Search PubMed⌕ Search

Biomedical subjects

H Inagaki

Publications and source records attributed to H Inagaki.

At least 91 records · Page 5Linked to original sources
PubMed1998

An active Ac-like transposable element in teleost fish.

The i4/i4 genotype of the medakafish, Oryzias latipes, exhibits a quasi albino phenotype due to insertion of a novel transposable element, Tol2, into the tyrosinase gene. Tol2 is 4681 bp in length, has short inverted terminal repeats, and contains four ORFs with the potential to encode a transposase protein. Excision activity of the element has been detected by PCR analysis. Genomic Southern of the Tol2 element revealed that about 20 copies are present in the diploid genome. Dot-matrix comparisons of amino acid sequences of ORFs show relatively high similarity with transposases from Ac of maize, hobo of Drosophila, and Tam3 of snapdragon, which are all active transposable elements. Tol2 is thus concluded to be an active Ac-like transposable element probably encoding a transposase protein. It should therefore find application as a unique material for establishing a gene tagging system in fish.

Journal Article↗
PubMed1997

Method for the analysis of the methylphosphonic acid metabolites of sarin and its ethanol-substituted analogue in urine as applied to the victims of the Tokyo sarin disaster.

An analysis method for the methylphosphonic acid metabolites of sarin in urine using trimethylsilyl derivatization and flame photometric detection is described in this report. Authentic reference standards of isopropyl methylphosphonic acid (IMPA) and ethyl methylphosphonic acid (EMPA) as well as methylphosphonic acid were employed to estimate the concentration in human urine. A sample pretreatment procedure was developed for urine using a column of cation-loaded ion-exchange resins (Ag+ -, Ba2+ - or H+ -Dowex) and adjusting the pH of the eluate from the column to 3.75-3.85 improved recovery of the target compounds. The eluate was evaporated to dryness under vacuum prior to trimethylsilylation, to remove water and any hydroxy- or amino-carrying volatile substances. The sarin metabolites, because of their low volatility, were concentrated and could be derivatized for analysis. The use of synthesized authentic sarin and ethylsarin metabolites, i.e., IMPA and EMPA, made it possible to establish the necessary sample pretreatment procedures for derivatization and gas chromatography-flame photometric detection (GC-FPD) analysis. The detection limits were 0.025 ppm both for EMPA and [MPA, and 0.625 microM for MPA, respectively. This method can be useful for estimating the exposure level to sarin by assaying the metabolites in urine and it is applicable to a large numbers of samples.

Chemical Warfare Agents↗
PubMed1997

Lack of association between variants of the angiotensinogen gene and the risk of coronary artery disease in middle-aged Japanese men.

The renin-angiotensin system is important in cardiovascular remodeling. Although a variant of the angiotensinogen gene is associated with an increased generation of angiotensinogen, it is unclear how this genetic variant might influence the activity of the renin-angiotensin system and thereby contribute to the predisposition to coronary artery disease (CAD). The relation between genetic polymorphisms in the gene-encoding angiotensinogen and the risk of CAD in middle-aged Japanese men was investigated. Two polymorphisms in exon 2 of the angiotensinogen gene, M235T and T174M, were analyzed in 327 patients with CAD and 352 matched control subjects. The genotype distribution of both polymorphisms did not differ between patients with CAD and control subjects. No combination of genotypes of the two polymorphisms was associated with CAD. Results indicate that the M235T and T174M variants of the angiotensinogen gene are not associated with CAD in Japanese men.

Angiotensinogen↗
PubMed1997

Lack of association of angiotensin converting enzyme gene polymorphism or serum enzyme activity with coronary artery disease in Japanese subjects.

The association of an insertion/deletion (I/D) polymorphism in the angiotensin converting enzyme (ACE) gene or the serum activity of ACE with coronary artery disease (CAD) was investigated in Japanese men and women. The ACE genotype of 947 CAD subjects who underwent coronary angiography and of 893 control subjects was determined by polymerase chain reaction analysis. No association of the DD genotype or the D allele with CAD was observed in men or women. In a low risk group (defined by a body mass index below the median value and the absence of a history of hypertension, diabetes mellitus, and hypercholesterolemia), there was also no association between the ACE gene polymorphism and CAD. No significant difference in serum ACE activity was detected between CAD subjects and controls of all genotypes or of the same genotype, whereas a significant association was apparent between serum ACE activity and ACE genotype for both CAD subjects and controls among both men and women. These results indicate that the ACE I/D polymorphism and genotype associated variation in serum ACE activity are not risk factors for CAD in Japanese men or women.

