Variations of volatile chlorinated hydrocarbons in ambient air at industrial areas in Niigata.
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Air Pollutants↗
Biomedical subjects
H Mukai
Publications and source records attributed to H Mukai.
Annual variation of insecticides in precipitation in rural Japan.
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Gas Chromatography-Mass Spectrometry↗
Protein interactions with particulate Teflon: implications for the foreign body response.
PURPOSE: This study examined the nature of protein interactions with particulate polytetrafluoroethylene (PTFE, Teflon) to elucidate possible mechanisms involved in the foreign body response directed against failed Proplast/Teflon implants. MATERIALS AND METHODS: Fifty milligrams PTFE prepared to particle sizes ranging from < 32 microns to > 300 microns was incubated with newborn bovine serum. The total amount of protein adsorbed to the PTFE particles was determined using a standard colorimetric assay. The structural and functional integrity of the proteins adsorbed to PTFE was also examined. For these studies, xanthine oxidase was substituted for serum, and the enzymatic activity of xanthine oxidase adsorbed to PTFE was determined. Finally, primary interactions between protein and PTFE particles were assessed in experiments using water, 2 or 8 mol/L urea, 1 mol/L Nacl, or 1% sodium dodecyl sulfate in an attempt to dissociate bound protein from the surfaces of PTFE particles. RESULTS: Serum proteins bind almost instantly to the surface of PTFE particles. The effective surface area of PTFE increases dramatically with reduction of the material to small particles, as does the total amount of protein adsorbed by the particulate PTFE. Proteins bind to PTFE principally by hydrophobic interactions, and their three-dimensional structure is significantly perturbed by this interaction. In the case of xanthine oxidase, adsorption to PTFE distorts protein structure to the extent that biologic activity is eliminated. CONCLUSIONS: The amount of serum protein adsorbed to PTFE particles varies inversely with particle size for a constant mass of material. It is believed that the foreign body response directed against this material is related to the amount and relative distortion of proteins adsorbed to its surface. If so, it appears that reduction of an implant to small particles (typically 50 micron or less) will dramatically increase the biologic signal to local cell populations. Thus, the severity of the biologic response to PTFE debris may be dependent largely on the size of the debris particles.
Adsorption↗
Further identification of neurokinin receptor types and mechanisms of calcium signaling evoked by neurokinins in the murine neuroblastoma C1300 cell line.
It has been suggested that murine neuroblastoma C1300 cells express endogenous neurokinin NK2 receptors with features that differ from those of NK2 receptors characterized in other systems. In this study, we have further characterized the neurokinin receptor types present in this cell line. RNA blots showed that mRNAs of NK2 and NK3 receptors, but not of NK1 receptors, were expressed in C1300 cells. The increase in the cytosolic calcium concentration ([Ca2+]i) induced by 0.33 microM neurokinin A was completely inhibited by SR 48968, an NK2 receptor antagonist, whereas the partial response to 0.33 microM neurokinin B was unaffected, and the response was completely inhibited by SR 142801, and NK3 receptor antagonist. In addition, the [Ca2+]i increase by 0.33 microM senktide, an NK3 receptor agonist, was inhibited by SR 142801 but not by SR 48968. These findings indicated that C1300 cells endogenously express functional NK2 and NK3 receptors. It was also demonstrated that NK2 and NK3 receptors can be activated independently by 3.3 microM neurokinin A in the presence of 1.0 microM SR 142801 or 1.0 microM senktide, respectively. Therefore, the mechanisms of Ca2+ signaling mediated by endogenous NK2 and NK3 receptors were investigated. The independent activation of NK2 or NK3 receptors induced not only the [Ca2+]i increase, but also stimulated the formation of inositol trisphosphates; both these responses were inhibited by U73122, a phospholipase C (PLC) inhibitor. In addition, NK2 and NK3 receptor-mediated [Ca2+]i increase was partially attenuated in the absence of extracellular Ca2+ or in the presence of nickel, an inorganic Ca2+ influx blocker, but was unaffected by nifedipine and omega-conotoxin, L- and N-type voltage-dependent Ca2+ channel blockers, respectively. Furthermore, the depolarization by 60 mM K+ did not affect the [Ca2+]i. These findings suggested that the NK2 and NK3 receptor-mediated [Ca2+]i increase was due to the activation of PLC and was dependent on the mobilization of internal Ca2+ and the entry of extracellular Ca2+ through voltage-independent channels. This study showed that the C1300 cell line is a useful system with which to investigate pharmacological functions and signaling pathways of endogenous NK2 and NK3 receptors.
