Biomedical subjects

K Kitajima

Publications and source records attributed to K Kitajima.

At least 163 records · Page 9Linked to original sources

PubMed1994

Identification, developmental expression and tissue distribution of deaminoneuraminate hydrolase (KDNase) activity in rainbow trout.

A deaminoneuraminosyl-glycohydrolase (KDNase), which catalyses the hydrolysis of alpha-ketosidic 2-keto-3-deoxy-D-glycero-D-galacto- nononic acid (or naturally occurring deaminated neuraminic acid; KDN) linkages in KDN-glycoconjugates, is required for their structural and functional studies since KDN residues are usually resistant to the action of known sialidases. A search for KDNase was initiated by examining various cells and tissues of rainbow trout because KDN-glycoconjugates were first found in this animal species. Tissue localization studies of KDNase activity showed it to be present in kidney, spleen and ovary. The highest KDNase activity was found in ovarian post-ovulatory follicles obtained from female fish at the time when the reproductive organ was undergoing natural effacement. Little if any activity was found in brain, heart, liver, muscle, mature eggs and testis. Developmentally, higher levels of KDNase were usually expressed 3-4 months before ovulation or spermiation. An exception to this was in the ovary (or ovarian follicles) where the most striking increase in KDNase occurred 1-2 months after the maturation of gamete cells. Enzyme extracts containing KDNase activity also contained sialidase activity. From the data based on a kinetic study using mixed substrates, both KDNase and sialidase activities were indicated to reside on a single enzyme protein. The KDN-sialidase displayed broad specificity, which could possibly limit its usefulness as a probe for KDN-glycoconjugates. Nevertheless, unlike sialidases, KDNase can selectively remove KDN residues, thus making it an important new reagent to identify KDN-glycoconjugates in vivo.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1994

Occurrence and biological roles of 'proximal glycanases' in animal cells.

Glycosylation of particular proteins and lipids has become generally acknowledged as being important for these molecules to express their functions in various biological events. However, much less attention has been paid to the biological significance of deglycosylation of such once-glycosylated molecules in the context other than catabolism and recycling in the lysosome. Recently, in various kinds of animal cells and tissues we found non-lysosomal peptide: N-glycanase (PNGase) activities. Before these findings, PNGase was only known in plants and bacteria, and our findings indicated that de-N-glycosylation reaction catalysed by PNGase occurred universally in bioorganisms, and might function as a certain biologically important modification, not as a degradative pathway. Now, we put forward and extend the concept to all the glycoconjugates that deglycosylation as well as glycosylation occur as a universal cellular system to modulate the function of the present molecules, and postulate 'proximal glycanases' (PROXIases) as enzymes that are responsible for the detachment of intact glycan from glycoconjugates and form free glycan and apo-glycoconjugates. In this article, we review the occurrence and possible function of proximal glycanases in animal cells.

Amino Acid Sequence↗

PubMed1994

Morphology of primary somatosensory cortical neurons receiving input from the tooth pulp.

