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Biomedical subjects

K Gomi

Publications and source records attributed to K Gomi.

At least 19 recordsLinked to original sources
PubMed2006

The 17-kDa sheath protein in enamel proteins induces cementum regeneration in experimental cavities created in a buccal dehiscence model of dogs.

BACKGROUND AND OBJECTIVE: Commercially available enamel proteins, such as Emdogain, are clinically used for periodontal regeneration. However, the real mechanisms behind the bioactivities of enamel proteins is still unclear, as enamel proteins have multicomponents. The purpose of this in vivo study was to identify the cementum regeneration-promoting factor in enamel proteins that is clinically used for periodontal regeneration to induce cementum-promotive and osteopromotive activities. MATERIAL AND METHODS: Cementum regeneration, which is an important part of periodontal regeneration, was examined in experimental cavities prepared on a buccal dehiscence model of dogs. The purification of enamel protein with cementum regeneration activity was carried out by gel filtration and ion exchange chromatographies of newly formed secretory enamel. RESULTS: Cementum regeneration activity was found in the aggregate comprising 13-17-kDa sheath proteins along with a small amount of amelogenins, found in the newly formed secretory enamel. In these proteins, cementum regeneration activity was detected upon application of the 17-kDa sheath protein, but not by other lower molecular-weight sheath proteins and amelogenins. However, the purified 17-kDa sheath protein induced cementum regeneration activity only in a small area, although the regenerated cementum was thick. The activity of the 17-kDa sheath protein was believed not to have been a result of contamination by growth factors such as transforming growth factor-beta1 (TGF-beta1) found in the enamel protein, as the application of TGF-beta1 induced weak cementum regeneration activity. CONCLUSION: It is concluded that the 17-kDa sheath protein itself exhibits cementum regeneration activity, although other factors may be needed to demonstrate its full ability.

Alveolar Bone Loss↗
PubMed2005

Enamel matrix derivative gel stimulates signal transduction of BMP and TGF-{beta}.

It has been shown that Emdogain Gel (Emd-Gel) containing enamel matrix proteins promotes biomineralization, such as osteogenesis and cementogenesis, during the regeneration of periodontal tissues. However, the growth factors involved in these activities of Emd-Gel remain unclear. In this study, Emd-Gel was fractionated into 22 sub-fractions by size exclusion chromatography. The osteoinductive factors, TGF-beta and BMP, were examined by a specific luciferase reporter gene assay. In the unfractionated Emd-Gel, TGF-beta-like activity was detected, while BMP activity was not. In contrast, in the fractionated Emd-Gel samples, TGF-beta-like activity was detected from fractions 8 to 13, and BMP-like activity was detected from fractions 4 to 6. Also, it was confirmed that the BMP-like activity in Emd-Gel was inhibited by authentic TGF-beta1 and TGF-beta-like activity. These results indicate that Emd-Gel contains both TGF-beta- and BMP-like growth factors that contribute to the induction of biomineralization during periodontal regeneration.

Animals↗
PubMed2004

Pharmacokinetic analysis of combination chemotherapy with carboplatin and etoposide in small-cell lung cancer patients undergoing hemodialysis.

BACKGROUND: The aim of this study was to use pharmacokinetic analysis to investigate the efficacy and toxicity of combined chemotherapy with carboplatin (CBDCA) and etoposide (ETP) in small-cell lung cancer (SCLC) patients with chronic renal failure undergoing hemodialysis (HD). PATIENTS AND METHODS: Three SCLC patients with chronic renal failure undergoing HD were treated with CBDCA (300 mg/m(2)) on day 1 and ETP (50 mg/m(2)) on days 1 and 3, followed by HD 1 h after completing the administration of anticancer agents on each day. The pharmacokinetic analysis of CBDCA and ETP was planned for at least the first two courses of the chemotherapy in each patient. RESULTS: Two complete responses and one partial response were achieved in the three patients. Two patients experienced grade 3/4 neutropenia and required blood transfusion due to thrombocytopenia and anemia. Non-hematological toxicities were moderate. The pharmacokinetic analysis revealed that the platinum and the ETP concentrations in the plasma were similar to those in patients with normal renal function during the first 24 h, while the platinum still remained in the plasma for over 90 h. CONCLUSIONS: Chemotherapy with CBDCA (300 mg/m(2) on day 1) and ETP (50 mg/m(2) on day 1, 3) as used in the present study may be a suitable regimen for SCLC patients undergoing HD, although careful attention should be given to hematological toxicities.

Aged↗
PubMed2003

Relative levels of mRNA encoding enamel proteins in enamel organ epithelia and odontoblasts.

