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T Tadakuma

Publications and source records attributed to T Tadakuma.

At least 73 records · Page 4Linked to original sources
PubMed1987

A simple and efficient liposome method for transfection of DNA into mammalian cells grown in suspension.

For a highly efficient plasmid transfection into mammalian cells grown in suspension, DNA was entrapped in liposomes prepared by the phosphatidylserine calcium-induced fusion method. Employing this technique, a transfection efficiency of about 2% was achieved, with 22 tk+-transformants obtained from 10(3) of mouse mammary carcinoma FM3Atk- cells transfected with a plasmid carrying the thymidine kinase (tk+) gene of the Herpes simplex virus. As compared with a previous report [Ayusawa et al., J. Biol. Chem. 258 (1983) 48-53], this transfection method was more than four orders of magnitude higher than the calcium phosphate method used for FM3Atk- cells. It was shown that the tk+ gene was integrated into the chromosomal DNA and was expressed in all the tk+-transformant clones tested. The described method could be applied to various types of DNA and cells, including those grown as monolayers.

Animals↗
PubMed1987

Application of liposomes to generation of monoclonal antibody to glycosphingolipid: production of monoclonal antibody to GgOse4Cer.

Liposomes were applied to the immunization with GgOse4Cer and screening for production of monoclonal antibody to GgOse4Cer. Four-week-old and 22-week-old Balb/c mice were immunized with GgOse4Cer and Salmonella minnesota R595 lipopolysaccharides incorporated liposomes which were composed of dipalmitoyl-phosphatidylcholine and cholesterol. Since antibody response to GgOse4Cer was higher in 22-week-old than 4-week-old Balb/c mice after immunization, 22-week-old Balb/c mice were used for the immunization prior to generation of the monoclonal antibodies to GgOse4Cer. The screening of monoclonal antibodies was performed by complement-dependent liposome immune lysis assay using GgOse4Cer-containing liposomes. Six kinds of monoclonal antibodies, AG-1, -2, -3, -4, -5, and -6, of the IgM class were established. The specificities of the monoclonal antibodies obtained were defined by complement-dependent liposome immune lysis assay using various glycosphingolipids incorporated in liposomes and by thin-layer chromatography (TLC) with immunostaining. All of the monoclonal antibodies reacted only with GgOse4Cer in the liposome immune lysis assay. In addition, the monoclonal antibodies reacted only with GgOse4Cer in the TLC immunostaining. However, none of the monoclonal antibodies obtained was capable of removing natural killer activity from C3H/He mice spleen cell suspensions in vitro. Liposomes may be useful in the procedures of immunization and screening for generation of antiserum and monoclonal antibody to GSLs.

Animals↗
PubMed1987

Efficient encapsulation of water-soluble materials into small-sized liposomes.

A large proportion of the aqueous phase was encapsulated into small-sized liposomes with diameters less than 100 nm, when multilamellar liposomes, prepared by vortexing a lipid film at an extremely high concentration of lipids up to 512 mM, were subsequently rendered to small vesicles by sonication. Sixty percent of adriamycin (8.6 mM) and 39% of sucrose (50 mM) presented were respectively entrapped into the vesicles prepared with 256 mM lipids. The vesicles formed were quite stable and retained a major portion of the entrapped materials. This simple procedure may be useful for encapsulation of valuable, water-soluble materials into small-sized liposomes.

Doxorubicin↗
PubMed1986

Cloned auto-Ia-reactive T cells elicit lichen planus-like lesion in the skin of syngeneic mice.

