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K Fukuhara

Publications and source records attributed to K Fukuhara.

At least 73 records · Page 4Linked to original sources
PubMed1996

Mutagenicity of nitrophenanthrene derivatives for Salmonella typhimurium: effects of nitroreductase and acetyltransferase.

To determine the mutagenicity of nitrophenanthrenes, three mononitrophenanthrenes (NPhs), 11 dinitrophenanthrenes (diNPhs) and eight trinitrophenanthrenes (tiNPhs) were synthesized, and their mutagenicity was investigated by using Salmonella typhimurium his- strains TA98, TA100, and TA98NR, nitroreductase-deficient, and TA98/1,8-DNP6, O-acetyltransferase-deficient mutants, and strains YG1021 and YG1026, nitroreductase-overproducing mutants of TA98 and TA100, respectively, and strains YG1024 and YG1029, O-acetyltransferase-overproducing mutants of TA98 and TA100, respectively. 1-, 3- and 9-NPhis induced 329, 620 and 438 revertants per nmol in strain TA100, respectively, and 4,839, 11,309 and 16728 revertants per nmol, respectively, in strain YG1029. Mutagenicity of 1,6-, 2,6-, 2,9-, 2,10-, 3,5-, 3,6- and 3,10-diNPh was elevated in strains YG1021, YG1024, YG1026 and YG1029. Among these derivatives, 1,6-, 2,6-, 3,6- and 3,10-diNPhs were more mutagenic in strains YG1024 and YG1029 than YG1021 and YG1026, and they showed a structure-activity relationship between mutagenicity and NO2-substitution. Nitro derivatives substituted at the 3 and 6 positions of their chemical structure strongly mutated both strains YG1024 and YG1029, whereas those substituted at the 9 and 10 positions showed weak mutagenicity. In addition, nitro substituents at positions 4 and 5 were perpendicular while those on positions 2,3,6 and 7 were nearly coplanar to the aromatic ring. Furthermore, 2,6,9-, 3,6,9- and 1,6,9-trinitrophenanthrenes (triNPhs) were mutagenic for strain TA100, and their mutagenicity was more enhanced in YG1024 and YG1029 than in YG1021 and YG1026. Of the eight triNPhs all except 1,5,10-triNP were mutagenic in TA98 and TA100, and their mutagenicity was more enhanced in YG1024 and YG1029 than in YG1021 and YG1026. These results suggest that these compounds are mutagens that are activated by O-acetyltransferase esterification following nitroreductase. The nitrated derivatives substituted at the 2(7) and 3(6) positions of the phenanthrene ring were highly mutagenic. The relationship between chemical structure and the mutagenicity of NPh derivatives is discussed.

Acetyltransferases↗
PubMed1996

Interrelations between sympathoadrenal system and hypothalamo-pituitary-adrenocortical/thyroid systems in rats exposed to cold stress.

The interrelations between sympathoadrenal (SA) system and hypothalamo-pituitary-adrenocortical (HPA) or hypothalamo-pituitary-thyroid (HPT) system during cold stress were examined by measuring plasma levels of dihydroxyphenylalanine (DOPA), catecholamine and their metabolites in adrenalectomized (ADX) and thyroidectomized (TX) rats exposed to cold stress (-3 degrees C). Plasma levels of adrenocorticotropic hormone (ACTH), corticosterone (CORT), thyroid-stimulating hormone (TSH) and thyroid hormones in cold-stressed rats were measured also. Plasma ACTH levels were increased transiently after 1 h of cold exposure, after which the circadian rhythm and plasma levels of ACTH were similar to those of normal rats. Plasma CORT levels were also elevated after 1 h of cold exposure; the increased levels of CORT tended to return to normal levels after 9 h of cold, but remained higher than those of normal rats during at least 24 h of cold exposure. Plasma ACTH levels of 5 day cold-stressed rats were no longer elevated above those of control rats and plasma CORT levels were only slightly higher than in control animals. However, plasma levels of TSH and free thyroid hormones were elevated after 1 day and remained elevated after 5 days of cold exposure. Thus, cold stress appears to activate chronically the HPT system, but only transiently activates the HPA system. ADX rats had higher basal plasma levels of dihydroxyphenylglycol (DHPG), methoxyhydroxyphenylglycol (MHPG), DOPA and homovanillic acid (HVA) than those of sham-operated (SHAM) rats, but norepinephrine (NE) levels were not significantly greater than in SHAM animals. TX rats had higher basal plasma levels of NE, epinephrine (EPI) and dopamine (DA), as well as much higher plasma levels of the metabolites. Exposure to cold increased plasma NE levels in both ADX and TX rats, but the increments in TX rats were much greater than in SHAM and ADX groups. Plasma EPI levels were not significantly elevated during cold exposure in SHAM rats, but were highly elevated in TX rats exposed to cold. TX rats had much larger increments in plasma levels of DHPG, MHPG, DA, dihydroxyphenylacetic acid (DOPAC) and HVA during cold exposure than those of SHAM and ADX rats. These results are consistent with the view that endogenous glucocorticoids restrain responses of catecholamine synthesis, release, reuptake, and metabolism in sympathetic nervous system of cold-stressed animals, but that in the absence of an effective HPT system, there is enhanced sympathoadrenal medullary function and augmentation of their responses to cold as a means for maintaining body temperature when the HPT thermogenesis system is impaired.

