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Biomedical subjects

H Kaji

Publications and source records attributed to H Kaji.

At least 73 records · Page 4Linked to original sources
PubMed1998

Comparison of the qualified occupational physician systems in the United Kingdom and Japan.

The British educational system of occupational medicine was compared to the Japanese system. Furthermore, a comparison was carried out between the certified occupational physician (COP) recognized by the Japan Society for Occupational Health in Japan and the Associateship of the Faculty of Occupational Medicine (AFOM) by the Faculty of Occupational Medicine (FOM) which is a part of the Royal College of Physicians in the United Kingdom. Judging from the comparison of the minimum total training period, the clinical training period, the occupational health training period, the method of examination and the success rate between COP and AFOM, it is suggested that the British system of occupational physicians may be better as a training system for occupational medicine and may regard occupational clinical training as more important than the Japanese system does. A comparison of a Diploma in Occupational Medicine (Dip Occ Med) approved by the FOM and the certification of occupational physicians by the Japan Medical Association has shown that the former has an examination but there is no test system in the latter. It should be discussed whether an examination system for the certification of occupational physicians should be introduced into the Japanese system in the near future.

Education, Medical, Continuing↗
PubMed1998

Molecular cloning, expression, and site-directed mutagenesis of inorganic pyrophosphatase from Thermus thermophilus HB8.

The genomic DNA encoding the inorganic pyrophosphatase from an extremely thermophilic bacterium, Thermus thermophilus HB8 (ATCC27634), was isolated by colony hybridization with a probe designed as a part of gene amplified by the PCR method, which was derived from the partial amino acid sequence of the enzyme. The DNA was cloned into a plasmid vector, pUC118, after digestion with BamHI. The inserted nucleotide fragment was about 1.8 kbp in length and the nucleotide sequence included a 525 bp open reading frame. The deduced amino acid sequence was completely identical with that of the enzyme determined by automated Edman analysis of peptide fragments isolated from digests obtained with Staphylococcus aureus V8 protease and Achromobacter protease I, and also from products obtained on chemical cleavage with cyanogen bromide and 70% formic acid. The subunit of this enzyme is composed of 174 amino acid residues with a calculated molecular weight of 19,084. Then, the gene was overexpressed in Escherichia coli BL21 (DE3) using a plasmid vector, pET15b, system. The recombinant enzyme was fully active, and exhibited higher thermostability than the E. coli enzyme. Amino acid residues located on the surface of the recombinant enzyme were determined by means of limited proteolysis, and the results revealed that the environment of Lys residues is almost the same as the crystal structure reported previously [Teplyakov, A. et al. (1994) Protein Sci. 3, 1098-1107]. Furthermore, the roles of two tryptophan residues were investigated by site-directed mutagenesis, which indicated that they may be responsible for the structural integrity and thermostability.

Amino Acid Sequence↗
PubMed1998

Pharmacologic profile of TA-606, a novel angiotensin II-receptor antagonist in the rat.

This study was carried out to characterize a novel angiotensin II-receptor antagonist, TA-606, (3-pentyloxy) carbonyloxymethyl-5-acetyl-2-n-propyl-3-[2'(1H-tetrazole-5-yl) biphenyl-4-yl]methyl-4,5,6,7-tetrahydro imidazo [4,5-c] pyridine-4-carboxylate hydrochloride, which is a newly synthesized prodrug of 606A. In anesthetized rats, 606A inhibited angiotensin II-induced pressor response with a median inhibitory concentration (IC50) of 6 microg/kg, i.v., and was 8 times more potent than EXP3174, an active metabolite of losartan. Bioavailability of TA-606 was 11 times higher than that of 606A in Sprague-Dawley rats, with consistent hypotensive potencies in spontaneously hypertensive rats (SHRs). In conscious renal hypertensive rats (RHRs) and conscious SHRs, TA-606 lowered the blood pressure without any effects on the heart rate, and its effective dose for 30 mm Hg (ED30) values were 0.14 and 0.21 mg/kg, p.o., respectively. The effect of TA-606 lasted > 10 h in both models. Moreover, the effect of TA-606 was approximately 30 and 10 times more potent than those of losartan in RHRs and SHRs, respectively. TA-606 did not affect the blood pressure in deoxycorticosterone acetate (DOCA)-salt hypertensive rats. TA-606 given for 12 weeks attenuated the development of hypertension in stroke-prone SHRs. These results indicate that TA-606 is a potent angiotensin II-receptor antagonist with antihypertensive efficacy. Thus TA-606 is suggested to be a possible useful agent in the treatment of hypertension.

