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PubMed1991

[The therapeutic effects of itraconazole, a new triazole antifungal agent, for experimental fungal infections].

The therapeutic efficacy of itraconazole (ITZ), and oral triazole antifungal agent, was studied using several experimental fungal infections in animals. The following results were obtained: 1. ED50 values of ITZ and ketoconazole (KCZ) in a murine model of systemic candidiasis produced by intravenous challenge of Candida albicans alls were 32.9 mg/kg and 224 mg/kg, respectively. ITZ suppressed the proliferation of Candida experimentally colonized in the GI-tract of mice and/or a secondary dissemination induced by prednisolone. 2. An oral dose of 40 mg/kg/day ITZ administered for experimental pulmonary cryptococcosis in mice inhibited the fungal proliferation in the lung and the dissemination to the brain. 3. ED50 values of ITZ and KCZ for experimental systemic Aspergillus infection in mice were 103.6 mg/kg and 882 mg/kg, respectively. 4. ITZ suppressed the development of local symptoms in guinea pigs with experimental dermatophytosis. Culture studies performed on cutaneous tissues from infected sites on day-19 postinfection revealed that ITZ treatment lowered the culture-positive rate to a greater extent than KCZ-treatment. 5. Plasma concentrations of ITZ after a single dose of 100 mg/kg in mice were determined using the bioassay method: Cmax was 11 micrograms/ml and T 1/2 was 24 hours. 6. These results show that oral ITZ is highly effective in the treatment of deep-seated and superficial fungal infections produced in experimental animals.

Animals↗
PubMed1983

Immobilization of proteins on partially hydrolyzed agarose beads.

Treatment of agarose beads with mild acid (0.2 M HCl, 55 degrees C, several hours) hydrolyzes some of the glycosidic bonds between D-galactosyl residues and 3,6-anhydro-L-galactosyl residues, and thus produces aldehydo-groups useful for immobilization of amino compounds by reductive amination with NaCNBH3. More than 20 mg (0.3 mumol) of bovine serum albumin could be coupled per gram of partially hydrolyzed agarose beads. Arthrobacter neuraminidase immobilized by this method was useful for desialylation of sialyl glycoconjugates, and was found not to leach from the gel and to be much more thermostable than the free enzyme.

Animals↗
PubMed1986

Effects of antibiotics on fibrinolytic activity.

Latamoxef, cefamandole, carbenicillin and cefotaxime were examined for their effects on fibrinolytic activity in vitro by means of the fibrin plate method, fibrin clot lysis time and euglobulin lysis time with human, rabbit and rat plasma. These antibiotics showed no fibrinolytic but weak antifibrinolytic activity at 1000 or 3000 micrograms/ml in some assay systems.

Animals↗
PubMed1996

[The Somatropin Reference Standard (Control 951) of the National Institute of Health Sciences].

Somatropin material was examined for preparation of the "Somatropin Reference Standard". The candidate material was evaluated by a domestic collaborative study in which eight laboratories participated. The protein content was determined to be 4.5 mg/Vial based on amino acid analysis. Because of the possibility of application as a chemical reference standard for assay by the HPLC method, a physico-chemical evaluation of the candidate material was also performed. By SE-HPLC, the content of polymer, dimer were determined to be 0.54%, 0.98%, respectively. By RP-HPLC, the early peak area ascribed to desamido and sulfoxide form was 1.07% of the total peak area. And for informational data, the potency of the candidate material, being estimated by three different biological methods, weight gain assay, tibia test and adiposeconversion assay is 14.8 IU/vial. Based on the above results, the candidate was authorized as the Somatropin Reference Standard of the National Institute of Health Sciences.

Amino Acids↗
PubMed1982

A retino-pulvinar projection in the macaque monkey as visualized by the use of anterograde transport of horseradish peroxidase.

A direct retino-pulvinar connection was found in the Japanese monkey (Macaca fuscata) by the anterograde horseradish peroxidase (HRP) method. HRP injected into the vitreous cavity of one eye labeled a few small clusters of optic fiber terminals in the medial border regions of the inferior pulvinar nucleus bilaterally, with a contralateral predominance.

