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Biomedical subjects

Y Kiso

Publications and source records attributed to Y Kiso.

At least 109 records · Page 6Linked to original sources
PubMed1994

Prostaglandin F2 alpha receptor is coupled to Gq in cDNA-transfected Chinese hamster ovary cells.

We examined intracellular signal transduction of prostaglandin F2 alpha (PGF2 alpha) receptors by transfection and stable expression of the receptor cDNA in Chinese hamster ovary (CHO-PGF2 alpha) cells. The binding to the membrane was specific for PGF2 alpha, and the Scatchard plot analysis showed a single high-affinity binding site (Kd = 25.2 nM). PGF2 alpha gradually elevated intracellular Ca2+ and stimulated phosphoinositide metabolism in CHO-PGF2 alpha cells. In whole-cell clamp recordings, PGF2 alpha induced an outward current in the presence of external Ca2+, but it induced a long-lasting inward Ca2+ current in a Na(+)-free solution containing K+ channel blockers. Gq alpha antibody applied intracellularly blocked both outward and inward currents induced by 1 microM PGF2 alpha. These results demonstrate that the PGF2 alpha receptor is coupled to phosphoinositide metabolism in CHO-PGF2 alpha cells via Gq.

Animals↗
PubMed1994

Plasma pharmacokinetics and urinary and biliary excretion of a new potent tripeptide HIV-1 protease inhibitor, KNI-272, in rats after intravenous administration.

The pharmacokinetic (PK) characteristics of KNI-272, a potent and selective HIV-1 protease inhibitor, were evaluated in rats after intravenous (IV) administration. The effect of dose on KNI-272 plasma kinetics, and the urinary and biliary elimination kinetics of KNI-272, were examined. After IV administration of 10.0 mg kg-1 KNI-272, the mean terminal elimination half-life, t1/2 lambda zeta, was 3.49 +/- 0.19 (SE) h, the total plasma clearance, CLtot, was 15.1 +/- 1.2 mL min-1 and the distribution volume at steady state, Vd,ss, was 3790 +/- 280 mL kg-1. On the other hand, after 1.0 mg kg-1 IV administration, t1/2 lambda zeta was 3.04 +/- 0.11 h, CLtot was 15.9 +/- 0.2 mL min-1, and Vd,ss was 6950 +/- 600 mL kg-1. The PK parameters of KNI-272 after IV administration showed that the disposition of KNI-272 in the rat plasma is linear within the dose range from 1.0 to 10.0 mg kg-1. Using an equilibrium dialysis method, the plasma binding of KNI-272 was measured in vitro. The free fractions were 17.7 +/- 0.6%, 12.1 +/- 1.5%, and 13.8 +/- 1.4% at the total concentration ranges of 9.898 +/- 0.097 microgram mL-1, 0.888 +/- 0.008 microgram mL-1, and 0.470 +/- 0.55 microgram mL-1, respectively. The percentages of the dose excreted into the urine and bile as the unchanged form were 1.20 +/- 1.06% and 1.61 +/- 0.32% at 1.0 mg kg-1 dose, and 0.164 +/- 0.083% and 1.42 +/- 0.26% at 10.0 mg kg-1 dose, respectively. The renal clearance (CLR) and the biliary clearance (CLB) were calculated to be 0.191 and 0.256 mL min-1 for 1.0 mg kg-1, and 0.0248 and 0.215 mL min-1 for 10.0 mg kg-1, respectively. When comparing these values with the CLtot values, the urinary and biliary excretion of KNI-272 are minor disposition routes.

Animals↗
PubMed1994

New disulfide bond-forming reactions for peptide and protein synthesis.