Amelogenin, enamelin, sheathlin (ameloblastin/ amelin), enamelysin (MMP-20), and KLK4 (EMSP-1) are the major structural proteins and proteinases in developing tooth enamel. Recently, odontoblasts were reported to express amelogenin, the most abundant enamel protein. In this study, we hypothesized that odontoblasts express all enamel proteins and proteases, and we measured their relative mRNA levels in enamel organ epithelia and odontoblasts associated with porcine secretory- and maturation-stage enamel by RT-PCR, using a LightCycler instrument. The results showed that amelogenin mRNA in secretory-stage EOE is 320-fold higher than in odontoblasts beneath secretory-stage enamel, and over 20,000-fold higher than in odontoblasts under maturation-stage enamel. Similar results were obtained for enamelin and sheathlin. Enamelysin mRNA levels were equivalent in these two tissues, while KLK4 mRNA was higher in odontoblasts than in secretory-stage EOE. These results support the conclusion that odontoblasts are involved in the formation of the enamel layer adjacent to enamel-dentin junction.

Ameloblasts↗
PubMed2001

Oxyluciferin, a luminescence product of firefly luciferase, is enzymatically regenerated into luciferin.

The activity regenerating luciferin from the luminescent product oxyluciferin was found in the protein fraction of a lantern extract from Photinus pyralis. The protein, luciferin-regenerating enzyme (LRE), was purified to homogeneity by ammonium sulfate precipitation followed by successive column chromatography on Ultrogel AcA34, S-Sepharose FF, Q-Sepharose FF, TSKgel super Q 5pw and TSKgel G3000 SW(XL). This enzyme was a single polypeptide with a molecular mass of 38 kDa. LRE converted oxyluciferin to 2-cyano-6-hydroxybenzothiazole and thioglycolic acid. In the presence of d-cysteine, 2-cyano-6-hydroxybenzothiazole was turned over into luciferin. The same activities were detected in the extracts from two Japanese fireflies, Luciola cruciata and Luciola lateralis. We have cloned a cDNA encoding LRE from poly(A)+ RNA of the lantern of P. pyralis using reverse transcription-polymerase chain reaction, 5'-RACE (rapid amplification of cDNA ends) and 3'-RACE. The primary structure of LRE from P. pyralis deduced from the nucleotide sequence was shown to consist of 308 amino acids with a molecular weight of 33,619. The cDNA was successfully expressed under the control of the tac promoter in Escherichia coli.

Amino Acid Sequence↗
PubMed2001

Cutting edge: Gln22 of mouse MD-2 is essential for species-specific lipopolysaccharide mimetic action of taxol.

MD-2 associates with the extracellular domain of Toll-like receptor 4 (TLR4) and greatly enhances LPS signaling via TLR4. Taxol, which mimics the action of LPS on murine macrophages, induces signals via mouse TLR4-MD-2, but not via human TLR4-MD-2. Here we investigated the molecular basis for this species-specific action of Taxol. Expression of mouse MD-2 conferred both LPS and Taxol responsiveness on human embryonic kidney 293 cells expressing mouse TLR4, whereas expression of human MD-2 conferred LPS responsiveness alone, suggesting that MD-2 is responsible for the species-specificity as to Taxol responsiveness. Furthermore, mouse MD-2 mutants, in which Gln(22) was changed to other amino acids, showed dramatically reduced ability to confer Taxol responsiveness, although their ability to confer LPS responsiveness was not affected. These results indicated that Gln(22) of mouse MD-2 is essential for Taxol signaling but not for LPS signaling.

Amino Acid Sequence↗
PubMed2001

Deletion analysis of the enolase gene (enoA) promoter from the filamentous fungus Aspegillus oryzae.

The enolase gene (enoA) is one of the most strongly expressed genes in Aspergillus oryzae. To elucidate the transcription regulatory element for this strong expression and the process of glucose induction, the transcription activity of a series of truncated enoA promoters was measured by using the Escherichia coli uidA gene as a reporter. Deletion of a 104-bp region located -224 nt to -121 nt upstream of the translation initiation site caused both a drastic decrease in the beta-glucuronidase (GUS) activity and a loss of glucose induction. Northern blot analysis confirmed that the decrease in GUS activity was achieved at the transcriptional level. In addition, electrophoretic gel mobility shift assays indicated that the 104-bp region contained a 15-bp element, to which one or more A. oryzae cellular factors specifically bind. These results suggest that the 15-bp element between -195 nt and -181 nt includes the sequence essential for the transcription regulation of the A. oryzae enoA gene.

Aspergillus oryzae↗
PubMed2001

Characterization of the amyR gene encoding a transcriptional activator for the amylase genes in Aspergillus nidulans.