Adult↗
PubMed1997

Structure-activity relationships of cyclic enediynes related to dynemicin A. I. Synthesis and antitumor activity of 9-acetoxy enediynes equipped with aryl carbamate moieties.

A series of the 9-acetoxy enediyne compounds, 6a-k which were simplified from natural dynemicin A, and designed to be equipped with various aryl carbamate moieties, was synthesized and evaluated for DNA-cleaving ability, in vitro cytotoxicity, and in vivo antitumor activity. As a result of this study of the structure-activity relationships (SAR) with regard to the Rt substituent, both compounds 6a and 6f with the phenyl carbamate and 4-chlorophenyl carbamate moiety, respectively, were found to exhibit significant activity (T/C > 200%) against murine P388 leukemia in mice, in spite of having IC50 values in the micromolar range. In particular, compound 6f showed the most potent activity with a maximum T/C of 256% at a daily dosage of 4.0 mg/kg for four days. Furthermore, both compounds 6a and 6f were effective against Meth A sarcoma in mice and inhibited 71 and 77% of the tumor growth at 2.0 and 3.0 mg/kg dosages, respectively. In contrast to 6f, compound 6i possessing the 2-nitrophenyl carbamate moiety showed only a slight in vivo activity, while it had about one order of magnitude higher in vitro cytotoxicity than 6f. For the stereochemistry-activity relationships at the C9 position, the (9R*)-isomers of 6c, 6g, and 6j were found to show higher in vitro and in vivo potencies than the corresponding (9S*)-isomers.

Animals↗
PubMed1997

Structure-activity relationships of cyclic enediynes related to dynemicin A. II. Synthesis and antitumor activity of 9- and 12-substituted enediynes equipped with aryl carbamate moieties.

Novel enediyne compounds 4-8, simple analogues of dynemicin A (1) equipped with the phenyl or 4-chlorophenyl carbamate moiety, were synthesized and evaluated for DNA-cleaving ability, in vitro cytotoxicity, and in vivo antitumor activity. As a result of the SAR study, it was revealed that the size and character of the substituents (R1 and R2) at the C9 position critically influenced both the stability and antitumor activity of the enediyne compounds. We found that the 9-deoxy compound 6a, a stable and less bulky enediyne having a hydrogen as the R1 and R2 substituents, showed a significant in vivo activity with a T/C of 215% at a daily dosage of 2.0 mg/kg for 4 days. The incorporation of an oxygen-containing functional group as the R3 substituent on a benzene ring resulted in considerable abolishing of both the in vitro and in vivo potencies. In a series of 9-acyloxy compounds, incorporation of the basic aromatic moiety such as 8e was effective for the in vitro activity, but it was ineffective for the in vivo activity. Furthermore, for the stereochemistry-activity relationships at the C9 position, the (9R*)-isomers of 8c, 8e, and 8f were found to show higher both in vitro and in vivo than the corresponding (9S*)-isomers. For the mechanistic studies, compound 6a underwent Bergman cycloaromatization via a diradical pathway under acidic conditions, whereas it scarcely showed DNA-cleaving activity due to the chemical stability of the aryl carbamate moiety under neutral conditions.

Animals↗
PubMed1997

Synthesis and antitumor activity of water-soluble enediyne compounds related to dynemicin A.