1. In order to solve the difficult problems for the synthesis of peptides or proteins containing several disulfide bonds, three new disulfide bond-forming reactions were developed. 2. Thallium(III) trifluoroacetate oxidation. This reagent, a mild oxidant with a soft acid character, cleaved various S-protecting groups of cysteine in trifluoroacetic acid, with spontaneous formation of cystine. Thus, some model peptides containing a disulfide bond were prepared by direct oxidative conversion of the respective S-substituted cysteine peptides. 3. Sulfoxide-directed disulfide bond-forming reaction. The disulfide bond was formed at the sulfur atom of Cys(R)(O) (sulfoxide) intermolecularly as well as intramolecularly following liberation of the SH group from the coreactant, Cys(R'), by a suitable acid. Thus, formation of the disulfide bond was found to be feasible at the position of Cys(R)(O). 4. Silylchloride-sulfoxide procedure. In the presence of diphenylsulfoxide, methyltrichlorosilane in trifluoroacetic acid cleaved various S-protecting groups to form cystine within 10 to 30 min at 4 degrees C. By combination of this new disulfide bond-forming reaction and others, some peptides containing two or three disulfide bonds were synthesized successfully.

Amino Acid Sequence↗
PubMed1994

Immunohistochemical analysis of beta 1-integrin receptors displayed by murine uterine natural killer cells over the course of successful pregnancy.

Granulated metrial gland (GMG) cells are a natural killer (NK) cell-like population present in large numbers in the pregnant rodent uterus. By day 8 of gestation GMG cells are large and granulated and localized to the mesometrial side of each implantation site. GMG cells appear to be highly migratory both in vivo and in vitro; however, little is known regarding their functions. Using indirect fluorescence immunohistochemistry, murine uteri and implantation sites were studied on successive days of gestation to characterize the extracellular matrix receptors of the VLA-integrin family displayed by GMG cells. On days 3 and 6 of gestation, double immunostaining using the monoclonal antibody LGL-1 was employed to recognize GMG cells because their morphology early in pregnancy resembles that of other lymphocytes. Between days 8-15 of gestation, GMG cells can be recognized by their unique morphology. The day 3 and day 6 LGL-1+ cells were positive for all antigens examined; that is, beta 1 plus alpha 1, alpha 3, alpha 4, alpha 5 and alpha 6. From days 8-15 of gestation, GMG cells were beta 1+, alpha 4+, alpha 5+ but alpha 1-, alpha 3-, alpha 6-. Thus, between days 6-8 of gestation, major changes occur in the uterine NK/GMG cell population which include the loss of the surface molecules VLA alpha 1, alpha 3 and alpha 6 or the rapid expansion of NK cells not expressing these proteins. It is postulated that major changes in the functions of uterine NK cells are likely to be associated with these alterations in cell surface phenotype and that functional studies of uterine NK cells should focus upon this relatively early gestational time point.

Animals↗
PubMed1994

Protein binding of human immunodeficiency virus protease inhibitor KNI-272 and alteration of its in vitro antiretroviral activity in the presence of high concentrations of proteins.

KNI-272 represents a peptide-based protease inhibitor having potent antiretroviral activity against human immunodeficiency virus (HIV) in vitro. The structure contains allophenylnorstatine [(2S,3S)-3-amino-2-hydroxy-4-phenylbutyric acid] with a hydroxymethylcarbonyl isostere. We asked whether this experimental anti-HIV agent could exert its activity in vitro in the presence of relatively high concentrations of fetal calf serum (FCS) and assessed its protein-binding properties by using fresh human plasma preparations. The 50 and 75% inhibitory concentrations of KNI-272 against HIV type 1 replication in vitro were 3- to 5-fold and 5-fold higher in the presence of 50% FCS and 15- to 25-fold and 25- to 100-fold higher in the presence of 80% ECS, respectively, than those with 15% FCS, whereas the antiviral activity of 2',3'-dideoxyinosine was not significantly affected by FCS concentrations in the culture. Detailed studies of the protein binding of KNI-272 suggest that in human plasma binding occurs predominantly to alpha 1-acid glycoprotein and that KNI-272 is probably extensively (approximately 98 to 99%) protein bound at concentrations likely to be achieved in the circulation. Thus, higher levels of KNI-272 in plasma may be required when this compound undergoes clinical trials relative to those inferred from in vitro data involving the use of 10 to 15% FCS-containing culture media. The current data may have a relevance to other antiretroviral drugs that are under development and that have a high protein-binding capacity.

Animals↗
PubMed1994

Racemization-free synthesis of C-terminal cysteine-peptide using 2-chlorotrityl resin.

