Biomedical subjects
Publications and source records attributed to K Yamaguchi.
The in vivo activity of olamufloxacin (HSR-903), a new fluoroquinolone, was evaluated and compared with ciprofloxacin, sparfloxacin and levofloxacin. Olamufloxacin was active against systemic infection in mice inoculated with both Gram-positive and -negative bacteria. Olamufloxacin had equal efficacy for experimental urinary tract infections in mice caused by Pseudomonas aeruginosa.
Azithromycin is an azalide with potent activity against Haemophilus influenzae including ampicillin-resistant strains. We evaluated the efficacy of azithromycin, clarithromycin and three beta -lactams when used for 1 day only and for 3 days for the treatment of a murine model of bronchopneumonia, using three strains of H. influenzae, two of which were ampicillin resistant. MICs of azithromycin (1-2 mg/L) and clarithromycin (4-8 mg/L) were similar for the three strains. The MICs of cefdinir and cefcapene for beta-lactamase-negative ampicillin-resistant (BLNAR) H. influenzae were 32 times higher than those for beta-lactamase-positive ampicillin-resistant and ampicillin-susceptible strains. The viable counts in the infected tissues of azithromycin-treated mice with bronchopneumonia caused by the susceptible strain TUM8, beta-lactamase-positive strain TUH36 and BLNAR strain TUH267 were less than the counts obtained with the other antibiotics used, irrespective of MIC. At a dose of 50 mg/kg, the area under the concentration curve and the half-life of azithromycin in the lungs were respectively three times higher and six times longer than those of clarithromycin. Our results indicate that azithromycin may be useful for both ampicillin-susceptible and ampicillin-resistant bronchopneumonial infections caused by H. influenzae.
The in vitro activity of ampicillin, cefotaxime, meropenem, panipenem, imipenem and biapenem was assayed using ampicillin-susceptible, beta-lactamase-positive and beta-lactamase-negative ampicillin-resistant (BLNAR) Haemophilus influenzae isolated recently in Japan. Against ampicillin-susceptible isolates, cefotaxime was the most potent (MIC(90) 0.016 mg/mL). Both cefotaxime and meropenem (MIC(90) of both, 0.5 mg/L) were the most potent against beta-lactamase-positive isolates. Against BLNAR isolates, meropenem (MIC(90) 0.5 mg/L) was the most potent. In murine bronchopneumonia caused by ampicillin-susceptible and BLNAR H. influenzae, cefotaxime showed the best efficacy, followed by meropenem. Our results indicate that meropenem could be a useful intravenous agent for infections caused by H. influenzae, including BLNAR strains.
Multiple endocrine neoplasia type 2 (MEN-2) is a hereditary syndrome characterized by medullary thyroid carcinoma (MTC), pheochromocytoma and hyperplasia or adenoma of the parathyroid gland with hyperparathyroidism. Recent genetic studies have identified the presence of germline missense mutations in the RET proto-oncogene in almost 100% of MEN-2 patients. We report here three generations of one MEN-2 family with rare missense mutation at codon 618 (Cys-->Arg) of the RET proto-oncogene. The first patient was surgically treated at the age of 63 years but died of bone metastasis. His two children (29-year-old daughter and 25-year-old son) were treated surgically for MTC and neck lymph node metastasis. Germline mutations of the RET proto-oncogene of these three MTC patients and two children of the 29-year-old daughter (9-year-old female and 7-year-old male) were examined. Three MTC patients and the 9-year-old female possessed the mutation. The phenotype of the family with this rare point mutation of the RET proto-oncogene is reported.
Plasmid pSC101 encodes a 37.5 kDa Rep (RepA) protein, which binds to three 21-base repeats (DR-1, DR-2, and DR-3) in the replication origin region (ori) of the plasmid to initiate replication. Rep also binds to two palindromic sequences (IR-1 and IR-2) which overlap the rep promoter. The binding of Rep to IR-2 represses the production of Rep itself. It is highly likely that the balance of these functions of Rep plays a major role in controlling the copy number of pSC101. In this study, we developed a positive-selection system for replication-deficient mutants of the initiator protein. This system can be applied to the study of other replication systems by changing ori and rep of pSC101 to the corresponding genes. Thirty-four replication-deficient (Ini(-)) mutants were isolated with this system, and analyzed as to the relation between the structure and function of the Rep protein. Seventeen of these 34 Ini(-) mutants were found to lack auto-repressor activity as well as initiator activity. DNA sequence analysis showed that one-third (from the C-terminus) of Rep is dispensable for the auto-repressor activity, while the initiator activity seems to require the whole protein.