The Aspergillus nidulans amyR gene and its cDNA were cloned and sequenced. The genomic gene comprised 2,092 bp, interrupted by two short introns, and encoded a cys-6 zinc transcriptional activator (AMYR) of 662 amino acid residues with a calculated molecular mass of 72,862 Da. Disruption of the amyR gene caused defects in the utilization of maltose and starch and abolished expression of the taaG2 gene encoding A. oryzae Taka-amylase A, which is inducibly and abundantly expressed in the wild-type A. nidulans. Expression of the amyR gene was under the control of the carbon catabolite repressor, CREA. The growth defect of the malA1 mutant on maltose was complemented by the amyR gene; and the amyR gene derived from the mutant possessed a single mutation, from A to T, at position 1,483, resulting in a substitution of His478 to Leu. These results indicate that the amyR gene is identical to the genetically defined malA gene. AMYR possessed five domains (Zn and MH1-MH4) homologous to Mal63p, a transcriptional activator for the genes involved in maltose utilization in Saccharomyces cerevisiae. The His478 to Leu substitution lay within the MH3 domain, corresponding to the negative regulatory domain of Mal63p which relieves the inhibitory effect on the activation function in response to maltose.

Amino Acid Sequence↗
PubMed2001

Comparative in-vitro activity of carbapenem antibiotics against respiratory pathogens isolated between 1999 and 2000.

We investigated the antibacterial activity of 12 antibiotics, inclusive of four carbapenems, against 167 strains of respiratory pathogens isolated between 1999 and 2000. Thirty strains of methicillin-susceptible Staphylococcus aureus (MSSA), 28 strains of methicillin-resistant S. aureus (MRSA), 11 strains of penicillin-susceptible Streptococcus pneumoniae (PSSP), 29 strains of penicillin-resistant S. pneumoniae (PRSP), 30 strains of Pseudomonas aeruginosa, 14 strains of Moraxella catarrhalis, and 25 strains of Haemophilus influenzae were examined. The minimum inhibitory concentration (MICs)50/90 (microg/ml) of imipenem, panipenem, meropenem, and biapenem against the clinical isolates obtained between 1999 and 2000 were: 0.06/0.25, 0.12/0.25, 0.12/0.25, and 0.12/0.25, respectively, against MSSA; 16/32, 16/32, 16/32, and 8/32 against MRSA; < or = 0.015/0.06, < or = 0.015/0.03, 0.03/0.12, and < or = 0.015/0.06 against PSSP; 0.12/0.25, 0.03/0.06, 0.25/0.5, and 0.12/0.25 against PRSP; 1/8, 2/8, 0.5/2, and 2/16 against P. aeruginosa; 0.06/0.06, 0.03/0.06, < or = 0.015/0.06, and 0.06/0.12 against M. catarrhalis; and 1/4, 1/4, 0.12/0.25, and 2/4 against H. influenzae. A comparison of the antibacterial activity of the four carbapenems with that found in our previous studies showed no significant difference in the susceptibility of clinical isolates, except for a slight decrease in the susceptibility of MSSA. Carbapenems have remained effective for severe infections. The MIC data showed that imipenem and panipenem were more active than meropenem and biapenem against gram-positive bacteria, and that meropenem and biapenem were more active than imipenem and panipenem against gram-negative bacteria. As only meropenem had an MIC90 below the breakpoint of pneumonia against all species except MRSA, meropenem was considered to be the most potent of the four carbapenems studied.

Anti-Bacterial Agents↗
PubMed2001

Chiral recognition of thiaheterohelicenes by alkyl beta-D-pyranoside micelles. Influence of extension of helix.

Chiral recognition of alkyl beta-D-pyranoside micelles toward [7] and [5] heterohelicenes possessing helical structures was investigated by 1H-NMR and CD (circular dichroism) spectroscopy. In dodecyl maltopyranoside micellar solution, P and M enantiomers of tetrathia[7]heterohelicene (7TH), which have rigid and stable helixes, manifested different chemical shifts in their 1H-NMR spectra due to differences in the diastereomeric interactions, implying that the micelles undergo stronger recognition toward the M enantiomer than the P enantiomer. On the other hand, in octyl glucopyranoside micellar solution, trithia[5]heterohelicene (5TH) and two kinds of its derivatives which are rapidly equilibrated between the enantiomers in solution, gave no distinctly resolved 1H-NMR peaks for either enantiomer even at a lowered temperature. However, these racemic [5]heterohelicenes in the micelles did develop induced CD absorptions owing to a displacement of the equilibrium, suggesting from the signs of their Cotton effects that the micelles prefer the M enantiomer to its antipode in conformity with the 1H-NMR results of 7TH.

