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Biomedical subjects

T Kinouchi

Publications and source records attributed to T Kinouchi.

At least 37 records · Page 2Linked to original sources

Genome analysis of Bacteroides by pulsed-field gel electrophoresis: chromosome sizes and restriction patterns.

The chromosomal DNAs of nine strains of seven Bacteroides species including B. fragilis, the type species of the genus Bacteroides, were digested with rare-cutting restriction enzymes I-Ceu I, Not I, and Asc I and analysed by pulsed-field gel electrophoresis. The genome sizes of B. fragilis, B. distasonis, B. eggerthii, B. ovatus, B. thetaiotaomicron, B. uniformis, and B. vulgatus were determined to be 5.3, 4.8, 4.4, 6.9, 4.8, 4.6, and 5.1 Mbp, respectively. B. distasonis and B. vulgatus, and also B. uniformis and B. eggerthii, showed similar I-Ceu I restriction profiles. I-Ceu I cut B. uniformis and B. eggerthii genomes into four, B. ovatus into five, B. fragilis and B. thetaiotaomicron into six, and B. distasonis and B. vulgatus into seven fragments. On the basis of genome size, restriction profile, and I-Ceu I fragment number, a phylogenetic tree of the Bacteroides species was proposed. This was in overall agreement with the previous phylogenetic tree obtained by 16S rRNA data, with the exceptions of B. distasonis and B. ovatus.

Bacteroides

Inhibitory effects of lemon grass (Cymbopogon citratus Stapf) on formation of azoxymethane-induced DNA adducts and aberrant crypt foci in the rat colon.

The 80%-ethanol extract of lemon grass (Cymbopogon citratus Stapf), a medicinal plant in Thailand, has been reported to be antimutagenic against various known mutagens in the Salmonella mutation assay. To investigate chemoprevention in an animal carcinogenesis model, we examined inhibitory effects of the lemon grass extract on the formation of azoxymethane (AOM)-induced DNA adducts and aberrant crypt foci (ACF) in the rat colon. One week after the start of the treatment with lemon grass extract at doses of 0.5 or 5 g/kg body wt by gavage, F344 rats received two s.c. injections of 15 mg of AOM per kg body weight at 1 week apart. For DNA adduct analysis of the colon and liver, the rats were killed 12 h after the second AOM injection. The DNA from the liver and colon were used for O6-methylguanine and N7-methylguanine analysis. For ACF analysis in the initiation stage, AOM-injected rats were continuously treated with lemon grass extract and were killed 3 weeks after the second AOM injection. For analysis in the promotion stage the treatment with the lemon grass extract (0.5 g/kg) started 2 weeks after the second AOM injection and continued for 12 weeks until the animals were killed. Lemon grass treatment significantly inhibited DNA adduct formation in both the colonic mucosa and the muscular layer but not in the liver. In addition, lemon grass extract treatment significantly inhibited ACF formation in both the initiation stage and the promotion stage. Especially in the promotion stage, lemon grass treatment inhibited the formation of larger ACF (with four or more crypts per focus), which was predictive of tumor incidence. Furthermore, lemon grass extract inhibited fecal beta-glucuronidase competitively and had antioxidant activity. These results suggest that the lemon grass extract inhibits the release of activated aglycon, methylazoxymethanol, from a glucuronide conjugate in the colon, and decreases the DNA adducts and ACF formation in the rat colon.

Animals

Deletion of an endosomal/lysosomal targeting signal promotes the secretion of Alzheimer's disease amyloid precursor protein (APP).

Alzheimer's disease amyloid precursor protein (APP) generates a beta-amyloid protein (A beta) that is a main component of the senile plaques found in the brains of Alzheimer's disease patients. APP is thought to undergo proteolysis via two different pathways, the amyloidogenic pathway which produces A beta, and the non-amyloidogenic pathway which releases a large N-terminal fragment into the medium. The proteases that mediate these processes remain unidentified. The physiological function of APP is not clear yet. Therefore, the cytoplasmic region of APP has attracted much interest, because this region is highly conserved among species, and members of the amyloid precursor-like protein (APLP) family. Several potentially functional sequences exist in the region, including signal sequences for protein sorting and a G0-protein binding sequence. We constructed two mutants, 695 deltaNPTY and 695 deltaGYEN. They lack potential endosome/lysosome targeting signals, NPTY and GY, in the cytoplasmic domain of APP695, respectively. The mutant APPs had longer half-lives and were secreted more easily into the medium than the wild type, suggesting that these sequences are important for the secretion and metabolism of APP.