Adrenal Glands↗
PubMed1996

Cloning and expression of an isovaleryl pepstatin-insensitive carboxyl proteinase gene from Xanthomonas sp. T-22.

Xanthomonas carboxyl proteinase (XCP), isolated from Xanthomonas sp. T-22, is the second example of the unique carboxyl proteinases [EC 3.4.23.33] which are insensitive to the classical aspartic proteinase inhibitor. The gene coding for XCP was cloned, sequenced, and expressed in Escherichia coli. The XCP gene contains an open reading frame of 2,481 base pairs encoding a protein of 827 amino acid residues with a M(r) of 83,677. The XCP was synthesized as a large precursor consisting of three regions: NH2-terminal prepro (N-Prepro) (237 amino acid residues); mature XCP (398 a.a.residues); and COOH-terminal pro (C-Pro) (192 a.a. residues). The N-Prepro and mature XCP regions had no sequence similarity to any other proteins reported so far, except the carboxyl proteinase from Pseudomonas sp. 101 [Oda, K., Takahashi, T., Tokuda, Y., Shibano, Y., and Takahashi, S. (1994) J. Biol. Chem. 269, 26518-26524]. The C-Pro region showed high similarity to COOH-terminal regions of other microbial proteinase precursors. E. coli carrying a plasmid containing the cloned wild-type XCP gene produced an 84-kDa protein. This protein was processed into a mature, active form under acidic conditions. This process was completely blocked by tyrostatin, an XCP-specific inhibitor from Kitasatosporia sp. 55, indicating an autocatalytic processing. The purified recombinant XCP had the same characteristics as authentic XCP except for the NH2-terminal amino acid sequence. When the mutant XCP gene truncated in the C-Pro region was expressed in E. coli, an expected 64-kDa protein was detected in the cells, and also processed into the 42-kDa active form under the acidic conditions. Thus, the C-Pro region was not essential for the formation of active mature XCP.

Amino Acid Sequence↗
PubMed1996

Structural and functional changes in the canine shoulder after cessation of immobilization.

The functional and structural consequences of remobilization of the glenohumeral joint after 12 weeks of immobilization were studied in 10 beagle dogs. The dogs were grouped in pairs: 2 were used as controls, and in 8 others 1 forelimb was immobilized. At the end of 12 weeks, 2 dogs in the experimental group were euthanized, as were the 2 dogs used as controls. The cast was removed from the other 6 dogs, and the remobilized animals were euthanized in pairs at the end of 4, 8, and 12 weeks. It was found that after 12 weeks of immobilization, the passive range of motion was markedly impaired, intraarticular pressure was raised during movements, and the filling volume of the joint cavity was reduced. Histologically, the capsule showed synovial lining hyperplasia and vascular proliferation in the wall. Immunohistochemical labeling for collagen Types I and III failed to show an increase of fibrous collagens in the capsular wall. Both the functional and structural changes were unaltered after 4 weeks of remobilization, but after 8 weeks they began to reverse, and they returned to normal levels after 12 weeks. Both functional and structural changes after 12 weeks of immobilization of the uninjured glenohumeral joint are reversible by remobilization. The collagen composition of the capsule seems unrelated to the degree of capsular contraction that occurs during 12 weeks of immobilization and subsequent remobilization.

Animals↗
PubMed1996

Effects of continuous and intermittent cold (SART) stress on sympathoadrenal system activity in rats.

