PubMed Health⌕ Search

Biomedical subjects

M Mizobuchi

Publications and source records attributed to M Mizobuchi.

At least 37 records · Page 2Linked to original sources
PubMed1996

Thyrotropin-releasing hormone release in normal and hyperthyroid rats as measured by microdialysis.

To measure the thyrotropin-releasing hormone (TRH) release from the hypothalamus in euthyroid and hyperthyroid states, we investigated the changes in TRH levels in the anterior pituitary of conscious male rats using an in vivo microdialysis technique. In the euthyroid rats (n = 18), TRH levels in the extracellular dialysates of the anterior pituitary varied from 1.0 to 101.0 pg/ml in a pulsatile fashion: 15.9 +/- 13.9 (mean +/- SD) pg/ml with 5.8 +/- 1.5 pulses/24 h. In the hyperthyroid rats (n = 5) who received L-thyroxine at 10 micrograms/100 g body weight for 7 days, TRH levels in the dialysates during 6 h was 3.6 +/- 1.7 pg/ml, and significantly lower (P < 0.05) than those of the control rats (15.5 +/- 14.1 pg/ml); the pulse frequency was unchanged. These findings demonstrated for the first time the existence of pulsatile TRH release from the hypothalamus, and showed that thyroid hormone inhibits TRH release by the reduction of pulse amplitude, but not of pulse frequency.

Animals↗
PubMed1995

Hampered expression of isoaspartyl protein carboxyl methyltransferase gene in the human cataractous lens.

Isoaspartyl protein carboxyl methyltransferase (PIMT) is implicated in the repair of age-damaged proteins by converting altered aspartic acid residues to normal L-aspartic acid residues. Northern blot and reverse transcription (RT)-PCR analyses have revealed that PIMT gene expression in the human lens is detected exclusively in epithelial cells, and that the mRNA levels in cataractous lens epithelia are significantly lower than those in normal age-matched lens tissue. These results suggest that PIMT may play a vital role in maintaining the clarity of the lens and preventing cataract formation.

Aspartic Acid↗
PubMed1995

Complete primary structure and phosphorylation site of the 65-kDa macrophage protein phosphorylated by stimulation with bacterial lipopolysaccharide.

There is ample evidence that intracellular protein phosphorylation is a mandatory event in the process of macrophage activation by LPS, yet how this event is initiated and what roles the phosphorylated proteins are assigned to are poorly understood. We previously isolated a 65-kDa cytosolic protein (pp65) that was phosphorylated specifically in LPS-stimulated murine macrophages. In the present study, the complete primary structure of pp65 was determined on the basis of the cDNA containing an open reading frame of 1881 bases. The sequence of pp65 revealed that it is a murine homologue of human L-plastin, recently identified as a novel transformation-induced polypeptide of neoplastic human cells, and that it contains a unique series of Ca2+, calmodulin, and actin binding domains. A single phosphorylated peptide was isolated from the tryptic digest of pp65 by reverse-phase HPLC. From the amino acid sequence of the dodecapeptide Gly-Ser-Val-Ser-Asp-Glu-Glu-Met-Met-Glu-Leu-Arg, the phosphorylation site of pp65 was located at the N-terminal region adjacent to the first Ca2+ binding domain. This sequence contains a repeat of the casein kinase II motif Ser-Xxx-Xxx-Glu/Asp and, together with the preceeding Arg residue, constitutes the consensus sequence Arg-Xxx-Ser for cAMP-dependent protein kinase (PKA) and protein kinase C (PKC), but not mitogen-activated protein kinase (MAPK)-specific motif is found. These results, taken together with previous observations on the process of macrophage activation by LPS, demonstrate that pp65 is phosphorylated by an LPS-induced protein kinase other than MAPK and exerts its function on the cytoskeleton in a Ca2+/calmodulin-dependent manner.

Amino Acid Sequence↗
PubMed1995

Characterization of three cDNAs encoding two isozymes of an isoaspartyl protein carboxyl methyltransferase from human erythroid leukemia cells.

