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Biomedical subjects

K Fujita

Publications and source records attributed to K Fujita.

At least 19 recordsLinked to original sources
PubMed1992

Mouse dopamine beta-hydroxylase: primary structure deduced from the cDNA sequence and exon/intron organization of the gene.

Genomic clones for mouse dopamine beta-hydroxylase (DBH) were isolated from two genomic libraries derived from DBA/2J and 129/SV mouse strains, by plaque hybridization with the human DBH cDNA probe. Subsequently, cDNA encoding mouse DBH was amplified with reverse transcription-polymerase chain reaction (RT-PCR) method using primers corresponding to 5'- and 3'-portions of the mouse DBH mRNA, subcloned into a plasmid vector, and subjected to nucleotide sequence analysis. The clone encoded a protein of 621 amino acids with a calculated molecular mass of 70,186 daltons. The predicted amino acid sequence of mouse DBH showed 87%, 80% and 79% identities with the rat, bovine and human enzymes, respectively. Several potential amino acid sequences that are involved in the posttranslational modification and catalytic function of DBH were identified in mouse DBH protein. Nucleotide sequence analysis of the overlapping genomic clones showed that the mouse DBH gene was composed of 12 exons about 17 kb in length. Typical TATA and CCAAT boxes were observed in the 5'-upstream region of the gene. Northern blot analysis of adrenal gland RNA detected a single size species of the mouse DBH mRNA.

Adrenal Glands↗
PubMed1992

Tissue-specific alternative splicing of the first exon generates two types of mRNAs in human aromatic L-amino acid decarboxylase.

Aromatic-L-amino-acid decarboxylase (AADC) is an enzyme that plays an essential role in synthesizing catecholamines and serotonin in neuronal and endocrine tissues. AADC has also been detected in other nonneuronal tissues including liver and kidney, although its physiological role in nonneuronal tissues has not yet been defined. Previously we have cloned a human AADC cDNA from a neuronal tissue (pheochromocytoma) [Ichinose, H., Kurosawa, Y., Titani, K., Fujita, K., & Nagatsu, T. (1989) Biochem. Biophys. Res. Commun. 164, 1024-1030] and the corresponding genomic DNA [Sumi-Ichinose, C., Ichinose, H., Takahashi, E., Hori, T., & Nagatsu, T. (1992) Biochemistry 31, 2229-2238]. Here we present isolation and characterization of AADC cDNA and genomic DNA from a nonneuronal tissue (human liver). The nonneuronal and neuronal AADC mRNAs differed only in the region corresponding to the untranslated first exon. The first exon for the nonneuronal-type mRNA was located 4.2 kilobases upstream to that for the neuronal-type mRNA and 22 kilobases from exon 2, to which it is spliced. Determination of the transcription initiation site indicated that the length of the nonneuronal-type exon 1 was 200 bp. A TATA box-like motif was located between positions -26 and -20 from the transcription initiation site. These results showed that an alternative usage of the first exon in the 5'-untranslated regions produces two types of mRNAs in AADC and suggested that alternative splicing would regulate the tissue-specific expression of AADC.

Adrenal Glands↗
PubMed1992

Multiple mRNA forms of human GTP cyclohydrolase I.

To isolate full length cDNA clones encoding human GTP cyclohydrolase I, the first and rate-limiting enzyme in tetrahydrobiopterin biosynthesis, a cDNA library generated from human liver was screened by plaque hybridization. Analysis of the clones, hybridized with rat cDNA fragment, by restriction mapping and partial sequencing showed the existence of three kinds of cDNAs. All three cDNAs were identical in their central and 5' regions. They were, however, found to diverge at 3' ends. Furthermore, the three species of mRNAs corresponding to the three cDNAs were detected in human liver by reverse transcription-polymerase chain reaction (RT-PCR) analysis. These results indicate that, in humans, GTP cyclohydrolase I molecules are encoded by at least three distinct mRNAs.

Amino Acid Sequence↗
PubMed1992

In rat brain amoxapine enhances dopamine metabolism: pharmacokinetic variations of the effect.

