Biomedical subjects

K Komori

Publications and source records attributed to K Komori.

At least 55 records · Page 3Linked to original sources

PubMed2001

Membrane-type 5 matrix metalloproteinase is expressed in differentiated neurons and regulates axonal growth.

Expression of membrane-type (MT) 5 matrix metalloproteinase (MMP) in the mouse brain was examined. MT5-MMP was expressed in the cerebrum in embryos, but it declined after birth. In contrast, expression in the cerebellum started to increase postnatally and continued thereafter. The cells expressing MT5-MMP were postmitotic neurons that showed gelatinolytic activities. Specific expression of MT5-MMP was observed in the neurons but not in the glial cells when embryonal mouse carcinoma P19 cells were differentiated in vitro by retinoic acid treatment. Neurons isolated from dorsal root ganglia also expressed MT5-MMP, and it was localized at the edge of growth cone. Proteoglycans inhibit neurite extension and regulate synaptogenesis. The inhibitory effect of the proteoglycans on neurite extension of dorsal root ganglia neurons was effectively eliminated by recombinant MT5-MMP. Thus, MT5-MMP expressed in neurons may play a role in axonal growth that contributes to the regulation of neural network formation.

Animals↗

PubMed2000

Mutational analysis of the Pyrococcus furiosus holliday junction resolvase hjc revealed functionally important residues for dimer formation, junction DNA binding, and cleavage activities.

The Holliday junction cleavage protein, Hjc resolvase of Pyrococcus furiosus, is the first Holliday junction resolvase to be discovered in Archaea. Although the archaeal resolvase shares certain biochemical properties with other non-archaeal junction resolvases, no amino acid sequence similarity has been identified. To investigate the structure-function relationship of this new Holliday junction resolvase, we constructed a series of mutant hjc genes using site-directed mutagenesis targeted at the residues conserved among the archaeal orthologs. The products of these mutant genes were purified to homogeneity. With analysis of the activity of the mutant proteins to bind and cleave synthetic Holliday junctions, one acidic residue, Glu-9, and two basic residues, Arg-10 and Arg-25, were found to play critical roles in enzyme action. This is in addition to the three conserved residues, Asp-33, Glu-46, and Lys-48, which are also conserved in the motif found in the type II restriction endonuclease family proteins. Two aromatic residues, Phe-68 and Phe-72, are important for the formation of the homodimer probably through hydrophobic interactions. The results of these studies have provided insights into the structure-function relationships of the archaeal Holliday junction resolvase as well as the universality and diversity of the Holliday junction cleavage reaction.

Amino Acid Sequence↗

PubMed2000

Hjc resolvase is a distantly related member of the type II restriction endonuclease family.

Hjc resolvase is an archaeal enzyme involved in homologous DNA recombination at the Holliday junction intermediate. However, the structure and the catalytic mechanism of the enzyme have not yet been identified. We performed database searching using the amino acid sequence of the enzyme from Pyrococcus furiosus as a query. We detected 59 amino acid sequences showing weak but significant sequence similarity to the Hjc resolvase. The detected sequences included DPN:II, HAE:II and Vsr endonuclease, which belong to the type II restriction endonuclease family. In addition, a highly conserved region was identified from a multiple alignment of the detected sequences, which was similar to an active site of the type II restriction endonucleases. We substituted three conserved amino acid residues in the highly conserved region of the Hjc resolvase with Ala residues. The amino acid replacements inactivated the enzyme. The experimental study, together with the results of the database searching, suggests that the Hjc resolvase is a distantly related member of the type II restriction endonuclease family. In addition, the results of our database searches suggested that the members of the RecB domain superfamily are evolutionarily related to the type II restriction endonuclease family.

Amino Acid Sequence↗

PubMed2000

Biochemical characterization of the hjc holliday junction resolvase of Pyrococcus furiosus.