Animals↗
Bradykinin B2 receptor antagonist increases chloride and water absorption in rat medullary collecting duct.
This study tested the hypothesis that intrarenal kinins play a regulatory role in electrolyte excretion by altering Cl- absorption in the collecting duct. We measured Cl- and insulin concentrations in tubular fluid samples obtained from medullary collecting ducts (MCD) of Dahl/Rapp salt-resistant (SR/ Jr) rats by microcatheterization of ducts of Bellini before and after treatment with the bradykinin receptor antagonist HOE-140. Tubular fluid was obtained from paired terminal inner medullary (t-IMCD) and outer medullary (OMCD) collecting duct sites of the left kidney. HOE-140 (n = 7) or vehicle (n = 5) was infused intravenously, and the collections were repeated. HOE-140 did not alter glomerular filtration rate but decreased urine flow rate (P < 0.05) and absolute and fractional Cl- excretion (P < 0.01). HOE-140 did not alter the fraction of filtered Cl- delivered (FDCl) to the OMCD but decreased FDCl to the t-IMCD from 2.3 +/- 0.3 to 1.3 +/- 0.3% (P < 0.05). The fraction of filtered Cl- absorbed per millimeter between the collection sites was increased from 0.2 +/- 0.1 to 0.6 +/- 0.1% (P < 0.05). Fractional absorption of water along the MCD was also increased (P < 0.05). No changes in excretory function or tubular Cl- or water absorption were observed in vehicle-treated rats. These studies show that kinin B2 receptor blockade enhances Cl- and water absorption in the MCD, a finding that supports a role of renal kinins in the regulation of NaCl and water excretion.
Absorption↗
Selective slow pathway ablation in atrioventricular nodal reentrant tachycardia--comparison of different methods and the site of slow pathway ablation.
The optimum potential of the slow pathway (SP) was investigated by determining the effectiveness and safety of high-radiofrequency catheter ablation to treat atrioventricular nodal reentrant tachycardia (AVNRT). The subjects consisted of 29 patients with AVNRT (11 men, with a mean age of 54 +/- 15 years). Three ablation methods were used: a) Method A used the earliest atrial activation site, which is retrograde to the slow pathway, b) Method SP used the SP potential, and c) Method SW, in which ablation was performed stepwise starting from the coronary sinus and moving toward the recording site of the His bundle potential. Five, 20, and 4 patients underwent Methods A, SP, and SW, respectively. The fewest number of applications was needed with Method SP (11 +/- 9, 6 +/- 4, and 13 +/- 9), and the delivered energy was also lowest with Method SP (9151 +/- 6119, 3712 +/- 2168, and 12183 +/- 4090 J, with Methods A, SP, and SW, respectively). In Method SP, the interval between the atrium and SP was significantly longer at sites which cured tachycardia, than at sites at which ablation was ineffective (88 +/- 26 vs 66 +/- 22 msec, p < 0.05). The SP potential showed a humped shape in 18 of 20 patients. Method SP was the most efficient ablation method for treating AVNRT.
Adult↗
[Reaction conditions of PCR].
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DNA Primers↗
[Hot start method].
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Polymerase Chain Reaction↗
[Hemorrhagic fever renal syndrome-associated virus].