Angiotensin Receptor Antagonists↗
PubMed1998

Short stature caused by a natural growth hormone antagonist.

Severe short stature in a male child due to a single mutation in the GH-1 gene was first reported in 1996 by Takahashi et al. [N Engl J Med 1996;334:432-436]. This missense mutation was predicted to convert codon 77 from arginine (R) to cysteine (C). The child's chronological age was 4 years and 11 months, and his bone age 2 years and 6 months, i.e., equal to only 51% of his chronological age. Body proportions were normal except for the prominent forehead and saddle nose. Pituitary size was normal on magnetic resonance imaging examinations. Serum IGF-1, IGFBP-3 and GHBP were all decreased or at the lower limit of the normal range. Nocturnal urinary growth hormone (GH) excretion was high. Isoelectric focusing analysis revealed the presence of an abnormal GH peak in addition to the normal one. The R77C mutant GH possessed a 6 times greater affinity to GHBP than the wild-type GH, and inhibited tyrosine phosphorylation in IM-9 cells 10 times more potently than the wild-type GH, showing an antagonistic or a dominant negative action. In agreement with the antagonistic property of the mutant GH exhibited, the child did not show any increase in serum IGF-1 levels after exogenous hGH administration. It should be noted that the child in this study is not a typical case of Kowarski syndrome in which endogenous GH is found to be simply bioinactive, as in the patient we recently described elsewhere. Therefore, this patient's condition should be categorized as a new syndrome of short stature caused by a natural GH antagonist.

Age Determination by Skeleton↗
PubMed1998

Growth hormone (GH) insensitivity syndrome with high serum GH-binding protein levels caused by a heterozygous splice site mutation of the GH receptor gene producing a lack of intracellular domain.

Most of the GH receptor (GHR) gene abnormalities causing GH insensitivity syndrome (GHIS) are located in the region coding the extracellular domain, and serum GH-binding protein (GHBP) levels, determined by ligand-mediated immunofunctional assay, are low in most of the patients with GHIS. We present here a heterozygous point mutation of the donor splice site in intron 9 of the GHR gene in two Japanese siblings with GHIS, whose serum GHBP levels were high. The same mutation was found in their mother as well. The analysis of ribonucleic acid from the peripheral leukocytes revealed complete skipping of exon 9 from one allele, but not the other, in the GHR complementary DNA and appearance of a premature stop codon in exon 10. The translated protein was truncated with deletion of 98% of the intracellular domain of the GHR, including boxes 1 and 2, which are critical for GH signal transduction and GHR internalization, respectively. Recently, it was shown that the truncated GHR lacking the intracellular domain was physiologically present in a minute amount, served as a negative regulator for GH signaling, and possessed increased capacity to generate GHBP. Therefore, the mutation found in our patients caused the pathogenetic production of the truncated GHR with a dominant negative effect on GH signaling, which is probably responsible for their short stature and high serum GHBP levels.

Adolescent↗
PubMed1998

Estrogen via the estrogen receptor blocks cAMP-mediated parathyroid hormone (PTH)-stimulated osteoclast formation.

Several lines of evidence indicate that estrogen inhibits parathyroid hormone (PTH)-induced bone resorption in vivo and in vitro. However, its precise mechanism remains unknown. The present study was performed to investigate whether osteoclast precursor cells possess the receptors for PTH/PTH-related protein (PTHrP) and/or estrogen and to clarify the mechanism by which estrogen affects PTH-induced osteoclast-like cell (Ocl) formation. The polymerase chain reaction (PCR) product corresponding in size to the mouse PTH/PTHrP receptor cDNA was detected in mouse hemopoietic blast cells supported by granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as in osteoblastic MC3T3-E1 cells. The nucleotide sequence of the PTH/PTHrP receptor PCR product of hemopoietic blast cells was found to be 95.4% identical to that of PTH/PTHrP receptor cDNA of rat osteoblastic ROS cells. The PCR product corresponding in size to the mouse estrogen receptor cDNA was detected in mouse hemopoietic blast cells supported by GM-CSF as well as in MC3T3-E1 cells. The nucleotide sequence of the estrogen receptor PCR product of hemopoietic blast cells was completely identical to that of mouse estrogen receptor cDNA. 17Beta-estradiol (17beta-E2) but not 17alpha-E2 dose dependently antagonized Ocl formation stimulated by human (h) PTH(1-34) at a minimal effective concentration of 10(-10) M in the hemopoietic blast cell culture. 17Beta-E2 also significantly inhibited Ocl formation stimulated by 10(-8) M hPTHrP(1-34), while it did not affect 1,25-dihydroxyvitamin D3-induced Ocl formation. However, 10(-8) M 17beta-E2 significantly inhibited Ocl formation stimulated by dibutyryladenosine cAMP (10(-4) M) and Sp-cAMPS (10(-4) M), an activator of cAMP-dependent protein kinase (PKA) as well as forskolin (10(-5) M). In contrast, 17beta-E2 did not affect Ocl formation by either phorbol myristate acetate (10(-7) M), an activator of protein kinase C (PKC), or A23187 (10(-7) M), a calcium ionophore. The pretreatment with 17beta-E2 significantly inhibited Ocl formation induced by the combined treatment with PTH and PKC inhibitors (H7 or staurosporine), while it did not affect Ocl formation stimulated by the combined treatment with PTH and Rp-cAMPS, a PKA inhibitor. The present data indicate that estrogen inhibits PTH-stimulated Ocl formation by directly acting on hemopoietic blast cells, possibly through blocking a PKA pathway but not a calcium/PKC pathway.

