Biomedical subjects
Publications and source records attributed to T Onoe.
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BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common inherited renal disorders in the world. Mutations in PKD1 are responsible for 80-95% of all autosomal dominant polycystic kidney disease (ADPKD). Although the need for linkage analysis of ADPKD is decreasing after the success of mutation detection at whole exons of PKD1, linkage analysis still has some advantages in detecting non-PKD1 families, thereby avoiding hopeless mutation analysis. METHODS: We evaluated ten microsatellite markers beside or inside PKD1 on chromosome 16p. Allele frequency and heterozygosity of each marker were calculated based on the 100 genotypes obtained from 50 normal Japanese. Automated microsatellite genotyping using ABI Prism 377 and GeneScan software was applied. Markers were mapped using radiation hybrid mapping. Finally, this strategy was applied in the linkage analysis of 6 independent Japanese ADPKD families. RESULTS: D16S3024, D16S3082, D16S3027 and D16S423 showed high heterozygosity (> 0.80) in a normal Japanese population and sufficient proximity to the PKD1 gene for linkage analysis. We could successfully analyze 144 genotypes within 7 hours. This strategy produced theoretically near-maximum LOD scores in 4 independent Japanese families inheriting ADPKD. CONCLUSIONS: Automated genotyping using microsatellite markers, D16S3024, D16S3082, D16S3027 and D16S423 are very useful in the linkage analysis of ADPKD.
Polycystic kidney disease (PKD) is one of the most common genetic disorders and a major cause of renal death or end-stage renal disease (ESRD) requiring regular hemodialysis. The responsible genes recently have been cloned; however, genetic factors influencing the rate of progression to ESRD in patients with PKD have yet to be defined. Several studies have shown increased activity of the renin-angiotensin system (RAS) in patients with PKD. In addition, genetic polymorphisms of the RAS have been associated with the development of cardiovascular diseases. Therefore, these polymorphisms are good candidates for disease-modifying genetic factors or markers in PKD. In two previous reports of white subjects with a cumulative survival analysis, it was suggested that patients with P:KD1 homozygous for the deletion allele of the angiotensin-converting enzyme (ACE) gene are at increased risk for early renal death. To confirm this hypothesis in Japanese subjects, 103 individuals with PKD were genotyped for several components of the RAS, ie, ACE insertion/deletion (I/D) polymorphism, angiotensinogen (AGT) M235T, and angiotensin II type 1 receptor (AT1) A1166C. Seventy-six of the 103 patients (73.8%) reached ESRD at an average age of 52.1 +/- 11.3 years. The frequencies of each genotype of the genes were similar to those expected from Hardy-Weinberg equilibrium. There was a tendency to an excess of patients homozygous for the D allele in patients with ESRD (DD in patients with ESRD, 11.8%; DD in patients without ESRD, 3.7%; chi-square, 1.505; P: = 0.22). Cumulative renal survival was significantly less in those with the DD genotype compared with ID/II genotypes. Estimated mean renal survival was 46.4 years (95% confidence interval, 39.5 to 53.3) in subjects with the DD genotype and 57.2 years (95% confidence interval, 54.2 to 60.2) in ID/II genotypes (chi-square, 7.76; P: = 0.0053). There was no association between age at onset of ESRD and either M235T or A1166C polymorphism. These findings suggest that Japanese patients with PKD homozygous for the D allele of the ACE gene are at increased risk for developing ESRD at an early age.
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OBJECTIVE: The purposes of this study were to examine the time-related changes in pH, oxidation-reduction potential, and concentration of chlorine of electrolyzed neutral water and to evaluate the bactericidal effect of electrolyzed neutral water against bacteria from infected root canals. STUDY DESIGN: Various properties of electrolyzed neutral water--pH value, oxidation-reduction potential, and concentration of chlorine--were measured at different times after storage of the water in the open state, the closed state, or the closed-and-dark state. The bactericidal effect of the various electrolyzed neutral water samples was then tested against 17 strains of bacteria, including 15 strains isolated from infected canals, as well as against 1 strain of fungus. Each bacterial or fungal suspension was mixed with electrolyzed neutral water, and the 2 substances were reacted together for 1 minute. After incubation for 1 to 7 days, the bactericidal effect of the electrolyzed neutral water was determined. RESULTS: The pH value and oxidation-reduction potential of electrolyzed neutral water remained almost unchanged when the water was stored in a dark, closed container. However, the concentration of chlorine decreased from 18.4 ppm to 10.6 ppm. Electrolyzed neutral water showed a bactericidal or growth-inhibitory effect against the bacteria. CONCLUSIONS: The results indicate that electrolyzed neutral water maintains a constant pH and oxidation-reduction potential when kept in a closed container without light and that it exhibits a bacteriostatic/bactericidal action against isolates obtained from infected root canals.