To explore the physiologic or pathologic roles of autoreactive T cells, we examined immunological functions of several autoreactive mouse T cell clones in vitro and in vivo. All of the T cell clones were Lyt-2-, L3T4+ and showed self-I region-restricted proliferative responses (one clone was self-I-E restricted, the other clones were self-I-A restricted). One clone derived from C57BL/6 mouse and reactive to the self-I-Ab product (clone bb1-2) showed cross-reactivity to the I-Ak product. Among four such auto-Ia-reactive T cell clones examined, one clone produced fairly large amounts of interleukin 2 (IL 2) in response to syngeneic stimulator cells, and mediated help for the in vitro cytotoxic T cell (CTL) responses of syngeneic thymocytes, whereas this clone did not mediate in vitro antibody responses of syngeneic B cells. The other three clones were producers of small amounts of IL 2 and did not mediate the in vitro CTL responses. Among the three clones, clone bb1-2 showed strong regulatory function, and clone kk-1 (B10.BR origin and self-I-Ak reactive) showed weak regulatory function in vitro antibody responses of syngeneic B cells. The physiologic or pathologic roles of autoreactive T cells in vivo were explored by injecting subcutaneously clone kk-1 T cells or clone bb1-2 T cells into the footpads of the respective syngeneic mice. Clone kk-1 T cells injected into syngeneic mice elicited swelling of the footpad and marked accumulation of mononuclear cells in the dermis, leaving the epidermis intact, as in the delayed-type hypersensitivity reaction. As a notable finding, clone bb1-2 T cells injected into syngeneic mice elicited marked swelling of the footpad and lichen planus-like skin lesions, i.e., infiltration of lymphocytes in the epidermis and epidermal cell damage. The lymphocytes infiltrating in the epidermis were evaluated, as were the injected clone bb1-2 T cells expressing the Lyt-1.2 phenotype, by examination of the skin lesions elicited in C3H/He mice (H-2k, Lyt-1.1, 2.1) by the clone T cells. Clone bb1-2 T cells exerted in vitro cytotoxicity against H-2b and H-2k target cells, whereas clone kk-1 T cells did not show any cytotoxic activity, indicating a correlation between the cytotoxic activity of clone bb1-2 T cells and their ability to elicit lichen planus-like lesions.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1985

Treatment of experimental salmonellosis in mice with streptomycin entrapped in liposomes.

Liposome-entrapped streptomycin (SM) was compared with free SM for therapeutic efficacy against experimental salmonellosis in mice. All of the mice infected with the virulent strain of Salmonella enteritidis 116-54 died between days 5 and 7, and a dose of 20 mg of free SM per kg administered 24 h after the bacterial inoculation did not prolong the survival. In contrast, the same dose of SM entrapped in liposomes prolonged paralleled the dose in the liposomes, and a dose as low as 1.2 mg of SM per kg in liposomes prolonged the survival. The advantage of using liposomes was more pronounced when a larger dose of SM was employed. The liposome-entrapped drug was less toxic than the free drug. A dose of 80 mg of free SM per kg caused convulsions, but the same dose entrapped in liposomes caused no side effects. Furthermore, two doses of liposome-entrapped SM further enhanced the therapeutic effect. The efficacy of the liposome-entrapped drug was still observed in mice infected with a large inoculum of S. enteritidis. A tissue distribution study on SM in various organs demonstrated that liposomal SM was selectively delivered to the spleen and liver with concentrations in these those in mice receiving the free drug. The prolongation of survival was due to suppression of the multiplication of S. enteritidis as demonstrated by viable cell counts in the spleens.

Animals↗
PubMed1985

Analysis of the mode of antigen presentation required for triggering of T cell proliferation by using various azobenzenearsonate-tyrosine derivatives.

Various azobenzenearsonate-tyrosine (ABA-Tyr) derivatives were synthesized by modifying amino and carboxyl groups at the alpha-carbon of tyrosine, with preservation of most of the ABA-Tyr moiety (ABA plus hydroxyphenyl portion of tyrosine). These derivatives were tested for the ability to stimulate ABA-L-Tyr specific T cell lines derived from B10.BR and B10.S mice. ABA-acetyltyramine, ABA-hydroxyphenylpropionic acid (ABA-PPr), and ABA-propylphenol, which lack either the carboxyl or amino group or both, could not induce T cell proliferation. The lack of stimulation by these derivatives was not due to their cytotoxic effects. A similar pattern of proliferation was obtained on stimulating lymph node T cells from B10.BR and B10.S mice primed with ABA-L-Tyr. Some differences were observed, however, between B10.BR and B10.S mice. ABA-L-Tyr-specific T cells from B10.BR mice could not respond well to ABA-D-Tyr in contrast to B10.S T cells. Furthermore, B10.BR mice primed with ABA-acetyltyramine or ABA-PPr in complete Freund's adjuvant could not induce ABA-L-Tyr-reactive T cells, whereas T cells from B10.S mice primed with these derivatives could proliferate in the presence of ABA-L-Tyr. The differences between B10.BR and B10.S mice were further investigated by using (B10.S X B10.BR)F1 mice. T cells from ABA-L-Tyr-immunized F1 mice responded poorly to ABA-D-Tyr when presented with B10.BR antigen-presenting cells (APC), but responded well when presented with B10.S APC. Similarly, T cells from ABA-PPr-primed F1 mice did not proliferate to ABA-L-Tyr in the presence of B10.BR APC, but could proliferate in the presence of B10.S APC. Our results clearly indicate that the presence of charged groups at the alpha-carbon of tyrosine plays a critical role in the triggering of ABA-L-Tyr-specific T cell proliferation. The significance of these results is discussed.