1. To elucidate the morphological and electrophysiological characteristics of tooth pulp-driven neurons (TPNs) in the primary somatosensory cortex (SI), we injected neurobiotin into TPNs whose electrophysiological characteristics had been identified. 2. TPNs, responsive to electrical stimulation of the tooth pulp, were recorded intracellularly and injected from areas 3a and 3b of SI. A total of 58 TPNs in SI were successfully injected and reconstructed. Nineteen of these TPNs were located in area 3a and 39 in area 3b. Three area 3a TPNs were identified in lamina II, eight in lamina III, seven in lamina V, and one in lamina VI. Five 3b TPNs were identified in lamina II, 19 in lamina III, 7 in lamina IV, 7 in lamina V, and 1 in lamina VI. 3. Thalamic and tooth pulp latencies of lamina III and IV TPNs were shorter than those of lamina II and V TPNs. On the other hand, lingual and masseteric nerve latencies of TPNs were not consistent with thalamic and tooth pulp latencies. 4. Three of 19 area 3a TPNs and 7 of 39 area 3b TPNs were classified as pulp-specific TPNs, which received only tooth pulp input. Thirteen of 19 area 3a TPNs and 24 of 32 area 3b TPNs were classified as low-threshold mechanoreceptive TPNs, which responded to nonnoxious mechanical stimulation of the receptive field, and only 2 area 3b TPNs were classified as wide-dynamic range TPNs. Six of the area 3a TPNs and 14 of the area 3b TPNs responded to electrical stimulation of the lingual and/or masseteric nerves. Nociceptive-specific TPNs were not recorded in this study. 5. Lamina II TPNs in areas 3a and 3b had small somata, and those in area 3a had dendrites spreading into laminae I-II. Two TPNs in area 3a had axon collaterals extending into area 4. In contrast, area 3b TPNs in lamina II have dendrites spreading into laminae I-III. Their axons did not extend deeply into the subcortical regions, and the axon collaterals reached into area 3a. 6. Lamina III TPNs were classified according to their morphological characteristics as pyramidal or nonpyramidal stellate TPNs. Pyramidal lamina III TPNs had typical pyramidal somata, like those of lamina V pyramidal cells. Furthermore, those in areas 3a and 3b had dendrites with numerous spines spreading into laminae I-III, and some of the area 3a TPNs have axons with collaterals projecting into area 4. Lamina III area 3b TPNs had morphological properties similar to those in area 3a.(ABSTRACT TRUNCATED AT 400 WORDS)

Afferent Pathways↗

PubMed1993

Structural diversity in the alpha 2-->8-linked polysialic acid chains in salmonid fish egg glycoproteins. Occurrence of poly(Neu5Ac), poly(Neu5Gc), poly(Neu5Ac, Neu5Gc), poly(KDN), and their partially acetylated forms.

alpha 2-->8-Linked polysialic acid (polySia) chains terminate O-linked oligosaccharide chains on Salmonidae fish egg polysialoglycoproteins (PSGPs). Expression of these surface PSGPs are developmentally regulated and the polySia epitope is functionally implicated in a number of distinct species-specific cell-cell recognition events during fertilization and early embryogenesis. To better understand the functional diversity of these PSGPs, structural studies of the polySia chains isolated from three genera and eight species of Salmonidae fish eggs were carried out by chemical, immunochemical, enzymatic, and 1H NMR methods. A remarkable degree of structural diversity was found, including differences in the N-acyl groups, i.e. N-acetylneuraminic acid (Neu5Ac) or N-glycolylneuraminic acid (Neu5Gc), and in the presence of either O-acetyl substitution at C4, C7, or C9 or O-lactyl substitution at C9. The presence of heteropolymers containing both Neu5Ac and Neu5Gc residues was also an unexpected finding. Accordingly, the different forms of alpha 2-->8-linked homo- and heteropolymers of these polySia structures include: poly(Neu5Ac), poly(Neu5Gc), poly(Neu5,chi Ac2), poly(Neu5Gc chi Ac), poly(Neu5Ac, Neu5Gc), poly-(Neu5Ac, Neu5,chi Ac2), poly(Neu5Ac, Neu5Gc chi Ac), poly(Neu5Gc,Neu5,chi Ac2), and poly(Neu5Gc, Neu5-Gc chi Ac), where chi represents the site of acetylation at carbon atom 4, 7, or 9. The significance of this new structural information, together with our recent finding of alpha 2-->8-linked polydeaminoneuraminic acid, poly(KDN), in the rainbow trout egg vitelline envelope, is that it demonstrates the natural occurrence of multiple forms of alpha 2-->8-linked polySia chains in Salmonidae fish glycoproteins that have not been previously described. The results also predict that a remarkable array of polysialylated glycoconjugates is yet to be discovered in animals other than teleost fishes.