Animals↗
PubMed1998

Carboxyl-terminal parathyroid hormone fragments stimulate type-1 procollagen and insulin-like growth factor-binding protein-5 mRNA expression in osteoblastic UMR-106 cells.

The effects of carboxyl-terminal (C-) PTH fragments, (35-84), (53-84) and (69-84), on the proliferation and function of osteoblastic UMR-106 cells were compared with those of amino-terminal (N-) (1-34) and intact (I-) (1-84) PTH. I-PTH as well as N-PTH at 10(-8)M significantly inhibited [3H] thymidine incorporation and stimulated alkaline phosphatase activity in UMR-106 cells. No C-PTH fragments affected them. In contrast, the expression of type-1 procollagen mRNA in these cells was stimulated by all C-PTH fragments, inhibited by N-PTH and not affected by I-PTH. All C-PTH fragments except (69-84) as well as N-PTH and I-PTH stimulated IGFBP-5 mRNA expression. The present study suggests that the C-portion of the PTH molecule exercises biological activities in mRNA expression of type-1 procollagen as well as IGFBP-5 in osteoblasts, and that it might be involved in the anabolic action of PTH on bone in vivo.

Cell Line↗
PubMed1998

The endogenous feeding suppressant, 2-buten-4-olide, impairs the pulsatile secretion of luteinizing hormone through endogenous opioid peptides.

OBJECTIVE: To clarify the mechanism of the suppressive effect of 2-buten-4-olide (2-B4O), an endogenous feeding suppressant, on the pulsatile secretion of luteinizing hormone (LH), by studying whether endogenous opioid peptides are involved in this suppressive effect. METHODS: Using ovariectomized (ovx) rats, blood samples were taken every 6 min for 2 h after administration of 2-B4O or saline into the third cerebroventricle (3V) and sequential i.v. injection of naloxone (0. 5 mg/kg per h) or saline. Rats were divided into three experimental groups: group 1: 3V saline + i.v. saline (control); group 2: 3V 2-B4O + i.v. saline; group 3: 3V 2-B4O + i.v. naloxone. Serum LH concentrations were determined by double-antibody RIA. To determine whether 2-B4O affected the biosynthetic activity of the opioidergic neurons within the ovx rat arcuate nucleus, we measured the concentrations of pro-opiomelanocortin (POMC) mRNA, a precursor of beta-endorphin, in the rostral arcuate nucleus using non-radioactive in situ hybridization and a computerized image-analysis system. RESULTS: 2-B4O significantly suppressed the pulse frequency of LH (group 2: 1.5+/-0.33 pulses/2 h, group 1: 2.43+/-0.2 pulses/2 h; P < 0.05), but naloxone blocked its suppressive effect and restored the pulse frequency (group 3: 3.29+/-0.36 pulses/2 h, group 2: 1.5+/-0.33 pulses/2 h: P < 0.01). There were no significant changes in the mean LH concentrations and amplitude. Furthermore, 2-B4O significantly stimulated the expression of POMC mRNA in the rostral arcuate nucleus. CONCLUSION: These results suggest that 2-B4O may impair the pulsatile secretion of LH by activating the opioid pathway within the hypothalamus.

4-Butyrolactone↗
PubMed1998

[GH receptor gene mutations and growth failure].

Abnormality of growth hormone (GH) receptor (GHR) and signal transduction system is a cause of primary GH insensitivity. Analysis of GHR gene have shown so far about 30 different kinds of mainly homozygous mutations in patients with Laron syndrome and heterozygous mutations in some patients with idiopathic short stature. In this paper, we review these diverse clinical and molecular characteristics of abnormal GHR gene in patients with GH insensitivity including our patient of Laron syndrome caused by novel compound heterozygous mutations and of familial short stature with high serum GHBP levels caused by a heterozygous splice site mutation of the GHR gene producing a lack of cytoplasmic domain.

