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Biomedical subjects

M Akashi

Publications and source records attributed to M Akashi.

At least 91 records · Page 5Linked to original sources
PubMed1994

Detection of interleukin-1 activity in human bladder cancer cell lines.

Detection of interleukin (IL)-1 activity was studied in two human bladder cancer cell lines, T24 and EJ1, and one rat bladder carcinoma cell line, 804G. Significantly high proliferation of mouse thymocytes in the assay of IL-1 activity was observed in the conditioned medium (CM) of T24 cells, indicating that the cells released IL-1-like activity. Enzyme-linked immunosorbent assay (ELISA) and Northern blot analysis showed the presence of both IL-1 alpha and IL-1 beta in the CM of T24 cells and expression of mRNAs of both cytokines in the cells. Interleukin-1 activity in EJ1 cells, which produced a little activity, was induced by E. coli lipopolysaccharide (LPS) while it was not induced in T24 by either LPS or other test substances. Conditioned medium of T24 increased proliferation of both T24 and EJ1 in cell-growth assay. Further investigation of the mode of action and role of cytokines, especially those from tumor cells themselves, is necessary in relation to BCG or photodynamic therapy.

Animals↗
PubMed1994

[The diagnosis of a mutation of the insulin receptor by non-radioisotropical RT-PCR-SSCP analysis].

We evaluated a 24 years old diabetes woman with type A insulin resistant (patient "Yakushima") who had some typical symptoms as acanthosis nigricans, hirsutism and virilization. Insulin binding to the patient's erythrocytes was significantly decreased to about 30% of the normal control. In order to determine the mutation of the insulin receptor, we used for reverse transcript-polymerase chain reaction-single strand conformation polymorphism (RT-PCR-SSCP) analysis without using radiolabeled materials. We also analysed the nucleotide sequence with non-isotropical probe. Our results suggested that the mutation was heterozygous, and the patient had a new missense mutation substituted Asn461 for Thr461 in the alpha-subunit.

Adult↗
PubMed1994

Radiosensitivity of CD45RO+ memory and CD45RO- naive T cells in culture.

Radiosensitivities of various human T-cell subsets were investigated by a proliferation assay and by a single-cell gel electrophoresis assay. Each T-cell subset was purified using a cell sorter and was induced to proliferate by ionomycin and interleukin 2. Unsorted T cells showed biphasic dose-survival curves, indicating the heterogeneity of T cells in terms of radiosensitivity. Purified CD4+ helper and CD8+ killer T cells showed similar biphasic dose-survival curves. Hence both T-cell subsets were composed of cells of different radiosensitivity. The T-cell subsets belonging to different activation stages such as CD45RO+ memory and CD45RO- naive T cells had different dose-survival curves. The former was more radiosensitive than the latter. The high radiosensitivity of CD45RO+ cells was also demonstrated by single-cell gel electrophoresis after irradiation. This is the first demonstration that a particular cell surface marker on T cells is correlated with greater radiosensitivity.

Antigens, CD↗
PubMed1993

Affinity gel electrophoresis of nucleic acids. Nucleobase-selective separation of DNA and RNA on agarose-poly(9-vinyladenine) conjugated gel.

Poly(9-vinyladenine) (PVAd) was immobilized within an agarose gel matrix to produce a novel affinity gel for the base-specific separation of nucleic acids by electrophoresis. The shape (single- or double-stranded) and base content of nucleic acids were specifically recognized by the affinity gel. Only single stranded DNA, of which the sequence is not regular enough to form a stable duplex hairpin structure, was selectively absorbed over double-stranded DNA. Among five polynucleotides having different bases such as poly(A), poly(G), poly(C), poly(U) and poly(I), poly(U) and poly(I) were base-specifically adsorbed by PVAd, probably by hydrogen bond formation. The effect of the molecular mass and size of poly(9-vinyladenine) was also examined.

Adenine↗
PubMed1993

Affinity gel electrophoresis of nucleic acids. Specific base- and shape-selective separation of DNA and RNA on polyacrylamide-nucleobase conjugated gel.