Two different sizes (approximately 1.0 and 1.6 kb) of transcripts of an isoaspartyl protein carboxyl methyltransferase (PIMT) were detected in eight cell lines derived from human hemopoietic cells on Northern blot analysis. We found three different sizes of cDNAs (907, 1,553, and 1,600 bp) in human erythroid leukemia cells (HEL) and a unique cDNA sequence corresponding to the 1.0 kb transcript was identified. These three cDNA sequences encoded two isozymes consisting of 226 (isozyme I) and 227 (isozyme II) amino acids. The 1.6 kb transcript was translated into two isozymes (isozyme I and II), while the 1.0 kb transcript was only translated into isozyme I. These results suggest that the two isozymes deduced from the cDNAs of the human erythroid leukemia cells may exist in normal human erythrocytes.

Amino Acid Sequence↗
PubMed1995

Growth hormone-releasing hormone immunoreactivity in mouse placenta, maternal blood, and amniotic fluid: molecular characterization and secretion from primary cell cultures in vitro.

The GH-releasing hormone (GRH) gene, along with those of many other hypothalamic hormones, is abundantly expressed in mouse and rat placenta. The presence of GRH immunoreactivity (GRH-IR) is described in mouse placenta, maternal blood, and amniotic fluid, and its molecular form has been characterized using HPLC. Two different molecular forms of mouse GRH-IR (mGRH-IR) were detected in the mouse hypothalamus and one in placenta. Twenty-five percent of mGRH-IR in the hypothalamus corresponded to mGRH(1-42)OH, whereas the remainder, and all of the mGRH-IR in placenta, had a retention time consistent with the GRH precursor. High levels of mGRH-IR were detected in both maternal plasma and amniotic fluid. In addition, a mouse placental cell primary culture system was established to study the regulation of mGRH-IR release. Turnover of mGRH in placental cells was rapid, resulting in a 24-h media content of 10 times that present in cells. Both 1-oleoyl-2-acetyl-sn-glycerol and 1,2-dioctanoyl-sn-glycerol significantly stimulated the release of mGRH-IR from cultured placental cells into the incubation media but had no effect on total peptide synthesis. These results suggest that the release of mGRH-IR from placental cells is mediated, at least in part, by the activation of protein kinase C. The HPLC elution profiles of mGRH-IR released from placental cells under basal and 1-oleoyl-2-acetyl-sn-glycerol-stimulated conditions were similar to those in placental tissue. Although the biological function of mGRH-IR in placental, maternal plasma, and amniotic fluid is not yet clear, the presence of mGRH-IR in these tissues and circulating fluids suggests the possibility that mGRH-IR may exert an important role in both fetal and maternal physiology.

Amniotic Fluid↗
PubMed1995

Differential gene expression of growth hormone (GH)-releasing hormone (GRH) and GRH receptor in various rat tissues.

Growth hormone (GH)-releasing hormone (GRH) acts on specific receptors in the anterior pituitary to stimulate the synthesis and release of GH. Recent reports suggest that GRH is also synthesized in extrahypothalamic tissues. To evaluate the potential roles of extrahypothalamic GRH, we studied the gene expression of GRH and GRH receptors in various rat tissues by reverse transcribed (RT)-polymerase chain reaction (PCR). Total RNA was extracted from twenty-three rat organs and RT-PCR was performed with GRH and GRH receptor primers. Highly-sensitive RT-PCR-Southern blotting showed that GRH and GRH receptor mRNA coexist in the widespread tissues (14 of 25 tissues). GRH mRNA was relatively abundant in the cerebral cortex, brain stem, testis, and placenta, while GRH receptor mRNA was abundant in renal medulla and renal pelvis. Northern blot hybridization using poly A+ RNA indicated that the transcript of GRH receptor gene found in the renal medulla was similar to the longer transcript (about 4 Kb) of pituitary GRH receptor in the size. These results suggest that GRH plays a potential role not only in the neuroendocrine axis, but also in the autocrine and paracrine systems in extrahypothalamic tissues.

Animals↗
PubMed1995

The orbital and medial prefrontal circuit through the primate basal ganglia.