The Hjc protein of Pyrococcus furiosus is an endonuclease that resolves Holliday junctions, the intermediates in homologous recombination. The amino acid sequence of Hjc is conserved in Archaea, however, it is not similar to any of the well-characterized Holliday junction resolvases. In order to investigate the similarity and diversity of the enzymatic properties of Hjc as a Holliday junction resolvase, highly purified Hjc produced in recombinant Escherichia coli was used for detailed biochemical characterizations. Hjc has specific binding activity to the Holliday-structured DNA, with an apparent dissociation constant (K:(d)) of 60 nM. The dimeric form of Hjc binds to the substrate DNA. The optimal reaction conditions were determined using a synthetic Holliday junction as substrate. Hjc required a divalent cation for cleavage activity and Mg(2+) at 5-10 mM was optimal. Mn(2+) could substitute for Mg(2+), but it was much less efficient than Mg(2+) as the cofactor. The cleavage reaction was stimulated by alkaline pH and KCl at approximately 200 mM. In addition to the high specific activity, Hjc was found to be extremely heat stable. In contrast to the case of SULFOLOBUS:, the Holliday junction resolving activity detected in P. furiosus cell extract thus far is only derived from Hjc.

Archaeal Proteins↗

PubMed2000

Domain analysis of an archaeal RadA protein for the strand exchange activity.

Archaeal RadA, like eukaryotic Rad51 and bacterial RecA, promotes strand exchange between DNA strands with homologous sequences in vitro and is believed to participate in the homologous recombination in cells. The amino acid sequences of the archaeal RadA proteins are more similar to the eukaryotic Rad51s rather than the bacterial RecAs, and the N-terminal region containing domain I is conserved among the RadA and Rad51 proteins but is absent from RecA. To understand the structure-function relationship of RadA, we divided the RadA protein from Pyrococcus furiosus into two parts, the N-terminal one-third (RadA-n) and the residual C-terminal two-thirds (RadA-c), the latter of which contains the central core domain (domain II) of the RecA/Rad51 family proteins. RadA-c had the DNA-dependent ATPase activity and the strand exchange activity by itself, although much weaker (10%) than that of the intact RadA. These activities of RadA-c were restored to 60% of those of RadA by addition of RadA-n, indicating that the proper active structure of RadA was reconstituted in vitro. These results suggest that the basic activities of the RecA/Rad51 family proteins for homologous recombination are derived from domain II, and the N-terminal region may help to enhance the catalytic efficiencies.

Adenosine Triphosphatases↗

PubMed2000

Both RadA and RadB are involved in homologous recombination in Pyrococcus furiosus.

RecA and Rad51 proteins are essential for homologous recombination in Bacteria and Eukarya, respectively. Homologous proteins, called RadA, have been described for Archaea. Here we present the characterization of two RecA/Rad51 family proteins, RadA and RadB, from Pyrococcus furiosus. The radA and radB genes were not induced by DNA damage resulting from exposure of the cells to gamma and UV irradiation and heat shock, suggesting that they might be constitutively expressed in this hyperthermophile. RadA had DNA-dependent ATPase, D-loop formation, and strand exchange activities. In contrast, RadB had a very weak ATPase activity that is not stimulated by DNA. This protein had a strong binding affinity for DNA, but little strand exchange activity could be detected. A direct interaction between RadA and RadB was detected by an immunoprecipitation assay. Moreover, RadB, but not RadA, coprecipitated with Hjc, a Holliday junction resolvase found in P. furiosus, in the absence of ATP. This interaction was suppressed in the presence of ATP. The Holliday junction cleavage activity of Hjc was inhibited by RadB in the absence, but not in the presence, of ATP. These results suggest that RadB has important roles in homologous recombination in Archaea and may regulate the cleavage reactions of the branch-structured DNA.

Adenosine Triphosphatases↗

PubMed2000

Crystal structure of an archaeal intein-encoded homing endonuclease PI-PfuI.