We compared sympathoadrenal responses to intermittent cold (SART) stress (in which cold exposure is interrupted by 4-hourly intervals daily at room temperature) with those to continuous cold (-3 degrees C) stress. Plasma levels of dihydroxyphenylalanine (DOPA), catecholamines and their metabolites as well as tyrosine hydroxylase (TH) activities in sympathetically innervated tissues were examined in rats exposed to each stressor for 1 day or for 5 days. Neither SART nor continuous exposure to cold for 1 day or 5 days altered plasma epinephrine (EPI) levels. However, norepinephrine (NE) and dihydroxyphenylglycol (DHPG) levels increased markedly during exposure to these stressors. On the first day of SART or continuous cold stress, NE levels were increased similarly, but the increments in DHPG levels were greater during SART stress. Since DHPG is formed in neurons, neural reuptake of NE may be more enhanced on the first day of SART stress than on the first day of continuous cold stress. After 5 days of SART stress plasma NE levels were significantly higher than those found after 5 days of continuous cold exposure. Plasma levels of DHPG were elevated to the same extent in both 5 days SART- and continuously cold-stressed rats, whereas plasma levels of methoxyhydroxyphenylglycol (MHPG) increased only by 5 days SART stress. Even at 1 h after the removal from 5 days SART stress, increased plasma levels of NE, DHPG and MHPG were still evident. These results suggest that 5 days SART stress elevates extraneuronal O-methylation of DHPG, and that NE turnover is more greatly increased by SART stress than by continuous cold stress. Plasma levels of DOPA, dopamine, dihydroxyphenylacetic acid and homovanillic acid also increased after either SART or continuous cold stress for 1 day and 5 days. Adrenal TH activities were significantly increased in rats exposed to SART or continuous cold stress for 1 day and 5 days, but in brown fat TH activity was elevated only in rats exposed to 5 days of continuous cold. Both SART and continuous cold stress are selective and potent stimuli for activation of the sympathoneural system, apparently without significant adrenomedullary EPI release. The increase of TH activity in the brown fat pad as well as of plasma NE and its metabolites is probably a result of adaptation to cold. It appears that even short intervals of return to a normal environmental temperature, as in SART, are sufficient to diminish sympathetic adaptation to cold.

3-Methoxy-4-hydroxyphenylethanol↗
PubMed1995

Sympathoadrenal system in stress. Interaction with the hypothalamic-pituitary-adrenocortical system.

Exposure of an organism to any of a variety of stressors markedly activates the sympathoadrenal and hypothalamic-pituitary-adrenocortical systems. Interactions of these major stress systems occur at several levels in the periphery and the brain. In the present study, we used sham-operated or adrenalectomized cortisol-treated conscious rats to examine glucocorticoid effects on indices of CA release, metabolism, and synthesis, and on CA biosynthetic enzyme activities and gene expression at baseline and during immobilization stress (IMO). Adrenalectomy (ADX) stimulated basal and stress-induced increments in norepinephrine release, reuptake, metabolism, turnover, and biosynthesis. Loss of adrenomedullary hormones after ADX did not appear to contribute to these increments. Cortisol treatment reversed the ADX effects on CA indices and suppressed catecholaminergic responses to IMO in intact rats. These results suggest that endogenous glucocorticoids restrain responses of catecholamine turnover, synthesis, release, reuptake, and metabolism during stress. In contrast, in intact rats, continuous administration of cortisol lasting for 7 days exaggerated the IMO-induced increases in plasma CA levels. Inhibition of DOPA conversion to dopamine elevated plasma DOPA levels in chronically cortisol-treated stressed rats compared to saline-treated ones, suggesting a cortisol-induced increase in tyrosine hydroxylation. Stress increases TH and PNMT activities and mRNA levels in the adrenal medulla. Hypophysectomy reduced adrenal PNMT but not TH mRNA levels in control and IMO rats. Pretreatment of hypophysectomized animals with ACTH fully restored the control and IMO-induced adrenal PNMT mRNA levels and augmented PNMT but not TH mRNA responses in intact rats. Long-term cortisol administration to intact rats also elevated adrenal PNMT but not TH mRNA levels. The results indicate a suppressive effect of endogenous glucocorticoids and a stimulatory effect of chronically elevated glucocorticoid levels on sympathoadrenal activity during stress. The results also suggest that a nonneuronal, nonpituitary factor contributes to TH gene expression during some forms of stress, whereas pituitary-adrenocortical factors play the essential role in the regulation of PNMT gene expression.