We investigated the effects of bases, resins, and S-protecting groups on the extent of racemization at the C-terminal cysteine during Fmoc-based(Fmoc=fluoren-9-yl-methoxy-carbonyl) solid phase peptide synthesis. The use of 2-chlorotrityl resin was most effective in suppressing the racemization caused by the base treatment for Fmoc-cleavage. Somatostatin was successfully synthesized with practically no racemization using 2-chlorotrityl resin by Fmoc-chemistry.

Chromatography, High Pressure Liquid↗
PubMed1994

Murine granulated metrial gland cell population in beige (bg/bg) and SCID (scid/scid) genotypes.

Morphology of granulated metrial gland (GMG) cell, a uterine natural killer (NK) cell, was reported to be normal in pregnant uteri of NK cell-deficient mice and T-cell- and B-cell-deficient mice, but little is known about the number of GMG cells. To determine whether the number of GMG cells is influenced by such mutations, their number in the bg mice (genotype C57BL/6J-bg/bg) and SCID mice (genotype C.B-17/Icr-scid/scid) was compared to that of control mice on day 12 of gestation. GMG cells in these mutant mice were normal in number. Thus, the present results support the previous reports that GMG cells can differentiate normally in bg mice and that the GMG cell differentiation is not affected by functional T- nor B-cells.

Animals↗
PubMed1993

Granulated metrial gland cells: a natural killer cell subset of the pregnant murine uterus.

The metrial gland develops in the uterus of many rodent species during normal pregnancy. It is a maternally-derived tissue that contains stromal and vascular elements plus a population of large cells, striking in their light microscopic appearance due to the presence of numerous cytoplasmic granules. These cells, which have become known in mice and rats as granulated metrial gland (GMG) cells, are derived from bone marrow precursors and recent work suggests they are a subset of lymphocytes belonging to the natural killer (NK) cell lineage. The functions of GMG cells during normal gestation have not been clearly defined. In vitro, GMG cells have been shown to produce cytokines and their cytokine profile is altered upon addition of medium containing the T cell growth factor interleukin-2 (IL-2). GMG cell granules contain the cytolytic protein perforin but GMG cells have a very limited capacity to kill in vitro unless they have been stimulated by IL-2 or interferon-gamma. Histological study of GMG cells has suggested they preferentially associate with fetal trophoblast. Since trophoblast appears resistant to immune lysis, except by IL-2-activated effector lymphocytes, and because resorbing murine embryos become infiltrated by lytic cells of the NK cell lineage, it is important to establish whether GMG cells are activated by pregnancy-associated events to play a major lytic role in vivo.

Animals↗
PubMed1993

Simulated quantitative and qualitative isotachophoretic indices of 73 amino acids and peptides in the pH range 6.4-10.

Qualitative and quantitative isotachophoretic indices of 73 amino acids, dipeptides and tripeptides were simulated under 24 leading electrolyte conditions covering the pH range 6.4-10. The RE values and time-based zone lengths are tabulated together with the absolute mobility (m0) and pKa values used. The leading electrolyte used was 10 mM HCl and the pH buffers were imidazole, tris(hydroxymethylamino)methane, 2-amino-2-methyl-1,3-propanediol and ethanolamine. The simulated indices will be useful in the assessment of the separability and determination of the listed and related compounds.

Amino Acid Sequence↗
PubMed1993

Pharmacokinetic study of a tripeptide HIV-1 protease inhibitor, KNI-174, in rats after intravenous and intraduodenal administrations.