Two types of affinity gels consisting of cross-linked polyacrylamide and affinity ligands possessing nucleic acid bases were prepared. One type of gel was polyacrylamide-poly(vinylnucleobase) conjugated gel, where the poly(vinylnucleobase) such as poly(9-vinyladenine) (PVAd) bearing a nucleobase in the side-chain was entrapped in the gel matrix. The other type of gel, in which a nucleobase such as adenine is chemically bonded to polyacrylamide gel, was prepared by copolymerization of acrylamide, cross-linker and 9-vinyladenine. These affinity gels, especially the former, demonstrated characteristic nucleobase- and shape-selective separation of nucleic acids. The gels showed high affinity for single-stranded DNA and both single- and double-stranded polynucleotides and could separate a double-stranded DNA in mixtures of double-stranded DNA and polynucleotides. The electrophoretic mobilities of poly(uridylic acid) and poly(inosinic acid) were selectively retarded even in the presence of 7 M urea. The electrophoretic behaviours of nucleic acids on the polyacrylamide-PVAd conjugated gels were compared with those on the agarose-PVAd conjugated gel. The effects of urea, temperature and concentration of PVAd were also examined. The polyacrylamide-PVAd conjugated gel served to elucidate interactions between PVAd and nucleic acids that could not be detected by usual spectroscopic methods.

Acrylic Resins↗
PubMed1993

Effect of urea concentration on the base-specific separation of oligodeoxynucleotides in capillary affinity gel electrophoresis.

Base-specific separations of oligodeoxynucleotides were achieved with high resolution by electrophoresis, using a urea-gel capillary, in which poly(9-vinyladenine) (PVAd) was utilized as an affinity ligand. The migration behaviour and the plate number of oligodeoxynucleotides were investigated as a function of urea concentration between 2 and 10 M in capillary gel electrophoresis (CGE) as well as capillary affinity gel electrophoresis (CAGE). The migration time in CGE separation increases as urea concentration increases. An increase in the viscosity of the gel buffer medium as well as a change in the conformation of oligodeoxynucleotides is found to be a predominant factor for an increase in the migration time. The migration time and the plate number of oligothymidylic acids, which interact with PVAd in CAGE, is manipulated by differing urea concentration, which leads to the change in the dissociation process of a specific hydrogen bonding between oligothymidylic acids and PVAd. The migration time of oligodeoxyadenylic acids, which do not interact with PVAd in CAGE, increases with an increase in urea concentration as in CGE separation. The plate number of oligodeoxyadenylic acid was not affected by the urea concentration.

Adenine↗
PubMed1993

Effect of human recombinant granulocyte colony-stimulating factor on induction of myeloid leukemias by X-irradiation in mice.

Hematopoietic suppression is one of the serious problems induced by whole body irradiation. Granulocyte colony-stimulating factor (G-CSF) stimulates the progenitors of granulocytes and accelerates their recovery from bone marrow suppression induced by cytotoxic chemotherapy or radiation. On the other hand, G-CSF stimulates proliferation of myeloid leukemia cells as well as normal granulocytes in vitro. We designed a method to determine if G-CSF affects the incidence of myeloid leukemias induced by irradiation and the types of leukemias induced according to the French-American-British (FAB) classification in RFM/MsNrs mice. Administration of G-CSF (2 micrograms/d for 7 days) after a single 3-Gy irradiation significantly increased the number of peripheral blood neutrophils as compared with those in control mice. Even after discontinuation of G-CSF, both the total leukocyte and neutrophil counts increased to day 10, and their levels remained elevated until day 14. The incidence of myeloid leukemia in mice exposed to a single 3-Gy irradiation was 18.6% (38 of 204), and treatment with G-CSF did not increase the incidence (15.7% [32 of 204]). In the mice with radiation-induced leukemia, those receiving G-CSF had a mean survival time of 357 days, whereas those not receiving the factor survived for 349 days. There was no significant difference of survivals between the two groups. Most of the radiation-induced leukemias in the two groups were M1 or M2, according to the FAB classification; no characteristic difference was observed among the types of leukemias. Although G-CSF stimulated the leukemia cells in vitro, G-CSF administration after irradiation did not increase the occurrence of radiation-induced myeloid leukemias. Our results show that administration of G-CSF effectively accelerates neutrophil recovery from irradiation-induced hematopoietic injury and does not enhance the induction of myeloid leukemia in RFM/MsNrs mice by irradiation.

Animals↗
PubMed1993

Lecithin-cholesterol acyltransferase and lipid transfer protein activities in liver disease.