Adrenalectomy↗
PubMed1995

Studies on the tumor-promoting activity of additives in biomaterials: inhibition of metabolic cooperation by phenolic antioxidants involved in rubber materials.

For the detection of tumor-promoting activities of phenolic antioxidants, the inhibitory activities on the intercellular gap-junctional communication were investigated using the V79 metabolic cooperation (MC) assay. Among eight antioxidants, 4,4'-butylidene-bis(3-methyl-6-tert-butyl-phenol), 2,2'-methylene-bis(4-methyl-6-tert-butylphenol) (MBMBP), and styrenated phenol (SP) showed stronger inhibitory activities than lithocholic acid, which is known to be a tumor promotor. However, 4,4'-thio-bis(3-methyl-6-tert-butylphenol), Irganox 1010, and 1330 did not inhibit at any concentrations. When the single-electron oxidation potentials were compared among antioxidants, the electrochemical ease estimated with the first oxidation potential was correlated with the cytotoxic potentials (r = 0.88), but not with the inhibitory activities in an MC assay. The tumor-promoting activity of MBMBP was also investigated using an in vitro, two-stage Balb/c 3T3 transformation assay. MBMBP did not show initiating activity, but significant promoting activity at concentrations of both 1 and 2.5 micrograms/ml were noted. These concentrations were close to the lowest effective inhibitory concentration (1.3 micrograms/ml) of MBMBP in an MC assay. In conclusion, there is a possibility that the phenolic antioxidants that show inhibitory activities in an MC assay contribute to the enhancement of tumor incidence induced by biomaterials.

Animals↗
PubMed1995

Regulation of plasma osteocalcin by corticosterone and norepinephrine during restraint stress.

Osteocalcin (OC), an extracellular calcium-binding protein of bone origin, is synthesized by osteoblasts and binds with high specificity to bone mineral crystals. A small, but relatively consistent portion of newly synthesized OC which is released to circulation has been well correlated with histological indices of osteoblastic activity. Synthesis of OC is regulated by numerous hormones including glucocorticoids. We previously reported that mild mental stressors such as cage change or cold exposure decreased rat plasma OC by up to 40% within 1 h. A similar response was induced in a time- and dose-related manner by injection of physiological levels of corticosterone (CS), the active glucocorticoid in rats. Prone immobilization by foot restraint of conscious rats for up to 2 h (IMMO) is a well-characterized model of classic "fight-or-flight" response. This model induces an immediate and prolonged elevation of CS, as well as the catecholamines epinephrine (E) and norepinephrine (NE). In marked contrast to milder stressors, immobilization induced an immediate increase of plasma OC, greater than 50% within 5-20 min, which returned toward normal after 2 h of restraint. Selective ablation of the hormones by adrenal medulectomy, adrenalectomy, or blockade of sympathetic ganglia did not abolish the initial rapid rise of plasma OC. Even before IMMO, plasma OC was increased by about 50% in the absence of sympathetic neural function or adrenal CS production. The presence of both CS and NE, but not E, was required to return plasma OC concentrations to basal levels. This strongly suggests interaction of CS and NE to regulate plasma OC and its release from bone. As expected, prior cold exposure lowered plasma OC, but did not abolish a subsequent increase in response to IMMO, nor did IMMO repeated daily for 7 days. The stimulus for the initial rapid elevation of OC is unknown, but likely to be of importance in the role OC plays in response to stress. Further investigation of the OC under mental stress should help to understand the function of this abundant and highly conserved bone protein.

Adrenalectomy↗
PubMed1995

The primary structure of pepstatin-insensitive carboxyl proteinase produced by Pseudomonas sp. No. 101.

A unique carboxyl proteinase [EC 3.4.23.33] from Pseudomonas sp. No. 101 is the first example of a prokaryotic enzyme which is insensitive to the classical inhibitor, pepstatin. The primary structure of the proteinase was determined by conventional methods. Pseudomonas carboxyl proteinase consists of 370 amino acid residues with one disulfide bond. This enzyme has no homologous sequence with any other known carboxyl proteinase, including carboxyl proteinase B from Scytalidium lignicolum, which is a pepstatin-insensitive carboxyl proteinase. In addition, Pseudomonas carboxyl proteinase lacks the Asp*-Thr-Gly, Glu*-Thr-Gly, and Asp*-Thr-Ser-Gly (*indicates the catalytic residue) sequences which are known as the motif sequences around a pair of catalytic residues in carboxyl proteinases reported so far. The results strongly indicate that Pseudomonas carboxyl proteinase is a new type of carboxyl proteinase.