Recently, as a new type of anti-AIDS drug, an HIV-1 protease inhibitor, KNI-174, has been synthesized; it shows a potent and selective HIV-1 protease inhibitory activity in vitro. In this study, we developed an HPLC assay system for KNI-174 in rat plasma and examined the pharmacokinetics of KNI-174 in rats using this assay method after both intravenous (i.v.) and intraduodenal (i.d.) administrations to obtain the disposition characteristics and bioavailability of this new anti-AIDS drug. This HPLC assay method is specific to KNI-174 and the standard curve was linear from 0.02 to 30 micrograms ml-1 plasma. After i.v. administration, 10.0 mg kg-1, KNI-174 disappeared from the rats' plasma in a three-exponential decay. The mean terminal elimination half-life, t1/2 lambda z, was 3.97 +/- 0.19 (S.E.) h, the total body clearance, CLtot, was 9.53 +/- 1.08 ml min-1 and the distribution volume at steady state, Vd,ss, was 7070 +/- 960 ml kg-1. In the case of the i.d. administration, 10.0 mg kg-1, the mean peak plasma concentration, Cmax, and the peak time, tmax, were 0.196 +/- 0.076 micrograms ml-1 and 0.444 +/- 0.193 h, respectively. The bioavailability of KNI-174 till infinity, BA(0-infinity), was 5.37 per cent. Because the IC50 of KNI-174 against HIV-1 in PHA-PBM was 138 ng ml-1, the time needed for maintaining the concentrations above IC50 after a single i.d. administration of KNI-174 is estimated to be 0.350 +/- 0.184 h.

Animals↗
PubMed1993

Comparison of a new orally potent tripeptide HIV-1 protease inhibitor (anti-AIDS drug) based on pharmacokinetic characteristics in rats after intravenous and intraduodenal administrations.

Recently, a series of KNI compounds such as KNI-227 and KNI-272 has been synthesized and shows potent and selective HIV-1 protease inhibitory activity in vitro. In this study, we developed an HPLC assay system for KNI-227 and KNI-272 in rat plasma and examined the pharmacokinetic characteristics in rats after both intravenous (i.v.) and intraduodenal (i.d.) administrations to obtain the disposition characteristics and bioavailabilities of these new anti-AIDS drugs. After i.v. administration of KNI-227, 10.0 mg kg-1, the mean terminal elimination half-life, t1/2 lambda zeta, was 0.808 +/- 0.161(SE) h, the total body clearance, CLtot, was 11.7 +/- 3.3 ml min-1 and the distribution volume at steady state (Vd,ss) was 1410 +/- 460 ml kg-1. On the other hand, after i.v. administration of KNI-272, 10.0 mg kg-1, t1/2 lambda zeta was 2.86 +/- 0.78 h, CLtot was 15.3 +/- 1.4 ml min-1 and Vd,ss was 3440 +/- 670 ml kg-1. In the case of the i.d. administration of drugs, the mean peak plasma concentrations, Cmax, of KNI-227 and KNI-272 were 0.374 +/- 0.110 microgram ml-1 and 0.900 +/- 0.093 micrograms ml-1, respectively. The bioavailabilities (BA) of KNI-227 and KNI-272 to infinity, BA(0-infinity), were 5.90% and 42.3%, respectively. As compared with the lead compound, KNI-174, the BA of KNI-272 was improved about 10 times. Although the anti-AIDS virus activity of these two drugs has not been investigated in vivo, KNI-272 is expected to be a better candidate for oral anti-AIDS therapies.

Animals↗
PubMed1993

An immunohistologic analysis of murine uterine T cells between birth and puberty.

Murine uterine T cells were analysed on the basis of surface phenotype expression from birth to adulthood. T cells were rare in the uterus from birth until 2 weeks of age. In genetically immunocompetent mice, mature T cells expressing either TCR alpha/beta or TCR gamma/delta were first present as a major cell population at 3 weeks of age. The ratio of TCR alpha/beta to TCR gamma/delta was 1:1 at 3 weeks of age and this ratio did not change during sexual maturation. Almost all uterine T cells were CD8+ and the majority of these cells expressed CD8 alpha/beta rather than CD8 alpha/alpha. Cells expressing Thy1.2 were less frequent than cells expressing CD3 while cells expressing CD5 were rare. No major changes in T cell subsets occurred at puberty. Further, the microbial flora of the mice did not alter the time of appearance, frequency or subset distribution of uterine TCR+ cells. In the uteri of immunodeficient mice of genotype scid/scid TCR+ cells were found in low numbers and the initial appearance of TCR+ cells was delayed until 5 weeks of age.

Animals↗
PubMed1993

Is TPO detectable in the circulation?

