Biomedical subjects
Publications and source records attributed to T Sawai.
On the basis of two case reports it is suggested that serious convulsions may occur in patients treated with flucoazole when serum trough concentrations exceed 80 microg/mL. The authors recommend monitoring fluconazole concentrations during high-dose therapy in patients with poor kidney function.
Pacemaker infection is one of the severe complication of pacemaker implantation. We report a case of pacemaker infection caused by Staphylococcus schleiferi which is a coagulase-negative staphylococcus, and its relation with human infection is not well characterized. In 1994, a 80-year-old male presented with a pacemaker pocket infection, cutaneous inflammation but no fever 2 months after insertion of a pacemaker. S. schleiferi was isolated from the pus. The patient was given cefazolin for 5 days. One month later he was readmitted because of cutaneous inflammation and the extruded generator was removed. S. schleiferi was isolated from the generator. After the patient was treated with cefazolin for 3 weeks, four consecutive wound cultures were all negative. A new generator was inserted on the same side. One month after re-insertion, the patient again presented a cutaneous inflammation, and S. schleiferi was isolated from the pus as well as the generator and the leads on their removal. Twenty-six days later, a new pacing system was inserted on the other side. There was no further recurrence of the infection. Removal of the entire pacing system was necessary to cure the infection. We expect further information of human infections caused by S. schleiferi.
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In the course of surveying for the carbapenem-hydrolyzing metallo-beta-lactamase gene bla(IMP) in pathogenic bacteria by the PCR method, we detected a gene encoding a variant metallo-beta-lactamase, designated IMP-3, which differed from IMP-1 by having low hydrolyzing activity for penicillins and carbapenems. PCR product direct sequencing of a 2.2-kb segment revealed that the gene bla(IMP-3) was located on a cassette inserted within a class I integron in the pMS390 plasmid. The 741-bp nucleotide sequence of bla(IMP-3) was identical to that of bla(IMP-1), except for seven base substitutions. Among these were two, at nucleotide positions 314 and 640, which caused amino acid alterations. Hybrid bla genes were constructed from bla(IMP-3) and bla(IMP-1) by recombinant DNA techniques, and beta-lactamases encoded by these genes were compared with those of the parents IMP-3 and IMP-1 under the same experimental conditions. The kinetic parameters indicated that the inefficient hydrolysis of benzylpenicillin, ampicillin, imipenem, and ceftazidime by IMP-3 was due to the substitution of glycine for serine at amino acid residue 196 in the mature enzyme. This alteration corresponded to the presence of guanine instead of an adenine at nucleotide position 640 of the bla(IMP-3) gene. This indicated that extension of the substrate profile in the metallo-beta-lactamase IMP-1 compared to IMP-3 is the result of a one-step single-base mutation, suggesting that the gene bla(IMP-3) is an ancestor of bla(IMP-1).
An R-plasmid-mediated metallo-beta-lactamase was found in Klebsiella pneumoniae DK4 isolated in Japan in 1991. The nucleotide sequence of its structural gene revealed that the beta-lactamase termed DK4 was identical to the IMP-1 metallo-beta-lactamase which was mediated by a chromosomal gene of Serratia marcescens TN9106 isolated in Japan in 1991 (E. Osano et al., Antimicrob. Agents Chemother. 38:71-78, 1994). The dose effect of DK4 beta-lactamase production on the resistance levels indicated a significant contribution of the enzyme to bacterial resistance to all the beta-lactams except monobactams. The enzymatic characteristics of the DK4 beta-lactamase and its kinetic parameters for nine beta-lactams were examined. The DK4 beta-lactamase was confirmed to contain 2 mol of zinc per mol of enzyme protein. The apoenzyme that lacked the two zincs was structurally unstable, and the activities of only 30% of the apoenzyme molecules could be restored by the addition of 1 mM zinc sulfate. The substitution of five conserved histidines (His28, His86, His88, His149, His210) and a cysteine (Cys168) for an alanine indicated that His86, His88, and His149 served as ligands to one of the zincs and that Cys168 played a role as a ligand to the second zinc. Both zinc molecules contribute to the enzymatic process. Mutant enzymes that lack only one of these retained some activity. Additionally, a conserved aspartic acid at position 90 was replaced by asparagine. This mutant enzyme showed an approximately 1,000 times lower k(cat) value for cephalothin than that of the wild-type enzyme but retained the two zincs even after dialysis against zinc-free buffer. The observed effect of pH on the activity suggested that Asp90 functions as a general base in the enzymatic process.
