Biomedical subjects

M Kai

Publications and source records attributed to M Kai.

At least 55 records · Page 3Linked to original sources

PubMed2000

Immune response against a cross-reactive epitope on the heat shock protein 60 homologue of Helicobacter pylori.

We previously established a monoclonal antibody (MAb), designated H9, which reacts with the heat shock protein 60 (HSP60) homologue of Helicobacter pylori as well as with other bacterial and human HSP60s. To determine the importance of a cross-reactive epitope on H. pylori HSP60 in H. pylori immunopathogenesis, we performed (i) mapping of an epitope on H. pylori HSP60 recognized by the H9 MAb, (ii) analysis of immunoglobulin G responses of patients with or without H. pylori infection to its epitope region, and (iii) studies of the protective effect of immunization with its epitope region on H. pylori infection in mice. The epitope recognized by the H9 MAb was mapped to the sequence of amino acids 189 to 203 (VEGMQFDRGYLSPYF) on the H. pylori HSP60 molecule. It was confirmed that the synthesized peptide designated pH9 was recognized by the H9 MAb. Enzyme-linked immunosorbent assay analysis showed that patients with H. pylori infection (n = 349) had significantly lower titers of pH9 antibody than did uninfected patients (n = 200) (P < 0.001), but this was not the case with purified H. pylori HSP60 recombinant Escherichia coli GroEL, or recombinant human HSP60. In C57BL/6 mice immunized with the pH9 peptide with Freund's complete adjuvant (FCA), the number of H. pylori organisms colonizing the stomach was significantly lower than that in mice immunized with pCont plus FCA (P < 0.0001) or FCA only (P < 0.005). The results suggest that the immune response to the cross-reactive epitope (pH9 region) on H. pylori HSP60 is unique and might be associated with protection against H. pylori infection.

Amino Acid Sequence↗

PubMed2000

Fission yeast Eso1p is required for establishing sister chromatid cohesion during S phase.

Sister chromatid cohesion is essential for cell viability. We have isolated a novel temperature-sensitive lethal mutant named eso1-H17 that displays spindle assembly checkpoint-dependent mitotic delay and abnormal chromosome segregation. At the permissive temperature, the eso1-H17 mutant shows mild sensitivity to UV irradiation and DNA-damaging chemicals. At the nonpermissive temperature, the mutant is arrested in M phase with a viability loss due to a failure to establish sister chromatid cohesion during S phase. The lethal M-phase arrest phenotype, however, is suppressed by inactivation of a spindle checkpoint. The eso1(+) gene is not essential for the onset and progression of DNA replication but has remarkable genetic interactions with those genes regulating the G(1)-S transition and DNA replication. The N-terminal two-thirds of Eso1p is highly homologous to DNA polymerase eta of budding yeast and humans, and the C-terminal one-third is homologous to budding yeast Eco1p (also called Ctf7p), which is required for the establishment of sister chromatid cohesion. Deletion analysis and determination of the mutation site reveal that the function of the Eco1p/Ctf7p-homologous domain is necessary and sufficient for sister chromatid cohesion. On the other hand, deletion of the DNA polymerase eta domain in Eso1p increases sensitivity to UV irradiation. These results indicate that Eso1p plays a dual role during DNA replication. The C-terminal region acts to establish sister chromatid cohesion, and the N-terminal region presumably catalyzes translesion DNA synthesis when template DNA contains lesions that block regular DNA replication.

Acetyltransferases↗

PubMed2000

Protection by polaprezinc, an anti-ulcer drug, against indomethacin-induced apoptosis in rat gastric mucosal cells.