Afferent Pathways↗
PubMed2001

Lipid peroxidation end products-responded induction of a preneoplastic marker enzyme glutathione S-transferase P-form (GST-P) in rat liver on admistration via the portal vein.

The in vivo induction mechanism of a preneoplastic marker enzyme, glutathione S-transferase P-form (GST-P), by a number of carcinogens and some noncarcinogens such as anti-oxidants [Proc. Natl. Acad. Sci. U.S.A. 85 (1984) 3964] has remained to be solved. Among the various administration routes tested, GST-P became immunohistochemically demonstrable in the liver centrilobular zone 3 after 24-48h on administration of prostaglandin J2's, 15-deoxy-Delta(12,14)-PGJ2, PGJ2 and Delta(12)-PGJ2 to male rats via the portal vein, whereby the animals had been pretreated with Soya oil intraperitoneally to exhaust fatty acid binding proteins. Unsaturated aldehydes, 4-hydroxynonenal, crotonaldehyde and acrolein, given by the same route induced putatively preneoplastic single cells positive for GST-P. As these lipid peroxidation end products are the substrates as well as inducers of the enzyme, its physiological function could be their detoxication. These results indicate that GST-P expression can be mediated through lipid peroxidation possibly accounting for induction observed with a wide variety of carcinogens. In addition, present method may also be of use as a direct, simple, rapid, and sensitive in vivo test in examination of other biological responses.

Animals↗
PubMed1997

Phosphodiester amidates of unsaturated nucleoside analogues: synthesis and anti-HIV activity.

The effect of introduction of a lipophilic phosphodiester amidate moiety on the HIV activity of inactive unsaturated nucleoside analogues was investigated. Phosphodiester alaninates 5a, 5b, and 6 derived from unsaturated nucleoside analogues 3b, 3c, and 4a were synthesized and investigated as inhibitors of cytopathic effect and replication of HIV-1 in ATH-8 cells. Compound 5a is an inhibitor of HIV-1 whereas analogue 6 is inactive with cytotoxicity appearing above 10 microM and 5b is both inactive and nontoxic. Alkaline or enzymic hydrolysis of 5a gave phosphomonoester alaninate 14, a putative product of intracellular metabolism. Compound 14 as well as adenallene derivative 15c were devoid of anti-HIV activity, and they also failed to inhibit HIV reverse transcriptase. A new regioselective method for preparation of (Z)-4-(benzoyloxy)-1-hydroxy-2-butene, 7, a key intermediate for the synthesis of unsaturated nucleoside analogues of cis configuration such as 3a, 3b, and 3c, is also described.

Animals↗
PubMed1978

Characterization of anti-ApApA antibodies.

Antibodies to a trinucleotide, ApApA, were prepared by injecting bovine serum albumin conjugated with ApApA into rabbits. The specificities of the antibodies were determined by estimating the inhibiton of the binding of [14C]ApApA to the antibodies by various nonradioactive mono-, oligo-, and polynucleotides, using the ammonium sulfate precipitation method. The antibodies were found to react with ApApN sequences and oligoadenylic acids, but also reacted slightly with polyadenylic acid, RNA and DNA. Significant crossreactions were observed with other oligonucleotides containing adenosine.

Adenine Nucleotides↗
PubMed1992

Isolation of adult pig islet. In vitro assessment and xenotransplantation.

This study was conducted to develop an isolation method for adult pig islets and to investigate in vivo function after xenotransplantation as well as in vitro function of isolated islets. From the splenic portion of the glands (n = 20), 3277 +/- 645 islets per gram of pancreas were isolated and recovered by a five-step dextran discontinuous density gradient method. Purity of the final preparation obtained from 20 different consecutive pig pancreata was 80-90%. In vitro incubation study revealed a significant insulin release from isolated adult pig islets in response to glucose stimulation. In vitro perifusion study demonstrated a biphasic insulin response to glucose stimulation from isolated islets. Xenotransplantation of approx 2000 isolated adult pig islets into the portal vein of diabetic Wistar rats (n = 8) significantly reduced serum glucose levels from 431 +/- 24 mg/dL to 173 +/- 18 mg/dL 24 h after transplantation. This study provides a development of an isolation method for adult pig islets and demonstrates that isolated islets are viable and functioning both in vitro study and in vivo study of xenotransplantation.