The ventral striatum is considered an interface between limbic and motor systems. We followed the orbital and medial prefrontal circuit through the monkey basal ganglia by analyzing the projection from this cortical area to the ventral striatum and the representation of orbitofrontal cortex via the striatum, in the globus pallidus and substantia nigra. Following injections of Lucifer yellow and horse radish peroxidase into the medial ventral striatum, there is a very densely labeled distribution of cells in areas 13a and 13b, primarily in layers V and VI, and in medial prefrontal areas 32 and 25. Injections into the shell of the nucleus accumbens labeled primarily areas 25 and 32. The reaction product in the globus pallidus and the substantia nigra supports previous studies demonstrating that efferent projections from the ventral striatum are represented topographically in the ventral pallidum and nontopographically in the substantia nigra, pars compacta. Tritiated amino acid or PHA-L tracer injections into orbitofrontal cortex produce dense patches of terminal labeling along the medial edge of the caudate nucleus and the dorsal part of the nucleus accumbens. These results demonstrate that the orbital prefrontal cortex projects primarily to the medial edge of the ventral striatum and to the core of the nucleus accumbens. The arrangement of terminals in the globus pallidus and substantia nigra show two different patterns. Thus, the orbital and medial prefrontal cortex is represented in a confined region of the globus pallidus but throughout an extensive area of the dorsal substantia nigra. Terminals are extensive throughout the region of the dopaminergic neurons, suggesting that this input may influence a wide area of both the striatum and frontal cortex.

Animals↗
PubMed1994

Tissue-specific expression of isoaspartyl protein carboxyl methyltransferase gene in rat brain and testis.

Isoaspartyl protein carboxyl methyltransferase (PIMT) is widely distributed in mammalian tissues. Using a polymerase chain reaction-generated 124-bp DNA fragment from brain cDNA as a probe, four different sizes (approximately 4.0, 2.5, 1.7, and 1.1 kb) of transcripts were detected with northern blot analysis. They were expressed predominantly in rat brain and testis. The major transcripts were 2.5 and 1.7 kb in the brain and 2.5 and 1.1 kb in the testis. One of the major transcripts specific to the testis (1.1 kb) was determined to study the structural difference of major transcripts in the two tissues. This testicular cDNA had neither the 5' (94 nucleotides) nor the 3' (594 nucleotides) end of previously reported brain cDNA corresponding to 1.7 kb. The mRNA levels and enzyme activities of different regions and developmental changes were examined in the brain. The mRNA levels and enzyme activities were concomitantly high in cerebral cortex and hippocampus. Although they increased rapidly approximately 30 days after birth in the testis and decreased in aged rats, they increased gradually after birth and remained high during the aging of the brain. Both structural and developmental studies show that the expression of the PIMT gene in brain and testis is regulated in a tissue-specific manner.

Aging↗
PubMed1994

Simple quantitative reverse transcribed-polymerase chain reaction (RT-PCR) method involving recombinant RNA generated by a false-priming PCR product.

A simple and efficient method for preparing rcRNA through competitive RT-PCR has been developed. The basis of this method is the use of a false-priming PCR product including the same primers as a specific product. A 290 bp fragment obtained by two-step PCR was subcloned into a plasmid vector and then the cloned DNA was transcribed into rcRNA. After competitive RT-PCR using sample RNA and rcRNA had been carried out, Southern blot hybridization was performed. The method was applied to determine the amounts of PIMT mRNAs in the rat pituitary. The quantitative analysis indicated that an at least 2-fold difference in PIMT mRNA level can be accurately determined with our method.

Animals↗
PubMed1994

Day-night variation of urine volume in Parkinson's disease.

Studies on the circadian rhythm of urine excretion in healthy men have demonstrated that the maximal urine flow occurs in the early afternoon and the minimal around midnight. In this study, an abnormality in the variation of urine volume was found in parkinsonian patients. Urine samples were collected during daytime (9:00-21:00) and nighttime (21:00-9:00). Fifteen healthy control subjects were examined and found to excrete 60% during the daytime and 40% during the nighttime of the total urine volume. Sixteen parkinsonian patients excreted 43% during the daytime and 57% during the nighttime. In contrast to the control subjects, the parkinsonian patients excreted a smaller volume of their urine during the daytime than during the nighttime. This finding might be related to the degeneration of dopaminergic and/or nondopaminergic neurons in the brain which control urinary excretion.