Polaprezinc [N-(3-aminopropionyl)-L-histidinato zinc] (PZ), an anti-ulcer drug, is a chelate compound consisting of zinc and L-carnosine. PZ has been shown to prevent gastric mucosal injury. In the present study, we investigated the inhibitory effect of PZ on indomethacin (IND)-induced apoptosis in a rat gastric mucosal cell line, RGM1. Pretreatment with PZ suppressed caspase-3 activation and subsequent apoptosis in the cells exposed to 500 microM IND in a dose-dependent manner, and 50 microM PZ exhibited the maximum inhibitory effect. Among PZ subcomponents, zinc but not L-carnosine played a pivotal role in this antiapoptotic function. PZ did not affect mitochondrial cytochrome c release upstream of caspase-3 activation in the IND-induced apoptotic signal pathway. Treatment with 500 microM IND evidently produced reactive oxygen species (ROS) in RGM1 cells. However, PZ did not scavenge ROS in IND-treated cells. Moreover, N-acetylL-cysteine, a potent antioxidant, inhibited ROS generation but did not suppress apoptosis in RGM1 cells exposed to IND. These observations demonstrate a novel pharmacological action of PZ; i.e., that PZ, and in particular its zinc subcomponent, inhibits apoptosis via inhibition of caspase-3 activation but not antioxidant activity.

Acetylcysteine↗

PubMed2000

Use of an instrument sandwiched between the hoof and shoe to measure vertical ground reaction forces and three-dimensional acceleration at the walk, trot, and canter in horses.

OBJECTIVE: To develop an instrument that could be sandwiched between the hoof and shoe of horses and that would reliably measure vertical ground reaction forces and three-dimensional acceleration at the walk, trot, and canter. ANIMALS: 5 clinically sound Thoroughbreds. PROCEDURES: The recording instrument (weight, 350 g) consisted of 2 metal plates, 2 bolts, 4 load cells, and 3 accelerometers. It was mounted to the hoof with a glue-on shoe and devised to support as much load exerted by a limb as possible. The load cells and accelerometers were wired to a 16-channel transmitter, and transmitted signals were received and amplified with a telemetry receiver. RESULTS: The recording instrument could measure in real time the 4 components of the ground reaction force or their resultant force along with acceleration in 3 dimensions as horses walked, trotted, or cantered on a treadmill. Patterns of force-time curves recorded for consecutive strides were similar to each other and to those previously reported, using a force plate. CONCLUSIONS AND CLINICAL RELEVANCE: The recording instrument developed for use in the present study allowed us to record vertical ground reaction force and acceleration in 3 dimensions in horses at the walk, trot, and canter.

Acceleration↗

PubMed2000

Cloning, sequencing, and transcriptional analysis of the dnaK heat shock operon of Listeria monocytogenes.

The complete dnaK operon of Listeria monocytogenes was isolated by chromosome walking using the previously cloned dnaK gene as a probe. Molecular analysis of the locus identified 6 genes in the order hrcA, grpE, dnaK, dnaJ, orf35, and orf29. Primer extension analysis revealed 3 transcription start sites-S1, S2, and S3-upstream of the hrcA, grpE, and dnaJ, respectively. The transcription from S1 was heat inducible. Analysis of the sequences revealed the consensus promoter sequences of gram-positive bacteria, P1 and P2 upstream of the hrcA and dnaJ, respectively. The hrcA gene and a regulatory sequence, designated CIRCE (controlling inverted repeat of chaperone expression), play a role in the regulation of expression of the dnaK locus in response to heat shock in several gram-positive bacteria. Their presence upstream of the dnaK locus in L. monocytogenes suggested a similar regulatory mechanism for the transcription initiated at the promoter, P1. Northern blot analysis led to the detection of 4 mRNA species of 4.9 kb, 3.6 kb, 3.6 kb, and 1.2 kb; the first 2 species were heat inducible. The current results indicate that 4 distinct transcripts directed by 3 promoters are involved in the expression of the dnaK operon of L. monocytogenes.

Amino Acid Sequence↗

PubMed2000

Extracorporeal shock wave lithotripsy: elimination of densely calcified gallstones and gallstones with calcified rims.

