Biomedical subjects

M Morikawa

Publications and source records attributed to M Morikawa.

At least 55 records · Page 3Linked to original sources

PubMed2001

Management of pregnancy with congenital antithrombin III deficiency: two case reports and a review of the literature.

Women with antithrombin (AT) III deficiency are prone to pregnancy-associated venous thromboembolism. We report 2 cases with genetically confirmed ATIII deficiency, one with a mutation in exon 3A and the other with an exon 4 deletion, in whom the pregnancies were successfully managed with prophylactic therapies for thrombosis. A 35-year-old pregnant woman was treated with intravenous infusions of ATIII concentrate alone, and the other 22-year-old pregnant woman was mainly treated with subcutaneous injections of heparin and oral low-dose aspirin therapy. Both pregnancies resulted in vaginal deliveries of healthy neonates. The literature concerning prophylactic therapies for thrombosis in ATIII deficiency-complicated pregnancy is reviewed, and the clinical problems, including the adverse effects of the therapies, are discussed.

Adult↗

PubMed2001

High NK cell activity in early pregnancy correlates with subsequent abortion with normal chromosomes in women with recurrent abortion.

PROBLEM: The aim of this study was to assess the role of natural killer (NK) cells in pregnant women with a history of recurrent spontaneous abortion (RSA). METHOD OF STUDY: Consecutive 66 pregnant women with a history of RSA were prospectively assessed for peripheral NK cell activity, percentage of the NK cell subsets, and subsequent pregnancy outcome. RESULTS: NK cell activity in women with subsequent live birth (group I) at 4-5 gestational weeks (GW) (mean +/- SD, 32.5 +/- 12.31%) significantly decreased at 6-7 GW (28.1 +/- 12.1%) and at 8 9 GW (28.0 +/- 11.8%). NK cell activity in women with subsequent abortion with normal chromosomes (group II) at 6 7 GW (41.2 +/- 19.0%) was significantly higher than that in group I women, while NK cell activity at 6-7 GW in women with subsequent abortion with abnormal chromosomes (group III) was the same as the level in group I women. CONCLUSIONS: High NK cell activity at 6-7 GW correlates with subsequent abortion with normal chromosomes.

Abortion, Habitual↗

PubMed2001

Active subtilisin-like protease from a hyperthermophilic archaeon in a form with a putative prosequence.

The gene encoding subtilisin-like protease T. kodakaraensis subtilisin was cloned from a hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. T. kodakaraensis subtilisin is a member of the subtilisin family and composed of 422 amino acid residues with a molecular weight of 43,783. It consists of a putative presequence, prosequence, and catalytic domain. Like bacterial subtilisins, T. kodakaraensis subtilisin was overproduced in Escherichia coli in a form with a putative prosequence in inclusion bodies, solubilized in the presence of 8 M urea, and refolded and converted to an active molecule. However, unlike bacterial subtilisins, in which the prosequence was removed from the catalytic domain by autoprocessing upon refolding, T. kodakaraensis subtilisin was refolded in a form with a putative prosequence. This refolded protein of recombinant T. kodakaraensis subtilisin which is composed of 398 amino acid residues (Gly(-82) to Gly(316)), was purified to give a single band on a sodium dodecyl sulfate (SDS)-polyacrylamide gel and characterized for biochemical and enzymatic properties. The good agreement of the molecular weights estimated by SDS-polyacrylamide gel electrophoresis (44,000) and gel filtration (40,000) suggests that T. kodakaraensis subtilisin exists in a monomeric form. T. kodakaraensis subtilisin hydrolyzed the synthetic substrate N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide only in the presence of the Ca(2+) ion with an optimal pH and temperature of pH 9.5 and 80 degrees C. Like bacterial subtilisins, it showed a broad substrate specificity, with a preference for aromatic or large nonpolar P1 substrate residues. However, it was much more stable than bacterial subtilisins against heat inactivation and lost activity with half-lives of >60 min at 80 degrees C, 20 min at 90 degrees C, and 7 min at 100 degrees C.

Amino Acid Sequence↗

PubMed2001

NK cell activity and subsets in women with a history of spontaneous abortion. Cause, number of abortions, and subsequent pregnancy outcome.