Animals↗
PubMed1984

Liposome immune lysis assay (LILA): a simple method to measure anti-protein antibody using protein antigen-bearing liposomes.

A new simple immunoassay technique using immune lysis of liposomes was developed to measure antibody against protein antigens. Multilamellar liposomes were composed of dipalmitoylphosphatidylcholine, cholesterol and phosphatidylethanolamine substituted with the hetero-bifunctional cross-linking reagent N-hydroxysuccinimidyl 3-(2-pyridyldithio)propionate (SPDP). The protein antigen (human IgG) was coupled to these liposomes after treatment with SPDP and mild reduction. As a release marker, carboxyfluorescein (CF) was entrapped in the liposomes. The CF release was specific to anti-human IgG antibody and depended on the presence of complement. This technique could detect 10(-15) mol of anti-human IgG antibody or human IgG. The liposomes were stable over 8 months at 4 degrees C under nitrogen gas.

Animals↗
PubMed1984

Further study of biological activities of chemically synthesized analogues of lipid A in artificial membrane vesicles.

In the previous paper [Eur. J. Biochem. 124, 405 (1982)], we demonstrated that chemically synthesized lipid-A analogues such as the 1-monophosphate or 1,4'-diphosphate of 6-O-(2-deoxy-2-tetradecanoylamino-6-O-tetradecanoyl-D-glucopyra nos yl)-2-deoxy-2-tetradecanolyamino-3,4-di-O-tetradecanoyl-D-gluco pyr anose enhanced immunogenicity of liposomal model membranes sensitized with amphipathic antigen when they were incorporated in the same liposomes. Here we extend the observation by testing the recently synthesized analogues including diglucosamine analogues carrying hydroxy and acyloxy fatty acids. Among the analogues tested, those which showed higher adjuvant and mitogenic activities in the liposomal system were N-acylated and O-acylated beta-1,6-linked D-glucosamine disaccharides carrying either amide-bound 3-hydroxytetradecanoic acids in addition to phosphate in position 1 of the reducing sugar or amide-bound 3-tetradecanoyloxytetradecanoic acids. The analogue carrying both amide-bound 3-hydroxytetradecanoic acids and phosphate in position 4 of the non-reducing sugar showed weak adjuvant activity and marginal mitogenic activity.

Adjuvants, Immunologic↗
PubMed1983

Interaction of hapten-sensitized liposomes with cells bearing hapten-specific receptors.

The binding of liposomes sensitized with 2,4-dinitrophenyl-6-N-aminocaproylphosphatidylethanolamine (DNP-Cap-PE) to MOPC-315 cells, which secrete and bear on their surfaces anti-DNP immunoglobulins, was studied. The binding was affected by cholesterol content, phospholipid composition and hapten density of liposomes: The binding of distearoylphosphatidylcholine liposomes sensitized with 5 mol% hapten to the cells increased with increasing cholesterol content in liposomes. The amount of liposomes composed of phospholipid with a higher transition temperature (such as distearoylphosphatidylcholine), which bound to MOPC-315 cells, was much higher than that of liposomes composed of phospholipid with a lower transition temperature (such as egg yolk phosphatidylcholine). The amount of distearoylphosphatidylcholine liposomes containing equimolar cholesterol, which bound to the cells at 0 degrees C, increased with increasing amount of the hapten in liposomes up to 2.5 mol%. The binding became maximum at 2.5 mol% and decreased with higher hapten density in liposomes. The immunogenicity of hapten-sensitized liposomes is known to be affected by the liposomal composition such as cholesterol content, phospholipid composition and hapten density. This model study suggests that the binding of liposomes to cells is important for expressing the immunogenicity of hapten-sensitized liposomes.