Genotype↗
PubMed1997

Biologically inactive growth hormone caused by an amino acid substitution.

Short stature caused by biologically inactive growth hormone (GH) is characterized by lack of GH action despite high immunoassayable GH levels in serum and marked catch-up growth to exogenous GH administration. We found a heterozygous single-base substitution (A-->G) in exon 4 of the GH-1 gene of a girl with short stature, clinically suspected to indicate the presence of bioinactive GH and resulting in the substitution of glycine for aspartic acid at codon 112. We confirmed the presence of mutant GH in the serum using isoelectric focusing analysis. The locus of mutation D112G was found within site 2 of the GH molecule in binding with GH receptor (GHR)/GH binding protein (GHBP). The expressed recombinant mutant GH tended to form a 1:1 instead of the 1:2 GH-GHBP complex normally produced by wild-type GH. The formation of a 1:2 GH-GHBP complex is compatible with the dimerization of GHRs by GH, a crucial step in GH signal transduction. Mutant GH was less potent than wild-type GH not only in phosphorylation of tyrosine residues in GHR, janus kinase 2 (JAK2), and signal transducers and activators of transcription 5 (STAT5) in IM-9 cells, but also in metabolic responses of BaF/GM cells, a stable clone transfected with cDNA of the chimera of the extracellular domain of human GHR, the transmembrane and the cytoplasmic domain of the human thrombopoietin receptor. These results indicate that the D112G mutation in the GH-1 gene causes production of bioinactive GH, which prevents dimerization of GHR and is therefore responsible for the patient's short stature.

Child, Preschool↗
PubMed1997

Mutational specificity of glyoxal, a product of DNA oxidation, in the lacI gene of wild-type Escherichia coli W3110.

To determine the mutation spectrum of glyoxal, which is produced from DNA by oxygen free-radicals, we analyzed the chromosomal lacI gene of mutants induced by treatment of a wild-type Escherichia coli strain with glyoxal. The cell death and the mutation frequency increased according to the concentration of glyoxal added to the culture medium. The majority of the spontaneous mutations (82%) and that of the glyoxal-induced mutations (50%) were the addition or deletion of a 5'-TGGC-3' sequence at positions 623-634, which was reported to be a mutational hot spot in the lacI gene. In the glyoxal-induced mutants, however, the ratio of base-pair substitutions was increased (35%). Although all types of base-pair substitutions were detected, 78% of the base-pair substitutions occurred at G:C sites. Among them, G:C-->A:T transitions were predominant, followed by G:C-->T:A transversions. These mutations appeared to be distributed randomly within the lacI gene. These results suggest that glyoxal-induced mutations may correlate to mutations induced by oxygen free-radicals.

Bacterial Proteins↗
PubMed1997

Inhibition of HIV type 1 RNA dimerization by antisense DNA corresponding to the 17-nucleotide sequence downstream from the splice donor site of HIV type 1 RNA.

HIV-1 RNA dimerization involves at least two key regions, one located upstream from the splice donor (SD) site, and the other located downstream from the SD site. To determine the precise location and the mechanism of action of the downstream region, we constructed a model system using a synthetic HIV-1 RNA fragment (HXB2, 455-1146), which dimerized at relatively low salt concentrations (100 mM KCl, 1 mM MgCl2). We tested in this system antisense DNAs that are complementary to both the upstream and downstream regions of HIV-1 RNA for their possible inhibitory effects on dimerization. Antisense DNAs complementary to nucleotides 773-789 located downstream from the SD site effectively inhibited dimerization of HIV-1 RNA. These inhibitory antisense DNAs hybridized with the dimer form of HIV-1 RNA, and dissociated the dimer into monomers. However, antisense DNAs complementary to the region upstream from the SD site did not hybridize with the dimer, although they inhibited RNA dimerization and also dissociated the preformed dimer.

Base Sequence↗
PubMed1997

Types of mutations induced by glyoxal, a major oxidative DNA-damage product, in Salmonella typhimurium.

We have analyzed the types of mutations induced by glyoxal, a major oxidative DNA-damage product, in Salmonella typhimurium. A set of six strains, TA7001 to TA7006, was used to detect base-pair substitutions, and the TA98 strain was employed to detect frameshift mutations. Glyoxal did not induce mutations at A:T base pairs. The majority of the mutations induced by glyoxal were base-pair substitutions at G:C base pairs, and a small level of frameshift mutations was detected in the TA98 strain.