Adult↗
PubMed1994

Acute effects of hypoglycemia and hyperglycemia on hypothalamic growth hormone-releasing hormone and somatostatin gene expression in the rat.

Although previous studies have indicated that changes in the plasma glucose concentration alter GH secretion in the rat, the roles of hypothalamic GH-releasing hormone (GRH) and somatostatin (SS) in the glucose modulation of GH secretion have not yet been elucidated. We investigated the acute effects of hypo- and hyperglycemia on hypothalamic GRH and SS mRNA in the rat. Long term atrial catheters were implanted in male rats, and sequential blood sampling was performed in conscious animals. Insulin and 50% glucose were administered iv to induce hypo- and hyperglycemia, respectively. At the end of the experiment, the hypothalamus was removed, and total RNA was extracted for Northern blotting. Hypoglycemia completely inhibited pulsatile GH secretion, whereas hyperglycemia partially inhibited GH secretion. Direct effects of insulin itself on GH secretion in the insulin-induced hypoglycemia were ruled out by glucose clamp studies. Hypothalamic SS mRNA levels were increased dramatically by hypoglycemia (266 +/- 37%) and moderately by hyperglycemia (183 +/- 45%). Hypothalamic GRH mRNA levels were increased only by hyperglycemia (159 +/- 18%). The GH response to GRH was decreased dramatically by hypoglycemia and moderately by hyperglycemia. These results indicated that acute changes in the plasma glucose concentration stimulated hypothalamic SS mRNA in the rat.

Animals↗
PubMed1994

Development of neuroepithelial tumors of the adrenal medulla in transgenic mice expressing a mouse hypothalamic growth hormone-releasing hormone promoter-simian virus-40 T-antigen fusion gene.

Expression of the mouse GH-releasing hormone (GRH) gene is restricted to neurons within the hypothalamus and to placenta. In an attempt to generate immortalized mouse hypothalamic neurons expressing GRH, the proximal 872-nucleotide segment of the 5'-flanking region of the hypothalamic mouse GRH gene was cloned by polymerase chain reaction and ligated to a 2.7-kilobase DNA sequence encoding the simian virus-40 (SV40) T-antigen, so that regulation of SV40 T-antigen expression was dependent on sequences within the mGRH 5'-flanking region. This region contains both TATA and CAAT boxes. The mouse GRH/SV40 T-antigen fusion gene was injected into 1-cell mouse embryos, and SV40 T-antigen incorporation in the mouse genome was found in 11 of 77 live births (3 males and 8 females). Although no evidence of hypothalamic tumors was found, all mice that expressed the transgene also developed tumors originating in the adrenal medulla. Gene copy number varied from 1-20 and was inversely proportional to survival, which ranged from 7-16 weeks. Corticosterone levels were normal. The male transgenic mice were fertile, and their progeny expressed the transgene and developed similar tumors. Microscopic examination of the tumors revealed a primitive neuroectodermal neoplasm that exhibited hematogenous and lymph node metastases and contained 100 ng norepinephrine, 2.85 ng epinephrine, and 1.1 ng dopamine/mg tumor tissue. Primary culture of dispersed tumor cells released norepinephrine into the medium (180 pg/ml.24 h). Cell lines from 2 tumors were established and exhibited characteristics similar to those of mixed neuroblastoma or primitive neuroectodermal tumors. In conclusion, the proximal 872 nucleotides of the hypothalamic mouse GRH promoter contain elements directing tissue-specific expression limited to early adrenal neuroectodermal cells. Other GRH DNA sequences appear to be required for restricted expression of mouse GRH within the hypothalamus.

Adrenal Gland Neoplasms↗
PubMed1994

Syndrome of inappropriate secretion of ADH (SIADH) due to small cell lung cancer with extremely high plasma vasopressin level.

A 76-year-old man with small cell lung cancer associated with the syndrome of inappropriate secretion of ADH (SIADH) visited our hospital. The serum Na level was normal on the first visit, but 2 weeks later it decreased to 114 mEq/L with an extremely high plasma vasopressin (VP) level of 1520 pg/ml. Serum Na was normalized after the reduction of the tumor size by chemotherapy, but the plasma VP level remained between 150 to 600 pg/ml. On gel filtration of plasma VP two peaks of immunoreactive VP were eluted at the positions of a larger molecule than authentic VP and authentic VP, and VP in urine gave only one peak compared to that of authentic VP. The dilution curve of plasma VP was almost parallel and that of urine was completely parallel to the standard curve. These findings suggest that a larger VP with low physiological activity was predominantly secreted in the present patient and manifested relatively mild symptoms despite the extremely high plasma VP level.