T(1), a mutant yeast lacking three regulatory proteins of F(1)F(o)ATPase, namely ATPase inhibitor, 9K protein and 15K protein, grew on non-fermentable carbon source at the same rate as normal cells but was less viable when incubated in water. During the incubation, the cellular ATP content decreased rapidly in the T(1) cells but not in normal cells, and respiration-deficient cells appeared among the T(1) cells. The same mutation was also induced in D26 cells lacking only the ATPase inhibitor. Overexpression of the ATPase inhibitor in YC63 cells, which were derived from the D26 strain harboring an expression vector containing the gene of the ATPase inhibitor, prevented the decrease of cellular ATP level and the mutation. Isolated T(1) mitochondria exhibited ATP hydrolysis for maintenance of membrane potential when antimycin A was added to the mitochondrial suspension, while normal and YC63 mitochondria continued to show low hydrolytic activity and low membrane potential. Thus, it is likely that deletion of the ATPase inhibitor induces ATPase activity of F(1)F(o)ATPase to create a dispensable membrane potential under the non-nutritional conditions and that this depletes mitochondrial and cellular ATP. The depletion of mitochondrial ATP in turn leads to occurrence of aberrant DNA in mitochondria.
To investigate the roles of peroxisomal membrane proteins in the reversible conversion of glyoxysomes to leaf peroxisomes, we characterized several membrane proteins of glyoxysomes. One of them was identified as an ascorbate peroxidase (pAPX) that is localized on glyoxysomal membranes. Its cDNA was isolated by immunoscreening. The deduced amino acid sequence encoded by the cDNA insert does not have a peroxisomal targeting signal (PTS), suggesting that pAPX is imported by one or more PTS-independent pathways. Subcellular fractionation of 3- and 5-d-old cotyledons of pumpkin revealed that pAPX was localized not only in the glyoxysomal fraction, but also in the ER fraction. A magnesium shift experiment showed that the density of pAPX in the ER fraction did not increase in the presence of Mg(2+), indicating that pAPX is not localized in the rough ER. Immunocytochemical analysis using a transgenic Arabidopsis which expressed pumpkin pAPX showed that pAPX was localized on peroxisomal membranes, and also on a unknown membranous structure in green cotyledons. The overall results suggested that pAPX is transported to glyoxysomal membranes via this unknown membranous structure.
UNLABELLED: Anaphylactic shock therapy includes the use of catecholamines but they may not always be effective. Because vasodilation during anaphylaxis is a result of the endothelial release of multiple mediators, we investigated the effects of epinephrine, vasopressin, and inhibitors of nitric oxide and prostanoid pathways on histamine-induced relaxation in human internal mammary artery. The vessel segments were obtained intraoperatively and were suspended in organ chambers to record isometric tension. Norepinephrine (10(-6) M) was used to precontract the rings followed by histamine (10(-6.5) M) to relax the vessels and mimic vascular collapse. Epinephrine, vasopressin, methylene blue, N(G)-monomethyl-L-arginine (L-NMA) and indomethacin were added in a cumulative fashion to reverse the histamine-induced vasodilation. The internal mammary artery segments exhibited greater contraction in the presence of the epinephrine (4.9 +/- 0.7 g) compared with vasopressin (2.6 +/- 0.7 g). Vasopressin (10(-11) to 10(-7) M), methylene blue (10(-7) to 10(-5) M), L-NMA (10(-6) to 10(-4) M), and indomethacin (10(-7) to 10(-5) M) were only partially effective. These findings suggest that vasopressin and methylene blue may offer a potential therapeutic option in the treatment of histamine-induced vasodilatory shock. IMPLICATIONS: Epinephrine only partially reverses histamine-induced vasodilation in human internal mammary arteries, whereas vasopressin, methylene blue, and drugs involved in the inhibition of nitric oxide and prostaglandin generation lead to a complete reversal of the vascular relaxation.
