A simple flow cytometric method for detecting humoral immunity against sheep red blood cells (SRBC) is described. The SRBC were incubated with the serum from SRBC-immunized mice, monoclonal anti-SRBC, or the supernatant which was obtained from the in vitro primary culture of spleen cells with SRBC. The antibodies which bound to SRBC were estimated by means of an immunofluorescence and a flow cytometry. When the channel number of the peak in the histogram of flow cytometry was measured as an index of fluorescence intensity of SRBC, the number significantly correlated with the concentration of IgM and IgG classes of anti-SRBC. The flow cytometry method and hemagglutination (HA) test, as a classic method, were compared in SRBC-immune sera and monoclonal anti-SRBC antibody. The sensitivity determined with flow cytometry was much higher than that with HA. The minimum detectable concentration of anti-SRBC antibody was found to be 3.4 ng/ml by the flow cytometry. The dose response of SRBC in in vitro primary culture was detected by the flow cytometry, not by HA, and the response increased with the dose of SRBC. Using this method, the effect of in vitro and in vivo concanavalin A (Con A)-induced spleen cells on humoral response against SRBC was examined in an in vitro culture system. Anti-SRBC response (IgM and IgG) was found to be suppressed by in vitro Con A-induced lymphocytes but enhanced by in vivo Con A-induced lymphocytes. Thus, this new approach is found to be a good method for detecting the in vitro primary humoral antibody response, which is known to have a low reactivity.

Animals↗

PubMedDate not supplied

Phosphorylation of glutaminyl-tRNA synthetase and threonyl-tRNA synthetase by the gene products of dnaK and dnaJ in Escherichia coli K-12 cells.

In Escherichia coli K-12, the heat shock protein DnaK and DnaJ participate in phosphorylation of both glutaminyl-tRNA synthetase and threonyl-tRNA synthetase since when cellular proteins extracted from the dnaK7(Ts), dnaK756(Ts) and dnaJ259(Ts) mutant cells labeled with 32Pi at 42 degrees C were analyzed by two-dimensional gel electrophoresis, no phosphorylation of glutaminyl-tRNA synthetase and threonyl-tRNA synthetase was observed while phosphorylation of both aminoacyl-tRNA synthetases was detected in the samples extracted from wild-type cells.

Amino Acyl-tRNA Synthetases↗

PubMedDate not supplied

In vivo function of VDR in gene expression-VDR knock-out mice.

Vitamin D exerts many biological actions through nuclear vitamin D receptor (VDR)-mediated gene expression. The transactivation function of VDR is activated by binding 1alpha,25-dihydroxyvitamin D3[1alpha,25(OH)2D3], an active form of vitamin D. Conversion from 25(OH)D3 is finely regulated in kidney by 25(OH)D3 1alpha-hydroxylase[25(OH)D 1alpha-hydroxylase], keeping serum levels of 1alpha,25(OH)2D3 constant. Deficiency of vitamin D and mutations in the genes like VDR (type II genetic rickets) are known to cause rickets like lowered serum calcium, alopecia and impaired bone formation. However, the molecular basis of vitamin D VDR system in the vitamin D action in intact animals remained to be established. In addition, the 1alpha-hydroxylase gene from any species had not yet been cloned, irrespective of its biological significance and putative link to the type I genetic rickets. We generated VDR-deficient mice (VDR KO mice). VDR KO mice grew up normally until weaning, but after weaning they developed abnormality like the type II rickets patients. These results demonstrated indispensability of vitamin D-VDR system in mineral and bone metabolism only in post-weaning life. Using a newly developed cloning system, we cloned the cDNA encoding a novel P450 enzyme, mouse and human 1alpha-hydroxylase. The study in VDR KO mice demonstrated the function of liganded VDR in the negative feed-back regulation of 1alpha,25(OH)2D3 production. Finally, from the analysis of type I rickets patients, we found missense genetic mutations in 1alpha-hydroxylase, leading to the conclusion that this gene is responsible for the type I rickets.