Inteins possess two different enzymatic activities, self-catalyzed protein splicing and site-specific DNA cleavage. These endonucleases, which are classified as part of the homing endonuclease family, initiate the mobility of their genetic elements into homologous alleles. They recognize long asymmetric nucleotide sequences and cleave both DNA strands in a monomer form. We present here the 2.1 A crystal structure of the archaeal PI-PfuI intein from Pyroccocus furiosus. The structure reveals a unique domain, designated here as the Stirrup domain, which is inserted between the Hint domain and an endonuclease domain. The horseshoe-shaped Hint domain contains a catalytic center for protein splicing, which involves both N and C-terminal residues. The endonuclease domain, which is inserted into the Hint domain, consists of two copies of substructure related by an internal pseudo 2-fold axis. In contrast with the I-CreI homing endonuclease, PI-PfuI possibly has two asymmetric catalytic sites at the center of a putative DNA-binding cleft formed by a pair of four-stranded beta-sheets. DNase I footprinting experiments showed that PI-PfuI covers more than 30 bp of the substrate asymmetrically across the cleavage site. A docking model of the DNA-enzyme complex suggests that the endonuclease domain covers the 20 bp DNA duplex encompassing the cleavage site, whereas the Stirrup domain could make an additional contact with another upstream 10 bp region. For the double-strand break, the two strands in the DNA duplex were cleaved by PI-PfuI with different efficiencies. We suggest that the cleavage of each strand is catalyzed by each of the two non-equivalent active sites.

Amino Acid Sequence↗

PubMed2000

Interrater reliability of the short-memory questionnaire in a variety of health professional representatives.

A study was performed to assess interrater reliability of the Japanese version of the Short-Memory Questionnaire (SMQ), which is an easily-administered, informant-based scale of cognitive function. The subjects were 18 consecutive patients with Alzheimer's disease who were outpatients of Department of Neuropsychiatry in Ehime University School of Medicine and their principal caregivers. One neuropsychiatrist (NP) administered the SMQ, and all sessions were videotaped. Then one nurse (Ns), one clinical psychologist (CP), one occupational therapist (OT), and one neurologist (NL) from another institution viewed the videotape and performed reassessments independently. Interrater reliability between the NP and Ns, CP, OT, or NL were all extremely good. Interrater reliability between the Ns and CP, between the Ns and OT, between the Ns and NL, between the CP and OT, between the CP and NL, and between the OT and NL were also extremely good. The SMQ is a convenient, quantitative scale, and in this study it showed good interrater reliability between personnel from different fields. Therefore, it is a very useful test for everyday medical consultations and for clinical research.

Aged↗

PubMed2000

Validity of the Short-Memory Questionnaire in vascular dementia.

PURPOSE: To determine whether the Short-Memory Questionnaire (SMQ) being administered by caregivers to patients with Alzheimer's disease (AD) is also valid when given to patients with vascular dementia (VaD). METHODS: Subjects were 58 patients with VaD, 26 patients with cerebrovascular disorders free of cognitive deficit (CVD) and 62 healthy controls. All subjects received the Mini-Mental State Examination (MMSE), and their primary caregivers (or family members with same household) received the SMQ. RESULTS: In the VaD patients, the SMQ score was highly correlated with the MMSE score. When 39/40 was defined as a cutoff point based on the results of previous study, the SMQ properly classified 55 of the 58 VaD patients and 61 of the 62 controls, but only about half of the 26 CVD patients, as cases. CONCLUSION: The SMQ, a simple quantitative rating test for memory disturbance, is useful for the assessment and screening of VaD patients as well as AD patients, although careful attention should be paid to the assessment of CVD patients.

Aged↗

PubMed2000

Increased plasma fibrinogen level and future risk of coronary artery disease after repair of abdominal aortic aneurysm.