OBJECTIVE: A noninvasive bronchoscopic microsampling (BMS) probe was developed to sample biochemical constituents of the epithelial lining fluid in small airways. DESIGN: Observational, controlled study. SETTING: Intensive care unit of academic medical center. PATIENTS AND PROCEDURE: BMS was applied in a control group of seven patients who had hemoptysis or small solitary peripheral nodules but no hypoxemia or other signs of acute inflammation and in four patients with acute respiratory distress syndrome (ARDS), to test whether BMS can ascertain the presence of acute pulmonary inflammation without complications. MEASUREMENTS AND RESULTS: Complications, including a significant decrease in arterial oxygen saturation, were observed neither during nor after BMS. In the ARDS group, albumin, lactate dehydrogenase, interleukin-6, basic fibroblast growth factor, and neutrophil elastase concentrations in epithelial lining fluid were significantly higher (p <.0001, p =.012, p <.0001, p <.0001, and p <.0001, respectively) than in the control group. Serial BMS was safely performed in one patient with ARDS, allowing us to observe a correlation between changes in the concentration of inflammation-related biochemical markers and clinical course of the disease. CONCLUSIONS: These results suggest that BMS is safe and useful to monitor pulmonary biochemical events in ARDS.
We established a new cell line, HPC-3P4a, with high peritoneal disseminated potential in nude mice. HPC-3P4a was derived from a human pancreatic carcinoma cell line (HPC-3) that had low capacity for peritoneal dissemination. HPC-3P4a developed peritoneal dissemination in 10 of 11 (90.9%) cases, whereas parental HPC-3 developed peritoneal dissemination in one of six (16.7%) cases. The metastatic foci in the peritoneum showed essentially the same histologic appearance of parental involvement. The tumorigenicity, motility, and adhesive activity of HPC-3P4a to the extracellular matrix were stronger than were those of the HPC-3. In FACS analysis, HPC-3P4a significantly increased the expression of alpha6 and alpha(v)beta5 integrins, while it decreased alpha2 integrin, hCD44H, and hCD44v 10, as compared with HPC-3. The VEGF production of HPC-3P4a was significantly lower than that of HPC-3. Analysis of gene macroarrays showed a variety of cytokines, interleukin, and other immunomodulatory, and their receptors were up-regulated and down-regulated on an mRNA level in HPC-3P4a cells, compared with HPC-3 cells. Intrasplenic injection of HPC-3P4a produced no liver metastasis. We named our original highly liver metastatic cell line HPC-3H4 (previously reported). This HPC-3H4 cell was established by repeated intrasplenic injection from parental cell HPC-3; thus, it developed high liver metastasis. Moreover, HPC-3H4 developed peritoneal dissemination by intra-abdominal injection. In contrast, HPC-3P4a did not develop liver metastasis by intrasplenic injection. These findings are very interesting and might suggest that the process of hematogenous metastasis differed from that of peritoneal dissemination. Thus, this cell line may be useful for investigating the mechanism of peritoneal dissemination in human pancreatic cancer.
A number of studies on mucin-producing cystic neoplasm of the pancreas have been reported since the first report of the tumor in 1982. There has been some controversy about nomenclatures and clinicopathologic entities of mucin-producing cystic tumor, mucinous cystic tumor, and intraductal papillary tumor of the pancreas. In 1996 and 1997, new classifications of pancreatic neoplasms were published by the World Health Organization (WHO) and Armed Forces Institute of Pathology (AFIP). According to the new WHO and AFIP classifications, mucin-producing cystic neoplasm of the pancreas corresponds mainly to intraductal papillary-mucinous tumor and mucinous cystic tumor of the pancreas. and these two diseases are independent conditions. Intraductal papillary-mucinous tumor is regarded as a unique clinical entity, but controversy remains about the term and clinicopathologic entity. Some confusion and problems remain betweeen the two lesions. In this review, we review their historical background, terminology, WHO and AFIP classification, and problems with classification.