The enediyne compounds 9-14, simple dynemicin A (1) analogues equipped with aryl carbamate moieties with various aliphatic amino or hydroxy groups at the C9 position, were synthesized and evaluated for DNA-cleaving ability, in vitro cytotoxicity, and in vivo antitumor activity. We found that the water-soluble compounds, in which the tert-amines such as the 2-(dimethylamino)ethyl (10b, 14b), 2-(pyrrolidino)ethyl (10c), or 1-azabicyclo[3.3.0]oct-5-ylmethyl (10d, 12d, 14d) group were attached, showed not only the enhanced in vivo antitumor activity but also the decreased toxicity compared to the corresponding 9-acetoxy enediyne compounds 6-8. In particular, compound 10c showed the most enhanced in vivo antitumor activity (T/C = 222% at a daily dose of 1.25 mg/kg for 4 days) at about half of the dose of 6. These results suggest that both the enhanced antitumor activity and the reduced toxicity might be due to the improved bioavailability or disposition of compounds 6-8 by their water-solubilization.

Animals↗
PubMed1997

Interferons suppress mitochondrial gene transcription by depleting mitochondrial transcription factor A (mtTFA).

Mitochondrial gene transcription activity in organello was suppressed after culturing HeLa cells with 1000 U/ml of interferon-alpha (IFN-alpha) or IFN-gamma for 18 h. The suppression was associated with reduced levels of mitochondrial gene transcripts. Northern blot analysis of HeLa cell RNA showed marked reduction of the mRNA encoding for mitochondrial transcription factor A (mtTFA). Immunoblotting with antiserum directed against recombinant mtTFA showed a reduced level of the protein as well. Consistently, gel-retardation assay of mitochondrial extract showed reduced level of functional mtTFA, which is known to bind to both heavy and light strand promoters of bidirectionally transcribed mitochondrial DNA. We suggest that depletion of mtTFA is a pathway through which IFNs depress the mitochondrial respiration.

DNA-Binding Proteins↗
PubMed1997

Neutrophil behavior in pancreas and liver and the role of nitric oxide in rat acute pancreatitis.

The behavior of neutrophils in a rat acute pancreatitis model was observed in the pancreas and liver using fluorescence microscopy with an image analyzing system after labeling with a specific fluorescent reagent. Nonviable cells of both organs were also labeled and quantified. The role of nitric oxide in neutrophil accumulation and organ damage was estimated by administering a relatively selective inhibitor of constitutive nitric oxide synthase, N-nitro-L-arginine (L-NNA). The animal model of acute pancreatitis was induced by cerulein injection (80 mg/kg). Two groups were created, one given and the other not given L-NNA (2.5 mg/kg) prior to the induction of pancreatitis. The number of accumulated neutrophils in the pancreas and liver increased in a time-dependent manner. There was a close relation between the distribution of the neutrophils and inviable acinar cells or hepatocytes. When pretreated with L-NNA, the numbers of accumulated neutrophils and nonviable cells increased significantly in the pancreas. In the liver, a more pronounced accumulation of neutrophils was observed after treatment with L-NNA. Although hepatocyte injury was mild despite the neutrophil accumulation in the control, such injury was marked in the group treated with L-NNA. This suggests that neutrophils serve an important role in exacerbating acute pancreatitis and that nitric oxide provides a defense mechanism against neutrophil accumulation in pancreas and liver.

Acute Disease↗
PubMed1997

Synthesis and biological evaluation of novel cyclic enediyne compounds related to dynemicin A as antitumor agents.

Novel cyclic enediyne compounds, which are simple functional analogs of dynemicin A (1) having the bicyclo-[7.3.1]tridec-4-ene-2,6-diyne system, were synthesized and evaluated for the DNA-cleaving ability, in vitro cytotoxicity and in vivo antitumor activity. All of the sulfones 19-24, which were equipped with a 2-(arylsulfonyl)-ethoxycarbonyl group or the 2-(methylsulfonyl)ethoxycarbonyl group as a triggering device, showed both potent DNA-cleaving activity and cytotoxicity against various tumor cell lines. However, these compounds were entirely inactive or only slightly active against murine P388 leukemia in mice. On the other hand, the enediyne 2a having a phenyl carbamate moiety as a stable N-protecting group showed effective antitumor activity both in vitro and in vivo. In particular, it exhibited significant antitumor activity against Lewis lung carcinoma in mice. These results show that the character of the carbamate moiety of the cyclic enediynes strikingly affects their biological activities, that is, the sulfonylethyl carbamate moiety is an effective triggering device for both DNA-cleaving activity and cytotoxicity, and the phenyl carbamate moiety is significant for antitumor activity in vivo. As part of a mechanistic study, the reactivities of 2a and 21 were examined under a weakly basic condition (pH 9.3); both compounds failed to give the Bergman cycloaromatization product.