OBJECTIVE: The aim of this study was to assess the role of NK cells in nonpregnant women with a history of spontaneous abortion. STUDY DESIGN: 113 nonpregnant women with a history of spontaneous abortion were assessed for peripheral NK cell activity and percentage of NK cell subsets, in relation to the cause of abortions, the number of spontaneous abortions, and subsequent pregnancy outcome (n = 56). RESULTS: Neither NK cell activity nor subsets showed a significant difference in relation to the cause or number of spontaneous abortions. NK cell activity in nonpregnant women who later experienced subsequent abortion with normal chromosomes (n = 10) (mean +/- SD: 42.8 +/- 15.8%) was relatively higher than that in women with subsequent live birth (control, n = 39) (32.1 +/- 13.7%) (p = 0.099). NK cell activity in women who later experienced subsequent abortion with abnormal chromosomes (n = 7) (28.7 +/- 21.4%) was the same as the level in the control. CONCLUSION: Peripheral NK cell activity or subsets during nonpregnant status were not related to the cause or number of previous spontaneous abortions. A relation between preconceptional NK cell activity and later experiencing abortion with normal chromosomes should be further studied.

Abortion, Habitual↗

PubMed2001

Elevated plasma homocysteine levels and risk of silent brain infarction in elderly people.

BACKGROUND AND PURPOSE: Silent brain infarction (SBI) on MRI is common in elderly people, and recent studies have demonstrated that SBI increases the risk of progression to clinically apparent stroke and cognitive decline. Therefore, an early and accurate detection of SBI and a search for potential treatable risk factors may have a significant impact on public health. METHODS: Community-dwelling elderly people aged >/=66 years who participated in the present study (n=153) underwent brain MRI and standardized physical and neuropsychological examinations as well as blood biochemistry determinations, including total plasma homocysteine (pHcy), renal function, vitamin status, and polymorphisms of the methylenetetrahydrofolate reductase gene. RESULTS: SBI was found in 24.8% of the participants. In the univariate analysis, the pHcy levels in subjects with SBI (13.6+/-4.1 micromol/L) were significantly higher (P=0.0004) than those in subjects without SBI (11.0+/-3.3 micromol/L). When pHcy levels were stratified into high (>/=15.1 mmol/L), moderate (11.6 to 15.0 mmol/L), and low (</=11.5 mmol/L) groups, age (P<0.0001), male sex (P<0.0001), the habits of cigarette smoking (P<0.0001) and of alcohol consumption (P=0.0002), and folate levels (P=0.01) were significantly associated with an elevation of pHcy levels. The elevated pHcy levels were significantly associated with SBI after individual adjustment for age, sex, hypertension, renal function, and the habits of smoking and alcohol consumption. CONCLUSIONS: pHcy level is associated with age and nutritional and other lifestyle factors, and it contributes to a risk for SBI.

Age Distribution↗

PubMed2001

Gene Cloning of an alcohol dehydrogenase from thermophilic alkane-degrading Bacillus thermoleovorans B23.

The gene encoding an alcohol dehydrogenase (Bt-ADH) was cloned from a newly isolated thermophilic alkane-degrading Bacillus thermoleovorans, strain B23. The gene conferred 1-tetradecanol dehydrogenase activity on Escherichia coli cells. Bt-ADH is composed of 249 amino acid residues and the calculated molecular mass is 27,196 Da. A tyrosine residue in the active site and a glycine-rich sequence (GGXXGI/LG) constituting probable nicotinamide adenine dinucleotide (NAD+) or nicotinamide adenine dinucleotide phosphate (NADP+) binding site were completely conserved in the Bt-ADH sequence at positions 155 and 11, respectively. A phylogenetic analysis of Bt-ADH suggested that the enzyme belongs to the zinc-independent ADH Group II. Its highest similarity (48% identical) was to a hypothetical oxidoreductase from a hyperthermophile, Thermotoga maritima.

Journal Article↗

PubMed2001

Stabilities of chimeras of hyperthermophilic and mesophilic glycerol kinases constructed by DNA shuffling.