When rats were given i.p. amoxapine, the drug was biotransformed to 7-hydroxyamoxapine, but not to 8-hydroxyamoxapine. The maximal concentrations of amoxapine and 7-hydroxyamoxapine in the serum and brain were found 30 min after the single injection, and the concentration of the former in the brain was higher than that of the latter. During the chronic treatment the concentration of amoxapine in the brain was much higher than that of 7-hydroxyamoxapine. A single administration of amoxapine increased the brain levels of dihydroxyphenylacetic acid and homovanillic acid. Their highest levels were observed 6 h after the injection. Repeated administration reduced the increases; chronic treatment caused tolerance to the enhancing effects on dopamine (DA) metabolism. Tolerance was observed in both striatum and hippocampus, but not in frontal cortex and hypothalamus. Single or chronic injection did not appear to change the level of DA in the brain. Amoxapine itself could be chiefly responsible for the enhancement of DA metabolism. In addition, the level of 3-methoxy-4-hydroxyphenylethyleneglycol in brain decreased transiently right after the injection of amoxapine, but the norepinephrine level did not seem to change following single or chronic administration of amoxapine.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1992

Genetic alteration of catecholamine specificity in transgenic mice.

Epinephrine-producing cells are characterized by the presence of phenylethanolamine N-methyltransferase (PNMT), which catalyzes the formation of epinephrine from norepinephrine. We generated a line of transgenic mice carrying a chimeric gene containing human PNMT cDNA fused to the 4-kilobase fragment of the human dopamine beta-hydroxylase (DBH) gene promoter, to switch catecholamine phenotype in the nervous and endocrine systems. Human PNMT transcripts and immunoreactivity were mainly detected in norepinephrine neurons in brain and sympathetic ganglion as well as in norepinephrine-producing cells in adrenal medulla of transgenic mice, indicating that the human DBH gene promoter of 4 kilobases is sufficient to direct expression of the gene in norepinephrine-producing cells. Analysis of catecholamines in the various tissues showed that the expression of human PNMT in transgenic mice induced the appearance of epinephrine in sympathetic ganglion and dramatic changes in norepinephrine and epinephrine levels in brain, adrenal gland, and blood. These results indicate that the additional PNMT expression in norepinephrine-producing cells can convert these cells to the epinephrine phenotype, and suggest that norepinephrine-producing cells normally possess the basic machinery required for the synthesis of epinephrine except for PNMT. Thus it appears that the only major difference between norepinephrine- and epinephrine-producing cells is the expression of PNMT. Our transgenic animals provide an experimental model to investigate the functional differences between norepinephrine and epinephrine.

Animals↗
PubMed1992

Predictive factors for intrahepatic recurrence of hepatocellular carcinoma after partial hepatectomy.

To establish useful predictors of the intrahepatic recurrence of hepatocellular carcinoma (HCC) after partial hepatectomy, retrospective analyses of clinical and pathologic factors were done in 112 of 206 patients treated by partial hepatectomy. The absence or presence of intrahepatic recurrence was confirmed by a follow-up study. Cancer-free survival rates after 1, 2, 3, and 5 years were 54.8%, 36.7%, 32.5%, and 25.6%, respectively. The significant factors affecting recurrence were tumor size, number of tumors, cancer cell infiltration of the fibrous capsule of the tumor, portal involvement, and stage of the tumor, but the grade of anaplasia according to Edmondson-Steiner's classification and the severity of associated liver cirrhosis did not show a correlation with the incidence of recurrence. According to Akaike's Information Criteria (AIC), tumor number is useful for predicting early prognosis, and capsular infiltration is a good indicator of long-term survival. However, portal involvement gives much prognostic information throughout the entire postoperative period.

Adult↗
PubMed1992

Development of the ability to detect visual motion in infant macaque monkeys.

A preferential looking technique was used to measure detection of visual motion by 12 infant macaque monkeys (10 Macaca fuscata and 2 Macaca mulatta at ages between 1 and 100 days). A 0.25 cycles/deg square-wave grating was presented at speeds from 40 to 0.4 deg/s. The threshold was determined by a two-alternative forced-choice preferential looking method, in conjunction with a staircase procedure. The threshold for detection of visual motion decreased exponentially with age over the first 100 days of life. At each age there was substantial variability among the individual infants tested, but each monkey's ability to detect visual motion improved with age.

Aging↗
PubMed1992

In vitro adherence of Staphylococcus saprophyticus, Staphylococcus epidermidis, Staphylococcus haemolyticus, and Staphylococcus aureus to human ureter.