Alzheimer Disease

Alanyl aminopeptidase from human seminal plasma: purification, characterization, and immunohistochemical localization in the male genital tract.

Alanyl aminopeptidase (AAP) was purified to homogeneity from human seminal plasma. The calculated molecular weight of the purified enzyme was approximately 137,000+/-5,000 from light scattering, 140,000 (main) and 137,000 (minor) from non-denatured PAGE and 153,000 from SDS-PAGE in the absence or presence of 2-mercaptoethanol (2-ME). These findings suggest that the enzyme is monomeric in form in human seminal plasma. The enzyme hydrolyzed several amino acid 4-methyl-coumaryl-7-amide (MCA) substrates. The order of Kcat/Km values of AAP at optimal pH (pH 7.5) was Ala- > Lys-Ala- > or = Met- > Leu- > Phe- > Arg- > or = Arg-Arg- > Lys- > Gly-MCAs. AAP was potently inhibited by bestatin, leuhistin, actinonin, amastatin, and 1,10-phenanthroline. These findings suggest that AAP is an aminopeptidase. We determined that the amino acid sequence of the first 22 residues of the enzyme was Ser1-Thr-Thr-Pro-Ser5-Ala-Ser-Ala-Thr-Thr10-Asn-Pro-Al a-Ser-Ala15-Thr-Thr-Leu-Asp-Gln20-Ser-Lys-. This sequence was completely coincident with that downstream of the transmembrane site of human intestinal alanyl aminopeptidase N (CD13). We also isolated cDNA encoding AAP from human prostate cDNA library, sequenced its structure, and confirmed human seminal plasma AAP to be identical with alanyl aminopeptidase N. We postulated that native human seminal plasma alanyl aminopeptidase is released into the seminal plasma after the specific site is cleaved by elastase or an elastase-like enzyme. The enzyme level in human seminal plasma determined by single radial immunodiffusion was 5.2+/-3.2 mg/100 ml (mean+/-SD, n=20) in individuals 20-47 years of age. AAP was immunohistochemically stained in the luminal site-cell membrane of epithelial cells in the prostatic gland and ductuli efferentes of the testis.

Adult

Intestinal anaerobic bacteria hydrolyse sorivudine, producing the high blood concentration of 5-(E)-(2-bromovinyl)uracil that increases the level and toxicity of 5-fluorouracil.

Sorivudine, 1-beta-D-arabinofuranosyl-5-(E)-(2-bromovinyl)uracil, is a potent antiviral agent against varicella-zoster virus and herpes simplex virus type 1. However, sorivudine should not be used in combination with anticancer drugs such as 5-fluorouracil (5-FU) because (E)-5-(2-bromovinyl)uracil (BVU), a metabolite of sorivudine, inhibits the degradation of 5-FU, resulting in its accumulation in the blood and marked enhancement of the toxicity of 5-FU. Since phosphorolytic enzymes generate BVU from sorivudine, we investigated the distribution of the enzyme activity in rats. High activity was found in the cecal and large intestinal contents, while very low or no detectable activity in the liver, kidney, stomach, cecum, large intestine, and the stomach and small intestinal contents. These results suggest that intestinal microflora play an important role in BVU production. Therefore, we measured the phosphorylase activity in cell-free extracts from 23 aerobes, 16 anaerobes and a fungus. Bacteroides species B. vulgatus, B. thetaiotaomicron, B. fragilis, B. uniformis and B. eggerthii, dominant members of intestinal microflora, had high activity to convert sorivudine to BVU. To elucidate the contribution of intestinal microflora to BVU production in vivo, we administered sorivudine to rats treated with several antibiotics and measured the BVU concentration in the serum of rats. When sorivudine was given to rats treated with ampicillin or a mixture of bacitracin, neomycin and streptomycin, which decreased the numbers of viable aerobes and anaerobes, only a small amount of BVU was found in the serum. BVU concentration in the serum of rats treated with metronidazole to decrease the number of intestinal anaerobes was also very low. In contrast, BVU concentration in the serum of rats treated with kanamycin, which was used to decrease the number of aerobes selectively, was higher than that of non-treated rats. These results also suggest that BVU is produced by intestinal anaerobic bacteria especially Bacteroides species in vivo.

Animals

Glioblastoma multiforme after radiotherapy for metastatic brain tumor of testicular cancer.

