Biomedical subjects

H Yoshimoto

Publications and source records attributed to H Yoshimoto.

At least 37 records · Page 2Linked to original sources

PubMed1999

Molecular mechanism of the multiple regulation of the Saccharomyces cerevisiae ATF1 gene encoding alcohol acetyltransferase.

The ATF1 gene encodes an alcohol acetyl transferase (AATase), that catalyses the synthesis of acetate esters from acetyl CoA and several kinds of alcohols. ATF1 transcription is negatively regulated by unsaturated fatty acids and oxygen. A series of analyses of the ATF1 promoter identified an 18 bp element essential for transcriptional activation. Ligation of the 18 bp element into a plasmid carrying the CYC1 promoter deleted UAS-activated transcription and conferred transcriptional repression by unsaturated fatty acids. The 18 bp element contains a binding sequence for Rap1p, which is a transcriptional repressor and activator. In vitro binding studies showed that Rap1p binds to the 18 bp element essential for transcriptional activation. The results of internal deletion studies of the promoter region suggested that there was also a region responsible for ATF1 oxygen regulation. This region contained the consensus binding sequence for the hypoxic repressor Rox1p. In vitro binding studies showed that Rox1p binds to the region responsible for oxygen regulation. To investigate the effect of the hypoxic repressor complex on transcription, ATF1 expression was measured in rox1, tup1 and ssn6 disruptant strains. It was found that rox1, tup1 and ssn6 disruption caused elevated expression of ATF1 under aerobic conditions. Thus, the activation of ATF1 transcription is dependent on Rap1p, and the Rox1p-Tup1p-Ssn6p hypoxic repressor complex is responsible for repression by oxygen. Furthermore, a study of ATF1 expression in a sch9 null mutant suggested that the Sch9p protein kinase is involved in ATF1 trancriptional activation.

Acetyltransferases↗

PubMed1999

Isolation and characterization of the ATF2 gene encoding alcohol acetyltransferase II in the bottom fermenting yeast Saccharomyces pastorianus.

The ATF2 gene encodes alcohol acetyltransferase II, which catalyses the synthesis of isoamyl acetate from acetyl coenzyme A and isoamyl alcohol. To characterize the ATF2 gene from the bottom fermenting yeast Saccharomyces pastorianus, the S. pastorianus ATF2 gene was cloned by colony hybridization using the S. cerevisiae ATF2 gene as a probe. When an atf1 null mutant strain was transformed with a multi-copy plasmid carrying the S. pastorianus ATF2 gene, the AATase activity of this strain was increased by 2.5-fold compared to the control. The S. pastorianus ATF2 gene has 99% nucleic acid homology in the coding region and 100% amino acid homology with the S. cerevisiae ATF2 gene. Southern blot analysis of chromosomes separated by pulse-field gel electrophoresis indicated that the ATF2 gene probe hybridized to chromosome VII in S. cerevisiae and to the 1100 kb chromosome in S. pastorianus. As S. pastorianus is thought to be a hybrid of S. cerevisiae and S. bayanus, the S. bayanus-type gene, which has a relatively low level of homology with the S. cerevisiae-type gene, is also usually detected. Interestingly, an S. bayanus-type ATF2 gene could not be detected. These results suggested that the cloned ATF2 gene was derived from S. cerevisiae. Analysis using an ATF2-lacZ fusion gene in S. pastorianus showed that expression of the ATF2 gene was relatively lower than that of the ATF1 gene and that it is repressed by aeration but activated by the addition of unsaturated fatty acids. The S. pastorianus ATF1, Lg-ATF1 and ATF2 Accession Numbers in the DDBJ Nucleotide Sequence Database are D63449, D63450 and D86480, respectively.

Acetyltransferases↗

PubMed1999

Serum macrophage colony-stimulating factor (M-CSF) level is elevated in patients with old cerebral infarction related to vascular damage.