OBJECTIVE: Until now, radiopaque gallstones have been excluded from extracorporeal shock wave lithotripsy (ESWL), because these stones in vivo are less sensitive to the forces that cause disintegration. In Japan there is a higher percentage of patients with radiopaque gallstones than in Western countries. Our purpose in working with patients in Japan was to warrant extensive indication of ESWL to radiopaque gallstones, especially densely calcified stones. DESIGN/METHODS: Retrospective analysis of clinical data. Patients were classified by computed tomography (CT) of stones. Group A consisted of 78 patients whose gallstones were densely calcified (CT attenuation values in Hounsfield units (HU), 473 +/- 323). Group B consisted of 22 patients whose stones had a calcified rim (CT attenuation values, 357 +/- 244). Ninety-eight patients received adjuvant dissolution therapy with ursodeoxycholic acid. Other recommendations, such as a glass of milk at night, were not given to the patients. SETTING: One university hospital and one general hospital. PATIENTS: One hundred consecutive Japanese patients with radiopaque stones in contractile gallbladder (CT attenuation values, > 150 HU, 447 +/- 310, mean +/- SD) were the subjects. With respect to the efficacy of ESWL, a degree of calcification for stones and its relationship with the rates of stone fragmentation and disappearance were assessed. Fragmentation to less than 3 mm in stone diameter was the aim, without limit of shock wave discharges and sessions. RESULTS: After ESWL sessions stones were fragmented successfully in 74 of the 100 patients (57 of the 78 patients in Group A, and 17 of the 22 patients in Group B). The mean number of discharges per patient was 10,435 +/- 8,726. The mean number of discharges for successful stone fragmentation of Group A (9,839 +/- 8,187) was not significantly different from that of Group B (11,376 +/- 6,344). One year after lithotripsy, 60 of the 100 patients were free of stones (45 in Group A, and 15 in Group B). CONCLUSION: It appears that patients with either densely calcified gallstones, or those in whom the stones have a calcified rim, are both suitable candidates for lithotripsy.

Adult↗

PubMed2000

Overexpression of S-adenosylmethionine decarboxylase (SAMDC) activates the maternal program of apoptosis shortly after MBT in Xenopus embryos.

Overexpression of S-adenosylmethionine decarboxylase (SAMDC) mRNA in 1- and 2-cell stage Xenopus embryos induces cell autonomous dissociation at the late blastula stage and developmental arrest at the early gastrula stage. The induction of cell dissociation took place "punctually" at the late blastula stage in the SAMDC-overexpressing cells, irrespective of the stage of the microinjection of SAMDC mRNA. When we examined the cells undergoing the dissociation, we found that they were TUNEL-positive and contained fragmented nuclei with condensed chromatin and fragmented DNA. Furthermore, by injecting Xenopus Bcl-2 mRNA together with SAMDC mRNA, we showed that SAMDC-overexpressing embryos are rescued completely by Bcl-2 and becometadpoles. These results indicatethat cell dissociation induced by SAMDC overexpression is due to apoptotic cell death. Since the level of S-adenosylmethionine (SAM) is greatly reduced in SAMDC-overexpressing embryos and this induces inhibition of protein synthesis accompanied by the inhibition of DNA and RNA syntheses, we conclude that deficiency in SAM induced by SAMDC overexpression activates the maternal program of apoptosis in Xenopus embryos at the late blastula stage, but not before. We propose that this mechanism serves as a surveillance mechanism to check and eliminate cells physiologically damaged during the cleavage stage.

Adenosylmethionine Decarboxylase↗

PubMed2000

Mycobacterium leprae typing by genomic diversity and global distribution of genotypes.

The genetic diversity and related global distribution of 51 Mycobacterium leprae isolates were studied. Isolates were obtained from leprosy patients from 12 geographically distinct regions of the world and two were obtained from nonhuman sources. Polymerase chain reaction (PCR) followed by DNA sequencing was performed targeting the rpoT gene of M. leprae. Isolates were classified into two groups based on the number of tandem repeats composed of 6 base pairs in the rpoT gene. Isolates from Japan (except Okinawa) and Korea belonged to one group, while those from Southeast Asian countries, Brazil, Haiti and Okinawa in Japan belonged to a second genotype. M. leprae obtained from two nonhuman sources (an armadillo and a mangabey monkey) revealed the latter genotype. These results demonstrate the genetic diversity of M. leprae and the related genotype-specific distribution in the world.