Amino Acid Sequence↗

PubMed1993

Structure of novel gangliosides, deaminated neuraminic acid (KDN)-containing glycosphingolipids, isolated from rainbow trout ovarian fluid.

Two acidic glycosphingolipids were isolated and purified from rainbow trout ovarian fluid. They were designated as ovarian fluid gangliosides ofg-2a and ofg-2b. Both of these glycolipids were found to contain glucose, galactose, and N-acetylgalactosamine in a molar ratio of 1:2:1, but they differ by the presence of 2 mol of deaminated neuraminic acid (KDN; 2-keto-3-deoxy-D-glycero-D-galacto-nononic acid) in ofg-2a and 1 mol each of KDN and 9-O-acetyl-KDN in ofg-2b. On the basis of composition analysis, methylation analysis, mild acid hydrolysis, fast atom bombardment mass spectrometry (FABMS), 400-MHz 1H nuclear magnetic resonance spectroscopy, and immunochemical analysis using a monoclonal antibody (mAb.kdn3G), the complete structures of these gangliosides were determined to be KDN alpha 2-->3Gal beta 1-->3GalNAc beta 1-->4(KDN alpha 2-->3)Gal beta 1-->4Glc beta 1-->Cer for ofg-2a [(KDN)GD1a] and 9-O-AcKDN alpha 2-->3Gal beta 1-->3GalNAc beta 1-->4(KDN alpha 2-->3)Gal beta 1-->4Glc beta 1-->Cer for ofg-2b [(KDN)GD1a(OAc+)]. The ceramide moieties (Cer) in both ofg-2a [(KDN)GD1a] and ofg-2b [(KDN)GD1a(OAc+)] were found by combining of the results from fatty acid analysis and FABMS measurements to be made up of 4-sphingenine and mainly a C24:1 fatty acyl chain (nervonate). The structures of ofg-2a and ofg-2b are novel, and they represent the second example of naturally occurring KDN-gangliosides. Mild acid hydrolysis of both ofg-2a and ofg-2b resulted in formation of (KDN)GM1a.

Animals↗

PubMed1993

Identification of peptide:N-glycanase activity in mammalian-derived cultured cells.

The recent finding of peptide:N-glycanase (PNGase) in medaka embryos (Seko, A., Kitajima, K., Inoue, Y., & Inoue, S., J.Biol. Chem. 266, 22110 (1991)) raised the question of how widespread is the occurrence of this type of de-N-glycosylating enzyme. In experiments designed to identify PNGase in the mammalian system, we searched for its activity in some cultured cell lines. Incubation of a 14C-labeled N-glycopeptide with extracts prepared from cultured cells resulted in producing the free glycan and the peptide. Detailed characterizations of the products, formed upon incubation of a 14C-labeled N-glycopeptide substrate with the enzyme preparation from C3H mouse loose connective tissue-derived L-929 cells, by HPLC, amino acid and carbohydrate composition analyses, and peptide sequence analysis unequivocally established the reaction products to be the free glycan having di-N-acetylchitobiosyl sequence at its reducing end and free peptide in which the originally glycan-linked Asn residue was converted to the Asp residue. This represents the first demonstration of PNGase in mammalian cells and thus PNGase appears to be a very common enzyme expressed in not only plants and bacteria but also a wide range of animals although its functional significance remains to be clarified.

Amidohydrolases↗

PubMed1993

Structural studies of a novel type of tetraantennary sialoglycan unit in a carbohydrate-rich glycopeptide isolated from the fertilized eggs of Indian Medaka fish, Oryzias melastigma.