Animals↗
PubMed1997

Quantitative immunohistochemical determination of 8-hydroxy-2'-deoxyguanosine by a monoclonal antibody N45.1: its application to ferric nitrilotriacetate-induced renal carcinogenesis model.

The DNA base-modified product 8-hydroxy-2'-deoxyguanosine (8-OHdG) is one of the most commonly used markers for the evaluation of oxidative DNA damage. A monoclonal antibody specific for 8-OHdG (N45.1) was characterized and applied in quantitative immunohistochemistry. N45.1 recognized both the modified base and deoxyribose structure of 8-OHdG and required a concentration two orders higher of 8-hydroxyguanosine as a competitor in the ELISA. In addition, N45.1 did not cross-react with the original four deoxyribonucleosides, other DNA base-modified products such as 8-hydroxy-2'-deoxyadenosine and O6-methyl-2'-deoxyguanosine, or urine components such as uric acid, creatine, and creatinine. A ferric nitrilotriacetate-induced rat renal carcinogenesis model was used for the evaluation of quantitative immunohistochemistry. The 8-OHdG index of quantitative immunohistochemistry, as analyzed by NIH image freeware, correlated reasonably well with the 8-OHdG amount determined by high-performance liquid chromatography with an electrochemical detector-except for a difference in peak time, which could be attributed to the selection of target location. The present method has advantages over the high-performance liquid chromatography/electrochemical detector, gas chromatography/mass spectrometry, and 32P-postlabeling methods in that it allows localization of 8-OHdG to be specified without the risk of artifactual production of 8-OHdG during the DNA extraction and hydrolytic processes.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1987

Analysis of ceramide and monohexaosyl glycolipid derivatives by high-performance liquid chromatography and its application to the determination of the molecular species in tissues.

Several molecular species of monohexaosyl glycolipids and ceramides were analyzed as their benzoyl derivatives by high-performance liquid chromatography on a silica column with mixtures of dioxane and hexane as eluents. Monohexaosyl glycolipid derivatives were classified according to their hydrophobic structures, and ceramides were separated based on the fatty acid groups and the long-chain bases. The method allowed us to observe the heterogeneities in the molecular species of cerebrosides and ceramides from the epidermis and dermis of the guinea pig, and clear differences in the molecular species were observed between the different regions.

Animals↗
PubMed2003

A randomized, placebo-controlled trial of the effect of antihistamine or corticosteroid treatment in acute otitis media.

OBJECTIVES: To determine whether the adjunctive drugs antihistamine and corticosteroid improve immediate and long-term outcomes of acute otitis media (AOM). STUDY DESIGN: Children with AOM (3 mos-6 y) were enrolled in a randomized, double-blind, placebo-controlled trial. All 179 children received one dose of intramuscular ceftriaxone and were assigned to receive either chlorpheniramine maleate (0.35 mg/kg/d) and/or prednisolone (2 mg/kg/day) or placebo for 5 days. Main outcome measures were rate of treatment failure during the first 2 weeks, duration of middle ear effusion, and rate of recurrences of AOM to 6 months. RESULTS: Clinical outcomes and recurrence rates did not differ significantly with treatment. Children who received antihistamine alone had significantly longer duration of middle ear effusion (median, 73 days) than subjects in other treatment groups (median, 23 to 36 days, P=.04). Temporary normalization of tympanometric findings on day 5 occurred more frequently in the corticosteroid-treated group (P=.04). CONCLUSIONS: Five-day treatment with antihistamine or corticosteroid, in addition to antibiotic, did not improve AOM outcomes. Antihistamine use during an acute episode of OM should be avoided, since the drug may prolong the duration of middle ear effusion. The efficacy of 7- to 10-day treatment of AOM with corticosteroid, in addition to antibiotic, deserves further investigation.

Acute Disease↗
PubMed1984

The value of ultrasonography in early detection of renal cell carcinoma.