Glycerol kinases from Thermococcus kodakaraensis KOD1 (Tk-GK) and Escherichia coli (Ec-GK) greatly differ in thermostability. The temperature (T(1/2)) at which the enzymes lose half of their activity upon incubation for 20 min is 50-55 degrees C for Ec-GK and approximately 95 degrees C for Tk-GK. To examine whether the amino acid substitutions that make Tk-GK more stable than Ec-GK are localized in a limited region, the chimeras of two parental genes encoding Tk-GK and Ec-GK were constructed by DNA shuffling. E. coli cells were transformed with a plasmid library harboring these chimeras and screened for those tht produce chimeric enzymes which are more stable than Ec-GK. Four chimeric enzymes were isolated and purified, and their biochemical properties characterized. Replacement of 83 or 93 residues in the C-terminus of Ec-GK with the corresponding ones of Tk-GK increased the T(1/2) value of Ec-GK by 25-30 degrees C. In contrast, replacement of 85 residues in the N-terminus of Ec-GK with the corresponding ones of Tk-GK reduced the T(1/2) value by 5-10 degrees C. In addition, replacement of 10 residues in the C-terminus of Tk-GK with the corresponding ones of Ec-GK reduced the T(1/2) value ot Tk-GK by approximately 15 degrees C. Measurement of the far-UV CD spectra indicates that the three-dimensional structures of the chimeric enzymes, as well as those of the parent enzymes, are similar to one another. These results suggest that the amino acid substitutions responsible for the high stability of Tk-GK are largely localized in the C-terminal region.

Journal Article↗

PubMed2001

Isolation and characterization of long-chain-alkane degrading Bacillus thermoleovorans from deep subterranean petroleum reservoirs.

Two extremely thermophilic alkane-degrading bacterial strains, B23 and H41, were respectively isolated from deep subterranean petroleum reservoirs in the Minami-aga (Niigata) and Yabase (Akita) oil fields. Both strains were able to grow at temperatures ranging from 50 to 80 degrees C, with optimal growth at 70 degrees C for B23 and 65 degrees C for H41. From 16S rRNA gene sequence analysis and physiological characterization, both strains were identified as Bacillus thermoleovorans (identities of 99.5% and 99.6% to strain DSM 5366, and 98.3% and 98.7% to the type strain LEH-1(TS), respectively). Strains B23 and H41 effectively (60%) degraded n-alkanes longer than C12 and C15, respectively, at 70 degrees C, while strain LEH-1(TS) degraded undecane (C11) most effectively. When B23 and H41 were cultivated in the presence of heptadecane, heptadecanoate and pentadecanoate were specifically accumulated in the cells. These results strongly suggest that the two strains degraded n-alkanes by a terminal oxidation pathway, followed by a beta-oxidation pathway.

Journal Article↗

PubMed2001

[A case of cerebral arteriovenous malformation complicated with intracerebral hemorrhage after endovascular embolization].

We reported a case of cerebral arteriovenous malformation (AVM), complicated with intracerebral hemorrhage (ICH), after endovascular embolization. A 51-year-old male suffered from intraventricular hemorrhage due to a rupture of an intranidal aneurysm on October 4, 1999. The first embolization procedure for the aneurysm and a part of the nidus was performed with 2-hydroxyethyl methacrylate-methyl methacrylate (HEMA-MMA) and Liquid coil on day 21 after admission. On day 28, a second embolization was carried out for the residual nidus. Although most of the nidus was obliterated, the patient became comatose 10 hours after the second embolization. Computed tomography revealed a massive ICH in the right parietal lobe, and he underwent emergency evacuation of the hematoma. During the surgery, HEMA-MMA was seen in a draining vein. This caused venous stasis. Although the patient gradually improved postoperatively, he became comatose again because of a recurrence of ICH on day 36. Evacuation of the hematoma and removal of the nidus were performed again. The operative specimen showed AVM embolized by HEMA-MMA with non-specific inflammation and partial inflammatory degeneration of the vascular wall. Hemodynamic change such as venous stasis or elevated pressure of the feeding artery seemed to be the cause of the hemorrhage. Multi-staged embolization with longer intervals and intraoperative flow control were regarded as crucial for avoiding delayed hemorrhage.

Aneurysm, Ruptured↗

PubMed2001

[A case of spinal dural AVF treated by endovascular embolization].

