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Biomedical subjects

N Hamasaki

Publications and source records attributed to N Hamasaki.

At least 19 recordsLinked to original sources
PubMed2005

Evidence for a novel racemization process of an asparaginyl residue in mouse lysozyme under physiological conditions.

We examined chemical reactions in mouse lysozyme after incubation under physiological conditions (pH 7 and 37 degrees C). After incubation for 8 weeks, racemization was observed specifically at Asn127 among the 19 Asp/Asn residues in mouse lysozyme. Furthermore, analysis of the primary structure showed that the racemized residue was not Asp, but Asn, which demonstrates that deamidation and isomerization did not occur. These results mean that this racemization occurs without forming a succinimide intermediate. This is the first example of D-asparaginyl formation in a protein occurring during the racemization process under physiological conditions.

Amino Acid Sequence↗
PubMed2004

Urine diacetylspermine as a novel tumour maker for pancreatobiliary carcinomas.

BACKGROUND: Serum carcinoembryonic antigen (highly specific) and carbohydrate antigen 19-9 (highly sensitive) have been used as tumour markers for pancreatobiliary cancers. A novel urine tumour marker, diacetylspermine, was compared with the two conventional serum tumour markers in 125 patients with pancreatobiliary diseases. RESULTS: When the diagnosis of benign or malignant condition was examined, the sensitivity of urine diacetylspermine (75%) was higher than that of serum carcinoembryonic antigen (44%; P=0.048) and the same as that of serum carbohydrate antigen 19-9 (75%). The specificity of urine diacetylspermine (81%) was lower than that of serum CEA (92%) and as high as that of serum carbohydrate antigen 19-9 (80%). The efficiency of urine diacetylspermine (79%) was higher than that of serum carcinoembryonic antigen (74%) and the same as that of serum carbohydrate antigen 19-9 (79%). CONCLUSION: These results suggest that urine diacetylspermine is a marker for pancreatobiliary carcinoma, which is as highly sensitive and specific as serum carbohydrate antigen 19-9.

Adult↗
PubMed2001

8-oxo-dGTPase, which prevents oxidative stress-induced DNA damage, increases in the mitochondria from failing hearts.

BACKGROUND: Reactive oxygen species (ROS) can cause an oxidative modification of nucleotides, such as 8-oxo-7,8-dihydrodeoxyguanosine triphosphate (8-oxo-dGTP), which can lead to defects in DNA replication. The misincorporation of 8-oxo-dGTP into DNA is prevented by 8-oxo-dGTPase, which hydrolyzes 8-oxo-dGTP into 8-oxo-dGMP. The changes in this defensive system have not yet been examined in failing hearts, in which the generation of ROS increases. METHODS AND RESULTS: Myocardial infarction (MI) was created in mice by ligating the left coronary artery. Four weeks later, the left ventricle was dilated and contractility was diminished on echocardiography. The generation of ROS, as measured by electron spin resonance spectroscopy with 4-hydroxy-2,2,6,6-tetramethyl-piperidine-N-oxyl, increased in the noninfarcted left ventricle from MI mice. The formation of thiobarbituric acid-reactive substances also increased in the mitochondria from MI mice. 8-Oxo-dGTPase was detected in the mitochondrial fractions isolated from MI mice using a Western blot analysis with an antibody to its human homologue (hMTH1). Immunohistochemistry showed positive staining for hMTH1 was localized in the cardiac myocytes. CONCLUSIONS: The level of 8-oxo-dGTPase increased in the mitochondria isolated from post-MI hearts as oxidative stress increased, thus suggesting that a preventive mechanism is activated against ROS-induced DNA damage. As a result, 8-oxo-dGTPase is considered a useful marker of mitochondrial oxidative stress in heart failure.

Animals↗
PubMed2001

Both heavy strand replication origins are active in partially duplicated human mitochondrial DNAs.