Animals↗
PubMed1982

Biological activity of chemically synthesized analogues of lipid A. Demonstration of adjuvant effect in hapten-sensitized liposomal system.

The adjuvant effect of synthetic analogues of lipid A was investigated in mice in vivo. Two of the five synthetic lipid A analogues showed adjuvant activity when they were incorporated into dipalmitoylglycerophosphocholine-cholesterol-dipalmitoylglycerophosphate liposomes sensitized with trinitrophenylaminocaproylglycerophosphoethanolamine. The minimum structure for an adjuvant effect in the liposomal immunogen system was 1-monophosphate of 6-0-(2-deoxy-2-tetradecanoylamino-6-0-tetradecanoyl-D-glucopyranosyl)-2-deoxy-2 -tetradecanoyl-amino-3,4-di-O-tetradecanoyl-D-glucopyranose.

Adjuvants, Immunologic↗
PubMed1982

Immunogenicity of liposomal model membranes sensitized with spin-labeled haptens and topographical distribution of haptens on the membranes.

The relation between the in vitro immunogenicity of phosphatidylcholine liposomes containing 2,4-dinitrophenyl-6-N-aminocaproylphosphatidylethanolamine (DNP-Cap-PE) as a hapten and the topographical distribution of the haptens on lipid membranes was studied. In distearoylphosphatidylcholine liposomes, the immunogenicity increased with increase of cholesterol content in the liposomal membranes. The electron spin resonance spectra of spin-labeled DNP-Cap-PE in distearoylphosphatidylcholine liposomes indicated that cholesterol affected the topographical distribution of spin-labeled DNP-Cap-PE on the membranes. In the absence of cholesterol, a considerable amount of haptens was clustered on the distearoylphosphatidylcholine membranes, but with increase of cholesterol, random distribution of the haptens on the membranes increased. The cholesterol-dependent change in the topographical distribution of the haptens on the membranes paralleled the change of immunogenicity, i.e., the immunogenicity was low when haptens were clustered on the liposomal membranes. Haptens arranged at a proper distance on the membranes may be required for optimum immune response.

Animals↗
PubMed1982

Effect of lipid A incorporation on characterization of liposomal model membranes as thymus-independent type 1 or type 2 immunogens.

We used the following criteria to determine whether liposomal model membranes are thymus-independent type 1 (TI-1) or type 2 (TI-2) immunogens: a) in vivo response of (CBA/N X BALB/c)F1 male and female mice; b) in vitro response of spleen cells from these mice; c) ontogeny of the response in cultures of (C57BL/6 X DBA/2)F1 male cells obtained from mice of different ages; d) ability of a monoclonal anti-delta antibody to block the in vitro response of C57BL/6 spleen cells. The liposomes were sensitized with N(2,4,6-trinitrophenyl-6-N-aminocaproyl)-dipalmitoylphosphatidylethanolamine (TNP-Cap-DPPE) and were prepared without and with incorporation of the B cell mitogen, lipid A. On the basis of the above criteria, liposomes prepared without lipid A can be characterized as a TI-2 immunogen and resemble TNP-Ficoll. In contrast, liposomes prepared with lipid A behave like a TI-1 immunogen such as TNP-BA (trinitrophenylated Brucella abortus). Conversion of liposomes from a TI-2 to a TI-1 immunogen by inclusion of lipid A in the bilayers was achieved under conditions of constant TNP-Cap-DPPE epitope density. Increasing the epitope density in liposomes made without lipid A does not result in transformation of immunogen type. These results, which bear on the features that may distinguish TI-1 from TI-2 immunogens, are discussed with regard to the involvement of macrophages and T cells in the response.

Animals↗
PubMed1981

A simple method to measure anti-glycolipid antibody by using complement-mediated immune lysis of fluorescent dye-trapped liposomes.

A simple, reproducible, and micro quantity method is described to measure the antibody against glycolipid antigens. The multilamellar liposomes containing carboxyfluorescein (CF), which is self-quenched at high concentration, are prepared by vortexing the dried lipid films consisting of egg lecithin, cholesterol, phosphatidic acid and Forssman glycolipid antigen. On addition of anti-glycolipid serum plus active complement, liposome lysis occurs, and trapped CF is released. The dilution of CF in the external volume abolishes the quenching, resulting in a high fluorescence signal. Experimental conditions to measure anti-glycolipid antibody is established in this paper.

Animals↗