BACKGROUND: Altered expression of blood group-related carbohydrate antigens such as sialyl Lewis (Le)(x) antigen in tumours is associated with tumour progression behaviour and subsequent prognosis. However, the prognostic value of the expression of Le-related antigens in colorectal tumours remains unclear. PURPOSE: To clarify the prognostic value of Le(a), sialyl Le(a), Le(x) and sialyl Le(x) expression in colorectal carcinomas as prognostic factors after surgery. PATIENTS AND METHODS: Colorectal carcinoma samples from 101 patients with primary colorectal carcinoma who underwent surgical resection were subject to immunohistochemical analyses for Lea, sialyl Lea, Lex and sialyl Le(x) expression with the respective monoclonal antibodies. RESULTS: Le(a), sialyl Le(a), Le(x) and sialyl Le(x) were expressed in 69 (68.3%), 73 (72.3%), 66 (65.4%) and 76 (75.3%) carcinomas, respectively. The patients with sialyl Lex-expressing tumours had more advanced cancer than those with nonsialyl Lex-expressing tumours (P=0.0029). The survival time after surgery of patients with Le(x)- or sialyl Le(x)-expressing tumours was significantly shorter than the survival time of those with non-Le(x)- or nonsialyl Le(x)-expressing tumours, respectively (P=0.023 and P=0. 0001, respectively). Cox's regression analysis revealed that Le(x) and sialyl Le(x) expression, separate from stage and histological type, were prognostic variables for patient survival (hazard ratio [HR] for sialyl Le(x)-positive expression to sialyl L(x)-negative expression 2.90; HR for Le(x)-positive expression to Le(x)-negative expression 12.76 in stage I/IV, 0.63 in stage II and 1.69 in stage III). CONCLUSIONS: Le(x) expression and sialyl Le(x) expression in colorectal carcinomas are each associated with poor prognosis. These variables should be considered in the design of future trials.
Colorectal tumors can be classified based on their growth pattern into the polypoid growth type (PG-type) and nonpolypoid growth type (NPG-type). We examined sialyl Tn antigen expression in advanced colorectal carcinomas that were classified as PG-type and NPG-type cancers in order to ascertain whether or not such expression correlates with other biologically and clinically important differences. A total of 94 advanced colorectal carcinomas were examined for sialyl Tn antigen expression, which was immunohistochemically detected by the monoclonal antibody TKH2. Univariate and multivariate analyses using logistic regression models were performed. Forty carcinomas (42.6%) were negative and 54 (57.5%) were positive for sialyl Tn antigen. Eighteen carcinomas (19.2%) were of PG type and 76 (80.8%) of NPG type. NPG-type cancers had a higher proportion of positive lymph node metastasis than PG-type cancers. Furthermore, sialyl Tn antigen was less often detected in NPG-type cancers (39 of 76; 51.3%) than in PG-type cancers (15 of 18; 83.3%; p = 0.0167). Multivariate analysis showed that two variables, lymph node metastasis and sialyl Tn antigen expression in carcinoma, were independently related to tumor growth patterns, that is NPG type and PG type. These data suggest that the difference in sialyl Tn antigen expression between two kinds of tumor growth patterns of advanced colorectal carcinomas, PG type and NPG type, may reflect different biological behaviors during tumor progression.
The carbohydrate structure of sialyl-Lewis X (SLe(x)) can function as a ligand for E- and P-selectin, which play important roles in mediating the initial interactions of leukocytes with the endothelium in inflammatory responses. In this study we evaluated the effects of inhibiting E- and P-selectin function with the SLe(x) molecule on the inflammatory response in an experimental murine model of hypersensitivity pneumonitis (HP). Antigen exposure induced marked interstitial and especially perivascular and peribronchiolar infiltration with lymphocytes and granuloma formation, in murine lung sensitized with Saccaropolyspora rectivirgula. These pathologic changes were significantly suppressed with SLe(x) ganglioside analogues through a reduction in the numbers of lymphocytes in bronchoalveolar lavage fluid, as evidenced by the lung index and histologic scores indicating the severity of the inflammatory response. Using specific antibodies, we also evaluated the immunohistochemical localization of SLe(x) in mononuclear cells in granulomas, and of E- and P-selectin in vascular endothelium. Our findings suggest that the molecular interaction between SLe(x), and E- and P-selectin mediates lymphocyte recruitment into the lung parenchyma, which is critical for the inflammatory response in experimental murine models of HP.