Animals↗

PubMedDate not supplied

A durable, non power consumptive, simple seal for rotary blood pumps.

One of the key technologic requirements for rotary blood pumps is the sealing of the motor shaft. A mechanical seal, a journal bearing, magnetic coupling, and magnetic suspension have been developed, but they have drawbacks such as wear, thrombus formation, and power consumption. A magnetic fluid seal was developed for an axial flow pump. A magnetic fluid seal is durable, simple, and non power consumptive. Long-term experiments and finite element modeling (FEM) analyses confirmed these advantages. The seal body was composed of a Ned-Fe magnet and two pole pieces; the seal was formed by injecting ferrofluid into the gap (50 microm) between the pole pieces and the motor shaft. To contain the ferrofluid in the seal and to minimize the possibility of ferrofluid making contact with blood, a shield with a small cavity was attached to the pole piece. While submerged in blood, the sealing pressure of the seal was measured and found to be 188 mm Hg with ferrofluid LS-40 (saturated magnetization, 24.3 kA/m) at a motor speed of 10,000 rpm and 225 mm Hg under static conditions. The magnetic fluid seals performed perfectly at a pressure of 100 mm Hg for 594 + days in a static condition, and 51, 39+, and 34+ days at a motor speed of 8,000 rpm. FEM analyses indicated a theoretical sealing pressure of 260 mm Hg. The state of the magnetic fluid in the seal in water was observed with a microscope. Neither splashing of magnetic fluid nor mixing of the magnetic fluid and water was observed. The specially designed magnetic fluid seal for keeping liquids out is useful for axial flow blood pumps. The magnetic fluid seal was incorporated into an intracardiac axial flow pump.

Heart-Assist Devices↗

PubMedDate not supplied

Perfusion chemotherapy of osteosarcoma. A clinical study on 75 cases.

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Adolescent↗

PubMedDate not supplied

Influence of ethyl 3-(4'-geranyloxy-3'-methoxyphenyl)-2-propenoate (EGMP) on early stage colon carcinogenesis in rats treated with azoxymethane (AOM).

The influence of ethyl 3-(4'-geranyloxy-3'-methoxyphenyl)-2-propenoate (EGMP) on the initiation and post-initiation stages of colon carcinogenesis was investigated in male F344 rats treated with azoxymethane (AOM). In experimental protocol 1, EGMP was given in the diet at 0.1 or 0.2% for 1 week together with two s.c. 15 mg/kg body weight injections of AOM on days 1 and 7 (initiation period). In protocol 2, the test compound was administered starting at week 3(post initiation stage), and in protocol 3, the test compound was given throughout the experimental period(whole stage). Sacrifice and quantitation of aberrant crypt foci (ACF) was performed at the end of week 5. Dose-dependent decreases in numbers of ACF were noted with both cases of post-initiation and whole period exposure (protocol 2 and 3), large size lesions considered most likely to be precursor lesions also being significantly reduced in the protocol 2(4-9 crypt size total with the 0.2% dose group, 48.9% and 59.6% of control values, respectively). No effects on body or liver weights were evident. The present results thus suggest that EGMP might find application as a chemopreventive agent against colon tumor development.

Animals↗

PubMedDate not supplied

Metabolites of feprazone in man.

Two male volunteers received 400 mg of nonlabeled feprazone, 1,2-diphenyl-3,5-dioxo-4-(3'-methyl-but-2'-enyl)pyrazolidine. Metabolites were extracted and isolated by chromatography on silica gel. Isolated metabolites were unchanged drug, E-1,2-diphenyl-3,5-dioxo-4-(3'-hydroxymethyl-but-2'-enyl)pyrazolidine, and feprazone (C4)-beta-glucuronide. Feprazone (C4)-beta-glucuronide was the main metabolite and it belongs to a novel type of conjugate, (C)-beta-glucuronide.

Feprazone↗