We measured the serum levels of macrophage colony-stimulating factor (M-CSF) in 37 patients with an old cerebral infarction who had been surmised to have a damaged vessel wall and who had been in a stable condition for over three months after stroke onset, and those of 41 healthy control subjects. The M-CSF levels in the patients were significantly higher (P < 0.01) than those of the controls at 1320.4 +/- 410.6 unit/ml and 853.9 +/- 180.3 unit/ml, respectively. The plasma levels of von Willebrand factor (vWF) antigen (P < 0.01) and thrombomodulin (TM) (P < 0.05), as well as those of thrombin-antithrombin III (TAT) complex (P < 0.05), prothrombin fragment 1+2 (F1+2) (P < 0.02), D-dimer products of crosslinked fibrin degradation products (D-dimer) (P < 0.01), and plasmin-antiplasmin (PAP) complex (P < 0.05) in the patients were also significantly higher than those in the controls. Significant positive correlations (P < 0.01) were found between these parameters and the M-CSF level, but there was no significant correlation between the M-CSF level and the white blood cell count, serum lipids, or blood pressure. Based on these results, we suggest that an increased M-CSF level indicates vascular damage or a thrombotic state in patients with an old cerebral infarction.

Aged↗

PubMed1999

Analysis of the genes activated by the FLO8 gene in Saccharomyces cerevisiae.

It is thought that the FLO8 gene encodes a transcriptional activator of the dominant flocculation gene FLO1 in Saccharomyces cerevisiae. To determine other genes which are regulated by FLO8, a detailed comparison of the transcripts from the FLO8 and Deltaflo8 strains was carried out. In addition to the FLO1 gene, it was found that transcription of the FLO11 and STA1 genes is positively regulated by FLO8. In flo8 strains, not only transcripts of the FLO11, STA1, and FLO1 genes but also invasive growth, extracellular glucoamylase production, and flocculation were undetected. From these results, it is suggested that FLO8 regulates these characteristics via the transcriptional regulation of the FLO11, STA1, and FLO1 genes.

Base Sequence↗

PubMed1999

Overexpression of insulin-like growth factor-1 (IGF-I) receptor and the invasiveness of cultured keloid fibroblasts.

Keloid is a dermal fibroproliferative tissue of unknown etiology. Protein tyrosine kinases (PTKs) play an important role in the regulation of cell growth and differentiation. Activation of PTK cascades in keloid fibroblasts is thought to be closely linked to abnormal cell proliferation and migration. We determined the expression profile of PTK genes in normal skin and keloid fibroblasts using the homology cloning method with a degenerated primer. Eight PTK genes were expressed among a total of 46 receptor-type clones. The most abundant type of PTK receptors was the platelet-derived growth factor receptor in both fibroblasts. However, insulin-like growth factor-I receptor (IGF-IR) was overexpressed only in keloid-derived fibroblasts (9 of 24). Immunohistochemical analysis confirmed the high expression of IGF-IR in keloid fibroblasts, but not in normal fibroblasts. To examine the functional properties of the IGF-I/IGF-IR pathway, we investigated cell proliferation and invasion activities of both types of fibroblasts. The mitogenic effect of IGF-I on both fibroblasts was very weak compared with serum stimulation. In contrast, the invasive activity of keloid fibroblasts was markedly increased in the presence of IGF-I, and inhibited by a neutralizing antibody against IGF-IR. Our results indicate the involvement of activated IGF-I/IGF-IR in the pathogenesis of keloid by enhancing the invasive activity of fibroblasts.

Adolescent↗

PubMed1999

A novel hemiacetal dehydrogenase activity involved in ethyl acetate synthesis in Candida utilis.

Acetate ester synthesis was studied in vitro with the ethyl acetate-producing yeast Candida utilis. The level of enzyme activity observed for the NAD+-dependent hemiacetal dehydrogenase acting on hemiacetal, which was produced non-enzymatically from an alcohol and an aldehyde, was much greater than that for the other enzyme involved in ester synthesis, alcohol acetyltransferase. The level of ethyl acetate synthesis in vivo approximately paralleled the hemiacetal dehydrogenase (HADH) activity. The results suggest that the main pathway for ethyl acetate synthesis in C. utilis involves a novel hemiacetal dehydrogenase activity.