Here we report a case of spinal dural arteriovenous fistula(AVF) treated by endovascular embolization. A 58-year-old female presented with progressive intermittent claudication and numbness of the lower extremities. MRI showed swelling of the spinal cord with intramedullary high signal intensity on T2-weighted image and intramedullary enhancement, suggested spinal cord myelopathy. Myelography demonstrated the dilated serpentine vessels in the subarachnoid space and focal filling defect. Angiography showed spinal dural AVF fed by bilateral lateral sacral artery. The draining vein was posterior spinal vein. Endovascular embolization using liquid material was performed under general anesthesia. The injection of glue included the distal feeding artery, the shunt itself and the initial part of draining vein. A complete cure was achieved, with a normal postoperative angiogram. MRI returned to normal with complete disappearance of T2 high signal, cord enlargement and enhancement by contrast medium. It was suggested that venous congestion induced the transient spinal ischemia, manifested as intermittent claudication. Endovascular embolization using liquid material was safe and quite effective for spinal dural AVF.

Central Nervous System Vascular Malformations↗

PubMed2001

[Long-term results of Rygg's monocusp ventricular outflow patch for the reconstruction of right ventricular outflow tract in tetralogy of Fallot].

Thirty surviving patients after corrective surgery for tetralogy of Fallot with right ventricular outflow tract reconstruction (RVOTR) using monocusp ventricular outflow patch (MVOP) were reviewed retrospectively to determine the long-term results. The age at operation ranged from 2 to 55 years with a mean of 19 years, and follow-up extended to 18.2 years (cumulative: 345.4 patient-year). There were 4 late deaths (1.2% per patient-year), and the cumulative survival rate was 85.3% at 18 years after the corrective surgery. Eight patients (2.3% per patient-year) required intracardiac reoperations mostly resulted from problems after RVOTR with MVOP, such as recurrent stenosis of right ventricular outflow tract (3 cases) or pulmonary valvular incompetence (4 cases). In addition, one patient underwent balloon angioplasty for the recurrent stenosis located in the distal end of MVOP. Freedom from surgical or catheter reintervention for the MVOP-related complication was 60.6% at 18 years after the corrective surgery. MVOP caused compression of the pulmonary artery at the distal end of the anastomosis and reoperation in a younger patients quite early after the corrective surgery. Like other transannular patches, tissue failure and degeneration of MVOP were inevitable, and resulted in severe pulmonary valvular incompetence that required the valve replacement in 4 patients (1.3% per patient-year). Freedom from pulmonary valve replacement was 71.2% at 18 years after the corrective surgery. As long-term results, our experiences emphasize the need for an innovative transannular patch that possesses significantly better long-term durability.

Adolescent↗

PubMed2000

A study on the structure-function relationship of lipopeptide biosurfactants.

Arthrofactin (AF) and surfactin (SF) are the most effective cyclic lipopeptide biosurfactants ever reported. Linear AF and linear SF were prepared by saponification of lactone ring. The oil displacement activities decreased to one third of their respective original values. When residues of both an aspartic acid and a glutamic acid of SF were methylated or amidated, the activity increased by 20%, although their water solubility was lost. When these amino acid residues were modified by aminomethane sulfonic acid, the activity was drastically decreased probably owing to charge repulsion and structural distortion inhibiting micelle formation. Both AF and SF expressed higher activity under alkaline conditions than acidic conditions. AF was more resistant to acidic conditions than SF and it kept high activity even under pH 0.5. Although SF drastically reduced its activity under acidic conditions, surfactin-Asp/Glu-amido ester and surfactin-Asp/Glu-methyl ester retained similar activities irrespective of the pH change. A couple of conformers of SF prepared by reverse-phase HPLC showed the same oil displacement activity but different surface tension-reducing activity. AF was produced as a series of different fatty acid chain lengths (from C8 to C12). Among them, AF with fatty acid chain length of C10, which was the main product of the strain, showed the highest activity.

Bacterial Proteins↗

PubMed2000

Catalysis by Escherichia coli ribonuclease HI is facilitated by a phosphate group of the substrate.