A novel carbohydrate-rich sialoglycopolyprotein of apparent molecular mass approximately 7000 Da was isolated from the fertilized eggs of the Medaka fish species, Oryzias melastigma. The glycoprotein was identified as a member of the L-hyosophorin family because it exhibited the following several distinctive features of L-hyosophorin molecules: (a) it contains a high proportion of carbohydrate (90% by weight), and (b) the amino acid sequence of the apopeptide was identical with that of the Oryzias latipes L-hyosophorin which has previously been demonstrated to be derived from a high molecular weight form of hyosophorin, i.e. H-hyosophorin, present in the cortical vesicles of unfertilized eggs. The apoprotein of H-hyosophorin is composed of tandem repeats of the L-hyosophorin apopeptide, i.e. it is a polyprotein. The structure of the carbohydrate portion of purified L-hyosophorin of O. melastigma was studied by composition and methylation analysis, selective chemical (periodate-Smith degradation; hydrazinolysis-nitrous acid deamination), and enzymatic (endo-beta-galactosidase; peptide:N-glycanase) degradation, together with instrumental methods (fast atom bombardment-mass spectrometry and 1H NMR). O. melastigma L-hyosophorin was found to contain two types of large, branched tetraantennary glycan units capped with sialic acids. The two glycans differ with respect to the branching pattern of the trimannosyl core (x = 4 or 6 in Eq. A). [formula: see text] The possible physiological significance of the hyosophorin family is discussed in the light of their unique structural features.

Animals↗

PubMed1993

Synthesis of CMP-deaminoneuraminic acid (CMP-KDN) using the CTP:CMP-3-deoxynonulosonate cytidylyltransferase from rainbow trout testis. Identification and characterization of a CMP-KDN synthetase.

The sugar nucleotide, cytidine 5'-(3-deoxy-D-glycero-D-galacto-2-nonulosonic phosphate) (CMP-KDN) is expected to serve as a donor of KDN residues in the synthesis of KDN-containing glycoconjugates. We report here the identification and characterization of CMP-KDN synthetase, a novel enzyme responsible for synthesis of CMP-KDN from KDN and CTP. The enzyme was partially purified from the testis of rainbow trout (Oncorhynchus mykiss), where KDN gangliosides were first discovered (Yu, S., Kitajima, K., Inoue, S., and Inoue, Y. (1991) J. Biol. Chem. 266, 21929-21935), and used to synthesize CMP-[14C]KDN, which was characterized by 1H NMR. Vmax/Km studies showed that KDN was a preferred nonulosonic acid substrate compared to N-acetylneuraminic acid (Neu5Ac) or N-glycolylneuraminic acid (Neu5Gc) (4.4 x 10(-3) min-1 for KDN versus 2.3 and 1.8 x 10(-3) min-1 for Neu5Ac and Neu5Gc, respectively). CMP-KDN synthetase activity was maximal at pH 9-10 and at 25 degrees C. The presence of either Mg2+ or Mn2+ was essential for CMP-KDN synthetase activity. 25 mM Mg2+ stimulated formation of CMP-KDN more than 10-fold, yet only stimulated formation of CMP-Neu5Ac and CMP-Neu5Gc 4-fold, relative to 1 mM Mg2+. A kinetic study using mixed substrates showed that both CMP-KDN and CMP-Neu5Ac synthetase activities in the partially purified enzyme were due to the same active site of a single enzyme. In contrast, Neu5Ac and Neu5Gc were the preferred nonulosonic acid substrates for the calf brain CMP-sialic acid synthetase. Thus, mammalian CMP-sialic acid synthetases recognizes similar, yet distinctively different, substrate specificity determinants. Thus, the trout testis enzyme was considered to synthesize activated sugar nucleotides required for synthesis of both (KDN)GM3 and (Neu5Ac)GM3. The expression of CMP-KDN synthetase was shown to be temporally correlated with development and to parallel the developmental expression of (KDN)GM3 in sperm.

Animals↗

PubMed1993

Monoclonal antibody specific to alpha-2-->3-linked deaminated neuraminyl beta-galactosyl sequence.