Journal Article↗

PubMed1999

Exogenous nitric oxide inhibits platelet activation in whole blood.

We examined the effect of NO on collagen-induced whole blood aggregation and platelet activation in whole blood by using impedance aggregometry and flow cytometry. For the extracellular generation of NO, we chose sodium nitroprusside dihydrate (SNP), and as intracellular generators of NO, L-arginine and isosorbide dinitrate (ISDN). The latter two significantly inhibited whole blood aggregation, whereas SNP had no such effect. The inhibitory effect of ISDN was diminished by addition of methylene blue (MB) or 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-l-oxyl 3-oxide (carboxyl-PTIO), and the inhibitory effect of L-arginine was diminished by addition of N(G)-monomethyl-L-arginine monoacetate (L-NMMA). Although the addition of ISDN increased the cyclic guanosine monophosphate (cGMP) level in whole blood and in the suspension of platelets and white blood cells (PLTs + WBCs), no increase was found in platelet-rich plasma (PRP). The P-selectin expression on the platelet surface in whole blood was reduced by ISDN and L-arginine. These findings suggest that the intracellular generation of NO inhibits whole blood aggregation, and this mechanism may play an important role in its antithrombotic effect in whole blood.

Arginine↗

PubMed1999

Coadministration of basic fibroblast growth factor and sucrose octasulfate (sucralfate) facilitates the rat dorsal flap survival and viability.

The effective use of local growth factors and cytokines may replace the lengthy staged surgical delay process. We tested the efficacy of basic fibroblast growth factor (bFGF) coadministered with sucralfate (sucrose octasulfate) on the rat dorsal flap model. A total of 76 male Wistar rats were used in this experiment. Four groups of the animals were divided. Group 1 (n = 5) was the vehicle control (saline soaked), group 2 (n = 5) was sucrose octasulfate soaked (100 microg/ml, 1 ml), group 3 (n = 5) was bFGF soaked (1 microg/ml, 1 ml), and group 4 (n = 5) was both bFGF and sucrose octasulfate soaked. All agents were soaked equally in Gelfoam. The flap survival measured by the quantitative computer-assisted morphologic analysis was significantly improved by day 5 postoperatively in the combined administration group compared with the vehicle control (81 and 53 percent, respectively; p < 0.05). In lead oxide-gelatin microangiography, there was enhanced pedicle vessel formation observed as well as the extended vessel sprouting up to very close to the distal end in combined group on day 5. The endogenous bFGF mRNA expressions shown by reverse transcriptase-polymerase chain reaction were detected in all four groups. The angiogenesis indicated by alpha-smooth muscle actin immunopositivity was significantly more enhanced in the combined group than the vehicle control (37.3 and 19.4, respectively; p < 0.01). In the combined group, there was stronger immunopositivity for bFGF in epidermis and hair follicles observed, and more notably bFGF-immunopositive dermal fibroblasts were evident. Thus, coadministration of bFGF and sucralfate markedly facilitates the rat dorsal flap survivability by enhancing the bFGF expression and angiogenesis.

Actins↗

PubMed1999

Transformation and expression of a cloned fimA gene in Porphyromonas gingivalis.

The Porphyromonas gingivalis fimbria is an important virulence factor involved in the adherence and colonization of the organism in the oral cavity. In this study, we transformed this organism with a gene, fimA381, encoding the fimbrial subunit of P. gingivalis 381 (fimbrillin) by using the host-vector system that we developed previously and examined expression of the cloned fimA381 gene. The recombinant plasmid pYHF2 was constructed by ligating a fragment containing the fimA381 gene into the plasmid vector pYH420 and transformed into the restriction-deficient P. gingivalis host YH522. pYHF2 was autonomously maintained in YH522 cells, and the fimbrillin polypeptide (recombinant fimbrillin) was fully expressed. The molecular mass of the recombinant fimbrillin was evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis as 41 kDa, which was identical to that of the native fimbrillin of strain 381. The amino acid sequences of the 20 amino-terminal residues of the recombinant fimbrillin and the native fimbrillin of the strain 381 were identical. In addition, characteristic long and thin fimbrial structures (recombinant fimbriae) that were distinguishable from the host's native fimbriae when examined by immunogold electron microscopy were observed around the cell surface of the transformants containing the fimA381 gene. These results suggested that transformation of fimA gene from a different strain of P. gingivalis followed by accumulation of the mature fimbrial subunit protein was sufficient for production of fimbrial structures that were observable by electron microscopy.