A gene upstream from fimA, the gene encoding fimbrilin, on the chromosome of Porphyromonas gingivalis was sequenced and shown to be the gene encoding an outer membrane protein in this organism based on homology and biochemical analyses. Therefore, the gene (formerly ORF5) was designated pgmA, the P. gingivalis outer membrane protein A gene. The gene product, PgmA, was sensitive to protease, and was detected as a 60-kDa protein from wild-type strains with trichloroacetic acid treatment, which was carried out to destroy intrinsic proteases, and from protease-deficient mutants without this treatment prior to electrophoresis. PgmA was indeed present in the membrane fraction. Its nature was determined to be that of outer membrane proteins in gram-negative bacteria based on attempts at differential extraction of inner membrane proteins with detergents. No evidence has been found thus far from functional analyses that this protein is related to fimbrial morphogenesis and functions or to serum resistance of this organism.
The patient, a female, aged 65 years, developed diffuse peritoneal calcification nine years after commencing CAPD therapy. No abdominal symptoms or evidence of peritonitis were discovered during this period. Before peritoneal calcification was detected, a dialysate with a high glucose concentration (3.86%) had been used once daily for 16 months. In the case of this patient, it was not possible to discover any of the previous indicated etiologies of peritoneal calcification such as significantly elevated values for the product Ca x P, overt secondary hyperparathyroidism, or relapsing peritonitis. It was realized that the use of a high-glucose dialysate in a patient on long-term CAPD treatment had been one causative factor. After peritoneal calcification had been confirmed, the calcium concentration of the dialysate changed from 3.5 mEq/l to 2.5 mEq/l and the patient was put on a regime of 2.0 g alumigel (aluminum-containing phosphate binders) a day. Eight months later, a CT scan was taken. The peritoneal calcification has clearly been mitigated. At present, CAPD therapy is being continued in the absence of any abdominal symptoms.
The downstream DNA region of the fimbrilin gene (fimA), which encodes the major subunit protein of Porphyromonas gingivalis fimbriae, was fully sequenced. Gene products, expressed from this region in Escherichia coli, were purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and their partial amino acid sequences were determined to verify open reading frames (ORFs) found in the region by DNA sequencing. Four ORFs, designated ORF1, ORF2, ORF3 and ORF4, were found in the 5.8-kb PstI fragment downstream from fimA, which was previously cloned and partially characterized by Yoshimura, Takahashi, Hibi, Takasawa, Kato, and Dickinson (Infect. Immun. 61: 5181-5189, 1993). The direction of transcription of all the ORFs was the same as that of fimA. The 50 and 80 kDa encoded proteins, ORF2 and ORF3, respectively, have been reported to be minor components associated with fimbriae. The 15 and 19 kDa proteins, ORF1 and ORF4, respectively, have been expressed in E. coli but not identified in P. gingivalis. However, all the gene products of the ORFs, expressed in E. coli, appeared to contain intact signal peptides based on their N-terminal amino acid sequences.
Fimbriae of Porphyromonas gingivalis have been shown to be important as one of the virulence factors for colonization on mucosal surfaces. The gene (fimA) encoding the fimbrial subunit (fimbrilin) was overexpressed in Escherichia coli by using a bacteriophage T7 promoter-polymerase expression vector system. Analysis of the resulting fimA gene product revealed that the prefimbrilin had a 46 amino acid leader peptide. This extremely long leader peptide was cleaved from the prefimbrilin by treatment with trypsin or P. gingivalis extracts containing trypsin-like protease activity, resulting in production of a mature fimbrilin. We also found that some transposon-induced trypsin-like protease deficient mutants of P. gingivalis exhibited deficiency in fimbriation and that one of the mutants accumulated a fimbrilin precursor possessing a 25 amino acid leader peptide in the cell. The presence of an extremely long leader peptide and the requirement for a leader peptidase with a substrate specificity similar to that of P. gingivalis trypsin-like protease for fimbrilin maturation indicate that P. gingivalis fimbrilin is a novel type that is different from fimbrilins of type I and IV families.
