereas control and ER+I mice did not. Labeling indices were reduced by ER in the lung as well as in other organs. The reduction of labeling indices was, however, almost recovered by increasing the housing temperature to 30 degrees C. CONCLUSIONS: ER which reduces cellular proliferation in various organs by torpor was shown to inhibit lung tumors. Inhibition of lung tumors by ER is likely to involve a decrease in cell proliferation.

Animals↗

PubMed1992

Potent antagonistic action of OPC-31260, a vasopressin V2 receptor antagonist, on [Arg8]vasopressin-induced vasoconstriction in isolated simian femoral arteries.

Using a perfusion technique with isolated vessels, we investigated the effect of OPC-21268 (a selective V1 receptor antagonist) and OPC-31260 (a V2 receptor antagonist) on arginine vasopressin (AVP)- or norepinephrine (NE)-induced vasoconstrictions of isolated simian femoral arteries. The dose-response curve for AVP was shifted to the right in parallel by OPC-31260 but not by OPC-21268. Neither antagonist significantly modified the NE-induced vasoconstriction. On the basis of these results, there are functionally constrictory vasopressin V2 receptors but no V1 receptors in isolated simian femoral arteries.

Animals↗