Staphylococcus saprophyticus adhered to human ureteral epithelium in vitro. The levels of adherence, which were determined quantitatively with the scanning electron microscope, correlated well with bacterial hemagglutinating activities with sheep erythrocytes (r = 0.9459, P < 0.01). Transmission electron microscopy revealed that the adhering bacteria and the hemagglutinating bacteria possessed similar pili-like structures on their cell surfaces. Staphylococcus epidermidis, Staphylococcus haemolyticus, and Staphylococcus aureus did not adhere to the epithelium. Only S. aureus adhered markedly to the connective tissue of the ureter, and adhesion of this organism was direct via its cell wall. This adherence test system clearly showed up differences in the abilities of these staphylococcal species to adhere to the urinary tract.

Bacterial Adhesion↗
PubMed1992

The association of hypopituitarism with small pituitary, invisible pituitary stalk, type 1 Arnold-Chiari malformation, and syringomyelia in seven patients born in breech position: a further proof of birth injury theory on the pathogenesis of "idiopathic hypopituitarism".

We report seven cases of hypopituitarism all having a history of breech delivery, asphyxia at birth, and syringomyelia. A small pituitary gland was found on MRI or CT in six cases, invisible pituitary stalk on MRI in five cases, and type 1 Arnold-Chiari malformation in six cases. A constellation of these abnormalities are best explained by traction of brain and spinal cord of the subjects exerted during breech delivery and further support the primary role of birth trauma in the genesis of "idiopathic hypopituitarism".

Adolescent↗
PubMed1992

Cytochrome b positive X-linked chronic granulomatous disease: a normal cell surface expression of cytochrome b.

The polymorphonuclear (PMN) cells from a patient with cytochrome b positive X-linked chronic granulomatous disease (Xb+ CGD) were studied using flow cytometry. Both the cell surface expression of monoclonal antibody defined cytochrome b and the superoxide production (intracellular 2',7'-Dichlorofluorescin Diacetate oxidation) were investigated at a single cell level. Flow cytometry clearly demonstrated the complete absence of superoxide production in the patient's PMN cells, the mosaicism in his mother's PMN cells and also indicated the normal cell surface expression of cytochrome b. The results obtained by Western blot analysis and reduced-minus-oxidized spectra confirmed the presence of functional and normal amounts of cytochrome b. We concluded that this is a case of Xb+ CGD with a normal cell surface expression of cytochrome b.

Adolescent↗
PubMed1992

Congenital chylothorax in a trisomy 21 newborn.

We here report a case of trisomy 21 with congenital chylothorax. The patient was a male newborn who had been diagnosed as having trisomy 21 with congenital chylothorax. This is the fifth case of the both conditions in English literature. Congenital chylothorax is very rare in a trisomy 21 patient. There, however, may be causal relationship between the two conditions.

Chylothorax↗
PubMed1992

Identification of the phenotypic modulation of rabbit arterial smooth muscle cells in primary culture by flow cytometry.

In atherosclerotic lesions, smooth muscle cells (SMC) change from a contractile to a synthetic phenotype. The in vivo and in vitro phenotypic transformations of SMC have been confirmed by transmission electron microscopy (TEM), but the relationship between this change and the cell cycle is still unknown. We demonstrated the structural modulation of rabbit arterial SMC in primary culture by TEM and immunocytochemistry and simultaneously studied changes in two-dimensional histograms of the relative DNA and RNA contents by flow cytometry. During the first day of primary culture, the cells exhibited the contractile phenotype and were composed of a population in the G0 phase characterized by low contents of DNA and RNA. On the second day of culture, some of the cells (18.2%) had started but not completed the transition into the synthetic phenotype and a cell population in the G1A phase with an RNA content above the G0 level appeared in almost the same proportion. This cell population could be categorized as an "intermediate" type. Moreover, after 3 days when about three-quarters of the cells had undergone structural transition, the same proportion of cells had entered into the cycling phase, while some cells still remained in the G0 and G1A phases. Thus, cell cycle analysis by flow cytometry corresponded well with the observations obtained by TEM and immunocytochemistry. These results show that flow cytometry can rapidly and relatively conveniently monitor the process of phenotypic modulation in SMC and is a useful method for the analysis of such transitions.

Animals↗
PubMed1992

Brugia pahangi: production of a monoclonal antibody reactive with the surface of infective larvae.