A patient with left testicular cancer and metastases to retroperitoneal lymph nodes, lung, and brain was treated by chemotherapy, radiotherapy and surgery, and obtained the state of no evidence of disease, but 10 years after radiotherapy, a glioblastoma multiforme tumor appeared in the brain. This is the first report of a glioma appearing after radiotherapy in a testicular cancer patient.

Adult

Mechanisms of CCK regulation of monitor peptide mRNA expression in pancreatic acinar AR42J cells.

We explored the mechanism(s) by which cholecystokinin (CCK) stimulation of AR42J rat pancreatoma cells results in increased mRNA expression of a CCK-releasing peptide [monitor peptide (MP)]. With the use of a newly established reverse transcription-polymerase chain reaction assay system, CCK was shown to increase the level of MP mRNA by about ninefold. When protein synthesis was blocked by addition of cycloheximide, the MP mRNA level remained unchanged in the presence of CCK. Inhibition of transcription with actinomycin D resulted in a half-life for MP mRNA of approximately 17 h, and this rate remained unchanged after CCK treatment, suggesting that CCK may regulate the MP mRNA level by influencing gene transcription. A-23187, bombesin, substance P, and carbachol increased the MP mRNA level. CoCl(2) abolished actions of both CCK and A-23187 on MP mRNA expression. Dibutyryl-adenosine 3',5'-cyclic monophosphate, forskolin, secretin, and vasoactive intestinal polypeptide had no effect on MP mRNA expression. 12-O-tetradecanoylphorbol-13-acetate and phorbol 12,13-dibutyrate also failed to increase MP mRNA. It was therefore proposed that CCK stimulates MP mRNA expression of AR42J cells in a Ca2+-dependent and protein kinase C-independent manner.

Animals

[Urethral recurrence of bladder cancer 16 years after total cystectomy. A case report].

A 74-year-old man was admitted to our hospital. Diagnosis of invasive bladder cancer was made and a total cystectomy was performed on January, 30, 1980. Histological examination showed transitional cell carcinoma, grade 2, pT1 is N0M0. After 16 years, when he suffered left femur, he had right inguinal lymphnode swelling. On the biopsy of the inguinal mass, histology was transitional cell carcinoma. The urethral washing cytology was positive. Lung CT scan dem onstrated multiple lesions. He was diagnosed of the urethral recurrence, right inguinal lymphnode metastasis and lung metastasis of bladder cancer. This case is the 5th case of urethral recurrence of bladder cancer following cystectomy in our hospital.

Aged

Coexistence of renal cell carcinoma and renal angiomyolipoma developing in a kidney: a case report.

Coexistence of renal cell carcinoma and angiomyolipoma in the same kidney is rare. A 54-year-old woman without tuberous sclerosis was admitted for further examination of incidental renal masses on ultrasonography. Computerized tomography revealed a 17-mm high density mass in the mediolateral portion of the right kidney and a 5-mm low density mass near the right lower pole. Because the former mass showed a typical tumor pattern on selective renal angiography and the latter mass was strongly hyperechoic on ultrasonography, a clinical diagnosis of renal cell carcinoma and angiomyolipoma was made. A right radical nephrectomy confirmed the preoperative diagnosis. She has been followed for 61 months with no recurrence.

Angiomyolipoma

Treatment of advanced renal cell carcinoma with a combination of human lymphoblastoid interferon-alpha and cimetidine.

PURPOSE: We examined whether cimetidine could augment the efficacy of interferon-alpha therapy for advanced renal cell carcinoma. MATERIALS AND METHODS: Of 31 male and 6 female patients treated with interferon-alpha and cimetidine 21 had metastases at diagnosis and 15 had a recurrence after nephrectomy. One patient had a primary tumor with thrombus in the inferior vena cava. Lymphoblastoid interferon-alpha was administered at 5 megaunits daily intramuscularly 5 to 7 days a week for at least 8 weeks, and cimetidine was administered orally at 800 mg. daily in 4 divided doses. The evaluable tumors included 30 in the lung, 6 lymph nodes, 5 bone, 4 kidney and 1 inferior vena cava. RESULTS: Combined therapy with interferon-alpha and cimetidine resulted in a complete response in 7 patients, a partial response in 8, stable disease in 12 and progression in 10. The objective response rate was 41%. The lung metastasis showed the best response to combined therapy. The 5-year survival rates for patients with and without response, and overall were 74, 20 and 41%, respectively. Histopathologically, high grade tumors had a better response to combined therapy than did low grade tumors. CONCLUSIONS: Combined therapy with interferon-alpha and cimetidine for advanced renal cell carcinoma resulted in a definitively good response. A prospective randomized trial should be performed to elucidate the efficacy of the combined therapy.