The replication of human mitochondrial DNA (mtDNA) is initiated from a pair of displaced origins, one priming continuous synthesis of daughter-strand DNA from the heavy strand (OH) and the other priming continuous synthesis from the light strand (OL). In patients with sporadic large-scale rearrangements of mitochondrial DNA (i.e., partially-deleted [Delta-mtDNA] and partially-duplicated [dup-mtDNA] molecules), the dup-mtDNAs typically contain extra origins of replication, but it is unknown at present whether they are competent for initiation of replication. Using cybrids harboring each of two types of dup-mtDNAs-one containing two OHs and two OLs, and one containing two OHs and one OL-we used ligation-mediated polymerase chain reaction (LMPCR) to measure the presence and relative amounts of nascent heavy strands originating from each OH. We found that the nascent heavy strands originated almost equally from the two OHs in each cell line, indicating that the extra OH present on a partially duplicated mtDNA is competent for heavy strand synthesis. This extra OH could potentially confer a replicative advantage to dup-mtDNAs, as these molecules may have twice as many opportunities to initiate replication compared to wild-type (or partially deleted) molecules.

Cell Line↗
PubMed2001

a4, a unique kidney-specific isoform of mouse vacuolar H+-ATPase subunit a.

The vacuolar-type H+ -ATPase (V-ATPase) translocates protons across membranes. Here, we have identified a mouse cDNA coding for a fourth isoform (a4) of the membrane sector subunit a of V-ATPase. This isoform was specifically expressed in kidney, but not in the heart, brain, spleen, lung, liver, muscle, or testis. Immunoprecipitation experiments, together with sequence similarities for other isoforms (a1, a2, and a3), indicate that the a4 isoform is a component of V-ATPase. Moreover, histochemical studies show that a4 is localized in the apical and basolateral plasma membranes of cortical alpha- and beta-intercalated cells, respectively. These results suggest that the V-ATPase, with the a4 isoform, is important for renal acid/base homeostasis.

Acid-Base Equilibrium↗
PubMed2001

Failure in the detection of aberrant mRNA from the heterozygotic splice site mutant allele for protein S in a patient with protein S deficiency.

A 29-year-old male patient with acute arterial obstruction and a medical history including thrombosis in the deep veins and pulmonary infarction presented with a reduced level of both protein S (PS) activity and free PS. Sequencing of the genomic PS gene in this patient revealed that the patient was heterozygous for the mutant PS allele, in which a nucleotide substitution occurred at the donor splice site in intron 12 (GT to GA). The patient was heterozygous for PS genes having dimorphic codons for Pro626 (CCA/CCG) and the aberrant allele in this patient was associated with the CCA form. Allelic exclusion of PS expression was demonstrated by use of Pro626 (CCA/CCG) dimorphism and only a normal mRNA sequence derived from the CCG-allele was identified in the patient. These findings suggested that the mutation at the splice site in the PS gene caused either defective production of mRNA or the gene may have produced extremely unstable RNA products, leading to reduced levels of PS activity and free PS in this patient.

Adult↗
PubMed2001

Mitochondrial DNA damage and dysfunction associated with oxidative stress in failing hearts after myocardial infarction.

Mitochondria are one of the enzymatic sources of reactive oxygen species (ROS) and could also be a major target for ROS-mediated damage. We hypothesized that ROS may induce mitochondrial DNA (mtDNA) damage, which leads to defects of mtDNA-encoded gene expression and respiratory chain complex enzymes and thus may contribute to the progression of left ventricular (LV) remodeling and failure after myocardial infarction (MI). In a murine model of MI and remodeling created by the left anterior descending coronary artery ligation for 4 weeks, the LV was dilated and contractility was diminished. Hydroxyl radicals, which originated from the superoxide anion, and lipid peroxide formation in the mitochondria were both increased in the noninfarcted LV from MI mice. The mtDNA copy number relative to the nuclear gene (18S rRNA) preferentially decreased by 44% in MI by a Southern blot analysis, associated with a parallel decrease (30% to 50% of sham) in the mtDNA-encoded gene transcripts, including the subunits of complex I (ND1, 2, 3, 4, 4L, and 5), complex III (cytochrome b), complex IV (cytochrome c oxidase), and rRNA (12S and 16S). Consistent with these molecular changes, the enzymatic activity of complexes I, III, and IV decreased in MI, whereas, in contrast, complex II and citrate synthase, encoded only by nuclear DNA, both remained at normal levels. An intimate link among ROS, mtDNA damage, and defects in the electron transport function, which may lead to an additional generation of ROS, might play an important role in the development and progression of LV remodeling and failure.

Animals↗
PubMed2001

A novel splice acceptor site mutation of protein S gene in affected individuals with type I protein S deficiency: allelic exclusion of the mutant gene.