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We have recently isolated from the gingival pockets, periapical lesions and saliva some anaerobic gram-negative, black-pigmented rods. Many of these isolates exhibited phenotypic characteristics similar to Prevotella intermedia (Bacteroides intermedius). However, several of these isolates, although resembling P. intermedia in most of the phenotypic expressions, were capable of fermenting lactose, a biochemical characteristic atypical of P. intermedia. These atypical clinical isolates (strains capable of fermenting lactose) and isolates exhibiting more typical phenotypic characteristics (i.e., lactose nonfermenting) were definitively identified as P. intermedia by DNA-DNA hybridization using a photoprobe biotin method. Quantitative hybridization of clinical isolates with labeled DNA of P. intermedia-type strains (ATCC 25611 and ATCC 33563) showed that almost all the clinical strains isolated from disease sites of adults belonged to the ATCC 25611 group, whereas strains isolated from the saliva of children belonged to the ATCC 33563 group. These data, together with the phenotypic characterization of the isolates, suggested that P. intermedia is a heterogeneous species both phenotypically and genetically.
Human papillomavirus (HPV) types 16, 18 and 33 were identified by means of the polymerase chain reaction using exfoliated cells from the uterine cervix in 361 patients. Of 261 patients without cervical lesions, 10(3.8%) patients had HPV DNA whereas 7(70.0%) of 10 patients with invasive cervical carcinomas had HPV DNA. The younger patients' group (29 year-old or less) without cervical lesions had a 6.5% HPV positive rate which was distinctly higher than the older patients' groups. No menopausal patient without cervical lesions had HPV DNA. In the cervical dysplasia group, the HPV DNA positive rate tended to be higher in the older patients. Type 16 was detected more often than types 18 or 33. However, the detectable incidence of type 16 in the follow up group was lower than in the cervical carcinoma groups. The younger patients without cervical lesions had a higher incidence of type 16 than the older patients. The younger patients with cervical neoplastic lesions had a lower incidence of type 16 than the older patients. These results suggest that type 16 has a higher frequency of cervical HPV infections than types 18 and 33. In addition, human papillomavirus is not the only causative factor in cervical carcinomas.
The oral condition of a 7 year old boy at the 12th and 13th remission induction therapy sessions since contracting acute lymphatic leukemia was investigated clinically and microbiologically. The findings obtained were as follows: 1) The appearance of gingivitis coincided with the decrease of white blood cells 7 to 9 days after administration of the anti-leukopenic drugs. 2) Gingivitis appeared on the same site of the oral cavity at both remission induction therapies sessions. 3) The bleeding from the gingival lesions corresponded to the decrease of blood platelet at both remission induction therapies sessions. Conversely, gingivitis disappeared with the improvement of peripheral blood conditions. 4) The total number of oral bacteria decreased after administration of the antibiotics, although the ratio of Candida in the oral microbial flora increased markedly. 5) Typical symptoms of oral candidiasis were not observed after the appearance of the microbisme substitute.
Production of 9 enzymatic activities of 527 strains freshly isolated from periodontal pockets in advancing periodontitis were investigated. Of these isolates, two strains showed lecithinase activity on egg yolk agar plate. Collagenase, plasmin and lipase were produced by 28 strains, 26 strains and 22 strains, respectively. Two lecithinase-producing strains were identified as Bacteroides intermedius. Nineteen strains of B. intermedius and 1 strain of Fusobacterium species produced lipase on egg yolk agar plate. All of the 28 collagenase-producing strains were B. gingivalis. B. gingivalis (20 strains) and non black-pigmented Bacteroides (6 strains) showed plasmin activity. These results indicate that Bacteroides species, mainly B. gingivalis and B. intermedius may exert an important influence on the exacerbation of the disease.
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