Recent reports have suggested that thyroid peroxidase (TPO) can be detected in the circulation of normal subjects and of patients with Graves' disease and we have attempted to confirm and extend these observations. A TPO radioimmunoassay with a sensitivity of 1 ng/mL was used to measure TPO in the sera from 20 normal subjects and 21 patients with Graves' disease. In addition, TPO was measured in serum samples from six normal subjects before and after oral TRH. We were unable to detect TPO in 46 out of the 47 sera studied (normals and autoimmune thyroid disease). In the one remaining serum (from a normal subject), low levels of TPO were apparently detected, but we demonstrated that this result was due to assay interference from TPO autoantibodies. Overall our studies suggest that (1) thyroid peroxidase is not detectable in normal subjects nor in TPO autoantibody negative patients with Graves' disease; (2) endogenous TPO autoantibodies can interfere in the TPO radioimmunoassay leading to false positive results; and (3) an acute increase of TSH in normal subjects does not result in TPO release into the circulation.

Adult↗
PubMed1993

Protective immunity and mast cell and eosinophil responses in mice infested with larval Haemaphysalis longicornis ticks.

WBB6F1-+/+ mice were infested with larval Haemaphysalis longicornis ticks twice at an interval of 14 days: apparent resistance against ticks was expressed in the second infestation. The first infestation induced degranulation of a small number of mast cells at the feeding sites within 6 days, and resulted in two-fold increases of mast cell numbers on day 14 with a significant elevation of total immunoglobulin E (IgE) levels in sera and high proportion of IgE-bound mast cells. The second infestation resulted in the intensive degranulation of the increased mast cells at the feeding sites. Eosinophils infiltrated into the feeding sites of ticks: the second infestation led to a greater maximal level of the infiltrating eosinophils. These data suggest that the resistance against larval H. longicornis ticks in mice may be expressed as a result of immediate hypersensitivity and many eosinophils infiltrating from the blood to the feeding sites might contribute to the tick rejection.

Animals↗
PubMed1993

Thyroxine 5-deiodinase in human brain tumors.

To determine whether human brains contain deiodinating pathways, we studied the activity of T4 5-monodeiodinase (5-D) in 20 human brain tumors obtained intraoperatively, including astrocytoma (10), meningioma (4), oligodendroglioma (2), glioblastoma (2), medulloblastoma (1), and malignant lymphoma (1). Mitochondrial-microsomal fractions prepared from these tumor tissues were used as the source of T4 5-D. Each sample was incubated with 32.2 nmol/L T4 and 30 mmol/L dithiothreitol at 37 C for 90 min. T4 5-D activity was measured by the production of rT3 from T4 with a RIA. T4 5-D activity was found in 6 of 10 astrocytomas, 2 oligodendrogliomas, 1 of 2 glioblastomas, and 1 malignant lymphoma. This activity depended on protein concentration, incubation time, incubation temperature, and pH of the incubation mixture. It was also heat labile. T4 5-D was not inhibited by 1 mmol/L propylthiouracil, but was inhibited by iopanoic acid and aurothioglucose in a dose-dependent manner. The apparent Km and maximum velocity for T4 5-D at 30 mmol/L dithiothreitol were 106.6 nmol/L and 22.7 pmol/mg protein.h, respectively. These data suggest that human gliomas (and probably malignant lymphomas) contain T4 5-D activity, which is similar to type III enzyme activity in the rat. T4 5-D may regulate the intracellular concentration of thyroid hormone in gliomas.

Adult↗
PubMed1993

Synthesis of rat brain natriuretic peptide 45 using tetrafluoroboric acid deprotection and silyl chloride disulfide formation.

A newly isolated rat brain natriuretic peptide 45 (rBNP45) was synthesized using the 9-fluorenylmethyloxycarbonyl (Fmoc)-based solid-phase method. Tetrafluoroboric acid (HBF4) deprotection was successfully applied for this synthesis, while the conventional trifluoroacetic acid (TFA)-thioanisole method gave unsatisfactory results. The disulfide bond of rBNP45 was constructed by using the silyl chloride method within 10 min, which was extremely advantageous to avoid the formation of Met(O)-rBNP45. The chick rectum relaxant activity of the synthetic rBNP45 was three times as potent as that of alpha-rat atrial natriuretic peptide (alpha-rANP).

Amino Acid Sequence↗