Sequencing studies of the protein S gene (PROS1) in a Japanese patient suffering from recurrent thrombosis revealed the following. The proband and his first daughter, but not the second daughter, were having the type I protein S (PS) deficiency due to a novel point mutation from A to G at the intronic acceptor splice site in intron 13 of the PROS1. In the affected daughter, exclusion of the aberrant allele was assessed by the BstX1 dimorphism of PROS1 at Pro626 (CCG/CCA). The reduced PS activities in the proband and his first daughter were apparently due to defective production of mRNA from the mutant allele.

Alleles↗
PubMed2001

Localization of human interleukin 13 receptor in non-haematopoietic cells.

Although the functional roles of interleukin (IL-)13 in haematopoietic cells are well investigated, those in non-haematopoietic cells remain to be addressed. IL-13 exerts its actions by binding to the IL-13 receptor (IL-13R) on target cells, which is composed of IL-13Ralpha1 and the IL-4 receptor alpha chain (IL-4Ralpha). However, there has been no study of localization of IL-13R in each tissue. To address this question, we generated monoclonal anti-IL-13Ralpha1 antibody, and performed immunohistochemistry using this antibody and anti-IL-4Ralpha antibody. Distribution of these two components was the same in all examined tissues. Staining was positive in keratinocytes, hair follicles, and sebaceous and sweat glands in skin; in ciliated respiratory epithelial cells in nasal tissue; in heart muscle cells; in foveola cells, gastric glands, and the smooth muscle layer in stomach; and in hepatocytes in liver. However, staining was undetectable in brain and bone marrow. Fibroblasts and endothelial cells were stained in some tissues. These results provide clues to elucidate the known pathological roles of IL-13 in atopic dermatitis and allergic rhinitis, as well as its unknown physiological roles.

Animals↗
PubMed2001

Expression of lysosome-associated membrane proteins in human colorectal neoplasms and inflammatory diseases.

The lysosome-associated membrane proteins (LAMPs)-1 and -2 are major constituents of the lysosomal membrane. These molecules are known to be among the most glycosylated proteins of several types of cells and cancer cells, and their expression in cancer cells is marked by a distinct difference in the structures of the oligosaccharides as compared to nonmalignant cells. We analyzed by immunohistochemistry the intensity and distribution of LAMP-1 and LAMP-2 in 9 human colorectal cancer cases and in 16 control cases, including inflammatory diseases (diverticulitis, ulcerative colitis, and Crohn's disease). LAMP proteins were expressed more intensely in the epithelium of colorectal neoplasms than in normal mucosa (P < 0.05), and no significant differences were found between adenoma and cancer cells (P > 0.05) in the same tissue section. Further, in sites of inactive inflammatory diseases and nonneoplastic areas in cancer specimens, no significant increases in epithelial LAMP proteins were observed, even in the proliferative zone of the lower crypt epithelium. Northern blot analysis showed increased expression of LAMP-1 and LAMP-2A in two of three colorectal cancers examined and increased LAMP-2B in all three cancers. Our findings suggest that LAMPs are related to neoplastic progression, but there is no direct association between the expression of LAMP molecules and cell proliferation.

Adenocarcinoma↗
PubMed2001

Activation of the lectin complement pathway by ficolins.

Mannose-binding lectin (MBL), a serum lectin specific for mannose or N-acetylglucosamine (GlcNAc), which contains both a collagen-like domain and a carbohydrate-recognition domain (CRD), plays a role in innate immunity by acting as an opsonin and activating complement in association with MBL-associated serine protease (MASP) via the lectin pathway. Another type of GlcNAc-binding lectins termed "ficolins" exist in serum. They are characterized by the presence of both a collagen-like domain and a fibrinogen-like domain. Investigations of two types of human serum ficolins, ficolin/P35 and Hakata antigen, revealed that they are associated with MASPs and sMAP, a truncated protein of MASP-2, and activate complement. These findings indicate that, like MBL, serum ficolins are collagenous lectins capable of activating the lectin pathway and thus have a role in innate immunity.

Carrier Proteins↗
PubMed2001

Standardization of laboratory data and establishment of reference intervals in the Fukuoka Prefecture: a Japanese perspective.