OBJECTIVES: Pancreaticobiliary maljunction, an anomalous union of the pancreatic duct with the common bile duct, is a risk factor for biliary carcinoma. We hypothesized that, in patients with pancreaticobiliary maljunction, persistent regurgitation of pancreatic juice into the biliary tract induces oxidative DNA damage. We assessed the expression of an oxidative DNA base-modified product, 8-hydroxy-2'-deoxyguanosine, in gallbladder epithelium. DESIGN: Eleven noncancerous gallbladders from patients with pancreaticobiliary maljunction, 12 gallbladder carcinomas from patients without pancreaticobiliary maljunction and 14 noncancerous gallbladders from patients without pancreaticobiliary maljunction (control) were studied. METHODS: Immunohistochemistry was performed for 4-hydroxy-2-nonenal-modified protein (as a marker for lipid peroxidation), 8-hydroxy-2'-deoxyguanosine and p53 gene product. RESULTS: Stronger cytoplasmic staining of 4-hydroxy-2-nonenal-modified protein was observed in the gallbladder epithelium from patients with pancreaticobiliary maljunction than in epithelium from gallbladder cancer patients or from control subjects with normal gallbladders. Clear, strong nuclear staining of 8-hydroxy-2'-deoxyguanosine was observed in the gallbladder epithelial cells from patients with pancreaticobiliary maljunction. Densitometric quantitation revealed significantly higher expression of 8-hydroxy-2'-deoxyguanosine in gallbladder epithelial cells from patients with pancreaticobiliary maljunction (index 27.3 +/- 3.1) than in cells from patients with gallbladder carcinoma (11.4 +/- 1.5; P < 0.05) or from control subjects with normal gallbladder (6.4 +/- 1.0; P < 0.05). Positivity of p53 was 27% in gallbladder epithelium associated with pancreaticobiliary maljunction, 75% in gallbladder carcinoma epithelium and 0% in control epithelium. CONCLUSIONS: These results suggest that reactive oxygen species are produced in the gallbladder of patients with pancreaticobiliary maljunction and that oxidative DNA injury is related to carcinogenesis in these patients.
Induction of IL-2 production and increased expression of CD25 were observed in C57BL/10 mice after weekly treatment with gold sodium thiomalate (GST). LP-BM5 murine leukemia virus (MuLV) infected mice treated with GST survived longer, had less cervical lymph node swelling, lower spleen weight, and fewer abnormalities in the expression of the cell surface markers, CD4, CD8a and CD45R/B220 on spleen cells than those that were not treated with GST. Thus, GST treatment may be beneficial through a decrease in disease progression via IL-2 induction in MuLV infected mice. This may have application in human immunodeficiency virus-infected individuals.
5-Fluorouracil (5FU) is most commonly used in chemotherapy for human malignancy. Over 80% of administered 5FU is metabolically degraded by dihydropyrimidine dehydrogenase (DPD), a primary and rate-limiting enzyme in the 5FU metabolic pathway. A DPD-deficient phenotype among cancer patients, which has posed a serious problem in 5FU-based chemotherapy, was reported to be in part ascribed to germline mutations in dihydropyrimidine dehydrogenase (DPYD) gene. Therefore, we for the first time examined the frequencies and types of germline mutations in the DPYD gene among a total of 107 Japanese cancer patients and healthy volunteers. Of 214 alleles examined among them, 181 alleles were of the same type, which was assigned as wild type; 21 alleles revealed a nucleotide substitution resulting in silent mutation; and the remaining 12 alleles showed five types of nucleotide deletion or substitutions resulting in one frameshift and four missense mutations. Three of them, A74G, 812delT and L572V, were novel mutations. None of the study subjects showed homozygous frameshift or missense mutated alleles. We also studied the association between toxic response to 5FU and heterozygous frame shift or missense mutation of the DPYD gene among eight cancer patients who had received 5FU-based chemotherapy. These patients did not show any adverse effects higher than grade 3, suggesting that heterozygotes are not associated with increased toxicity to 5FU. Our results indicate that a very small percentage, about 0.2%, of the Japanese population seems to carry homozygous mutations in DPYD gene, mutations which possibly indicate genetically increased toxicity of 5FU-based chemotherapy.
We determined antimicrobial susceptibilities of 23 strains of Legionella spp. clinically isolated in Japan, between September 1994 through November 1999 by using Etest. The MICs of all isolates against macrorides, fluoroquinolones and rifampicin showed 1 microgram/ml or less. Especially rifampicin showed the most potent activity, of which the MICs of all isolates showed 0.064 microgram/ml or less. Although we could not find resistant strains against any antibiotics tested in this study, we think it is important to follow up the susceptibilities of clinical isolates of Legionella.