BACKGROUND: Cardiovascular disease such as coronary artery disease is a major cause of late death after repair of abdominal aortic aneurysm (AAA). But risk factors are not well known. So, we investigated the incidence of cardiovascular events after surgery and examined the prognostic factors. STUDY DESIGN: We retrospectively reviewed 270 patients who underwent elective surgery for AAA from 1985 to 1995. Kaplan-Meier survival analysis was used to estimate survival rates and the probability of coronary, cerebrovascular, and cardiovascular events. The risk factors for each endpoint were investigated using multivariate analysis. RESULTS: The overall survival rate was 87.3% at 3 years, 76.4% at 5 years, and 52.3% at 10 years. Current cigarette use, renal insufficiency, advanced age (> or = 70 years old), and higher plasma fibrinogen level (> or = 300 mg/dL) were significant factors influencing survival. The probability of a coronary event was 4.9% at 3 years, 7.1% at 5 years, and 20.7% at 10 years. Plasma fibrinogen level and cerebrovascular disease were significant prognostic factors for coronary events. The probability of a cerebrovascular event was 5.3% at 3 years, 7.6% at 5 years, and 18.0% at 10 years. No significant prognostic factors for cerebrovascular events existed. The probability of a cardiovascular event was 10.3% at 3 years, 14.9% at 5 years, and 33.6% at 10 years. Plasma fibrinogen level was a significant risk factor for cardiovascular events. But the presence of coronary artery disease did not affect survival or the incidence of coronary, cerebrovascular, or cardiovascular events. CONCLUSIONS: Plasma fibrinogen level is an independent risk factor of future coronary events after surgery for AAA, and the increased risk of coronary artery events contributes to the impaired survival. Patients with higher plasma fibrinogen level need careful surveillance for cardiovascular disease after surgery.

Adult↗

PubMed2000

Low novelty-seeking differentiates obsessive-compulsive disorder from major depression.

OBJECTIVE: To make a direct comparison of patients with obsessive-compulsive disorder (OCD) and major depression (MD) and a normal control group in terms of the Temperament and Character Inventory (TCI) personality dimensions. METHOD: Additionally to 43 patients with primary OCD, 43 MD patients and 43 normal subjects who were matched against the OCD patients for sex and age filled out the TCI. RESULTS: Compared to the controls, the OCD and MD patients scored significantly higher on harm avoidance and significantly lower on self-directedness and co-operativeness. The OCD patients scored significantly lower on novelty-seeking than the MD patients and the controls. CONCLUSION: Whereas OCD and MD share similar personality deviations on harm avoidance, self-directedness and co-operativeness, OCD is distinguishable from MD in terms of low novelty-seeking. Low novelty-seeking may have a profound relationship to the specific aetiology of OCD.

Adolescent↗

PubMed2000

Expression of topoisomerase II alpha, Ki-67 and p53 in early stage laryngeal carcinomas not featuring vocal cord fixation.

AIM: To determine whether topoisomerase II alpha (topoIIa) expression is an additional prognostic marker for less advanced stage laryngeal cancers first treated without surgery. Ki-67 and p53 protein levels were also assessed for comparison. EXPERIMENTAL DESIGN: Formalin-fixed, paraffin-embedded tumor material from 63 cases of squamous cell carcinoma (SCC) of the larynx (glottis, stages 0,1,2) was immunohistochemically stained for topoIIa, Ki-67 (MIB-1) and p53 (DO-7) and the results were compared with clinicopathologic findings. RESULTS: There were 7 stage 0 (TisN0M0), 33 stage I (T1N0M0), and 23 stage II (T2N0M0) SCCs with the TNM classification. Significant differences between carcinomas and normal mucosa were found for the topoIIa-LI, Ki-67-LI, and topoIIa-to-Ki-67 ratio. Regarding histologic grade, a significant difference in topoIIa-to-Ki-67 ratio was evident between well or moderately and poorly differentiated lesions. There were 19 cases of recurrence and 44 cases of nonrecurrence, but no significant differences were found for either of the indices or their ratio. No significant variation with p53 positivity was evident with reference to histologic differentiation, T-factor, clinical course, or proliferation. CONCLUSIONS: The results demonstrate that the topoIIa-to-Ki-67 ratio is a more sensitive parameter reflecting proliferation, for histologic grading of less advanced laryngeal SCCs, than topoIIa- or Ki-67-LIs.

Adult↗

PubMed2000

Modulation of platelet aggregation after percutaneous transluminal angioplasty of the iliac artery for atherosclerosis obliterans.