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Animals↗
[Results of radiotherapy combined with CDDP (BAI) plus VP-16 in the treatment of inoperable non-small cell lung cancer].
Twenty-six patients with unresectable non-small cell lung cancer were treated by chemotherapy alone or chemotherapy followed by radiotherapy from June 1988 to May 1990. CDDP (80 mg/m2) was given by bronchial arterial infusion (BAI) on day 1 in each course, VP-16 (80 mg/body) was given by drip intravenous infusion on days 1 and 2, and VP-16 (150 mg/body) was given orally on days 3 and 5. The patients were irradiated after this regimen with a single dose of 1.8-2.0 Gy, five times each week, to a volume which encompassed only the primary tumor. A total dose of 60 Gy or more was delivered. There were 10 squamous cell carcinomas, nine adenocarcinomas, five large cell carcinomas, one adenosquamous cell carcinoma and one poorly differentiated carcinoma. Median survival time (MST) was 354 days in stage III B patients and 280 days in stage IV patients. MST was 155 days in adenocarcinoma and 310 days in squamous cell carcinoma. MST was 372 days in PS 0 to 1 patients and 140 days in PS 2 patients. With chemo-radiotherapy in 17 cases resulted in no complete remissions and 10 partial remissions. The overall response rate was 58.8%. It is concluded that CDDP via BAI combined with VP-16 administration and sequential irradiation is useful to acquire a high response rate in non-small cell lung cancer. However, further studies on this modality are required.
Aged↗
[Appearance of cytoplasmic processes of megakaryocytes in the peripheral blood in a Munchausen syndrome with factitious anemia].
Many cytoplasmic processes of megakaryocytes were seen in a 45-year-old male patient of Munchausen syndrome with sustained severe anemia due to repeated self-blood drawing. He had a past history of repeated infection and removal of skin-graft transplanted for giant congenital melanocytic nevus due to self-infliction (later confessed by the patient). On the admission, he presented with high fever (39 approximately 40 degrees C) and severe sustained anemia refractory to repeated blood transfusions. Any specific clinical data indicating bleeding or hemolysis were not found. Self-blood drawing was discovered by a nurse on his 27th hospital day. Syringes and needles for blooddrawing were also found. He recovered from anemia under intensive watching without any specific treatment. He confessed that the high fever was artificial. It was of interest that cytoplasmic processes of megakaryocytes were seen in the peripheral blood film until he recovered from anemia for one month. The serum level of erythropoietin was elevated (1540 mU/ml), but not significantly was that of thrombopoietin (1.54 fmol/ml). This case was considered to be valuable to understand the mechanism of platelet-production by megakaryocytes at persistent bleeding.
Anemia↗
[Long and accurate PCR (LA PCR)].
PCR (Polymerase Chain Reaction) has become a standard and essential tool used in disciplines such as molecular genetics, genome mapping and sequencing, and forensics. It has proven to be easier, quicker, and less costly than traditional techniques of molecular cloning. While its use in these fielde has already contributed to great advancements and revolutionary changes, current PCR using Taq DNA polymerase is usually limited to amplification up to 5kb. Recently, "long" PCR conditions have been identified which allow the amplification of DNA templates up to 40kb with higher fidelity. In this report, we describe the amplification of longer human genomic fragments (up to 27kb) using LA PCR technology.
Cloning, Molecular↗
Identification of Schizosaccharomyces pombe gene psk1+, encoding a novel putative serine/threonine protein kinase, whose mutation conferred resistance to phenylarsine oxide.
We have identified a novel putative protein kinase-encoding gene from Schizosaccharomyces pombe (Sp), designated psk1+, by using a highly conserved amino acid (aa) sequence motif to design amplification of DNA fragments using PCR. The putative translation product of psk1+ contains 436 aa, with a molecular mass of 49,317 Da. A single psk1+ was identified by genomic Southern blot analysis, and the genomic mapping indicated that psk1+ was localized in Sp chromosome III. Growth of wild-type Sp cells was inhibited by 0.5 microM phenylarsine oxide, a protein tyrosine phosphatase inhibitor, but psk1- cells were relatively resistant to this drug.