Recently, Vancomycin-resistant enterococci (VRE) have become important nosocomial pathogens in the world. In Japan, the VRE-infection was first reported in 1996. However, an epidemiological study on VRE has not been aggressively done in Japan. We conducted a survey study to explore the incidence and antimicrobial susceptibility of vancomycin-resistant enterococci isolated from fecal samples at 45 hospitals in the east area of Japan (Kanto, Koshinetsu, Tohoku, and Hokkaido) during June 1998 to March 1999. The Enterococcosel agar containing vanocomycin (BBL) was used for screening VRE from fecal samples in each hospital. The susceptibilities of the isolates to 8 antimicrobials were determined by the broth microdilution method and the definitions of resistance were based on current standards of the NCCLS standards. The VRE genotypes (vanA, vanB, vanC1, and van C2/3) were confirmed by amplifying the respective genes by PCR. Eight hundred and ninety four strains of enterococci were tested by the microtiter plates hybridization method (WAKUNAGA SEIYAKU, Japan). One thousand five hundred eighty three strains of enterococci were collected from 6,914 patients in 45 hospitals. These strains included 72 (4.5%) strains Enterococcus faecalis, 33 (2.1%) strains Enterococcus faecium, 17 (1.1%) strains Enterococcus avium, 1,040 (65.7%) strains Enterococcus gallinarum, 386 (24.4%) strains Enterococcus cassliflavus, and 35 (2.2%) strains Enterococcus flavescens. These strains of vancomycin-resistant E. faecalis were isolated from 3 patients, two of these 3 strains had van A gene and other one had van B gene. Those 3 strains were in the Kanto area, and 2 of 3 strains were in Tokyo, Generally, though van A type VRE was highly resistaant to both vancomycin and teicoplanin. In our study, two strains of van A type E. faecalis were highly resistant to vancomycin (MICs > 128 micrograms/ml) and susceptible to teicoplanin with MICs 4 micrograms/ml. Those two strains were different in susceptibilities of minocycline and ofloxacin. The result of the analysis of PFGE had also different patterns. VanB type E. fecalis was highly resistant to vancomycin and susceptible to teicoplanin (MICs 0.25 microgram/ml). For ampicillin and imipenem, 3 strains of E. faecalis were susceptible (MIC < or = 1 microgram/ml). One of 562 strains of E. gallinarum had vanB and vanC1 genes and was moderately resistant to vancomycin and susceptible to teicoplanin. All strains of E. casseliflavus and E. flavescens had vanC2/C3 gene only. All strains of E. faecium and E. avium did not detect van genes. From this result, it was supposed that VRE were very rare in the east of Japan.
We investigated the drug susceptibilities of Neisseria gonorrhoeae by using 98 strains of clinical isolates at Toho University Omori Hospital from 1994 to 1998. Minimum Inhibitory Concentrations (MICs) of 15 antimicrobial agents were determined with agar dilution methods according to the guidelines of NCCLS. Among these isolates, only 4 strains (4.1%) were found to be penicillinase producing N. gonorrhoeae. Ceftriaxone showed the most potent activity of which MICs of all strains were 0.06 microgram/ml or less. Macroride antimicrobial agents and minocycline also showed strong activities of which MICs of most of the strains were 0.06 microgram/ml or less. With the criteria of NCCLS, 10 strains (10.2%) were found to be resistant to ciprofloxacin and these 10 strains also showed cross resistance to other fluoroquiolones we tested. Our results also revealed that the number of resistant strains against fluoroquiolones abruptly increased from 1996 and indicate the needs of further surveillance of antimicrobial resistance in clinical isolates of N. gonorrhoeae.
L-084 (a prodrug of LJC 11,036 [L-036]) is a new oral carbapenem. Here we compared the in vitro and in vivo antibacterial activities of L-036 with those of imipenem, faropenem, ceditoren-pivoxil, cefdinir, amoxicillin, and levofloxacin. The MICs at which 90% of the isolates were inhibited of L-036 against methicillin-susceptible staphylococci, Streptococcus pneumoniae including penicillin-resistant organisms, Escherichia coli, Klebsiella pneumoniae, Haemophilus influenzae including ampicillin-resistant organisms, Legionella pneumophila, and Moraxella catarrhalis were equal to or less than 1 microg/ml. In pharmacokinetics studies of L-084 in lungs of mice, the maximum concentration in serum, half-life, and area under the concentration-time curve of this drug were 9.09 microg/g of tissue, 6.18 h, and 31.0 microg. h/ml, respectively. In murine respiratory infection models of penicillin-susceptible and -resistant S. pneumoniae and H. influenzae, the efficacies of L-084 were better than those of reference drugs. Our results indicate that the in vitro high potency and good distribution in the lungs might be the underlying mechanisms of its efficacy in the murine model of pneumonia.