Standardization of 22 clinical chemistry analytes and five serum protein constituents has been performed in the Fukuoka Prefecture, which has a population of approximately five million. The standardization project was established to determine reference intervals for these analytes by educating physicians, medical technologists and staff of medical institutions, and by daily or monthly monitoring the use of common control samples through e-mail. Standardization extended to 97% of the institutions in the prefecture. Results for 14 of the 22 clinical chemistry analytes have become highly reliable and differences between institutions decreased. Standardization of other analytes is now in progress. Regional collaboration based on international guidelines led to a significant improvement in interlaboratory comparability. Areas where further improvements are needed have been identified.

Adolescent↗
PubMed2001

[Molecular biology techniques as clinical laboratory tests].

The quality of results obtained with molecular biology techniques depends on the control of preanalytical and analytical error associated with such techniques. Preanalytical error can be introduced during the isolation of DNA and RNA. The type of detergent used in cell lysis can affect the amplification of DNA by techniques such as the polymerase chain reaction(PCR). Ribonuclease(RNase) contamination is a serious problem in the isolation of undegraded RNA, and, thus, this enzyme should be inhibited. Anticoagulants used for blood collection can affect the quality of results with molecular biology techniques. The control of contamination from the working environment is essential to the minimization of preanalytical, analytical and postanalytical error. Molecular biology techniques for a wide range of clinical laboratory tests have been established in hospitals such as clinical laboratory tests for infections, molecular diagnoses of leukemia and aberrant genes in metabolism.

Blood Coagulation Factors↗
PubMed2001

[Genetic polymorphism and risk of thromboses].

We have attempted to establish a systematic pathogenetic analysis of thrombophilia by including assays of antithrombin III(AT III), protein C(PC), protein S(PS), fibrinogen, plasminogen and heparin cofactor II by both functional and immunological methods as well as detecting lupus anticoagulants. Such a comprehensive scheme was instrumental in systematically identifying and confirming the pathogenesis of 164 cases which otherwise would have escaped detection since 1994 in our laboratory (Kyushu University Hospital). The analysis was conducted on 485 consecutive patients with venous thrombosis, arterial thrombosis and disorders in which small vessel thrombosis were implicated. Hundred and sixty four patients, (40% of the examined patients), were found to have low activities of PS, PC, ATIII etc. Among them, seventy five patients(46%) had low PS activity, and twenty nine(18%) had low PC activity. Genetic analyses performed on specimens with low PS/PC activities resulted in the confirmation of 24 genetic abnormalities. Such genetic abnormalities, however, does not solely lead to the pathogenesis of thromboses. We have found that some genetic polymorphisms, such as PS Tokushima, factor XII 46C allele, were also additional risk factors for thromboses.

Antithrombin III↗
PubMed2001

[Primary thalamic hemorrhage penetrating to the lateral ventricle in an infant with abnormal protein S(protein S-Tokushima)].

We experienced a 3 months infant with thalamic hemorrhage penetrating to lateral ventricle with abnormal Protein S. Although the coagulation factor and fibrinogenolysis factors were evaluated, there were no remarkable abnormal laboratory data except for slightly decline of Protein S. The DNA analysis was performed for Protein S, and a missense mutation(A to G transmission) was found, which was resulting in Lys-155 to Glu. The total Protein S antigen was normal level, but co-factor activity for activated Protein C was declined. That mutation is named Protein S-Tokushima, and the patient who has abnormal Protein S tends to suffer recurrent coagulopathy. In our patient, it was interesting that any thrombotic disease had not occurred, but cerebral hemorrhage had occurred.

Cerebral Hemorrhage↗
PubMed2001

[Clinical laboratory data and reference intervals standardized in Fukuoka].

In Fukuoka whose population is approximately five million inhabitants, surveys on the accuracy of laboratory data have been performed by the Fukuoka Prefecture Medical Association for the last 30 years. We have been attempting to evaluate the data for routine use since 1988, and it has become possible to share laboratory data between all institutions in Fukuoka prefectures. As a result, reference intervals for 23 clinical chemistry analytes were established in 1995, to which were added in 1996 five serum protein constituents that have been utilized for clinical examinations. Methods for documentations and monitorings the data obtained in the prefecture were also established, standardization of the above analytes extended to 97% of the institutions in the prefecture. Results for 14 of the 23 clinical chemistry analytes have become highly reliable and clinically useful as differences between institutions in terms of results have narrowed. Standardization of other analytes is now in progress.

Aged↗