Bacterial Proteins↗

PubMed2000

The role of cyclosporine A and interleukin-2 in obliterative airway disease in a rat tracheal transplant model.

The pathogenesis of obliterative bronchiolitis (OB) following lung and heart-lung transplantation remains unclear. We evaluated the role of CsA and IL-2 on the development of obliterative airway disease (OAD) by administrating exogenous IL-2 in a CsA-treated rat tracheal transplant model. Tracheal grafts were implanted into the peritoneal cavity from Brown Norway (BN) to BN rats or to Lewis (LEW) rats. Allotransplant: No treatment was given in group 1. Short-term CsA (25 mg/kg, i.m. on POD 2 and 3) was used in group 2. Group 3 was treated with long-term CsA (25 mg/kg, i.m. on POD 2 and 3, followed by 5 mg/kg on POD 4 to 27). Administration of IL-2 (300, 000 IU/kg, i.p. on POD 15 to 19 and 22 to 26) was performed to long-term CsA treated rats in group 4. Isotransplant: No treatment was given to group 5, group 6 was treated with IL-2 (same regimen as in group 4). Grafts were harvested at different time points after Tx for histological assessment. No luminal obliteration was observed in group 5 and 6. Complete luminal obliteration was noted 4 weeks after Tx in group 1. In group 2 and 3, obliterative lesion occurred 4-6 weeks after CsA withdrawal. IL-2 increased epithelial loss, lymphocytic infiltration, and obliterative changes in group 4. Our results suggest that OAD is an immune mediated disorder. Furthermore, administration of exogenous IL-2 might be able to abrogate the protection from OAD by CsA therapy.

Animals↗

PubMed1999

Diaminodiphenylsulfone resistance of Mycobacterium leprae due to mutations in the dihydropteroate synthase gene.

The nucleotide sequence analysis of the dihydropteroate synthase (DHPS) gene of six diaminodiphenylsulfone-resistant Mycobacterium leprae strains revealed that the mutation was limited at highly conserved amino acid residues 53 or 55. Though the mutation at amino acid residue 55 or its homologous site has been reported in other bacteria, the mutation at residue 53 is the first case in bacteria. This is the first paper which links the mutations in DHPS and sulfonamide resistance in M. leprae. This finding is medically and socially relevant, since leprosy is still a big problem in certain regions.

Dapsone↗

PubMed1999

Structure-function studies of an anti-asialo GM1 antibody obtained from a phage display library.

Although gangliosides elicit human autoantibodies, they are extremely weak immunogens in mice. We obtained a monoclonal antibody Fab fragment (clone 10) that is specific for asialo GM1 (GA1), from a phage display library. The Vkappa domain of clone 10 could be replaced by two different Vkappa domains without changing the specificity of the antibody. Mutagenesis of the third hypervariable regions of the heavy and light chains of clone 10 yielded three mutants that exhibited a 3 to 4 times increase in avidity for GA1. A molecular model of clone 10 indicated that the putative antigen-binding site contained a shallow surface pocket. These data illustrate the use of recombinant DNA techniques to obtain anti-ganglioside antibodies, and to explore the molecular basis of their antigen-binding activity.

Amino Acid Sequence↗

PubMed1999

Characterization of a 1200-kb genomic segment of chromosome 3p22-p21.3.

We previously determined the nucleotide sequence and characterized the 685-kb proximal half of CEPH YAC936c1, which corresponds to a portion of human chromosome 3p21.3. In the study reported here, we characterized the remaining 515-kb of this YAC clone corresponding to the telomeric half of its human insert. The newly sequenced region contained a total of ten genes including six reported previously: phospholipase C delta 1 (PLCD1), human activin receptor type IIB (hActR-IIB), organic cation transporter-like 1 (OCTL1), organic cation transporter-like 2 (OCTL2), oxidative stress response 1 (OSR1), and human xylulokinase-like protein (XYLB). The remaining four genes present in the telomeric region included two known genes, MyD88 and ACAA, and two novel genes. One (designated ENGL) of the novel sequences was found to encode an amino-acid sequence homologous to the family of DNA/RNA endonucleases, especially endonuclease G. The other gene F56 revealed no significant homology to any known genes. These results disclosed complete physical and transcriptional maps of the 1200-kb region of 3p present in YAC 936c1.