The medicolegal issues surrounding telepathology and telecytology in Japan have been partly resolved since the government declared that telediagnosis in these fields using a microscope is essentially a medical practice between doctors. Thus it does not contravene article 20 of the law pertaining to medical practice in Japan, which states that there must be an actual face-to-face encounter between the doctor performing any diagnostic or treatment episode and the patient. It is, however, only beginning to be recognized that the medicolegal issues surrounding telepathology and telecytology depend on factors such as the type of system used (e.g. active versus passive). In an active diagnostic system, the telepathologist or telecytopathologist can control a robotic microscope at the remote site, while in a passive diagnostic system the telepathologist or telecytopathologist makes the diagnosis based on the microscope images that have been selected, and transmitted, by someone in the remote hospital. In the former case the interpreting telepathologist or telecytopathologist must assume most of the responsibility for the diagnostic process, while in the latter more responsibility must be assumed by those at the remote site. The duties and responsibilities of all participants in the telediagnostic process, whatever the system employed, must be determined. We have established working guidelines for telepathological and telecytological diagnoses to help achieve this and hence ensure safe and effective clinical practice.
Current Japanese clinical practice involves the usage of large amounts of new macrolides such as clarithromycin and roxithromycin for the treatment of diffuse panbronchiolitis, Helicobacter pylori and Mycobacterium avium complex infections. In this study, the phenotypes, genotypes, and macrolide resistance mechanisms of macrolide-inactivating Escherichia coli recovered in Japan from 1996 to 1997, were investigated. The isolation rate of erythromycin A highly-resistant E. coli (MIC > or = 1,600 microg/ml) in Japan slightly increased from 0.5% in 1986 to 1.2% in 1997. In six macrolide-resistant strains, recovered from the strains collected for this study during 1996 to 1997, the inactivation of macrolide could be detected with or without added ATP in the assay system. The appearance of erythromycin A-inactivating enzyme independent of ATP was novel from Japanese isolates, and the 1H NMR spectra of oleandomycin hydrolyzed by the three ATP-independent isolates were examined. It was clearly shown that the lactone ring at the position of C-13 was cleaved as 13-H signal in aglycon of oleandomycin upper shifted. These results suggested the first detection of macrolide-lactone ring-hydrolase from clinical isolates in Japan. These results suggested the first detection of an ATP-independent macrolide-hydrolyzing enzyme from Japanese clinical isolates. Substrate specificity of the macrolide-hydrolyzing enzyme was determined with twelve macrolides including the newer members of this group and it was found that not only erythromycin A but also the new macrolides, such as clarithromycin, roxithromycin, and azithromycin were inactivated. The NMR data, broad spectrum of activity, and independence of co-enzyme supported our naming of the enzyme as a macrolide esterase. PCR methodology was employed to detect an ereB-like gene from the 3 isolates producing macrolide esterase, and one of these was subsequently shown to contain both ereB-like and ermB-like genes. It was also clearly shown that the other three isolates, which inactivated macrolide in the presence of ATP, had an mphA-like gene.
Clinical isolates collected from clinical facilities across Japan in 1998 were tested against five aminoglycosides and three beta-lactams. The resistance of 50 strains each of methicillin sensitive Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Escherichia coli, Citrobacter freundii, Klebsiella pneumoniae, Enterobacter sp., Serratia sp., Pseudomonas aeruginosa and Proteus sp. (P. mirabilis 25 strains and P. vulgaris 25 strains) to the aminoglycosides isepamicin (ISP), amikacin (AMK), gentamicin, tobramycin and dibekacin, and to the beta-lactams imipenem, ceftazidime and piperacillin (all three known to be effective against P. aeruginosa) were investigated using a micro liquid dilution method with the following results: 1. ISP was effective against all strains except for 14% of MRSA, 2% of Proteus sp., and 4% of P. aeruginosa. 2. Six strains of MRSA were resistant to all eight drugs; however, in these cases ISP exhibited a relatively low minimum inhibitory concentration (MIC) compared to the other compounds. 3. Four strains of MRSA were resistant to all drugs except ISP. MRSA was the only isolate to demonstrate a resistance to seven or more drugs. 4. Twenty-one strains of MRSA and 1 strain of P. aeruginosa were resistant to six drugs; however, all of these were susceptible to both ISP and AMK. 5. Against all strains tested, ISP generally exhibited a lower MIC compared to AMK. These results suggest that, even ten years after its entering the market, ISP is still an aminoglycoside having a high anti-bacterial activity against a wide range of clinical isolates.