Adult

[Clinical outcome of radical prostatectomy for stage C prostate cancer: comparison with other treatment modalities].

Of the 69 patients with clinical stage C prostate cancer under 75 years old and with good performance status between 1986 and 1995, 29 underwent radical prostatectomy combined with endocrine therapy, 21 underwent radiation therapy combined with endocrine therapy and remaining 19 patients were treated by endocrine therapy alone. The median followup was 44 months (range 4 to 122). Radical prostatectomy resulted in progression-free rates of 79% and 61% at 5 and 10 years, respectively. Progression-free rates were lower in patients with lymph node metastasis or positive surgical margins. In patients with clinical stage T3a-c and well or moderately differentiated tumor, radical prostatectomy resulted in a progression-free rate of 100% at 5 years. However, in patients with clinical stage T4a or poorly differentiated tumor, radiation therapy resulted in a better progression-free rate than radical prostatectomy. These findings suggest that patients with clinical stage T3a-c and well or moderately differentiated tumor will benefit from radical prostatectomy combined with endocrine therapy and that radiation therapy well be effective for advanced diseases.

Aged

[A case of scrotal cancer with inguinal lymph node metastasis treated by multidisciplinary modalities including chemotherapy with methotrexate, bleomycin and cisplatin].

We report a case of scrotum with inguinal lymph node metastasis which was successfully treated by multidisciplinary modalities including combination chemotherapy and radiotherapy. A 60-year-old man was admitted with ulcerative induration of the scrotum and inguinal lymph node swelling. Biopsy of the scrotal skin showed squamous cell carcinoma. He received 4 courses of combination chemotherapy with methotrexate, bleomycin and cisplatin. The primary lesion disappeared macroscopically and metastatic lymphadenopathy showed 50% reduction in size. Both lesions were further treated with radiotherapy (60 Gy). Because the primary lesion became ulcerative 7 months after irradiation, partial resection of the scrotum was performed. He has been free of recurrence 22 months after chemotherapy.

Antineoplastic Combined Chemotherapy Protocols

Mutagenicity of 1-methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid treated with nitrite in the presence of alcohols.

The mutagenicity of a product produced from 1-methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid (MTCCA), which is a component in soy sauce, after treatment with 50 mM nitrite at pH 3, 37 degrees C, for 60 min in the presence of 7.5% ethanol was much higher than that in the absence of ethanol during the nitrite treatment. The enhancement of the mutagenicity of nitrite-treated MTCCA by ethanol required simultaneous treatment of MTCCA with nitrite and ethanol. The mutagenicity of MTCCA treated with nitrite in the presence and absence of ethanol was detected in the same fractions on HPLC and was highest for Salmonella typhimurium strain YG1029 possessing elevated O-acetyltransferase activity among the several Salmonella test strains, suggesting that the same mutagen belonging to aromatic compounds was produced both in the presence and absence of ethanol. Methanol, n-propanol and isopropanol as well as ethanol were also observed to have an augmenting effect. However, the sugars glucose and sucrose had no effect. When MTCCA was treated with nitrite in the presence of commercial alcoholic beverages equivalent to 1.25-10% ethanol, Japanese 'sake' and 'shochu' were demonstrated to have a highly augmenting effect and beer, wine, whisky and brandy to have a mildly augmenting effect.

Alcoholic Beverages

In vivo formation of mutagens by intraperitoneal administration of polycyclic aromatic hydrocarbons in animals during exposure to nitrogen dioxide.