Circular Dichroism↗
PubMed2001

High-level secretory production of phospholipase A1 by Saccharomyces cerevisiae and Aspergillus oryzae.

Phospholipase A1 (PLA1) is a hydrolytic enzyme that catalyzes the removal of the acyl group from position 1 of lecithin to form lysolecithin. The PLA1 gene, which had been cloned from Aspergillus oryzae, was expressed in Saccharomyces cerevisiae and A. oryzae. Through the modification of the medium composition and the feeding conditions of substrate, the production level of PLA1 by S. cerevisiae was increased to a level fivefold higher than that indicated in a previous report. In the case of A. oryzae, introduction of multicopies of PLA1 expression units, and the morphological change from the pellet form to the filamentous form were effective for the enhancement of PLA1 production. We succeeded in producing 3,500 U/ml of PLA1 using an industrial-scale fermentor.

Aspergillus oryzae↗
PubMed2001

[Hepatitis C virus genotyping by restriction fragment length polymorphism of polymerase chain reaction products generated with a HCV detection kit].

Highly conserved sequence in the 5' untranslated region(UTR) of hepatitis C virus(HCV) genome have been targeted by most nucleic acid amplification-based detection assays, such as Amplicor HCV test, a commercially available assay kit. In this study, we classified HCV genotypes by direct sequencing determination for 5' UTR of nested-PCR after Amplicor HCV test. Then, based on the results of sequence, RFLP analysis after digestion of the nested PCR fragments with Hae III or Sau 3AI to classify HCV genotype was evaluated. RFLP analysis distinguished the type 1, 2a and 2b. Only one of 29 samples was not classified by RFLP analysis due to the point mutation of Hae III recognition site. HCV genotypes commonly found in JAPAN were classified into three types, 1b, 2a, and 2b. Also, RFLP analysis requires fewer resources than serotype grouping test. Hence, the present method provides an adaptable and rapid HCV genotyping in clinical laboratory in JAPAN.

Base Sequence↗
PubMed2001

[Creatine kinase and its isozymes].

Creatine kinase (CK; EC 2.7.3.2) is found in a variety of striated and smooth muscles, and the brain, and is an important enzyme regulator of high-energy phosphate production and utilization within contractile tissues. Serum CK activity is measured routinely as a sensitive indicator of injuries to the skeletal muscle and myocardium. CK has three isozymes (CK-MM, CK-MB and CK-BB) in cytoplasm and two isozymes (non-sarcomeric and sarcomeric) in mitochondria. CK isozymes provide more specific information about injured tissue because of their tissue distribution. CK-MM is useful in skeletal muscle diseases, such as muscle dystrophy, CK-MB in acute myocardial infarction (AMI), and CK-BB in brain damage and malignant tumor of the gastro-intestinal tract. CK-MB is measured either by enzyme activity or mass concentration and is measured as a marker not only in the diagnosis of AMI but also in suspected AMI and unstable angina. Mitochondrial CK, a useful indicator for pinpointing the severeness of muscle injuries, and CK-linked immunoglobulin are recognized as macro-CK because of their large molecular size.

Angina, Unstable↗
PubMed2000

Prostaglandin E2 selectively enhances the IgE-mediated production of IL-6 and granulocyte-macrophage colony-stimulating factor by mast cells through an EP1/EP3-dependent mechanism.

PGE(2) is an endogenously synthesized inflammatory mediator that is over-produced in chronic inflammatory disorders such as allergic asthma. In this study, we investigated the regulatory effects of PGE(2) on mast cell degranulation and the production of cytokines relevant to allergic disease. Murine bone marrow-derived mast cells (BMMC) were treated with PGE(2) alone or in the context of IgE-mediated activation. PGE(2) treatment alone specifically enhanced IL-6 production, and neither induced nor inhibited degranulation and the release of other mast cell cytokines, including IL-4, IL-10, IFN-gamma, and GM-CSF. IgE/Ag-mediated activation of BMMC induced the secretion of IL-4, IL-6, and GM-CSF, and concurrent PGE(2) stimulation synergistically increased mast cell degranulation and IL-6 and GM-CSF, but not IL-4, production. A similar potentiation of degranulation and IL-6 production by PGE(2), in the context of IgE-directed activation, was observed in the well-established IL-3-dependent murine mast cell line, MC/9. RT-PCR analysis of unstimulated MC/9 cells revealed the expression of EP(1), EP(3), and EP(4) PGE receptor subtypes, including a novel splice variant of the EP(1) receptor. Pharmacological studies using PGE receptor subtype-selective analogs showed that the potentiation of IgE/Ag-induced degranulation and IL-6 production by PGE(2) is mediated through EP(1) and/or EP(3) receptors. Our results suggest that PGE(2) may profoundly alter the nature of the mast cell degranulation and cytokine responses at sites of allergic inflammation through an EP(1)/EP(3)-dependent mechanism.