Fusion of spleen cells from a BALB/c mouse immunized with KDN alpha 2-->3Gal beta 1-->4Glc beta 1-->1Cer ((KDN)GM3) with P3-X63 Ag8.U1 (P3U1) mouse myeloma cells yielded a hybrid cell line that produced monoclonal antibody that bound to (KDN)GM3, but not to Neu5Ac alpha 2-->3Gal beta 1-->4Glc-beta 1-->1Cer ((Neu5Ac)GM3). The specificity of the monoclonal antibody was determined chiefly by the enzyme-linked immunosorbent assay procedure. This antibody was found to react most strongly with (KDN)GM3 and less strongly with a glycoprotein containing a number of KDN alpha 2-->3Gal beta 1-->3-GalNAc alpha 1-->3[8KDN alpha 2-->)n-->6]GalNAc alpha 1-->chains (< n > av = approximately 3). The results indicated that the monoclonal antibody (designated mAb.kdn3G) specifically and effectively recognized a disaccharide structure, KDN alpha 2-->3Gal beta 1-->, and specifically discriminated (KDN)GM3 from (Neu5Ac)GM3. The mAb.kdn3G was used to localize (KDN)GM3 in rainbow trout sperm by the indirect immunofluorescence procedure and the antigen was shown to be mostly, if not completely, associated with the external surface of the entire plasma membrane of rainbow trout sperm. The potential utility of mAb.kdn3G is addressed in searching for KDN-glycoconjugates which contain glycan units having the KDN alpha 2-->3Gal beta 1-->epitope structure.

Animals↗

PubMed1993

Treatment of arteriosclerotic obstruction by LDL adsorption.

According to the authors' clinical analysis, about half of the patients who suffer from arteriosclerotic obstruction (ASO) in the lower extremity(-ies) with clinical manifestation are dyslipidemic (total cholesterol > or = 220 mg/dL or LDL cholesterol > or = 140 mg/dL). As suggested by clinical success in regression of ASO in the coronary arteries as a result of aggressive removal of LDL, LDL adsorption utilizing an extracorporeal circulation technique with a dextran sulfate/cellulose adsorbent column was applied in 33 patients (22 men and 11 women) with ASO. Clinical results obtained after a series of 10 LDL adsorption procedures as a standard showed encouraging success. Improvement in subjective symptoms was achieved as follows: 88.5% for cold lower extremity, 87.1% for intermittent claudication, 53.8% for leg/toe pain at rest, and 60% for disappearance/size diminution of ulcer/necrosis. Improvements in objective examination findings supported subjective ones: 85.7% by plethysmography, 81% by thermography and 70% by ankle pressure index. No serious complications or untoward effects were observed during or after the adsorption procedures. In conclusion, LDL adsorption appears to be a useful and safe tool in treatment of ASO patients with dyslipidemia.

Adsorption↗

PubMed1993

The inhibitory effect of noradrenaline on thyrotrophin-stimulated 3,5,3'-tri-iodothyronine and thyroxine release is mediated through a Ca(2+)-dependent process in the thyroid gland of the mouse.

We examined the effect of noradrenaline on the release of 3,5,3'-tri-iodothyronine (T3) and thyroxine (T4) from perifused mouse thyroid. Noradrenaline suppressed the thyrotrophin (TSH)-stimulated release of T3 and T4. The addition of prazosin, which is a specific alpha 1 antagonist, or the depletion of Ca2+ from the perifusion buffer completely abolished the inhibitory effect of noradrenaline on TSH-stimulated T3 and T4 release. Noradrenaline did not inhibit TSH-stimulated cyclic adenosine 3',5'-monophosphate (cAMP) release in the presence of 3-isobutyl-1-methylxanthine (IBMX), which inhibits both cAMP-specific and calmodulin-sensitive phosphodiesterases. Noradrenaline significantly suppressed the TSH-stimulated release of T3 and T4 in the presence of IBMX. These results suggest that the inhibitory effect of noradrenaline on TSH-stimulated T3 and T4 release is not mediated through a cAMP-dependent process or the activation of a calmodulin-sensitive phosphodiesterase, and that this inhibition is mediated through a Ca(2+)-dependent process regulated by the alpha 1-adrenergic system in the mouse thyroid.