Consumption of fossil fuels has increased indoor and outdoor concentrations of polycyclic aromatic hydrocarbons (PAHs) and nitrogen dioxide (NO2). To study the combined effect of PAH administration and NO2 exposure on mutagenicity of urine from animals we injected 400 mg/kg body wt i.p. one of five kinds of PAH (pyrene, fluoranthene, fluorene, anthracene and chrysene) into ICR mice, Wistar rats, Syrian golden hamsters or Hartley guinea pigs after exposure to 20 p.p.m. NO2 gas for 24 h and then exposed the animals to NO2 gas for an additional 24 h. During the latter 24 h we collected the urine and assayed its mutagenicity with the Ames Salmonella strains after treatment with beta-glucuronidase and arylsulfatase and extraction with dichloromethane. The urine from mice treated with both PAH and NO2 showed high mutagenicity for Salmonella typhimurium strains TA98 and TA100, whereas the urine from mice treated with PAH and air showed almost no mutagenic activity. The mutagenicity was decreased in nitroreductase- and acetyltransferase-deficient strains TA98NR and TA98/1,8-DNP6 respectively. Treatment with a mixture of 20% of each of the five kinds of PAH and NO2 augmented the urinary mutagenicity of mice 1.5-fold. The urine from hamsters treated with pyrene or fluoranthene and NO2 was also highly mutagenic, but that from rats or guinea pigs was not very mutagenic. The mutagenicity was also decreased in strains TA98NR and TA98/1,8-DNP6. These results suggest that the urine contains nitro compounds and that the nitration of PAHs occurs in the body of animals under exposure to NO2 gas. Actually, the nitrated metabolites of pyrene, 1-nitro-6/8-hydroxypyrene and 1-nitro-3-hydroxypyrene, were detected in the urine from mice treated with pyrene under exposure to NO2 gas. To elucidate the mechanism of in vivo nitration, NO2 (20 p.p.m.) was bubbled through 50 mM Tris-HCl buffer (pH 7.4) or dichloromethane solution containing pyrene or 1-hydroxypyrene (10 microg/ml). Pyrene was not nitrated by NO2 in either aqueous or organic solutions. However, 1-hydroxypyrene was changed to nitrohydroxypyrenes by NO2 in the Tris-HCl buffer, but not in the organic solution. Ascorbic acid, alpha-tocopherol, glutathione oleic acid and hemoglobin were found to inhibit the nitration of 1-hydroxypyrene in aqueous solution. The urinary mutagenicity of mice treated with both pyrene and NO2 was also decreased by oral administration of ascorbic acid and alpha-tocopherol. These results suggest that 1-hydroxypyrene is nitrated by an ionic reaction in the animal body after hydroxylation of pyrene in the liver.

Animals

Role of intestinal bacteria in ileal ulcer formation in rats treated with a nonsteroidal antiinflammatory drug.

The role of intestinal bacteria in induction and repression of ulcer formation in the ileum of rats treated with one of the nonsteroidal antiinflammatory drugs (NSAIDs), 5-bromo-2-(4-fluorophenyl)-3-(4-methylsulfonylphenyl) thiophene (BFMeT), was examined in this study. BFMeT was administered by intragastric gavage once at doses of 500-1,500 mg/kg of body weight to Wistar rats treated with and without antibiotics (bacitracin, neomycin, streptomycin), germ-free rats and gnotobiotic rats, and 72 hr later their gastrointestinal tracts were examined for ulcer formation. A single oral administration of BFMeT induced ileal ulcers in specific pathogen-free rats. However, the rats given antibiotics to reduce the intestinal bacteria had no ulcers. BFMeT-treated germ-free rats and gnotobiotic rats mono-associated with Bifidobacterium adolescentis or Lactobacillus acidophilus also had no intestinal ulcers. However, the drug induced ileal ulcers in gnotobiotic rats mono-associated with Eubacterium limosum or Escherichia coli. An overnight culture of B. adolescentis or L. acidophilus or yogurt containing Bifidobacterium breve and Streptococcus thermophilus, when given as drinking water, inhibited ulcer formation in the ileum of rats treated with BFMeT. Gram staining of the ileal contents of normal rats revealed that 97.4% of the stained microorganisms were Gram-positive rods and only 1.2% were Gram-negative rods. In the group of rats with ulcers induced by BFMeT, the Gram-positive rods decreased by 56.4% and the Gram-negative rods including Escherichia coli, Klebsiella, Proteus and Bacteroides increased by 37.3%. However, in the group of rats administered the Bifidobacterium culture, the Lactobacillus culture or yogurt, the percentages of the Gram-negative rods were decreased. Although Lactobacillus was a major bacterium in the ileum of normal rats, the Gram-negative facultatively anaerobic rods E.coli, Klebsiella and Proteus were increased in the ulcerated ileum of rats treated with BFMeT, suggesting that these bacteria are associated with ulcer formation in rats treated with NSAIDs, and that Lactobacillus and Bifidobacterium inhibit it by repressing the growth of ulcer-inducing bacteria.

Animals

Primary adenocarcinoma in an ileal conduit.

Reports of primary small intestine malignancies are rare. Even more uncommon is primary carcinoma in an ileal conduit. Here, we report a case of primary adenocarcinoma in an ileal conduit that developed 14 years after radical cystectomy and diversion to an ileal conduit for transitional cell carcinoma of the bladder. To our knowledge, only one case of primary adenocarcinoma developing in an ileal conduit after a radical surgery for bladder cancer has been reported previously.

Adenocarcinoma