Monoclonal antibodies against infective third-stage larvae (L3) of Brugia pahangi were generated from mice immunized with L3 antigens. The monoclonal antibodies were L3 stage-specific or stage-nonspecific. A BpG1 monoclonal antibody (IgG1 subclass) showing L3 stage-specificity was examined in detail. BpG1 recognized the surface of B. pahangi L3 and also reacted with the surface of Brugia malayi L3 but not with the surface of filarial worms of other genera, such as Acanthocheilonema viteae and Litomosoides carinii. BpG1 promoted cellular adhesion to the surface of B. pahangi L3. BpG1 bound on living L3 was shed but the shedding rate was relatively slow. The surface antigen recognized by BpG1 had a molecular weight of 58 kDa. It was stable to heat and periodate treatments but sensitive to trypsin digestion and was released from living L3 by SDS but not by Triton X-100 or CTAB. Preincubation of L3 with BpG1 significantly reduced the recovery rate of worms compared with the preincubation with a monoclonal antibody (IgG1 subclass) against the inner tissues of B. pahangi L3 or control supernatant of P3U1 myeloma cells. This result suggests that the antigen containing the BpG1 epitope may be one of the targets of a protective immune response against Brugia infection.

Animals↗
PubMed1992

Effect of anticancer agents on cell cycle of regenerating hepatocytes in rats.

The effect of three anticancer agents, cisplatin, doxorubicin, and mitomycin, on liver regeneration after 70% partial hepatectomy in rats was investigated by total DNA content of the liver and flow cytometric analysis of hepatocyte nuclei using two-color staining of anti-bromodeoxyuridine monoclonal antibody and propidium iodide. Total DNA content of regenerating liver 7 days after hepatectomy showed significant suppression of regeneration by these agents (P less than 0.01). The inhibitory effects of the agents on the cell cycle of hepatocytes by flow cytometric analysis were (1) a delay of the peak or a decrease in the proportion of S phase nuclei and/or (2) polyploidization of the nuclei, demonstrated by accumulation of 8c and occasionally 16c nuclei, of which the DNA contents were four and eight times as much as that of diploid (2c) cell nuclei, respectively. The former (1) suggests G0 or G1 phase block, and the latter (2) G2 phase block. In terms of total DNA content of regenerating liver, the inhibitory effect was most prominent in the cisplatin-administered groups. The polyploidization of nuclei was most remarkable in the mitomycin-administered groups. Although the total DNA content recovered to the level of control at 6 weeks after hepatectomy, the polyploidization effect persisted in the drug-administered groups. These agents had no cytocidal action on proliferating hepatocytes as can be seen from the aspartate aminotransferase and alanine aminotransferase serum levels. We conclude that in a short-term observation, the anti-cancer agents significantly inhibit liver regeneration, although the inhibitory effect on DNA synthesis does not last long.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

The activity of dipeptidyl peptidase II and dipeptidyl peptidase IV in mice immunized with type II collagen.

We investigated the activity of peptidases in the serum of mice with experimental polyarthritis that was induced by the injection of type II collagen, an experimental model of human rheumatoid arthritis. The activity of dipeptidyl peptidase II (DPP II) was increased and that of dipeptidyl peptidase IV (DPP IV) was decreased resulting in the significant increase of the serum DPP II/DPP IV ratio in the polyarthritic mice compared with that of controls. These results indicate that the DPP II/DPP IV ratio is a novel index of disease activity in mice with collagen-induced polyarthritis and may be useful in assessing the activity of rheumatoid arthritis in humans.

Animals↗
PubMed1992

Neuron-specific enolase reduction in irradiated salivary glands of the rat--an immunohistochemical study.

Neuron-specific enolase (NSE) in the salivary glands of normal and irradiated rats was studied by immunohistochemical methods. The normal salivary gland of Sprague-Dawley rats (465 g) showed positive staining for NSE in striated ducts and granular convoluted tubule (GCT) cells. A single radiation, TDF (time, dose, fractionation factor) 100 and 200 (18.82 Gy and 27.97 Gy, respectively) was done, and 4 groups (1, 2, 3 and 4 weeks after radiation) of 5 rats each were used. Irradiated salivary glands indicated a remarkable reduction of NSE staining in GCT cells and a reduction but to a lesser degree in the striated duct of the submandibular gland. Immunohistochemical deposition of NSE was not changed in the sublingual glands of irradiated rats. The reduction of NSE immunodeposition was irradiation dose dependent.

Animals↗