Bacterial Proteins↗

PubMed1999

Platelet aggregating activity of plasma in patients with thrombotic thrombocytopenic purpura evaluated by a particle counting method using light scattering.

The platelet aggregating activity of plasma obtained from five patients with thrombotic thrombocytopenic purpura (TTP) was evaluated by a particle counting method using light scattering. When normal platelets were suspended in the plasma obtained from TTP patients, small aggregate formation was observed after stirring at 1000 rpm without the addition of platelet aggregating agents; no aggregate was observed, however, in the plasma obtained from healthy donors. Since the inhibitory effect of the addition of normal plasma to TTP plasma on this reaction was dose-dependent and not additive, the efficacy of plasma therapy was not confirmed. Small aggregates were formed in the high molecular weight fraction (HMWF) of TTP plasma but not in the low molecular weight fraction (LMWF), suggesting that the platelet aggregating activity existed in HMWF. Among the antiplatelet agents usually used for TTP, dipyridamole was more effective for the inhibition of this reaction than aspirin. This spontaneous platelet aggregation reaction by a particle counting method using light scattering could be useful for evaluating the platelet aggregating activity in patients with TTP.

Adolescent↗

PubMed1999

[Evaluation of direct identification testing from positive blood culture bottles with MicroScan rapid identification panels].

We evaluated the direct identification method from growth-positive blood culture bottles using MicroScan Rapid ID panels (DADE BEHRING) for the purpose of rapid identification. The inoculum for Rapid ID panels were prepared using an isolation method from blood culture bottles by VACUTAINER (BD). McF 1.0 had a better result than McF 0.5 as the inoculum concentration for Rapid ID panels. Rapid ID panel identification results were effected by blood contamination for > or =0.3% of S. aureus and 0.9% of a strain of E. coli. Blood contamination from the bottle may cause an issue to the identification results. The accuracy of this direct identification testing was 72.0% (36 out of 50) for gram positives organisms and 88. 9% (80 out of 90) for gram negatives organisms. Although some strains including S. pyogenes, coagulase-negative staphylococci and non-Fermentative Gram Negative Rods had not identified correctly, this method provides a preliminary result within 3 hours and provides a fast turn around time. In conclusion, this method was considered as an effective method for routine testing.

Bacteriological Techniques↗

PubMed1999

Functional properties of nociceptive neurons in the nucleus centralis lateralis of the cat thalamus.

We studied functional properties of nociceptive neurons in the thalamic intralaminar nuclei, especially the nucleus centralis lateralis (CL) in urethane-chloralose anesthetized cats. Nociceptive neurons were found in the intralaminar nuclei, i.e. CL, nucleus centralis medialis (CeM) and nucleus parafascicularis (Pf). One third of these nociceptive neurons had their receptive fields throughout the body, including the orofacial area. Namely, they received convergent nociceptive input from the trigeminal and spinal nerve territories. These neurons with widely complex receptive fields were similar to those we found in the middle third of the caudal bulbar reticular formation (RF). Electrical stimulation of the CL antidromically activated about half of the caudal bulbar RF neurons tested. Furthermore, electrical stimulation of the cingulate gyrus antidromically activated some of CL neurons. These findings suggest that most of the CL neurons receiving nociceptive input from throughout the body receive information directly or indirectly via the caudal bulbar RF, and that certain of this information is then conveyed to the cingulate gyrus.