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Telepathology is the performance of pathology at distance using available telecommunication links such as optic fiber, communication satellite, and integrated services digitized network (ISDN). The main applications of telepathology are to provide frozen section service, consultation between pathologists at a distance, and conducting conferences using displays. These activities are required due to the shortage and disproportional distribution of pathologists. Telepathology for frozen section service is fairly effective in providing indications for surgical procedures by discriminating malignant tumor from benign tumor, confirming metastasis to distant organs or lymph nodes, and decisions regarding the surgical margin. However, there are several public insurance problems in the spread and practice of telepathology. To solve these problems, not only from a medical approach but also from the development of mechanical engineering, economic and legal issues must be considered.
Frequent gains of 20q have been identified recently in many neoplasias, including breast, bladder, ovarian, pancreatic, and colon cancers. A high level of 20q13.2 amplification is associated with poor prognosis in breast cancer. We investigated the copy number of the 20q13.2 region including the ZNF217 oncogene in 17 nonmetastatic colorectal cancers (CRCs), 18 primary CRC tumors with liver metastasis, and 18 metastatic lesions by two-color fluorescence in situ hybridization to evaluate the significance of an increased copy number of 20q13.2 in CRC, especially in those cases with liver metastasis. The frequency of increased relative copy number of the 20q13.2 region was higher in primary and liver metastatic lesions of CRC than in CRC lesions without liver metastasis. In particular, a high-level increase (>3.0-fold) in the relative copy number of 20q13.2 was observed in 2 of 18 (11%) primary CRC lesions with liver metastasis, 7 of 18 (39%) liver metastatic lesions, and in none of the cases of primary CRC without liver metastasis. The absolute and relative copy number of chromosome 20q13.2 was higher in CRCs with metastasis than in CRCs without metastasis. The percentage of cells with high-level 20q13.2 amplification was also higher in both lesions with metastasis per specimen than without metastasis. Our results suggest that the level of 20q13.2 amplification correlates with the metastatic potential and tumor progression of CRC. The results also suggest that 20q13.2 amplification with ZNF217 is associated with increased metastatic potential.
The cleavage signal transferred to the future cleavage cortex during anaphase has been proposed as "cleavage stimulus," but no signal has proved to induce cleavage furrows. The local Ca2+ transient along the cleavage furrow has been reported, but the Ca2+ source has remained unknown. To address these questions, we studied functions of Ca2+ stores in dividing newt eggs and found that microinjection of the Ca2+ store-enriched microsome fraction to the dividing newt egg induced a local extra-cleavage furrow at the injection site in 64-67% of the injected newt eggs while coinjection with inositol 1,4, 5-trisphosphate receptor (IP(3)R) antagonists heparin or anti-type 1-IP(3)R antibody clearly suppressed this induction (5 and 11% in induction rates, respectively). Injection of cerebellar microsomes from the type 1-IP(3)R-deficient mice induced extracleavage furrows albeit at a low rate (19%). Our observations strongly suggest that Ca2+ stores with IP(3)R induce and position a cleavage furrow via IP(3)-induced Ca2+ release (IICR) as Ca(2+)-releasing machinery and putative cleavage stimulus itself.
Three mammalian hyaluronan synthase genes, HAS1, HAS2, and HAS3, have recently been cloned. In this study, we characterized and compared the enzymatic properties of these three HAS proteins. Expression of any of these genes in COS-1 cells or rat 3Y1 fibroblasts yielded de novo formation of a hyaluronan coat. The pericellular coats formed by HAS1 transfectants were significantly smaller than those formed by HAS2 or HAS3 transfectants. Kinetic studies of these enzymes in the membrane fractions isolated from HAS transfectants demonstrated that HAS proteins are distinct from each other in enzyme stability, elongation rate of HA, and apparent K(m) values for the two substrates UDP-GlcNAc and UDP-GlcUA. Analysis of the size distributions of hyaluronan generated in vitro by the recombinant proteins demonstrated that HAS3 synthesized hyaluronan with a molecular mass of 1 x 10(5) to 1 x 10(6) Da, shorter than those synthesized by HAS1 and HAS2 which have molecular masses of 2 x 10(5) to approximately 2 x 10(6) Da. Furthermore, comparisons of hyaluronan secreted into the culture media by stable HAS transfectants showed that HAS1 and HAS3 generated hyaluronan with broad size distributions (molecular masses of 2 x 10(5) to approximately 2 x 10(6) Da), whereas HAS2 generated hyaluronan with a broad but extremely large size (average molecular mass of >2 x 10(6) Da). The occurrence of three HAS isoforms with such distinct enzymatic characteristics may provide the cells with flexibility in the control of hyaluronan biosynthesis and functions.