Amino Acid Sequence↗
Purification and characterization of a fatty acid-activated protein kinase (PKN) from rat testis.
PKN, a novel protein kinase with a catalytic domain homologous to that of the protein kinase C (PKC) family and unique N-terminal leucine-zipper-like sequences, was identified by molecular cloning from a human hippocampus cDNA library [Mukai and Ono (1994) Biochem. Biophys. Res. Commun. 199, 897-904]. Recently we partially purified recombinant PKN from COS7 cells transfected with the cDNA construct encoding human PKN, and demonstrated that the recombinant PKN was activated by unsaturated fatty acids and limited proteolysis [Mukai, Kitagawa, Shibata et al. (1994) Biochem. Biophys. Res. Commun. 204, 348-356]. The present work has focused on the further purification and characterization of PKN from native rat tissue. Immunochemical measurement revealed that PKN was found in every tissue, and was especially abundant in testis, spleen and brain; subcellular fractionation of rat brain showed that half of the PKN was localized in the soluble cytosolic fraction. PKN was purified approx. 8000-fold to apparent homogeneity from the cytosolic fraction of rat testis by DEAE-cellulose chromatography, ammonium sulphate fractionation and chromatography on butyl-Sepharose, heparin-Sepharose, Mono Q and protamine-CH-Sepharose. The enzyme migrates as a band of apparent molecular mass 120 kDa. Using serine-containing peptides based on the pseudosubstrate sequence of PKC-delta as phosphate acceptors, the kinase activity was stimulated several-fold by 40 microM unsaturated fatty acids or by detergents such as 0.04% sodium deoxycholate and 0.004% SDS. In the absence of modifiers, protamine sulphate, myelin basic protein and synthetic peptides based on the pseudosubstrate site of PKCs or ribosomal S6 protein were good substrates for phosphorylation by the kinase. In the presence of 40 microM arachidonic acid the kinase activity of PKN for these phosphate acceptors was increased 2-18-fold. The autophosphorylation activity of purified PKN was partially inhibited by pretreatment with alkaline phosphatase. These properties appear to distinguish PKN from many protein kinases isolated previously.
Amino Acid Sequence↗
Functional characterization of the promoter region of the mouse protein kinase C gamma gene.
Promoter activity of protein kinase C (PKC) gamma gene was analysed by chloramphenicol acetyltransferase (CAT) assay using extracts from the cells transfected with various fusion constructs containing the 5'-flanking region of the mouse PKC gamma gene and CAT gene. Transient expression experiments in PC12 cells revealed that the upstream region of 87 bp from the transcriptional initiation site was sufficient for promoter activity. The region containing nucleotides 87 upstream from the transcriptional initiation site was shown to silence CAT activity in Balb/c3T3 cells, in which mRNA of PKC gamma was not detected, suggesting that this region might contain a transcriptional regulatory element for the cell type-specific expression of the PKC gamma gene.
Animals↗
Xenopus PKN: cloning and sequencing of the cDNA and identification of conserved domains.
cDNA clone encoding Xenopus laevis PKN has been isolated from Xenopus kidney library. Sequencing of this clone has revealed a single open reading frame encoding a protein of 901 amino acids. Immunoprecipitate from cytoplasmic fraction of COS7 cells transfected with this cDNA construct using antiserum against bacterially expressed Xenopus PKN revealed arachidonic acid-dependent autophosphorylation activity. Comparison of the closely related sequences of human and rat PKN with a protein from evolutionarily distant Xenopus, revealed several highly invariant domains in the NH2-terminal regulatory regions, suggesting that they participate in binding interaction with arachidonic acid.
Amino Acid Sequence↗