Amino Acid Sequence↗
PubMed1995

Decreased stress responsivity of central and peripheral catecholaminergic systems in aged 344/N Fischer rats.

We investigated the effects of stress on central and peripheral sympatho-adrenal and sympatho-neural functions in healthy, intact young (3-4 mo) and aged (24 mo) male Fischer 344/N rats. Extracellular fluid (ECF) levels of the catecholamines norepinephrine (NE), dihydroxyphenylglycol (DHPG), methoxyhydroxyphenylglycol (MHPG), and dihydroxyphenylacetic acid (DOPAC) were obtained by microdialysis in the paraventricular nucleus (PVN) of the hypothalamus at baseline and during immobilization (IMMO). The baseline levels of these substances were similar in both age groups, and their concentrations increased significantly in response to IMMO. The IMMO-induced increases of NE and MHPG, however, were significantly smaller in old than in young rats. Plasma levels of the catecholamines NE, DHPG, MHPG, DOPAC, dihydroxyphenylalanine (DOPA), epinephrine (EPI), dopamine (DA), and HVA were also determined in young and old rats during IMMO. Basal levels of these substances were significantly higher in old than in young rats. The magnitude of the IMMO-induced increases in the majority of these compounds however, was significantly smaller in old than in young rats. We conclude that, at the basal state, aging in the Fischer rat is associated with normal PVN ECF, but high plasma catecholamine levels; at stress state, however, old rats have substantially lesser activation of their central and peripheral catecholaminergic systems than young rats.

Adrenergic Fibers↗
PubMed1995

Establishment of a new extrahepatic bile duct carcinoma cell line, TFK-1.

A new human extrahepatic bile duct carcinoma cell line (TFK-1) was established from a surgically resected tumor specimen, which was histologically diagnosed as partly papillary adenocarcinoma and partly differentiated tubular adenocarcinoma. The tumor cells cultured in RPMI-1640 medium supplemented with 10% FBS grew as monolayers showing epithelial-like morphology with a population doubling time of 37 hr during exponential growth at passage 40. Chromosome number was distributed in the range of 72 to 76, with a modal number of 73. Tumor markers (CEA, CA19-9, ST-439, DUPAN-2) were negative in culture supernatant and plasma of SCID mice grafted with TFK-1 cells. Though no point mutation at 12 codon of K-ras was detected, expression of c-erb B-2 product and MUC1 antigen was positive. TFK-1 is the third cell line established from extrahepatic bile duct carcinomas in the world literature, and should provide useful information on various aspects of this type of neoplasm.

Adenocarcinoma↗
PubMed1994

Induction of tyrosine hydroxylase gene expression by a nonneuronal nonpituitary-mediated mechanism in immobilization stress.

Stress stimulates the sympathoadrenal system, causing activation of the catecholamine biosynthetic enzymes. Here we examine the changes of gene expression of tyrosine hydroxylase (TH; EC 1.14.16.2), the initial enzyme of catecholamine biosynthesis, with stress. A single immobilization of rats led to a large transient elevation in TH mRNA and a small elevation in TH immunoreactive protein and activity. Repeated daily immobilizations triggered more sustained changes in TH mRNA levels. After two immobilizations, the levels remained elevated even 3 days later. The rise in TH mRNA was followed by increased immunoreactive protein but only a small elevation in activity. With seven repeated immobilizations, the animals did not appear to adapt and still manifested a further rise in TH mRNA. TH activity was markedly elevated and returned to control levels 7 days after the immobilization. The rise in TH mRNA with a single immobilization occurred even in adrenals of hypophysectomized rats that underwent splanchnic nerve section. Immobilization for 30 min was sufficient to increase TH mRNA. The effect was abolished by the transcriptional inhibitor actinomycin D. Mobility gel-shift assays revealed increased binding of c-Fos and c-Jun to the AP-1 transcription factor site after a single immobilization, and the binding was not further elevated with repeated stress. This study shows that a single immobilization can activate TH gene expression by a nonneuronal nonpituitary-mediated pathway associated with increased binding of AP-1 transcription factors.

Adrenal Glands↗