Alkynes↗
PubMed1997

Long-term culture of primary rat hepatocytes on heparin- or lambda carrageenan-containing collagen gels.

The interactions of glycosaminoglycans (GAGs) with collagen are thought to be important in cell adhesion and cell differentiation. To investigate whether the interactions of GAG or sulfated polysaccharide with collagen can maintain the functions of cultured primary rat hepatocytes, GAG- or sulfated polysaccharide-containing collagen gels were reconstituted in vitro and used for culture of hepatocytes. Among the GAGs and sulfated polysaccharides examined, heparin- and lambda carrageenan-containing collagen gels were found to be able to stimulate and sustain albumin synthesis, while the other GAG- or sulfated polysaccharide-containing collagen gels had almost no effect on maintenance of albumin synthesis. In the cultures using collagen gels that contained 400 micrograms/ml heparin or 100 micrograms/ml lambda carrageenan, albumin synthesis by rat hepatocytes was prolonged to about 4 and 5 weeks, respectively, but albumin synthesis was kept up for only one week in the cultures using conventional collagen gels. These results suggest that the interactions of heparin or lambda carrageenan might be of importance for long-term maintenance of hepatocyte functions.

Albumins↗
PubMed1997

Apoptosis, bcl-2 protein, and Fas antigen in thymic epithelial tumors.

We examined 35 thymic epithelial tumors by immunohistochemical techniques to investigate the extent of apoptosis detected by in situ end-labeling of fragmented DNA and the expression of bcl-2 and Fas. There were 22 thymomas (4 medullary, 9 mixed, 3 predominantly cortical, 6 cortical), 3 well-differentiated thymic carcinomas (WDTCs), and 10 high-grade thymic carcinomas (HGTCs), according to the Müller-Hermelink histologic classification. The HGTC showed the highest apoptotic index of the tumor cells. The apoptotic indices of the thymomas and WDTCs were significantly lower than those of the HGTCs (P < .001), but there was no significant difference among each type of thymoma and the WDTCs. bcl-2 protein was expressed in the tumor cells of the medullary-type thymomas and of the HGTCs but not in the other types of thymoma or WDTC. Fas antigen was negative in the HGTCs but was predominantly expressed in thymomas and WDTCs with advanced clinical stages (P < .05). From the immunostaining pattern, WDTC might be more likely to belong to the class of thymoma than to the category of HGTC.

Adult↗
PubMed1997

[Research on development of hybrid artificial liver utilizing reconstituted extracellular matrix].

We investigated culture of adult rat hepatocytes on reconstituted extracellular matrix, glycosaminoglycans containing collagen gel and on a porous gelatin sponge (gelform) for the maintenance of the liver-specific functions of hepatocytes. In the culture on reconstituted extracellular matrix, only heparin containing collagen cultures could significantly sustain albumin synthesis for over 3 weeks. On the other hand, in the culture of rat hepatocytes on a porous gelatin sponge, the morphology of the hepatocytes immobilized on the support was close to that in vivo and the secretion of albumin and bile acids was stable for over 12 days. These results demonstrate that culture on two- and three-dimensional culture with reconstituted extracellular matrix is one of the promising hepatocytes cultures for development of artificial liver.

Albumins↗
PubMed1996

Interleukin (IL)-1 beta, IL-2, IL-4, IL-6 and transforming growth factor-alpha levels are elevated in ventricular cerebrospinal fluid in juvenile parkinsonism and Parkinson's disease.