The activities of lecithin-cholesterol acyltransferase (LCAT) and lipid transfer protein (LTP) were assayed using sensitive radioassay methods in controls (n = 113) and in patients with various liver diseases (n = 72). Plasma LCAT activity decreased with progression of hepatocellular damage. Plasma LTP activity in controls was 216 +/- 68 nmol/mL/h, and there were no significant differences between controls and patients with chronic hepatitis ([CH], 193 +/- 70), compensated liver cirrhosis (LC) with or without hepatocellular carcinoma ([HCC], 197 +/- 48 and 193 +/- 62, respectively), or decompensated liver cirrhosis ([dLC], 182 +/- 65). In acute viral hepatitis, LTP activity decreased significantly; however, the degree of reduction was not as dramatic as that for LCAT. There was no correlation between LCAT and LTP activity both in controls and patients with various liver diseases. LCAT activity was positively correlated with serum albumin (r = .52, P < 0.1) and cholinesterase (r = .37, P < .01) levels, and inversely correlated with serum bilirubin level (r = -.38, P < 0.1); there was no correlation between plasma LTP activity and these parameters of liver function. That plasma LTP activity did not change with hepatocellular damage may indicate that the liver in humans may not be the primary site of LTP production.

Acute Disease↗
PubMed1993

A novel biomaterial: poly(dimethylsiloxane)-polyamide multiblock copolymer I. Synthesis and evaluation of blood compatibility.

Aramid-silicone resins (PASs) consisting of aromatic polyamide (aramid) and poly(dimethyl-siloxane) (PDMS) segments were synthesized by low temperature solution polycondensation. For the evaluation of blood compatibility in vitro, two kinds of experiments were carried out. One was the thromboxane B2(TXB2) release test from platelets attaching to PAS and Biomer. The other was the observation of the platelet adhesion on the surfaces of PAS by scanning electron microscopy (SEM). The results indicated that PAS was bio-inert in vitro. The surface chemical composition of PAS films was investigated by means of electron probe micro analysis (EPMA), X-ray photoelectron spectroscopy (XPS), and dynamic contact angle measurements. The relationship between blood compatibility and surface composition of PAS is discussed.

Biocompatible Materials↗
PubMed1993

Nucleotides bearing a cleavable genotoxic group on the phosphate.

A new class of nucleotide derivatives in which N-nitroso-pyrrolidine (NPYR) is linked to the phosphate group through an ester linkage at the alpha-carbon of the nitrosamine were prepared by a reaction between a nucleoside monophosphate and alpha-acetoxy-NPYR. The NPYR derivatives were prepared from dpT, dpC, dpA, dpG, pA and dpCpT. Treatment of the NPYR-pX with acid, snake venom phosphodiesterase, or near ultraviolet light caused cleavage of the NPYR group, regenerating pX. All of these nucleotide derivatives were directly mutagenic towards Salmonella typhimurium TA1535. When phage M13mp2 double-stranded covalently closed circular DNA was treated with NPYR-dpT with near ultraviolet irradiation, single strand breaks of the DNA took place.

Bacteriophage M13↗
PubMed1993

Affinity gel electrophoresis of nucleic acids. The interaction between water soluble polymers having malachite green and double-stranded DNAs.

Copolymers prepared by the free radical copolymerization of acrylamide and vinyl malachite green were immobilized in an agarose gel matrix to produce a novel affinity gel for double-stranded DNAs. Relative electrophoretic mobilities of calf thymus DNA and poly(dA-dT)2 strongly depended on the amount of vinyl malachite green to show the strong interaction between DNAs and the copolymers. But any affinity interaction was not observed between poly(dG-dC)2 and the copolymers.

Acrylamide↗
PubMed1993

[A method for measurement of circulating blood volume with fluorescein isothiocyanate labeled red blood cells].

We developed a new method to measure the total circulating blood volume using fluorescein isothiocyanate (FITC) labeled autologous red blood cell as an indicator. Red blood cells were labeled by incubation with 0.1 mg.ml-1 FITC in PBS (pH 7.2) for 30 min at 37 degrees C. Eighteen dogs were given an intravenous injection of FITC labeled red blood cells, which were approximately 0.1% of the total circulating red blood cells estimated from the body weight. The ratio of labeled cells to the normal circulating red blood cells was determined by a flow cytometry. The total blood volume was calculated from its dilution rate. The average circulating blood volume determined by this method was 76.8 +/- 9.2 ml.kg-1 (mean +/- SD). This result coincides well with the values reported previously. This method using non radioactive tracers enables a reproducible measurement of circulating blood volume and its change with time.

Animals↗
PubMed1992

Irradiation increases levels of GM-CSF through RNA stabilization which requires an AU-rich region in cancer cells.