Amino Acid Sequence↗

PubMed1999

Molecular cloning of homologs of RAS and RHO1 genes from Cryptococcus neoformans.

We cloned and sequenced homologs of RAS(CnRAS) and RHO1(CnRHO1) genes from Cryptococcus neoformans. The proteins encoded by the CnRAS and CnRHO1 genes contained 216 and 197 amino acids, respectively. The deduced amino acid sequence of the CnRAS gene shared a high degree of sequence identity with the Ras proteins in other fungal species: Coprinus cinereus(76%), Lentinula edodes(74%), Saccharomyces cerevisiae RAS2(72%), and Schizosaccharomyces pombe(68%). The deduced amino acid sequence of the CnRHO1 gene shared a high degree of sequence identity with the Rho1 proteins in other fungal species: Candida albicans(78%), S. pombe(77%) and S. cerevisiae(76%). The deduced proteins contained GTP-binding and GTP-hydrolysis domains, and the prenylation site that are conserved among the small G protein superfamily. The synthetic peptides that contained the C-terminal amino acid sequence of the CnRas and CnRho1 proteins were geranylgeranylated.

Amino Acid Sequence↗

PubMed1999

A predominant increase of arterial beta-hydroxybutyrate concentration during partial hepatectomies in patients with impaired indocyanine green clearance test.

BACKGROUND: It has been reported that a concentration ratio of two ketone bodies, [acetoacetate]/[beta-hydroxybutyrate], reflects the redox state in the liver mitochondria, but there are few reports which examined each ketone body concentration during hepatectomy. Materials and Methods. (1) Ketone body levels in the radial artery and the hepatic vein were measured simultaneously during hepatectomies in 12 patients. (2) Arterial ketone body levels were measured during hepatectomies (at three points) in 73 patients. RESULTS: (1) Both ketone body levels in the radial artery well reflected those in the hepatic vein (P < 0.001). (2) There was no change in the arterial acetoacetate level. The level of beta-hydroxybutyrate increased significantly from 30.4 to 76.5 micromol/L (P < 0.005) only in the group of 25 patients with seriously impaired indocyanine green clearance test. CONCLUSIONS: Ketosis during hepatectomy was caused by increased beta-hydroxybutyrate, and this phenomenon was observed only in patients with seriously impaired liver function.

3-Hydroxybutyric Acid↗

PubMed1999

Molecular characterization of the type 2 phosphatidic acid phosphatase.

Phosphatidic acid phosphatase (PAP) converts phosphatidic acid to diacylglycerol, thus regulating the de novo synthesis of glycerolipids and also signal transduction mediated by phospholipase D. We initially succeeded in the cDNA cloning of the mouse 35 kDa PAP bound to plasma membranes (type 2 enzyme). This work subsequently led us to the identification of two human PAP isozymes designated 2a and 2b. A third human PAP isozyme (2c) has also been described. The cloned enzymes are, in common, N-glycosylated and possess six transmembrane domains. The transmembrane dispositions of these enzymes are predicted and the catalytic sites are tentatively located in the 2nd and 3rd extracellular loops, thus suggesting that the type 2 PAPs may act as ecto-enzymes dephosphorylating exogenous substrates. Furthermore, the type 2 PAPs have been proposed to belong to a novel phosphatase superfamily consisting of a number of soluble and membrane-bound enzymes. In vitro enzyme assays show that the type 2 PAPs can dephosphorylate lyso-phosphatidate, ceramide-1-phosphate, sphingosine-1-phosphate and diacylglycerol pyrophosphate. Although the physiological implications of such a broad substrate specificity need to be further investigated, the type 2 PAPs appear to metabolize a wide range of lipid mediators derived from both glycero- and sphingolipids.

Amino Acid Sequence↗

PubMed1999

Hydrogen peroxide-induced apoptosis in HL-60 cells requires caspase-3 activation.