Afferent Pathways↗

PubMed1998

Transcriptional co-regulation of Saccharomyces cerevisiae alcohol acetyltransferase gene, ATF1 and delta-9 fatty acid desaturase gene, OLE1 by unsaturated fatty acids.

The ATF1 gene encodes an alcohol acetyl transferase which catalyzes the synthesis of acetate esters from acetyl CoA and several kinds of alcohols. ATF1 expression is repressed by unsaturated fatty acids or oxygen. Analysis using ATF1-lacZ fusion plasmid revealed that ATF1 gene expression is widely repressed by a variety of unsaturated fatty acids, and the degree of ATF1 transcriptional repression varies according to the structure of the unsaturated fatty acids. Interestingly, it was noted that the degree of ATF1 transcriptional repression was related to the melting point of unsaturated fatty acids added to the medium. The OLE1 gene, which encodes delta-9 fatty acid desaturase, has been reported to be repressed by unsaturated fatty acids. Transcription of OLE1 was also repressed by a wide variety of unsaturated fatty acids under anaerobic conditions. The degree of transcriptional repression of OLE1 was also related to the melting point of the added unsaturated fatty acids. Therefore, it is considered that ATF1 and OLE1 transcription are regulated in response to cell membrane fluidity. As has been reported for OLE1, the repression of ATF1 by unsaturated fatty acids was relieved in a disruptant carrying a faa1 and faa4 double mutation, two fatty acid activation genes. However, the ATF1 transcript in this double gene disruptant was repressed by oxygen. These results suggested that ATF1 transcription was co-regulated by the same mechanism as the OLE1 gene and that unsaturated fatty acids and oxygen repressed the ATF1 transcript by a different regulation pathway.

Acetyltransferases↗

PubMed1998

Co-existence of two types of chromosome in the bottom fermenting yeast, Saccharomyces pastorianus.

The bottom fermenting yeasts in our collection were classified as Saccharomyces pastorianus on the basis of their DNA relatedness. The genomic organization of bottom fermenting yeast was analysed by Southern hybridization using eleven genes on chromosome IV, six genes on chromosome II and five genes on chromosome XV of S. cerevisiae as probes. Gene probes constructed from S. cerevisiae chromosomes II and IV hybridized strongly to the 820-kb chromosome and the 1500-kb chromosome of the bottom fermenting yeast, respectively. Five gene probes constructed from segments of chromosome XV hybridized strongly to the 1050-kb and the 1000-kb chromosomes. These chromosomes are thought to be S. cerevisiae-type chromosomes. In addition, these probes also hybridized weakly to the 1100-kb, 1350-kb, 850-kb and 700-kb chromosome. Gene probes constructed from segments including the left arm to TRP1 of chromosome IV and the right arm of chromosome II hybridized to the 1100-kb chromosomes of S. pastorianus. Gene probes constructed using the right arm of chromosome IV and the left arm of chromosome II hybridized to the 1350-kb chromosome of S. pastorianus. These results suggested that the 1100-kb and 1350-kb chromosomes were generated by reciprocal translocation between chromosome II and IV in S. pastorianus. Three gene probes constructed using the right arm of chromosome XV hybridized weakly to the 850-kb chromosome, and two gene probes from the left arm hybridized weakly to the 700-kb chromosome. These results suggested that chromosome XV of S. cerevisiae was rearranged into the 850-kb and 700-kb chromosomes in S. pastorianus. These weak hybridization patterns were identical to those obtained with S. bayanus. Therefore, two types of chromosome co-exist independently in bottom fermenting yeast: one set which originated from S. bayanus and another set from S. cerevisiae. This result supports the hypothesis that S. pastorianus is a hybrid of S. cerevisiae and S. bayanus.

Chimera↗

PubMed1998

Effects of transjugular intrahepatic portosystemic shunt (TIPS) on esophageal motor function and gastroesophageal reflux.