1-Methyl-3-isobutylxanthine↗

PubMed1993

Intraoral pressure in the evaluation of laryngeal function.

The difference in peak intraoral pressure between the consonants /p/ and /b/ (abbreviated as Pr./p-b/) showed a significant correlation with the transglottal pressure during consonant /b/ production. The values of Pr./p-b/ in subjects with vocal fold palsy were smaller than those in subjects with a cancer or polyp. The values of Pr./p-b/ divided by the estimated subglottal pressure in the subjects with cancers showed different distributions from those in subjects with palsies, polyps and in normal subjects. These parameters have potential benefit for the evaluation of pathological conditions of the glottis, especially for insufficient closure of the glottis and high stiffness of the vocal folds.

Adult↗

PubMed1993

Interplay between differentiation and cell cycle arrest at G1 in phorbol diester-treated human myelocytic leukemia cells.

The cell cycle arrest at G1 of myelocytic leukemia cells by incubation with phorbol diesters was examined in HL-60 cells and more mature THP-1 cells. We found: (1) induction of phenotypic parameters of differentiation prior to G1 arrest; (2) a long induction period in HL-60 cells compared to zero in THP-1 cells; and (3) strong inhibition of G1 arrest with a protein kinase C inhibitor in THP-1 cells. We conclude, from these lines of evidence and some additional information, that differentiation induces G1 arrest. At present, the most plausible causative molecule is Mac-1.

Cell Differentiation↗

PubMed1992

Fish egg glycophosphoproteins have species-specific N-linked glycan units previously found in a storage pool of free glycan chains.

Recent findings (Ishii, K., Iwasaki, M., Inoue, S., Kenny, P. T. M., Komura, H., and Inoue, Y. (1989) J. Biol. Chem. 264, 1623-1630; Inoue, S., Iwasaki, M., Ishii, K., Kitajima, K., and Inoue, Y. (1989) J. Biol. Chem. 264, 18520-18526) of a relatively large quantity of complex-type free sialo-oligosaccharides in the unfertilized eggs of freshwater fish, Plecoglossus altivelis and Tribolodon hakonensis, prompted us to search for their progenitor glycoproteins. First we demonstrated a third occurrence of free sialoglycans in the unfertilized eggs of Medaka fish (Oryzias latipes). Next, in all three species studied, a uniformly high level of glycophosphoproteins (GPP) was identified and found to possess N-linked glycan units. The carbohydrate structures of the GPP were determined to be identical with those of the free glycans isolated from the unfertilized eggs of the respective fish species. Thus, the most likely candidate for the progenitor of free sialoglycans appeared to be the oocyte GPPs. This implies that the liberation of the free glycans by a putative peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase may represent a necessary biochemical event during vitellogenesis or oogenesis. The present results may provide insight into a new concept of a "protein N-glycosylation/de-N-glycosylation system" recently proposed by us (Seko, A., Kitajima, K., Inoue, Y., and Inoue, S. (1991) J. Biol. Chem. 266, 22110-22114).

Amino Acids↗

PubMed1992

Detection, isolation, and characterization of oligo/poly(sialic acid) and oligo/poly(deaminoneuraminic acid) units in glycoconjugates.