Apoptosis has been associated with oxidative stress in biological systems. Caspases have been considered to play a pivotal role in the execution phase of apoptosis. However, which caspases function as executioners in reactive oxygen species (ROS)-induced apoptosis is not known. The present study was performed to identify the major caspases acting in ROS-induced apoptosis. Treatment of HL-60 cells with 50 microM hydrogen peroxide (H2O2) for 4 h induced the morphological changes such as condensed and/or fragmented nuclei, increase in caspase-3 subfamily protease activities, reduction of the procaspase-3 and a DNA fragmentation. To determine the role of caspases in H2O2-induced apoptosis, caspase inhibitors, acetyl-Tyr-Val-Ala-Asp-chloromethyl ketone (Ac-YVAD-cmk), acetyl-Asp-Glu-Val-Asp-aldehyde (Ac-DEVD-CHO) and acetyl-Val-Glu-Ile-Asp-aldehyde (Ac-VEID-CHO), selective for caspase-1 subfamily, caspase-3 subfamily and caspase-6, respectively, were loaded into the cells using an osmotic lysis of pinosomes method. Of these caspase inhibitors, only Ac-DEVD-CHO completely blocked morphological changes, caspase-3 subfamily protease activation and DNA ladder formation in H2O2-treated HL-60 cells. This inhibitory effect was dose-dependent. These results suggest that caspase-3, but not caspase-1 is required for commitment to ROS-triggered apoptosis.

Amino Acid Chloromethyl Ketones↗

PubMed1999

Influence of training on autonomic nervous function in horses: evaluation by power spectral analysis of heart rate variability.

We studied the influence of training on autonomic nervous function in the horse. For this purpose, Holter electrocardiogram (ECG) was recorded before and after training from 24 Thoroughbred horses (2-year-olds) and autonomic nervous function was evaluated by the power spectral analysis of heart rate (HR) variability. We obtained HR, low-frequency (LF) power, high-frequency (HF) power, and LF/HF ratio from recording. We set LF at 0.01-0.07 Hz and HF at 0.07-0.6 Hz. The HF power is thought to reflect primarily parasympathetic nervous function. Both the sympathetic and parasympathetic nervous systems have been shown to contribute to the LF power. The LF/HF ratio is considered as an index of the cardiac sympathovagal balance. Second degree atrioventricular blocks were found in the ECG of 3 and 5 horses before and after the training period, respectively. Ventricular premature depolarisations were detected in a horse after the training period. Heart rate decreased at night. The lowest HR values had a tendency to appear in the early morning. Both the LF and HF power tended to be higher at night. However, the LF/HF ratio was almost the same throughout the day. The HR was significantly decreased by training. Although the LF power and LF/HF ratio were significantly increased, the HF power was not changed by training. These results suggest that parasympathetic nervous activity may be fully activated even before training in this species.

Animals↗

PubMed1999

Influence of rough track surfaces on components of vertical forces in cantering thoroughbred horses.

No kinetic data are available to indicate the influence of a rough track surface and the resultant loss of footing on the components of hoof vertical forces. The aim of the present study was to evaluate the influence of rough track surfaces on the components of hoof vertical forces. Seven clinically sound 2-year-old Thoroughbred horses had instrumented shoes attached to both of the fore hooves to obtain the magnitudes of consecutive forces exerted in the hoof. All horses were cantered at approximately 10 m/s on straight tracks of smooth and rough surfaces of woodchip and dirt. The instrumented shoe indicated in real time 4 component forces acting on the hoof via 4 transducers positioned on the lateral and medial sides of the toe (LT, MT) and the heel (LH, MH). The peak vertical component forces of the heel were about twice that of the toe. The variances of the 4 components of peak vertical forces significantly increased on the rough track surface of both the woodchip (Leading: LH, MH and MT; Trailing: MH and LT) and dirt (Leading: LH and LT; Trailing: MH and MT) tracks. The trajectories of the position of the resultant force acting on the hoof also tended to increase the variance on rough track surfaces. Therefore, running on a rough track changes the vertical hoof forces and the balance of the resultant hoof forces. These results suggest a mechanism by which the rough track surfaces may influence the soundness of the horse.

Animals↗