To investigate the role of the phosphate group 3' to the scissile phosphodiester bond of the substrate in the catalytic mechanism of Escherichia coli ribonuclease HI (RNase HI), we have used modified RNA-DNA hybrid substrates carrying a phosphorothioate substitution at this position or lacking this phosphate group for the cleavage reaction. Kinetic parameters of the H124A mutant enzyme, in which His(124) was substituted with Ala, as well as those of the wild-type RNase HI, were determined. Substitution of the pro-R(p)-oxygen of the phosphate group 3' to the scissile phosphodiester bond of the substrate with sulfur reduced the k(cat) value of the wild-type RNase HI by 6.9-fold and that of the H124A mutant enzyme by only 1. 9-fold. In contrast, substitution of the pro-S(p)-oxygen of the phosphate group at this position with sulfur had little effect on the k(cat) value of the wild-type and H124A mutant enzymes. The results obtained for the substrate lacking this phosphate group were consistent with those obtained for the substrates with the phosphorothioate substitutions. In addition, a severalfold increase in the K(m) value was observed by the substitution of the pro-R(p)-oxygen of the substrate with sulfur or by the substitution of His(124) of the enzyme with Ala, suggesting that a hydrogen bond is formed between the pro-R(p)-oxygen and His(124). These results allow us to propose that the pro-R(p)-oxygen contributes to orient His(124) to the best position for the catalytic function through the formation of a hydrogen bond.

Alanine↗

PubMed2000

Enhancement of the enzymatic activity of ribonuclease HI from Thermus thermophilus HB8 with a suppressor mutation method.

A genetic method for isolating a mutant enzyme of ribonuclease HI (RNase HI) from Thermus thermophilus HB8 with enhanced activity at moderate temperatures was developed. T. thermophilus RNase HI has an ability to complement the RNase H-dependent temperature-sensitive (ts) growth phenotype of Escherichia coli MIC3001. However, this complementation ability was greatly reduced by replacing Asp(134), which is one of the active site residues, with His, probably due to a reduction in the catalytic activity. Random mutagenesis of the gene encoding the resultant D134H enzyme, followed by screening for second-site revertants, allowed us to isolate three single mutations (Ala(12) --> Ser, Lys(75) --> Met, and Ala(77) --> Pro) that restore the normal complementation ability to the D134H enzyme. These mutations were individually or simultaneously introduced into the wild-type enzyme, and the kinetic parameters of the resultant mutant enzymes for the hydrolysis of a DNA-RNA-DNA/DNA substrate were determined at 30 degrees C. Each mutation increased the k(cat)/K(m) value of the wild-type enzyme by 2.1-4.8-fold. The effects of the mutations on the enzymatic activity were roughly cumulative, and the combination of these three mutations increased the k(cat)/K(m) value of the wild-type enzyme by 40-fold (5.5-fold in k(cat)). Measurement of thermal stability of the mutant enzymes with circular dichroism spectroscopy in the presence of 1 M guanidine hydrochloride and 1 mM dithiothreitol showed that the T(m) value of the triple mutant enzyme, in which all three mutations were combined, was comparable to that of the wild-type enzyme (75.0 vs 77.4 degrees C). These results demonstrate that the activity of a thermophilic enzyme can be improved without a cost of protein stability.

Amino Acid Substitution↗

PubMed2000

Identification of the histidine and aspartic acid residues essential for enzymatic activity of a family I.3 lipase by site-directed mutagenesis.

A lipase from Pseudomonas sp. MIS38 (PML) is a member of the lipase family I.3. We analyzed the roles of the five histidine residues (His(30), His(274), His(291), His(313), and His(365)) and five acidic amino acid residues (Glu(253), Asp(255), Asp(262), Asp(275), and Asp(290)), which are fully conserved in the amino acid sequences of family I.3 lipases, by site-directed mutagenesis. We showed that the mutation of His(313) or Asp(255) to Ala almost fully inactivated the enzyme, whereas the mutations of other residues to Ala did not seriously affect the enzymatic activity. Measurement of the far- and near-UV circular dichroism spectra suggests that inactivation by the mutation of His(313) or Asp(255) is not due to marked changes in the tertiary structure. We propose that His(313) and Asp(255), together with Ser(207), form a catalytic triad in PML.

Amino Acid Substitution↗

PubMed2000

Overproduction in Escherichia coli, purification and characterization of a family I.3 lipase from Pseudomonas sp. MIS38.