We have evaluated methods for separation, preparation, and characterization of alpha-2----8-linked oligomers of sialic acids (Neu5Ac and Neu5Gc) and deaminated neuraminic acid (KDN; 2-keto-3-deoxy-D-glycero-D-galacto-nononic acid) recently found as a naturally occurring novel type of sialic acid analogue. (A) We examined preparative anion-exchange chromatography for fractionation and preparation of oligo(Neu5Ac), oligo(Neu5Gc), and oligo(KDN). (B) We also examined the TLC method for separation and differentiation of the partial acid hydrolysates of colominic acid, as well as polysialoglycoproteins (PSGP) and poly(KDN)-glycoproteins (KDN-gp) isolated from rainbow trout eggs, and for discrimination of lower oligomers of Neu5Ac, Neu5Gc, and KDN. (C) We developed the high-performance adsorption-partition chromatographic method for (a) separation of monomers and oligomers of three nonulosonates according to the difference in substituents at C-5 and the presence or absence of 9-O-acetyl groups in oligo(KDN) and (b) separation of three homologous series of lower oligomers according to the degree of polymerization. (D) We examined and compared high-performance anion-exchange chromatographic separation of 3H-labeled oligo(Neu5Ac), oligo(Neu5Gc), and oligo(KDN) alditols by using Mono-Q HR 5/5 resin. (E) We examined a method of selective and quantitative microprecipitation for separation and purification of oligomers and polymers of Neu5Ac by treating them with cetylpyridinium chloride. We also used PSGP and KDN-gp to test both the sensitivity and the selectivity of this method.

Animals↗

PubMed1992

Analytical methods for identifying and quantitating deamidated sialic acid (2-keto-3-deoxy-D-glycero-D-galactonononic acid) and alpha 2----8-linked poly(oligo)nonulosonate residues in glycoconjugates.

In 1986 we reported the natural occurrence of deaminated neuraminic acid (2-keto-3-deoxy-D-glycero-D-galactonononic acid, KDN) in fish egg glycoprotein. Subsequently, we have shown that many types of sialic acid as well as KDN occur in polymeric chains, poly(oligo)-Sia and poly(oligo)KDN in nature. In this study we demonstrate that the conventional colorimetric and gas-liquid chromatographic methods used in the analysis of sialic acid can be applied to analysis of these new nonulosonate and poly(oligo)nonulosonates. We report that the thiobarbituric acid reaction can be used to analyze both free and bound KDN, but gives lower extinction values when applied to poly(oligo)KDN without prior hydrolysis. Further, the published hydrolytic and/or methanolytic procedures are suitable to release the terminal sialic acid residues, but are not appropriate for quantitative release of the nonulosonic acids from poly(oligo)nonulosonates. A new gas-liquid chromatographic procedure for the identification-quantitation of nonulosonates in poly(oligo)meric forms is described.

Animals↗

PubMed1992

Determination of transcriptional activities of typical gene promoters in HL-60 cells.

We developed a highly sensitive procedure for assaying chloramphenicol acetyltransferase (CAT) enzyme activity in extracts of eukaryotic cells transfected with the CAT gene expression vector, by modification of the partition extraction procedure described by Sleigh [Anal. Biochem. 156, 251-256 (1986)]. The sensitivity of the new method was improved 100-fold on commercial purified enzyme. In routine measurements with cell extracts a CAT activity as low as 1.3 x 10(-4) unit could be measured within an error of less than 30%. The CAT enzyme expressions in undifferentiated human promyelocytic leukemic cell line HL-60 from typical gene promoters could be measured by the new method and compared to select a stronger promoter. Similar measurements were made with more mature monocytic THP-1 cells to evaluate the change in the promoter activity with cell maturation. Differentiation induction with 12-O-tetradecanoylphorbol-13-acetate (TPA) activated transcription from the human immunodeficiency virus (HIV) promoter about 10-fold in HL-60 cells, as expected, but the level was less than that in untreated THP-1 cells. In addition, a similar activation was observed in THP-1 cells as well.

Cell Differentiation↗

PubMed1992

Absence of HTLV-II co-infection in HTLV-I-associated myelopathy patients.

Syncytium inhibition assay to distinguish between HTLV-I and HTLV-II infection was performed with sera and cerebrospinal fluid of 15 HTLV-I-associated myelopathy (HAM) patients. Also, genome analysis of HTLV-II proviral DNA was performed in some HAM patients by use of the polymerase chain reaction method. All of the HAM patients were negative for co-infection with HTLV-II.

Adult↗