The effects of transjugular intrahepatic portosystemic shunt (TIPS) placement on esophageal motor function and gastroesophageal reflux were investigated in patients with esophageal varices. In six men with esophageal varices, esophageal manometry and upper gastrointestinal endoscopy were performed before and 15-20 days after TIPS placement. Intraesophageal pH monitoring was performed in the four patients with severe esophageal varices (defined as the largest sized varices) following TIPS placement. Findings were compared with those in six healthy men (controls) who underwent esophageal manometry and intraesophageal pH monitoring. The esophageal varices resolved or were reduced after TIPS placement. Resting lower esophageal sphincter (LES) pressures were similar in the study group before and after TIPS placement and in the control subjects. The incidence and progression of esophageal contractions were similar in the study group before and after TIPS placement and in the control subjects. At 3 cm above the LES, the amplitude of esophageal contraction after TIPS placement was significantly higher than that before TIPS placement. At 3 and 8 cm above the LES, the amplitude of esophageal contraction in the control subjects was significantly higher than that in the study group before and after TIPS placement. Esophageal acid exposure time after TIPS placement was similar to that in the controls. TIPS placement is a useful treatment that improves esophageal motor function without the occurrence of pathologic gastroesophageal reflux.

Esophageal and Gastric Varices↗

PubMed1998

Increased proliferative activity of osteoblasts in congenital hemifacial hypertrophy.

Congenital hemifacial hypertrophy is expressed in facial asymmetry resulting from unilateral overgrowth. We examined proliferative activity of primary cultured osteoblasts taken from the hemifacial hypertrophic side of an 18-year-old woman compared with the normal side. Both fetal calf serum and basic fibroblast growth factor stimulated osteoblast DNA synthesis, and cultured osteoblast proliferation was markedly increased in the hypertrophic side. Neutralizing antibody against basic fibroblast growth factor was effective to partially block fetal calf serum-induced DNA synthesis. These results suggest that fibroblast growth factor and its receptor signal transduction axis may be selectively involved in affected osteoblasts, leading to progression of congenital hemifacial hypertrophy.

Adolescent↗

PubMed1998

Hemiacetal dehydrogenation activity of alcohol dehydrogenases in Saccharomyces cerevisiae.

Some methylotrophic yeasts produce methyl formate from methanol and formaldehyde via hemiacetal formation. We investigated Saccharomyces cerevisiae to find whether this yeast has a carboxylate ester producing pathway that proceeds via hemiacetal dehydrogenation. We confirmed that the purified alcohol dehydrogenase (Adh) protein from S. cerevisiae can catalyze the production of esters. High specific activities were observed toward the hemiacetals corresponding to the primary alcohols when ether groups were substituted for methylene groups, resulting in the formation of formate esters. Both ADH and methyl formate synthesizing activities were sharply reduced in the delta adh1 delta adh2 mutant. The ADH1 and ADH2 genes encode the major Adh proteins in S. cerevisiae. Thus, it was concluded that the S. cerevisiae Adh protein catalyzes activities for the production of certain carboxylate esters.

Acetals↗

PubMed1998

[Architectural changes with duration of hypertension in the retinal capillary network of spontaneously hypertensive rats].

Narrowing of the retinal artery in hypertension has been considered as an indicator of peripheral vascular resistance, which is largely caused by contraction or narrowness of retinal arterioles. But the relationship between peripheral vascular resistance and the retinal capillary network is little known. To study this relationship, three groups of age-matched rats were used: normotensive Wistar-Kyoto rats 6, 15, and 20 weeks, of age, and spontaneously hypertensive rats (stroke-resistant spontaneously hypertensive rats and stroke-prone spontaneously hypertensive rats) 6, 15, and 20 weeks of age. The size of the retinal capillary network in a given area was measured in these groups. There was no difference in the size of the retinal capillary network at any point in the Wistar-Kyoto rats. There was no difference in the size of the retinal capillary network among the three groups at 6 weeks of age, but there was a significant increase in spontaneously hypertensive rats at 15 weeks. We could not measure the size of the retinal capillary network in part of the spontaneously hypertensive rats at 20 weeks. These results suggested that retinal capillary network becomes larger with duration of hypertension and it acts to increase peripheral vascular resistance.

Animals↗