BACKGROUND: Platelet aggregation is modulated by blood flow. We investigated whether platelet function is altered during percutaneous transluminal balloon angioplasty in patients with atherosclerosis obliterans. METHODS: Blood samples were obtained from the iliac artery in 9 lower limbs of 7 patients undergoing percutaneous balloon angioplasty of the iliac artery. An agonists-induced platelet aggregation test was performed with an aggregometer. Femoral blood flow was measured with a Doppler velocimeter before and after the procedure. RESULTS: Before dilatation, the maximum platelet aggregation rates (+/- SEM) induced by adenosine phosphate, epinephrine, and arachidonic acid were 54.7% +/- 5.8%, 64.8% +/- 4.3%, and 60.5% +/- 6.1%, respectively. After angioplasty, these values reduced to 36.7% +/- 4.1%, 36.1% +/- 8.6%, and 40.1% +/- 5.0%, respectively (P < .05). The pre-procedural ankle-brachial pressure index, mean flow rate, mean velocity, and shear stress variation were 0.63 +/- 0.1, 218.1 +/- 32.1 mL/min, 9.4 +/- 1.1 cm/sec, and 60.6 +/- 17.7 dyne/cm2, respectively. The mean velocity at the stenotic lesion was 215.1 +/- 83.9 cm/sec, which was significantly greater than those of the distal artery or after angioplasty (P < .01). Both ankle-brachial pressure index and shear stress variation increased after angioplasty to 0.99 +/- 0.07 (P < .05) and 139.8 +/- 17.0 (P < .05) dyne/cm2, but the mean flow rate and the mean velocity (198.3 +/- 24.5 mL/min and 8.8 +/- 1.2 cm/sec after angioplasty) did not change significantly. CONCLUSIONS: These results indicate that activated platelet function at a stenosed artery was decreased after angioplasty, possibly because of normalized blood flow with reduction of stenotic lesion.

Aged↗

PubMed2000

Functional interdependence of DNA polymerizing and 3'-->5' exonucleolytic activities in Pyrococcus furiosus DNA polymerase I.

Pyrococcus furiosus DNA polymerase I (Pol BI) belongs to the family B (alpha-like) DNA polymerases and has a strong 3'-->5' exonucleolytic activity, in addition to its DNA polymerizing activity. To understand the relationship between the structure and function of this DNA polymerase, three deletion mutants, Delta1 (DeltaLeu746-Ser775), Delta2 (DeltaLeu717-Ser775) and Delta3 (DeltaHis672-Ser775), and two substituted mutants of Asp405, D405A and D405E, were constructed. These substitutions affected both the DNA polymerizing and the 3'-->5' exonucleolytic activities. The Delta1 mutant protein had DNA polymerizing activity with higher specific activity than that of the wild-type Pol BI, but retained only 10% of the exonucleolytic activity of the wild-type. The other two deletion mutants lost most of both activities. These results suggest that the DNA polymerizing and exonucleolytic activities are closely related to each other in the folded structure of this DNA polymerase, as proposed in the family B DNA polymerases.

DNA Polymerase I↗

PubMed2000

Inhibition of 12(S)-hydroxyeicosatetraenoic acid (12-HETE) production suppressed the intimal hyperplasia caused by poor-runoff conditions in the rabbit autologous vein grafts.

The efficacy of OPC-29030, a newly developed inhibitor of 12(S)-hydroxyeicosatetraenoic acid (12-HETE) production, was evaluated on intimal hyperplasia of experimental autologous vein grafts in a distal poor-runoff model and a hyperlipidemic model in rabbits. First, rabbits were divided into two groups, the distal poor-runoff group (PR group) and the hyperlipidemic group (HL group). After 4 weeks preparing the PR model and the HL model, the femoral vein was implanted into the ipsilateral femoral artery. Then they were subdivided into two groups, depending on the diet provided; diet group with 0.1% OPC-29030 (OPC-29030 group) and normal diet group (control group). At 4 weeks, the grafts were harvested, and intimal hyperplasia of the graft was measured with an ocular cytometer. Intimal cell proliferation was determined by bromodeoxyuridine (BrdU) incorporation at 2 weeks after surgery. In addition, the effect of OPC-29030 on the proliferation or migration of rat aortic smooth muscle cells in culture was investigated. In the in vivo study in the PR group, the intimal hyperplasia and the plasma 12-HETE levels in the OPC-29030 group were significantly inhibited, compared with those of the control group. However, in the HL group, the intimal hyperplasia in both the OPC-29030 and control groups showed a remarkable degree of intimal hyperplasia. There was no significant difference between those two groups. Furthermore, there was no significant difference in the plasma 12-HETE levels in the HL group irrespective of the presence of OPC-29030. The BrdU labeling index at 2 weeks after grafting was significantly lower in the OPC-29030 group compared with that in the control group in the PR group. In the in vitro study, OPC-29030 did not inhibit smooth muscle cell proliferation; however, OPC-29030 inhibited the migration. These results demonstrate the efficacy of OPC-29030 in reducing the degree of intimal hyperplasia under PR conditions, but not under hyperlipidemic conditions. The mechanism of reducing the intimal hyperplasia may be that OPC-29030 inhibited 12-HETE production, which did not inhibit proliferation while inhibiting migration of the smooth muscle cell. These results suggested the possible involvement of 12-HETE with the intimal hyperplasia under PR conditions.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗

PubMed2000

Amino acid regulation of gene transcription of rat insulin-like growth factor-binding protein-1.

To investigate the molecular mechanisms of increased transcription of the insulin-like growth factor-binding protein-1 (IGFBP-1) gene in dietary protein-deprived animals, the cis-acting sequence that is involved in this regulation was analyzed. We first showed that IGFBP-1 gene transcription was up-regulated by amino acid deprivation in cultured liver cell lines: H4IIE and HuH-7. Since HuH-7 cells showed a greater increase in IGFBP-1 mRNA in response to amino acid deprivation, this cell line was used in further experiments. Using a promoter function assay, we found that up-regulation of promoter activity responding to amino acid deprivation was abolished by deleting the region between -112 and -81 bp from the cap site from the gene construct. This cis-acting region includes the insulin-responsive element (IRE) and glucocorticoid responsive element (GRE) of IGFBP-1. In summary, the present observation suggests that the 32-bp (-112 to -81) in the IGFBP-1 gene 5' promoter region is involved in the induction of the IGFBP-1 gene in response to amino acid deprivation.

Amino Acids↗

PubMed2000

[Color Doppler ultrasonography in the diagnosis of the acute scrotum].

PURPOSE: To determine the value of color Doppler ultrasonography (CDUS) in the diagnosis of acute scrotum. MATERIALS AND METHODS: 10 patients referred to our hospital with acute scrotal pain were included in this study. All patients were evaluated with CDUS after the initial clinical examinations. Blood flow of the involved testis was compared semiquantitatively to that of the opposite testis. Patients with a diagnosis of testicular torsion by CDUS underwent surgical exploration. Patients with CDUS diagnosis of epididymitis were treated with intravenous antibiotics. RESULTS: Of the 10 patients evaluated, CDUS diagnosed 5 patients with testicular torsion and 5 patients with epididymitis. All cases of torsion were confirmed intraoperatively. 2 cases with no intratesticular blood flow on CDUS had necrotic testes and underwent orchiectomy with orchiopexy of the contralateral testes. A case with absent flow and 2 cases with decreased flow had bilateral orchiopexy. CDUS findings of normal or increased flow were present in all patients with epididymitis. No cases of testicular atrophy were encountered on long-term follow up in patients with epididymitis. CONCLUSION: CDUS is helpful in detecting the perfusion of the testis as well as in getting anatomical information. CDUS is a very useful device which causes a minimal burden to the patient with acute scrotum. In most cases it will rapidly provides us the correct information although it may not be regarded as the definitive adjunct.

Acute Disease↗

PubMed2000

[Giant bladder stone: a case report].

An 88-year-old woman was hospitalized with the chief complaints of lower abdominal pain, miction pain and pollakisuria. Radiographs showed a giant bladder stone shadow, 12.0 x 9.0 cm in size. Cystolithotomy was performed under the diagnosis of bladder stone. The extirpated stone weighed 510 g, and measured 10.0 x 7.5 x 6.0 cm in size. The stone had four compartments of stratified lamellae composed of calcium phosphate and magnesium ammonium phosphate. The postoperative course was uneventful and the bilateral hydronephrosis improved markedly on DIP.

Aged↗