Granulocyte/macrophage colony stimulating factor (GM-CSF) is a hematopoietic growth factor that stimulates a wide range of myeloid hematopoietic cells; RNAs coding for many oncogenes and cytokines including GM-CSF have a very short half-life. The motif of AUUUA is a highly conserved sequence in the 3'untranslated regions (3'UTR) of these transcripts and is repeated a number of times in these short-lived cytokines and oncogenes. These sequences play a major role in controlling stability of these transcripts. Human cancer cells were transfected with a chimeric rabbit beta-globin gene linked to either a 58 bp sequence of the AT-rich region from GM-CSF or a control sequence. We have found that irradiation stimulates accumulation of GM-CSF, interleukin-6 (IL-6), and IL-1 beta RNAs. In addition, this accumulation of GM-CSF was at least, in part, a result of increased stabilization of GM-CSF transcripts. Further experiments showed that irradiation increased levels of the chimeric beta-globin transcripts containing AUUUA sequences from GM-CSF, but not those containing the control sequences. Our results suggest that irradiation increases expression of GM-CSF RNA and that posttranscriptional stabilization requiring AUUUA sequences probably is in part one of the mechanisms producing the increased levels of GM-CSF RNA by irradiation.

Base Sequence↗
PubMed1992

Specific base recognition of oligodeoxynucleotides by capillary affinity gel electrophoresis using polyacrylamide-poly(9-vinyladenine) conjugated gel.

Poly(9-vinyladenine) was synthesized and utilized as an affinity macroligand entrapped within the gel matrix. Base-specific separation of oligodeoxynucleotides was achieved with high resolution and high speed by electrophoresis, using capillaries filled with conjugated polyacrylamide-poly(9-vinyladenine) gel. Oligothymidylic acids were selectively separated from the mixture of oligothymidylic and oligodeoxyadenylic acids by utilizing a specific hydrogen bonding between poly(9-vinyladenine) and oligothymidylic acids. Migration time and resolution of oligodeoxynucleotides were influenced by several parameters, such as the size of poly(9-vinyladenine), capillary temperature, and concentrations of poly(9-vinyladenine) and urea. Some guidelines are presented, based on the theoretical formulation of the effect of these parameters, in order to find optimum electrophoretic conditions. Analytical capillary affinity gel electrophoresis was developed for the selective and sensitive base recognition of oligodeoxynucleotides with efficiencies as high as several 10(6) plates/m by using a urea-gel capillary with poly(9-vinyladenine) and temperature-programming.

Adenine↗
PubMed1992

High-performance affinity chromatography of oligonucleotides on nucleic acid analogue immobilized silica gel columns.

The nucleic acid analogues poly(9-vinyladenine) (PVAd), poly(9-adenylethyl methacrylate) and poly(thymylethyl methacrylate) (PTM) were chemically bonded to porous silica gel, which had been pretreated with 3-trimethoxysilylpropyl methacrylate, by free radical copolymerization to produce novel packing materials for affinity chromatographic columns. The columns separated nucleosides and nucleotide dimers on the basis of hydrophobic interaction using an aqueous buffer and complementary hydrogen bonding interaction in methanol as an eluent. The PVAd- and PTM-silica gel columns gave a nucleobase-selective separation of oligonucleotides differing in length from mixtures of oligoadenylic and oligouridylic acids. On the PVAd-silica gel column terminal phosphate isomers of oligouridylic acid up to seven mer were resolved and the elution order of the isomers was different from that on an ODS column.

Chromatography, Affinity↗
PubMed1992

Spectroscopic study of the interaction between poly-(9-vinyladenine) and single or multistrand RNA.

The interaction between poly(9-vinyladenine) (PVAd) and poly[r(U)] was investigated by means of uv, CD, 1H-, and 31P-nmr spectroscopies. The interaction was dependent on the molecular weight of PVAd determined by uv and CD spectroscopies. Based on imino proton nmr, it was clearly found that PVAd formed the complex with poly[r(U)] by complementary hydrogen bonding. The interaction of PVAd with double- and triple-stranded helices of RNA was also investigated by uv melting behavior and 31P-nmr spectroscopy. The results suggested that PVAd could not interact with the double-stranded poly[r(A)].poly[r(U)] but did with the triple-stranded RNA.

Adenine↗
PubMed1992

Recognition of (d(TTTATT) and d(TTATTT) by capillary affinity gel electrophoresis (CAGE) using poly(9-vinyladenine)-polyacrylamide conjugated gel.

Sequence-specific recognition of oligodeoxynucleotide isomers (TTTATT and TTATTT) was achieved by using polyacrylamide-poly(9-vinyladenine) conjugated gel filled capillary affinity gel electrophoresis. It was found that the interaction between poly(9-vinyladenine) and the isomers was dependent on the sequential thymidylic acids of them.

Adenine↗