Determination of the nucleotide sequence of the gene encoding a lipase from Pseudomonas sp. MIS38 (PML) revealed that PML is a member of the lipase family I.3 and is composed of 617 amino acid residues with a calculated molecular weight of 64510. Recombinant PML (rPML) was overproduced in Escherichia coli in an insoluble form, solubilized in the presence of 8 M urea, purified in a urea-denatured form and refolded by removing urea in the presence of the Ca(2+) ion. Gel filtration chromatography suggests that this refolded protein is monomeric. rPML showed relatively broad substrate specificities and hydrolyzed glyceryl tributyrate and olive oil with comparable efficiencies. rPML was active only in the form of a holo-enzyme, in which at least 12 Ca(2+) ions bound. These Ca(2+) ions bound too tightly to be removed from the protein upon dialysis, but were removed from it upon EDTA treatment. The resultant apo-enzyme was fully active in the presence of 10 mM CaCl(2), but was inactive in the absence of the Ca(2+) ion. PML has a GXSXG motif, which is conserved in lipases/esterases and generally contains the active-site serine. The mutation of Ser(207) within this motif to Ala completely inactivated PML, suggesting that Ser(207) is the active-site serine of PML.

Amino Acid Sequence↗

PubMed2000

Blocking the CD28-B7 T-cell costimulatory pathway abrogates the development of obliterative bronchiolitis in a murine heterotopic airway model.

BACKGROUND: CTLA4IgG that binds to B7 effectively inhibits the signaling of CD28/B7 pathway and induces antigen-specific T-cell unresponsiveness in vitro and in vivo. We examined whether the development of obliterative bronchiolitis in a murine heterotopic airway transplantation model is T cell dependent and whether CTLA4IgG abrogates the development of obliterative bronchiolitis. METHODS: Tracheae with main bronchi from C3H/He (H2k), BALB/C (H2d), or C57BL/6 (H2b) mice were transplanted heterotopically into subcutaneous pockets on the backs of BALB/C or BALB/C nu/nu mice on day 0. Recipient mice were untreated or intraperitoneally treated with either CTLA4IgG or human IgG with different time and dose schedules. RESULTS: The development of obliterative bronchiolitis, which leads to luminal obliteration by fibrous tissue in a murine heterotopic airway transplantation model, was T cell dependent and the development of obliterative bronchiolitis was significantly abrogated by the CTLA4IgG treatment. However, the normal ciliated columnar respiratory epithelial cells in allografts were lost and replaced by flattened attenuated epithelial cells even after the CTLA4IgG treatment. We further demonstrated that CTLA4IgG treatment did not result in the induction of donor-specific unresponsiveness. CONCLUSIONS: We demonstrated that the development of obliterative bronchiolitis in a murine heterotopic airway model involves both CD28/B7-dependent and -independent processes. The luminal obliteration by fibrous tissue is clearly CD28/B7 dependent and can be inhibited by CTLA4IgG. The luminal obliteration of allografted trachea by fibrous tissues and the loss of ciliated columnar respiratory epithelial cells represent distinct disease processes.

Abatacept↗

PubMed2000

Loss as a determinant of PTSD in a cohort of adult survivors of the 1988 earthquake in Armenia: implications for policy.

OBJECTIVE: To study the relationship of post-traumatic stress disorder (PTSD) to severity of the disaster experience. METHOD: A sample of 1785 adult participants of an epidemiological study initiated in the immediate aftermath of the 1988 earthquake in Armenia were interviewed about 2 years following the disaster based on the NIMH DIS-Disaster Supplement. All 154 cases of pure PTSD were compared with 583 controls without symptoms satisfying psychiatric diagnoses of interest. RESULTS: PTSD cases included more persons from areas with the worst destruction. Having the highest level of education compared to lowest (OR 0.6 [95% CI 0.4-0.9]), being accompanied at the moment of the earthquake (OR 0.6 [95% CI 0.4-0.9]) and making new friends after the earthquake (OR 0.6 [95% CI 0.5-0.8]) were protective for PTSD. PTSD risk increased with the total amount of loss to the family (OR for highest level of loss 4.1 [95% CI 2.3-7.5]). CONCLUSION: Based on this large population sample, we believe that early support to survivors with high levels of loss may reduce PTSD following earthquakes.

Adolescent↗