Interleukin (IL)-1 beta , IL-2, IL-4, IL-6, epidermal growth factor (EGF), and transforming growth factor (TGF)-alpha were measured for the first time in ventricular cerebrospinal fluid (VCSF) from control non-parkinsonian patients, patients with juvenile parkinsonism (JP) and patients with Parkinson's disease (PD) by highly sensitive sandwich enzyme immunoassays. All cytokines were detectable in VCSF from control and parkinsonian patients, and the concentrations were much higher than those in lumbar CFS. The concentrations of IL-1 beta, IL-2, IL-4 and TGF-alpha in VCSF were higher in JP than those in controls (P < 0.05). In contrast, the concentrations of IL-2 and IL-6 in VCSF from patients with PD were higher than those from control patients (P < 0.05). These results agree with our previous reports, in which the cytokine levels were elevated in the striatal dopaminergic region of the brain from patients with PD. Since VCSF is produced in the ventricles, the alteration of cytokines in VCSF may reflect the changes of cytokines in the brain. Because cytokines play an important role as mitogens and neurotrophic factors in the brain, the increases in cytokines as a compensatory response may occur in the brain of patients of JP or PD during the progress of neurodegeneration. Increase in cytokines may contribute not only as a compensatory response but as a primary initiating trigger for the neurodegeneration.

Adolescent↗
PubMed1996

A large DNA-binding nuclear protein with RNA recognition motif and serine/arginine-rich domain.

cDNA species encoding a large DNA-binding protein (NP220) of 1978 amino acids was isolated from human cDNA libraries. Human NP220 binds to double-stranded DNA fragments by recognizing clusters of cytidines. Immunofluorescent microscopy with antiserum directed against NP220 revealed a punctate or "speckled" pattern and coiled body-like structures in the nucleoplasm of various human cell lines. These structures diffused in the cytoplasm during mitosis. Western blot analysis showed that NP220 is enriched in the lithium 3,5-diiodosalicylate-insoluble fraction of nuclei. The domain essential for DNA binding is localized in C-terminal half of NP220. Human NP220 shares three types of domains (MH1, MH2, and MH3) with the acidic nuclear protein, matrin 3 (Belgrader, P., Dey, R., and Berezney, R. (1991) J. Biol. Chem. 266, 9893-9899). MH1 is a 48-amino acid sequence near the N terminus of both human NP220 and rat matrin 3. MH2 is a 75-amino acid sequence homologous to the RNA recognition motifs of heterogeneous nuclear RNP I and L. It is repeated three times in NP220 and twice in matrin 3. MH3 is a 60-amino acid sequence at the C terminus of both NP220 and matrin 3. NP220 has an arginine/serine-rich domain commonly found in pre-mRNA splicing factors. Close to the domain essential for DNA binding, there are nine repeats of the sequence LVTVDEVIEEEDL. Thus, NP220 is a novel type of nucleoplasmic protein with multiple domains.

Amino Acid Sequence↗
PubMed1996

Evaluation of cross-sensitization among dye-intermediate agents using a modified lymphocyte transformation test.

We evaluated cross-sensitization between p-phenylenediamine (pPDA) and p-aminophenol (pAP) or m-phenylenediamine (mPDA) by a modified lymphocyte transformation test. Guinea pigs were sensitized with pPDA using the maximization test procedure. Lymph node cells from the animals were then cultured with pPDA, pAP or mPDA in the presence or absence of epidermal cells (EC). Transformed lymphocyte counts were evaluated by means of 3H-thymidine uptake. Non-sensitized guinea pigs were used as controls. Blastogenesis in lymphocytes from sensitized guinea pigs was enhanced when cultured with pPDA, pAP or mPDA in the absence or presence of EC than without the sensitizers, and the extent of response depended on the concentration of pPDA, pAP or mPDA added to the cultures. Blastogenesis in lymphocytes from control animals was not significantly enhanced in response to pPDA, pAP or mPDA in the presence or absence of EC. The extent of the response to pPDA was greater than that to pAP, which in turn was greater than that to mPDA. In contrast, because pPDA, pAP and mPDA are color developing agents, cross-sensitization between pPDA and pAP or mPDA could not be evaluated by the results of an in vivo challenge due to pigmentation in the patch application sites. The results suggested that there is cross-sensitization between pPDA and pAP or mPDA, and that the modified lymphocyte transformation test is a useful predictive means of detecting cross-sensitization among chemicals, especially for color developing agents.

Aminophenols↗