Aged↗
PubMed1994

Biotransformation of a new synthetic retinoid, 4-[(5,6,7, 8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)carbamoyl] benzoic acid (Am-80), in the rat. Structure elucidation of the metabolites by mass and nmr spectrometry.

1. The major metabolites of Am-80 in rat bile were examined by tlc-radiochromatography and hplc-radiochromatography after the intravenous administration of 14C-Am-80, and > 15 metabolites were detected. 2. Intact and Glusulase-treated bile were analysed by both tlc-radiochromatography and hplc-radiochromatography. As both samples gave similar patterns in both chromatography systems, we have concluded that neither glucuronide nor sulphate conjugates were present. 3. 2H3-Am-80 and 2H6-Am-80 were administered to rats, and the major metabolites in bile were detected and their structures elucidated by GC-MS. 4. The chemical structures of seven major biliary metabolites, including unchanged Am-80, were identified by mass and two-dimensional nmr spectrometry, and included 6-hydroxy-Am-80 (M-3), 7-hydroxy-Am-80 (M-4), 6-oxo-Am-80 (M-5), unchanged Am-80 (M-0) and the taurine conjugates of M-3 (M-1), M-4 (M-2) and M-0 (M-6).

Animals↗
PubMed1994

[Adhesion molecules and immune-mediated diseases of the central nervous system].

Expression of adhesion molecules in immune-mediated diseases of the central nervous system was reviewed. In multiple sclerosis and experimental allergic encephalomyelitis (EAE), endothelial cells of active lesions increase expression of the adhesion molecules such as ICAM-1, VCAM-1 and inflammatory cells including memory T cells and macrophages express high levels of adhesion molecules such as LFA-1, VLA-4. Astrocytes also express CD44, ICAM-1, VCAM-1 and E-selectin in response to cytokine stimuli. In EAE, the majority of infiltrating cells are not MBP-specific memory T cells, thus it is speculated that the up-regulation of the adhesion molecules in the endothelial cells plays a decisive role in the development of immune-mediated diseases of the central nervous system. Therapeutic potency of clinical usage of anti-adhesion molecule antibodies has been explored.

Animals↗
PubMed1993

High levels of quinolinic acid in brain of epilepsy-prone E1 mice.

Quinolinic acid (QUIN) may act.as an excitotoxin when it is abundant in the brain. We have shown previously that the activity of 3-hydroxyanthranilate 3,4-dioxygenase, a QUIN-synthesizing enzyme, was abnormally high in the brains of epilepsy-prone E1 mice as compared with that of ddY mice. Here, we estimated the QUIN contents in the brains of these mice. The results showed that the basal QUIN content in the cerebral cortex of E1 mice was twice as high as that of ddY mice. Systemic injection of 400 mumol/kg body weight of L-tryptophan (L-Trp) increased the cortical levels of QUIN in both E1 mice and ddY mice by 189% and 118%, respectively. Administration of 400 mumol/kg each of L-threonine and D,L-methionine had no appreciable effect on the L-Trp-caused increase in the cortical QUIN levels. Co-administration of 5-fluorotryptophan or 5-methyltryptophan, tryptophan analogs, with L-Trp did not reduce but rather enhanced the cortical QUIN levels (by 18% and 92%, respectively). No significant change in the cortical QUIN concentrations was observed with injection of 2 mg/kg body weight of E. coli lipopolysaccharide (LPS) in E1 mice. However, injection of L-Trp in the LPS-treated E1 mice produced a more marked increase in the cortical QUIN levels than that injected with L-Trp alone. These results suggest that the brain QUIN contents of E1 mice are dependent not only on the activity of QUIN-synthesizing enzyme but also on the rate of flux of its substrate, L-Trp or its metabolite(s), in the brain.

Aging↗