Base Composition↗
PubMed1997

Tetrahydronaphthalene lignan compounds as potent anti-HIV type 1 agents.

Anti-HIV-1 activity of tetrahydronaphthalene (THN) derivatives of lignan compounds was studied. THN derivatives prevented cell death caused by HIV-1 infection in MT-4 cells. They also inhibited giant cell formation by HIV-1 in Sup-T1 cells, and p24 production in HIV-1-infected H9 cells. The 50% effective concentration (ED50) of the most active compound, 1737 [5,6,7-trimethoxy-4-(3,4,5-trimethoxyphenyl)-1,3,3a,4,9,9a-hexahydron aphtho[2,3-c]thiophene], for inhibition of the cytopathic effects of HIV-1 infection ranged from 0.15 to 0.8 microM. The 50% cytotoxic concentration (CC50) of compound 1737 measured by the viability of MT-4 cells was 58 microM, indicating a selective index (CC50/ED50) of 70-400. Substitution of the phenyl ring with other structures markedly decreased cytotoxicity but did not affect the antiviral activity of the compounds. This resulted in compounds with a high selective index. One such compound was 1738 [7-methoxy5,6-methylenedioxy-4-(4-benzyloxy-3-methoxyphenyl)1,3,3a ,4,9,9a-hexahydronaphtho[2,3-c]thiophene], with a selective index higher than 770. The time-of-addition experiment indicated that these compounds acted at or near the reverse transcription step of the HIV-1 life cycle. THN derivatives inhibited HIV-1 reverse transcriptase (RT) in vitro at a concentration of 1 microM. Resistant viruses selected in the presence of THN derivatives showed some degree of cross-resistance to other nonnucleoside RT inhibitors, but not to the nucleoside RT inhibitor, AZT. THN derivatives failed to inhibit replication of pyridinone- and nevirapine-resistant HIV strains. However, compound 1737 inhibited replication of a TIBO-resistant strain more effectively than the wild-type HIV-1. Consistent with this result, compound 1737 also inhibited TIBO-resistant RT more effectively than the wild-type RT in vitro. These results suggested that THN derivatives interact with RT in a manner similar to but slightly different from that of other nonnucleoside HIV-1 RT inhibitors.

Anti-HIV Agents↗
PubMed1997

Leptin induces mitogen-activated protein kinase-dependent proliferation of C3H10T1/2 cells.

Leptin, secreted by adipocytes, regulates satiety and energy expenditure. Several forms of leptin receptors produced by alternative mRNA splicing are found in many tissues, including the hypothalamus, liver, lung, kidney, hematopoietic cells, and gonads, suggesting that leptin exerts effects in these tissues. In accordance with the distribution of leptin receptors, there is accumulating evidence that leptin plays various roles in reproduction, hematopoiesis, and the immune systems in addition to the regulation of food intake and energy expenditure. In the present study, we examined the in vitro effects of leptin on proliferation of a mouse embryonic cell line, C3H10T1/2, and its mechanism of action. Leptin caused a dose- and time-dependent increase in mitogen-activated protein kinase (MAPK) activity that was accompanied by an increase in C3H10T1/2 cell number. The MAPK kinase-1-specific inhibitor PD98059 completely blocked the increases in both MAPK activity and cell proliferation caused by leptin. These findings indicate that leptin stimulates the proliferation of C3H10T1/2 cells via the MAPK cascade.

Animals↗
PubMed1997

Glyoxal, a major product of DNA oxidation, induces mutations at G:C sites on a shuttle vector plasmid replicated in mammalian cells.

Glyoxal is a major product of DNA oxidation in which Fenton-type oxygen free radical-forming systems are involved. To determine the mutation spectrum of glyoxal in mammalian cells and to compare the spectrum with those observed in other experimental systems, we analyzed mutations in a bacterial suppressor tRNA gene (supF) in the shuttle vector plasmid pMY189. We treated pMY189 with glyoxal and immediately transfected it into simian COS-7 cells. The cytotoxicity and mutation frequency increased according to the dose of glyoxal. The majority of glyoxal-induced mutations (48%) were single-base substitutions. Eighty three percent of the single-base substitutions occurred at G:C base pairs. Among them, G:C-->T:A transversions were predominant, followed by G:C-->C:G transversions and G:C-->A:T transitions. A:T-->T:A transversions were also observed. Mutational hotspots within the supF gene were detected. These results suggest that glyoxal may play an important role in mutagenesis induced by oxygen free radicals.

Animals↗