Renal cell carcinoma is the third most frequent cause of urological cancer death in Japan. The first is bladder cancer and the second is prostatic cancer. However, there has been no clinical trial of mass screening for early detection of renal cell carcinoma. With a view to discovering a method of early detection for renal cell carcinoma, we analyzed the procedures for discovering renal cell carcinoma in 33 patients who were treated at our hospital. In 10 patients, carcinoma had been discovered accidentally without gross hematuria, abdominal mass or flank pain. Seven of these patients were first suspected of having renal cell carcinoma by ultrasonography, two by intravenous pyelography (IVP) and one by computed tomography. Seven out of nine patients operated on were surgically diagnosed as having stage I and two as having stage II. Ultrasonography is far better than IVP from the points of view of noninvasiveness, ease of performance and accuracy. In our study, ultrasonography was the most useful method for early detection of renal cell carcinoma. However, for the mass screening of renal cell cancer by ultrasonography, it is necessary to make its specificity and sensitivity clear and to debate its cost effectiveness.

Adult↗
PubMed2004

An improved method for detection of replication-competent retrovirus in retrovirus vector products.

Contamination by replication-competent retrovirus (RCR) is one of the most important safety issues of retrovirus vector products for gene therapy clinical research. To improve the sensitivity of RCR detection and to shorten the assay period, we have developed a novel RCR detection method (infectivity RT-PCR method) based on real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) in combination with virus infection and a novel virus concentration method using polyethyleneimine (PEI)-conjugated magnetic beads. In this method, permissive cells were infected with RCR samples, and amplified RCR in the culture supernatants was adsorbed by PEI-beads. Then RCR RNA extracted from PEI-beads was quantified by real-time RT-PCR. We demonstrated that 1 infectious unit (iu) of RCR spiked in 10(6) cfu/ml of vector products could be detected within 3 days, and the sensitivity for viral detection was increased 3- to 10-fold compared with the direct S+L- assay. By this method, the presence of retroviral vector interfered with RCR detection only slightly. In conclusion, infectivity RT-PCR conducted in conjunction with virus concentration using PEI-beads can detect RCR more sensitively and rapidly than the conventional infectivity assay.

Animals↗
PubMed2002

15-deoxy-delta 12,14-prostaglandin J2. A prostaglandin D2 metabolite generated during inflammatory processes.

Prostaglandin D(2) (PGD(2)), a major cyclooxygenase product in a variety of tissues, readily undergoes dehydration to yield the cyclopentenone-type PGs of the J(2) series, such as 15-deoxy-Delta(12,14)-PGJ(2) (15d-PGJ(2)), which have been suggested to exert anti-inflammatory effects in vivo. Meanwhile, the mechanism of these effects is not well understood and the natural site and the extent of its production in vivo remain unclear. In the present study, we raised a monoclonal antibody specific to 15d-PGJ(2) and determined its production in inflammation-related events. The monoclonal antibody (mAb11G2) was raised against the 15d-PGJ(2)-keyhole limpet hemocyanin conjugate and was found to recognize free 15d-PGJ(2) specifically. The presence of 15d-PGJ(2) in vivo was immunohistochemically verified in the cytoplasm of most of the foamy macrophages in human atherosclerotic plaques. In addition, the immunostaining of lipopolysaccharide-stimulated RAW264.7 macrophages with mAb11G2 demonstrated an enhanced intracellular accumulation of 15d-PGJ(2), suggesting that the PGD(2) metabolic pathway, generating the anti-inflammatory PGs, is indeed utilized in the cells during inflammation. The activation of macrophages also resulted in the extracellular production of PGD(2), which was associated with a significant increase in the extracellular 15d-PGJ(2) levels, and the extracellular 15d-PGJ(2) production was reproduced by incubating PGD(2) in a cell-free medium and in phosphate-buffered saline. Moreover, using a chiral high performance liquid chromatography method for separation of PGD(2) metabolites, we established a novel metabolic pathway, in which PGD(2) is converted to 15d-PGJ(2) via an albumin-independent mechanism.

Animals↗