Adjuvants, Immunologic↗
PubMed2000

Novel arbekacin- and amikacin-modifying enzyme of methicillin-resistant Staphylococcus aureus.

An aminoglycoside-modifying enzyme in arbekacin-resistant methicillin-resistant Staphylococcus aureus (MRSA), exhibiting 4'''-N-acetylation, was examined. Although the MRSA strain with AAC(4''') had no AAC(6')-APH(2") activity, a DNA fragment of the AAC(6')-APH(2") gene was amplified by PCR and the purified N-terminal 30-amino acid sequence of this AAC(4''') was identical to AAC(6')-APH(2"). Direct DNA sequencing of this 'silent' AAC(6')-APH(2") gene revealed a single point mutation leading to a substitution of Gly for Asp80, through which the secondary structure is affected. A change in protein conformation could lead to a cleavage and a change of the enzymatic activity. We propose a new aminoglycoside-resistance mediated by AAC(4''') is caused by a mutation-modified AAC(6')-APH(2").

Acetyltransferases↗
PubMed2000

Phosphatidylinositol 3-kinase inhibitors, Wortmannin or LY294002, inhibited accumulation of p21 protein after gamma-irradiation by stabilization of the protein.

Expression of the cyclin kinase inhibitor, p21, is regulated both transcriptionally and posttranscriptionally by the ubiquitin-proteasome degradation pathway. Recently, we reported that DNA damage is required for efficient p21 expression by demonstrating that enhanced p21 mRNA expression induced by DNA damage results in increased p21 protein, but enhanced p21 mRNA without DNA damage does not. In addition, we demonstrated that DNA damage suppressed the ubiquitination of p21. In this study, we analyze the link between p21 stabilization and DNA damage. Enhanced p21 protein expression in ML-1 cells resulting from 15 Gy gamma-irradiation was diminished by Wortmannin or LY294002 pretreatment of cells. However, the levels of p21 mRNA were not affected by inhibitor pretreatment. Wortmannin or LY294002 pretreatment reduces p53 expression after gamma-irradiation to a lesser degree than that of p21. In addition, we examined the involvement of DNA-PK, whose activity is inhibited by Wortmannin or LY294002, in p21 stabilization using the SCID fibroblast cell line and a DNA-PK targeting ML-1 cell line. Accumulation of p21 protein by gamma-irradiation was similar to that of DNA-PK intact cells and was reduced by Wortmannin or LY294002 pretreatment. Involvement of another DNA damage detecting enzyme, the ATM gene product, whose activity is also inhibited by Wortmannin or LY294002, was evaluated. ATM deficient cells induced p21 after gamma-irradiation, gamma-irradiation-induced p21 protein was diminished by pretreatment of cells with Wortmannin or LY294002. We conclude that the p21 stabilization mechanism functions after gamma-irradiation, was sensitive to Wortmannin or LY294002, and required neither DNA-PK nor ATM gene product for activity.

3T3 Cells↗
PubMed2000

High galactosylation of oligosaccharides in umbilical cord blood IgG, and its relationship to placental function.

N-linked oligosaccharides on human serum IgGs have been reported to modulate IgG function. We studied umbilical cord blood to determine whether neonatal IgGs have characteristic structures related to developmental and pathological status. Oligosaccharide patterns of serum IgG from 45 umbilical cord blood samples were characterized by HPLC, and compared with those of serum IgG from 11 normal adults. Oligosaccharyl amines from purified IgG were released by recombinant N-glycanase, labeled with fluorescence reagent FMOC (9-fluorenylmethyl chloroformate), and analyzed quantitatively by high-pressure liquid chromatography (HPLC). Increased galactosylation was observed in cord blood. The ratio of galactosylated to non-galactosylated oligosaccharides on IgG was 7.90+/-3.92 (mean+/-S.D.) in cord blood, significantly higher than the ratio in adults (1.60+/-0.62, P<0.0001). There were weak but not significant correlations between the ratio and birth weight, gestation period, mother's age, and no correlation with serum IgG concentration. The ratio was lower for premature or intra-uterine growth retarded neonates. Our results, in conjunction with previous reports that galactosylated IgG stimulates Fc-mediated phagocytosis of monocytes, suggest that increased galactosylation of IgG enhances neonatal immunity.

Adult↗